Mature B-Cell Lymphoma And Leukemia Study III
Phase: PHASE2
Conditions: Mature B-Cell Lymphoma
Sponsor: St. Jude Children's Research Hospital

SJBC3
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
St. Jude
SJBC3-NCT01046825
Initial version, dated:  04-07-2009, Resubmitted to CPSRMC 05-22-2009 and 7-2-2009.  
Resubmitted to IRB 7-23-2009 (IRB approved: 8-3-2009)
Amendment 1.0, dated: 11-10-2009. Resubmitted to IRB 12-23-2009
Amendment 2.0, dated 7-27-2011, Resubmitted to IRB on 11-15-2011 (IRB approved: 11-20-2011)
Activation date: 3-15-2012
Revision 2.1, dated: 08-02-2012 (IRB approved: 8-8-2012)
Revision 2.2, dated: 01-28-2013 (IRB approved: 1-29-2013)
Activation date: 02-08-2013 
Revision 2.3, dated 04-22-2013 (IRB approved: 05-07-2013) Activation date: 05-15-2013
Revision 2.4, dated 04-17-2014 (IRB approved: 05-09-2014)
Revision 2.5, dated 10-1-2014 (consent form change only).
Amendment 3.0, dated 05-25-2016. Resubmitted to IRB 02-09-2017 and 04-07-2017 (IRB approved: 04-14-2017) 
Activation date: 04-27-2017
Amendment 4.0, dated 11-04-2019 (IRB approved: 02-03-2020)
Amendment 5.0, dated 3-27-2020 (IRB approved:06-02-2020             ) Activation date: 06-10-2020 
Amendment 6.0, date 04-22-2022 (IRB approved:   )
MATURE B-CELL LYMPHOMA AND LEUKEMIA STUDY III
Principal Investigator
Raul C. Ribeiro, MD
Department of Oncology
Sub-Investigators
Oncology
Hiroto Inaba, MD
Kim E. Nichols, MD
Stephanie Dixon, MD
Rose B. McGee, MS, CGC
Pharmaceutical Sciences
Jennifer Pauley, PharmD
Pathology
Charles G. Mullighan,   
MBBS(Hons), MSc, MD
Lu Wang, MD, PhD
Diagnostic Imaging
M. Beth McCarville, MD
Sue Kaste, DO
Zoltán Patay, MD, PhD
Wilburn E. Reddick, PhD
Barry Shulkin, MD, MBA
Biostatistics
Cheng Cheng, PhD
Yinmei Zhou, MS
Psychology
Heather Conklin, PhD
Lisa M. Jacola, PhD
Cancer Prevention and Control
Melissa M. Hudson, MD
Daniel M. Green, MD
Vijaya M. Joshi, MD
Biologic Collaborators
Gerard Zambetti, PhD - Biochemistry
Claire Sample, PhD – Virology, Hershey Medical Center (External 
Collaborator)
St. Jude Children's Research Hospital
262 Danny Thomas Place
Memphis, Tennessee38105-2794
Telephone: (901) 595-3300
Contents of this document may not be extracted without permission from the Principal Investigator.
St. Jude Children's Research Hospital
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Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
External Collaborating Sites
Sites Participating in Therapeutic and Biological Objectives
Deborah Schiff, MD
Pediatric Hematology/Oncology/Blood and Marrow Transplant
Rady Children's Hospital, San Diego
3020 Children's Way
San Diego, CA92123
Sites Participating in Biological Objectives Only
Dr. Francisco Pedrosa
IMIP/CEHOPE
Rua Joaquim Inacio
187, Recife, Brazil
Pernambucco 50702070
Dr. Federico Antillon
Unidad Nacional de Oncologia Pediatrica
A Partirdel 20 de Marzo
Neustras Instalaciones
Estaran Ubicades en: 9
Avenida 8-00, zona 11
Tercer Nivel Edificio
UNOP, Guatemala
Dr. Roberto Vasquez
Hospital Nacional de Ninos Benjamin Bloom
Hospital Centro
Pediatrico Local No 15
Final Diag
San Salvador, El Salvador
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Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
External Collaborating Sites
Dr. Armando Pena
Universidad Nacional Autonoma de Honduras (UNAH)
Hospital Escuela Bloque Maerno Infantil
Blvd Suyapa, Teqgucigalpa
Honduras/Central America
Prof Allen Yeoh
Nuhkids University Children’s Medical Institute
Viva-University
Children’s Cancer Centre, 5 Lower Kent Ridge Road
Kent Ridge Wing, Level 8
Singapore 119074
Prof Cristina Stefan
Tygerbert Hospital/Stellenbosch University
Cape Town, Wester Cap
South Africa
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Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
Protocol summary
SJBC3:  MATURE B-CELL LYMPHOMA AND LEUKEMIA STUDY III
Principal Investigator: Raul C. Ribeiro, M.D.
IND holder: Not applicable, non-IND study. St. Jude is the study sponsor.
Brief overview: This is a phase II/III clinical trial using risk-adapted therapy. Treatment 
outcomes for children with B-cell NHL are excellent. Further improvements in outcome will 
likely be achieved through more focused study of the biology of the tumors and prospective 
studies of the late effects of treatment. Toward this end, this study features a spectrum of 
prospective biologic and late effect studies performed in patients treated with a modified regimen 
derived from the very successful LMB-96 regimen.
Intervention: Chemotherapy based on assignment of group (A, B, or C). St. Jude and Rady 
Children’s Hospital San Diego are currently the centers participating in both therapeutic and 
biological portion of study. International and other external sites will submit biological 
specimens to St. Jude for research and banking, after obtaining consent locally and enrolling 
participants through St. Jude Clinical Trials Office (CTO).
Brief outline of treatment plan: 
Overview - the chemotherapy regimen used varies with grouping based on extent of disease (see 
Appendix II for definition of Group A, B, and C)
    Group
  Pre-Phase
   Induction
      X 2
Consolidation
     X 2
Continuation
      A
    COPAD
      B
    COP
    COPAD-M3
     CYM
      C
    COP
    COPAD-M8
     CYVE
    Seq. No 1,2,3,4
Study design: Multicenter, phase III study utilizing risk-adapted multi-agent treatment.
Sample size: Target accrual for the biological studies is 68 evaluable tumor samples from several 
participating international institutions and for the late effects and outcome studies 100 newly 
diagnosed pediatric B-cell lymphoma patients treated at St. Jude.
Data management: Data management and statistical analysis will be provided by the 
Comprehensive Cancer Center clinical research staff (Leukemia/Lymphoma Division) and 
Biostatistics Department at St. Jude Children’s Research Hospital.
Human subjects: The risks to participants will be related to the toxicity of multi-agent 
chemotherapy. Participants will be informed of this and other potential side effects during 
informed consent. Adverse events will be monitored and reported and treated appropriately.
St. Jude Children's Research Hospital
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Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
Table of Contents
1.0
OBJECTIVES........................................................................................................1
1.1
Biological Primary Aims...........................................................................................................1
1.2
Exploratory Aims ......................................................................................................................1
2.0 
BACKGROUND AND RATIONALE .................................................................3
2.1
Background ...............................................................................................................................3
2.2 
Rationale for this Study...........................................................................................................12
3.0 
ELIGIBILITY CRITERIA AND STUDY ENROLLMENT...........................13
3.1
Inclusion Criteria.....................................................................................................................13
3.2
Exclusion Criteria....................................................................................................................14
3.3
Research Participant Recruitment and Screening ...................................................................14
3.4
Enrollment on Study................................................................................................................15
4.0
TREATMENT PLAN..........................................................................................16
4.1
Treatment Overview................................................................................................................16
4.2
Intrathecal Treatment ..............................................................................................................17
4.3
Chemotherapy Details.............................................................................................................18
4.4
Supportive Care Guidelines ....................................................................................................32
4.5
Dosage Modifications for Toxicity .........................................................................................34
4.6 
Surgical Guidelines .................................................................................................................36
4.7
Radiation Therapy Guidelines.................................................................................................37
5.0
DRUG INFORMATION.....................................................................................37
5.1
Cytarabine (Ara-C) (Cytosar-U) ...........................................................................................37
5.2
Cytarabine (High Dose Ara-C)................................................................................................38
5.3
Cyclophosphamide..................................................................................................................39
5.4
Doxorubicin (Adriamycin)....................................................................................................39
5.5
Etoposide (VP16) (Vepesid) .................................................................................................40
5.6
Etoposide phosphate (Etopophos®).........................................................................................41
5.7
Filgrastim (G-CSF, Neupogen).............................................................................................41
    5.8     Peg-filgrastim (pegylated filgrastim, PEG-filgrastim, Neulasta®)…………………………..42
5.9
Hydrocortisone, Intrathecal (Cortef, Solu-Cortef)…………………………………………..42
5.10
Leucovorin (folinic acid).........................................................................................................43
    5.11     Methotrexate…………………………………………………………………………………44
5.12
Prednisone (prednisolone).......................................................................................................45
5.13
Rituximab (Rituxan)..............................................................................................................45
5.14
Vincristine ...............................................................................................................................46
6.0 
EVALUATIONS, TESTS, AND OBSERVATIONS........................................47
6.1
Suggested Evaluations Before and During Therapy – for Good Medical Care**..................47
6.2
Suggested Evaluations for Research .......................................................................................48
6.3
Advanced MR Imaging Methods ............................................................................................49
6.4
Suggested Long-Term Follow-up Evaluations........................................................................49
6.5
Modifications for Collaborating Sites Participating in Therapeutic & Biological Objectives50
7.0 
EVALUATION CRITERIA ...............................................................................51
7.1
Definition of Evaluable Disease..............................................................................................51
7.2
Definition of Response............................................................................................................51
7.3
Response Evaluations According to Treatment Group...........................................................51
8.0
OFF PROTOCOL TREATMENT AND OFF-STUDY CRITERIA ..............52
8.1
Off Treatment Criteria.............................................................................................................52
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8.2
Off Study Criteria....................................................................................................................53
9.0
SAFETY AND ADVERSE EVENT REPORTING REQUIREMENTS........53
9.1
Reporting Adverse Experiences (AEs) and Deaths to the St. Jude IRB .................................53
9.2
Recording AEs and SAEs .......................................................................................................55
9.3
Reporting AEs to and from St. Jude and Collaborating Sites/Affiliates.................................56
10.0
DATA COLLECTION, STUDY MONITORING, AND CONFIDENTIALITY
57
10.1
Data Collection........................................................................................................................57
10.2
Data Collection Instructions for Collaborating Sites ..............................................................57
10.3
Study Monitoring ....................................................................................................................57
10.4
Confidentiality.........................................................................................................................58
11.0
STATISTICAL CONSIDERATIONS...............................................................58
11.1
Biological Aims.......................................................................................................................58
11.2
Exploratory Aims ....................................................................................................................64
12.0 
OBTAINING INFORMED CONSENT ............................................................67
12.1
Consent/Assent at Enrollment.................................................................................................67
12.2
Consent at Age of Majority.....................................................................................................67
12.3
Consent When English is Not the Primary Language.............................................................67
12.4
Collection of Collaborating Institution Consent Forms ..........................................................68
13.0
REFERENCES ....................................................................................................69
APPENDICES
Appendix I:
Pediatric Rituximab Infusion Guidelines
Appendix II:
Group Classification/Murphy Stage
Appendix III:  Biological Specimen Collection and Submission Guidelines
Appendix IV:  Psychological Testing
Appendix V: Data & Specimen Collection Forms for Biology Studies
Appendix VI: Clinical Data for Participants at Collaborating Sites Participating in Biological 
Objectives Only
Appendix VII: Sub-study for Collaborating Sites Participating in Biological Objectives Only
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1.0
OBJECTIVES
1.1
Biological Primary Aims
1.1.1
To perform genomic analysis of childhood newly diagnosed mature B cell lymphomas 
(e.g. Burkitt lymphoma/leukemia, DLBCL, and MLBCL) obtained from different parts 
of the world.
1.1.2
To describe the types and frequency of mutations in the ARF-HDM2-TP53 pathway, in 
B-cell lymphomas in the United States and those found in selected geographic regions 
of the world.
1.1.3
To describe the expression of ARF-HDM2-TP53 and PUMA-associated pathways in B-
cell lymphomas in the United States and that found in B-cell lymphomas of other 
selected geographic regions of the world.
1.1.4
To describe the pattern and frequency of XLP gene mutations presenting with B-cell 
lymphomas in the United States and selected geographic regions.
1.1.5
To describe the frequency of EBV-positive B-cell lymphomas in the United States and 
selected geographical regions of the world.

To describe the pattern of EBV protein and gene expression (e.g., EBNA 3) in 
EBV-positive lymphomas.

To compare patterns of EBV protein and gene expression with clinical, laboratory 
and outcome data.
1.2
Exploratory Aims
1.2.1
Neuroimaging
1.2.1.1 In participants who develop acute, clinically symptomatic neurotoxicity during 
treatment:  

Obtain data, especially during the early acute phase of events in order to improve 
our understanding of the likely sequential pathophysiological processes in acute 
methotrexate-induced leukoencephalopathy using diffusion-based MRI surrogate 
biomarkers (ADC, FA) and other advanced MR techniques (MRS, DSC-perfusion 
MRI)
1.2.1.2 In participants who do not develop overt, clinically symptomatic neurotoxicity 
during treatment:

Determine the incidence of measurable ADC and FA changes in white matter (in 
particular, in normal appearing white matter) in various brain regions during the 
course of treatment, and

Describe the topographical and temporal predilections of subclinical white matter 
lesions in order to propose a clinically feasible screening strategy for future patients 
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receiving such treatment with the ultimate goal to develop preventive measures to 
reduce the long-term burden of neurotoxicity
1.2.1.3 To describe cerebral artery perfusion rates at baseline and at various time points during 
therapy.
1.2.2
Neuropsychology Late Effects
To establish the prevalence and type of neuropsychological sequelae associated with a 
regimen derived from the LMB-96 regimen, and to compare them with neuro-imaging 
findings.
1.2.3
Bone Mineral Density (BMD)
1.2.3.1 Prospectively estimate BMD at diagnosis and correlate with risk factors for potential 
BMD deficits in pediatric patients with NHL.
1.2.3.2 Prospectively estimate BMD at end of therapy and correlate with risk factors for 
potential BMD deficits in pediatric patients with NHL.
1.2.4
Fertility Late Effects
To establish the prevalence of gonadal and germ cell dysfunction in children treated 
with this regimen. 
1.2.5
Cardiac Late Effects
To establish the prevalence and degree of cardiac toxicity associated with this regimen.
1.2.6
Tumor Bank
To establish a tumor bank of mature B-cell lymphoma samples for future molecular 
studies aimed to clarify the role of c-MYC oncogene and its associated pathways, and 
other pathways found to harbor genetic alterations from the genomic profiling studies 
described in 1.1.1.
1.2.7
To determine the feasibility of administration and toxicity profile of the SJBC3 
protocol, which features the use of rituximab for selected higher-risk Group B and all 
Group C participants.
1.2.8
To describe the relationship between treatment outcome and clinical, biological, and 
radiological features (including day 7 PET).
1.2.9
To estimate the complete response rate, event-free survival, and overall survival rates in 
patients with Burkitt lymphoma (BL), Burkitt leukemia/B-cell acute leukemia (B-ALL) 
and diffuse large B-cell lymphoma (DLBCL) treated with a stage-adapted regimen 
based on the SJBCII protocol.
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2.0 
BACKGROUND AND RATIONALE
2.1
Background
2.1.1
Genome-Wide Profiling of Genetic Alterations in Lymphoma
Genome-wide profiling of the gene expression and genetic alterations, including DNA copy 
number abnormalities (CNA) and loss of heterozygosity (LOH), and genome sequencing has 
provided valuable insights into the pathogenesis of lymphoid neoplasms.1,2 Gene expression 
profiling has been found to be an effective tool in differentiating BL from other mature B-cell 
lymphomas like DLBCL in adults.3,4 These studies have shown that some cases that cannot be 
classified unequivocally by classical morphological or cytogenetic modalities express a clear 
genetic signature that allows for their classification. Three major DNA microarray studies have 
looked into the biological and clinical heterogeneity of adult DLBCL.5-7 The initial DLBCL 
gene expression profiling study used genes that define the germinal center stage of B-cell 
differentiation to identify 2 prominent DLBCL subgroups. The “germinal center B-cell-like” 
DLBCL (GCB DLBCL) expressed genes characteristic of normal germinal center B cells (e.g. 
CD 10, BCL6, A-myb), whereas the “activated B cells “ DLBCL subgroup (ABC DLBCL) 
expressed genes that are induced during mitogenic activation of peripheral blood B cells (e.g. 
BCL2, IRF-4, cyclin D2).5 These two different patterns of differentiation were associated with 
significantly different outcomes. Another larger gene expression profiling study of DLBCL 
cases confirmed the existence of these two subgroups as well as a third one called “type 3” that 
did not resemble either and may represent an additional subgroup.6 Genome-wide profiling of 
CNA in DLBCL has identified multiple recurring regions of DNA gain and loss, many of 
which are associated with the DLBCL subtypes identified by gene expression profiling. These 
include amplification of a region of chromosome 19 harboring the SPIB transcription factor, 
deletion of CDKN2A/B and trisomy 3 in ABC DLBCL; and amplification of the mir-17-92 
microRNA cluster and deletion of PTEN and GCB DLBCL. To date, there are limited genome-
wide profiling data of genetic alterations pediatric B-NHL. Our aim is to perform integrated 
analysis of genomic data to identify genetic pathways that are altered in this disease, to identify 
tumor subtypes, to examine variation in genetic features according to geographic region, and to 
perform a comparative analysis with existing data from adult cases.8,9
Techniques for studying the genomic profile of patient tumors (somatic and germline) continue 
to advance. These approaches to be used, depending on sample availability and quality include 
RNA (transcriptome) sequencing, exome sequencing, single nucleotide polymorphism array 
analysis and whole genome sequencing. These will be done using standard established 
approaches in the Mullighan laboratory and the Genomic Sequencing Facility of the Hartwell 
Center at St. Jude.
2.1.2
ARF-HDM2-TP53 and PUMA
Burkitt lymphoma (BL) and mature B-cell leukemia (B-ALL) are characterized by one of three 
reciprocal chromosomal translocations [t(8;14);t(2;8);t(8;22)]. The unifying feature of these 
translocations is the juxtaposition of the c-MYC proto-oncogene and one of the immunoglobulin 
genes. These translocations result in the constitutive over-expression of c-MYC, a transcription 
factor which is necessary and sufficient for promoting cell proliferation and for provoking 
angiogenesis. In normal B-cells, c-MYC expression is tightly regulated and rapidly responds to 
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mitogenic or growth inhibitory signals. These controls are lost in BL/B-ALL, resulting in 
aberrantly high levels of c-MYC protein. The c-MYC translocations that occur in BL/B-ALL 
have been extensively studied with respect to their breakpoint patterns and their mechanism of 
de-regulation of c-MYC expression. However, recent insights into the molecular pathogenesis of 
BL/B-ALL have come from a detailed analysis of E-myc transgenic mice, a model which 
accurately mimics the chromosomal abnormality and pathology of BL/B-ALL.10 In normal 
cells, including B lymphocytes, c-Myc expression activates the Arf-Mdm2-p53 pathway 
resulting in apoptosis.11-13 Arf (mouse p19Arf /human p14ARF) is encoded by an alternative 
reading frame of the Ink4a locus and limits lymphomagenesis by antagonizing Mdm2 (murine 
double minute-2; HDM2 in man), a negative regulator of the tumor suppressor p53.14,15 The c-
Myc transgenic animal studies have shown that 80% of the lymphomas that arise in these mice 
carry mutations affecting apoptotic checkpoints, from either deletions of Ink4a/Arf (25%), 
mutation of p53 (30%) or overexpression of Mdm2 (50%).13,16-20 Although data suggest that 
one-third of human BL/B-ALL may have mutations of TP5321,22 many of the studies performed 
have been anecdotal and/or limited to the study of BL/B-ALL cell lines.23,24 Therefore a 
detailed study of alterations in this pathway is currently lacking in human mature B cell 
malignancies.
Although these studies established that c-Myc activates Arf and p53, and that functional loss of 
this pathway impairs Myc-induced apoptosis, Myc activation can still trigger cell death in the 
absence of Arf and/or p53.13 Activation of c-Myc in normal B cells suppresses the expression of 
Bcl-2 or Bcl-XL, which are anti-apoptotic proteins that protect the cells from a variety of insults, 
including chemotherapeutic drugs. Importantly, an analysis of lymphomas arising in E-Myc 
transgenic mice has established that this pathway is also bypassed during lymphoma 
development and that this also occurs at an equal frequency in tumors having alterations in the 
ARF-Mdm2-p53 pathway.
Recent studies have shown that about one third of BL in humans harbor c-MYC mutations, and 
two recurrent mutant c-MYC alleles derived from BL have been shown to be able to evade the 
p53 tumor surveillance network and promote rapid lymphoma development in mice.25 Another 
possible mediator in the altered apoptotic pathways of BL is PUMA (p53-up-regulated 
modulator of apoptosis). PUMA, a pro-apoptotic BH3-only Bcl-2 family protein, is an essential 
mediator of p53- dependent and independent cell death,26 and recent observations have 
indicated that 40% of primary BLs (n=19) fail to express detectable levels of PUMA, and that 
in some cases PUMA is inactivated by epigenetic silencing.27
Understanding the exact mechanism by which the apoptotic pathways are affected in these 
lymphomas will provide an opportunity for the development of novel therapies. The 
development of more rational therapies with a selective effect in specific tumor pathways while 
sparing normal tissue can have a great impact in the survival of cancer especially in the 
developing world, where children are at high risk of dying of the toxic effects of aggressive 
chemotherapy due to suboptimal hospital infrastructure.
2.1.3
Epstein-Barr Virus
Epstein–Barr virus (EBV) is a B lymphotropic herpes virus linked to several human diseases 
including B-cell malignancies.28 Greater than 90% of the world’s population is persistently 
infected with EBV. It is a ubiquitous γ-herpes virus that, upon primary infection, causes a brief 
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and confined replication of the virus at the site of infection in the immunocompetent host.29 In 
industrialized societies, infection with EBV generally occurs later in life, compared to early 
childhood infection common in countries with limited resources. EBV is transmitted orally from 
oropharyngeal secretions of asymptomatic carriers and patients with infectious mononucleosis.28 
While EBV infects B lymphocytes early in the course of primary infection, it normally does not 
replicate in them, but rather establishes a latent infection characterized by the limited expression 
of a subset of latency-associated genes.30 The latency genes consist of six nuclear antigens 
(EBNAs 1, 2, 3A, 3B, 3C and LP) and three membrane proteins (LMPs 1, 2A and 2B).31 There 
have been three different latency-gene expression programs described after EBV infection. In 
BL/B-ALL only a very limited repertoire of the latency-associated genes (EBNA-1, EBV-
encoded RNA (EBER)-1 and EBER-2 and the BamHI-A rightwards transcripts [BARTs]) are 
routinely expressed (Latency I) contributing to immune evasion.32 The Latency II program 
includes additional expression of LMP1 and LMP2A/B, and is commonly observed in EBV-
positive Hodgkin lymphoma, some types of T and NK-cell lymphomas and some cases of B-cell 
lymphoma.33 Latency III is expressed in lymphoblastoid cell lines and newly infected B cells in 
vivo, and includes the entire array of EBV latency-associated genes. These cells would normally 
be very susceptible to killing by EBV-specific T-cells, and, therefore, this pattern of expression 
is only seen in the EBV-associated lymphoproliferative disease (EBV-LPD) associated with 
primary or secondary immunodeficiency.33 The EBER RNAs are present in all previously 
described programs of EBV latency, and their presence within tumor samples is evidence of a 
latent EBV infection.34
The EBNA3 proteins (3A, 3B and 3C) are not normally expressed in classic endemic BL (eBL); 
however, recent studies have revealed a subset of eBLs (~ 15%) that express these EBV proteins 
as a consequence of a deletion in the EBNA2 gene.35,36 These tumors are particularly resistant to 
apoptotic stimuli. Dr. Clare Sample’s laboratory has determined that, in a SCID mouse model of 
tumorigenicity, these variant EBNA-3+ eBLs are particularly aggressive relative to the eBLs that 
do not carry the EBNA2 deletion (C. Sample, personal communication) and additional 
unpublished data suggests that the EBNA-3 proteins may contribute directly to proliferation of 
the tumor cells. Thus, it is important to determine whether the pattern of EBV gene expression is 
associated with biologic and clinical and outcome features. 
EBV has not been shown to be directly involved in BL pathogenesis, but has rather been 
suggested to increase the target pool of cells susceptible to malignant transformation. However, 
an interesting study by Ruf, et. al has demonstrated that the malignant phenotype of a group I 
BL line requires the EBV genome, and that this is associated with EBV’s ability to impair the 
c-MYC apoptotic program.37 Therefore, at least in this scenario, malignant transformation 
requires EBV gene expression altering apoptotic pathways. These observations suggest that 
EBV may influence c-MYC’s ability to trigger apoptosis in BL and possibly other pathways as 
well. Hence, we propose to evaluate and correlate EBV gene expression with alterations in the 
ARF-Mdm-p53 and Bcl-2/Bcl-X apoptotic pathways.
There are two main clinical presentations of BL clustering in different geographic regions. The 
eBL, which affects younger children and often involves the jaw, is found mainly in equatorial 
Africa and New Guinea.38-40 The majority of patients with eBL have been infected with 
malaria. By contrast, non-endemic or sporadic BL (sBL) occurs worldwide and mainly affects 
young adults. Both forms of BL differ in their association with EBV: eBL is almost always 
associated with EBV, whereas sBL has a more irregular association, ranging from 10% to 
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30%.41 However, our knowledge of other clinical and biological forms of BL, particularly in 
developing countries outside of Africa, is very limited. A study examining EBV association 
with BL in temperate countries of South America, found a 50% EBV association with 
predominantly abdominal presentation,42 suggesting an intermediate form. The pattern of EBV 
gene expression has not been determined in these BLs. The rate of South American patients 
with BL infected with malaria is extremely low.  Studies in Northeastern Brazil and Northern 
Africa have shown about 85-90% positivity for EBV (like eBL), but have a predominant 
abdominal presentation of the tumors (90%) similar to sBL, suggesting that other still unknown 
genetic or environmental factors might modify the clinical presentation, and possibly response 
to therapy of BL in these regions.43,44
2.1.4
X-Linked Lymphoproliferative Syndrome
X-linked lymphoproliferative (XLP) syndrome is a sex-linked immunodeficiency syndrome. 
Boys with this syndrome are at increased risk for the development of B-cell lymphomas and 
fatal infectious mononucleosis. Most are diagnosed by the time they are 5 years of age, 
although some have gone undiagnosed until much later in life (i.e. 5th or 6th decade). There may 
be an associated hypogammaglobulinemia; however, there is generally not a significant history 
of infections. As a result, patients are usually undiagnosed until they develop a B-cell 
lymphoma or die from infectious mononucleosis. Currently, boys with newly diagnosed high 
grade B-cell NHL are not screened for the XLP gene mutation. Hence the actual frequency of 
these mutations in boys with primary high grade B-cell lymphoma is not known.
We have identified five boys with B-cell lymphoma who have subsequently been found to have 
an XLP gene mutation. These cases were only identified after their clinical course suggested 
the possibility of XLP. Now that we have an accurate and efficient assay to screen for the XLP 
gene mutations, we believe that boys with BL and other B-cell lymphomas should routinely be 
screened for XLP mutations. It is not known whether different XLP mutations result in the 
same clinical phenotype or have prognostic importance. 
2.1.5
Treatment
Mature B cell lymphomas (e.g. Burkitt lymphoma/ leukemia, DLBCL, and MLBCL) are 
aggressive B-cell neoplasms that account for about 15% of cases of childhood cancer in the 
developed world but make up to 50% of all childhood cancers in some parts of Africa. 
Dramatic improvement in the treatment outcome for children with advanced stage BL has been 
achieved since design of the COMP regimen.45-50 The St. Jude Total B regimen incorporated 
sequential high dose methotrexate and escalating continuous infusion Ara-C.46 This resulted in 
excellent outcome for patients with stage III disease; however those with stage IV disease had 
only a 20% long term event free survival. The POG 8617 protocol, derived from the Total B 
regimen which incorporated early administration of high dose pulse Ara-C resulted in improved 
outcome for patients with stage IV disease. During this same time period, Magrath (NCI) 
reported an equally improved result for patients with advanced stage disease using a CHOP-
HDMTX regimen (NCI-7704).45 Further improvement in treatment outcome has been reported 
by Patte (SFOP),51 and the BFM group.49 These regimens incorporated the above mentioned 
agents (COMP, high-dose MTX, and high-dose Ara-C) and VP-16 and/or ifosfamide.
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Patte et al, have reported excellent results for children with mature B-cell lymphomas using the 
LMB 89 regimen, with approximately 87% of B-ALL patients achieving long term event-free 
survival.51,52 For patients with CNS disease, a group with a poorer prognosis, they reported 
70% long term event-free survival.  B-ALL and the BL are rapidly growing and are often 
associated with tumor lysis syndrome (including hyperuricemia). Carefully attention to the 
control of metabolic abnormalities during induction greatly reduced early mortality associated 
with initial treatment. Future directions with this treatment strategy should include efforts to 
reduce morbidity while improving outcome for children with CNS disease.
In light of the outstanding results reported by Patte et al,51 in the SJBCII we adopted the LMB-
89 regimen as a standard of care with the following 3 modifications: 1) deletion of cranial 
radiation (CR RT) for those with CNS disease at diagnosis, (2) placement of an Ommaya 
reservoir in patients with CNS disease at diagnosis, and (3) inclusion of G-CSF after 
myelosuppressive therapy. Treatment was tailored based on disease stage and degree of tumor 
resection.  Group A included only those with completely resected limited stage disease. Group 
C included those with either >70% replacement of marrow with lymphoma cells or those with 
involvement of the central nervous system at diagnosis. Group B included all those not meeting 
criteria for Group A or C and represents the largest and most heterogeneous patient cohort. 
Treatment intensity was the greatest for Group C and mildest for Group A. There were 64 
evaluable patients enrolled on study. Among these, 62 (~97%) have achieved a complete 
remission. There have been only 6 adverse events including relapse in 4 and induction failure in 
2. Fifty-nine of 64 (92%) are alive and disease-free. The international LMB 96 study (CCG, 
SFOP, and UKCCG) randomized patients with Group B and C to standard LMB doses versus 
stepwise reduced doses. Efficacy data show that in group B patients the 3-year EFS are greater 
than 90% in all arms.53 Thus, in Group B patients, non-escalation of cyclophosphamide in the 
second course of COPADM3 and deletion of the maintenance sequence, did not compromise 
the excellent outcome. In contrast, dose reduction for Group C patients was associated with a 
poorer outcome: 3-year EFS was 90% for full dose vs. only 79% for the dose reduced arm, 
respectively. Finally, group C CNS positive patients had an inferior outcome with 3-year EFS of 
71% (vs. 87% for CNS negative group C patients).54 One other group which is associated with a 
poorer treatment outcome is those patients with mediastinal large B-cell lymphoma (MLBCL). 
In the LMB-96 study, the 3 year EFS for these patients was ~70%, a finding consistent with 
other studies. It appears therefore that they require novel or more intensive therapy.
In summary, the intensity of conventional chemotherapy regimens for mature B-cell lymphomas 
is reaching maximal intensity for many of the patients. To further increase the intensity of 
current regimens may lead to unacceptable toxicities. The introduction of compounds with 
diverse mechanisms of action such as rituximab into the current armamentarium of mature B-
cell malignancies is being studied in large randomized clinical trials. For this study we propose 
to use the backbone of LMB-96 study. Specifically, the cyclophosphamide dosing is not 
escalated in the second COPADM block and the maintenance sequence one is deleted in Group 
B patients. Patients with CNS positive Group C will be considered for intra-Ommaya 
chemotherapy administration. Higher-risk patients (e.g., those with MLBLC, Stage III with 
LDH > 2 x ULN, and/or bone marrow/CNS involvement) will be eligible to receive rituximab 
in addition to protocol specified chemotherapy based on disease group.
The only currently available targeted drug for B-cell lymphomas is the anti-CD20 monoclonal 
antibody (rituximab) which has been approved for treatment of adult B-cell lymphomas. 
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Rituximab has also been used in various immune-related diseases such as immune 
thrombocytopenic purpura, myasthenia gravis, and rheumatoid arthritis. In transplant recipients 
it has been used for treatment of post-transplant lymphoproliferative disease, and prevention 
and treatment of acute rejection. There are few data on its use in children with relapsed CD20+ 
lymphomas. The most recent Children's Oncology Group (COG) front-line trial for B-cell 
lymphomas tested the feasibility of incorporating rituximab into Groups B and C of an LMB-96 
based regimen. The Group B arm was completed uneventfully; however, the Group C arm had 
enrollment suspended to evaluate early infectious complications – this arm of the study is now 
being re-opened.
2.1.6 Amendment 3.0 update
Rituximab has long been known to be active in adult CD20+ B-cell lymphomas (low grade and 
high grade).55,56 Initial case reports of its activity in pediatric high grade B-cell lymphomas 
were followed by clinical trials which examined both its activity and safety.57-59 The BFM are 
conducting a trial that features a rituximab window (prior to receiving conventional B-cell 
chemotherapy) that demonstrates both activity and a reasonable/acceptable toxicity profile.57  
The COG performed pilot studies of rituximab incorporating it into both Group B and Group C 
LMB-based therapy.58,59 This safety data were used in part to justify the current use of 
rituximab for Group B primary mediastinal large B-cell lymphomas (PMLBCL) in the current 
SJBC3 protocol. In an effort to determine whether the incorporation of rituximab would 
improve outcome for children with high grade mature B-cell lymphomas, the COG is currently 
conducting a randomized trial (ANHL1131) in which children with Stage III disease and an 
LDH >2x ULN (institutional adult value), stage IV or mature-B-cell leukemia (e.g., Burkitt 
leukemia) are randomized to receive (or not receive) 6 doses of rituximab (Groups B and C).  
This features the addition of two doses of rituximab during the two courses of induction 
therapy (COPADM; total of 4 doses) and one dose of rituximab during the two courses of 
consolidation therapy (CYM and CYVE for Groups B and C, respectively; total of 2 doses) for 
a total of 6 doses of rituximab. The oversight committee for the COG study recently advised 
that the randomization be stopped because of an apparent outcome advantage for those who 
received rituximab (please see attached memo). We would therefore like to expand the use of 
rituximab in SJBC3 to include these higher risk patients studied in the COG study. By way of 
clarification, our institutional adult upper limit of normal for LDH is 260; therefore, stage III 
patients with an LDH > 520 will receive rituximab. Additionally, any patient with Group B 
disease who does not meet the criteria for rituximab incorporation but is found to have high risk 
disease as reflected in a poor COP response (<20% shrinkage; moved to Group C), will also 
have rituximab added to their Group C therapy (same schedule as other Group C patients) and 
any patient who is not already receiving rituximab but is found to have residual disease follow 
CYM #1 and moved to Group C, will receive rituximab with their two courses of CYVE 
therapy (one dose in each course as all Group C patients). We will also permit the incorporation 
of rituximab for other individual patients who in the judgment of the primary physician have 
unique clinical features which they believe put the patient at higher risk. We will include 6 
doses of rituximab to some Group B participants (e.g., those with MLBLC, LDH > 2 x ULN, 
and/or bone marrow involvement) and all Group C participants, with 4 total doses to be given 
during induction (COPADM) and two doses given during consolidation (CYM and CYVE for 
Groups B and C, respectively). This dosing schedule is already in place in SJBC3 for MLBCL.
A challenge for future studies includes the identification of the 10% of patients at the time of 
diagnosis who are at increased risk to fail therapy. We have now started an exciting era where 
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the interest in targeted therapy is blooming; however, without an understanding of the exact 
pathways involved in lymphomagenesis we will be limited in our ability to specifically target 
therapy. Therefore, it is critical to study the various factors and pathways that influence tumor 
cell proliferation and death. St. Jude is positioned to unravel these pathways because of our 
basic science orientation and international connections. Hence, we will provide patients with 
the best treatment available (cure more than 90% of patients with acceptable toxicity) and 
investigate tumor biology in the process.
2.1.7
Psychological Testing
Psychological testing will be performed at baseline, one, three and five years post treatment. 
All selected measures have age-specific norms from large, representative standardization 
samples. Measures also have appropriately demonstrated reliability and validity. Given the 
longitudinal study design, measures were chosen with appropriate test-retest reliability and 
negligible practice effects for the proposed interval between testing time points. An effort was 
made to select measures that could assess the widest age range possible given the likely age 
distribution at presentation and longitudinal design. Nonetheless, no single intelligence measure 
exists that covers this entire age spectrum, such that a separate IQ measure is needed for 
children less than three years of age. Children less than three years of age will be administered 
the Mullen Scales of Early Learning.60 Parent questionnaires will be the same for this age group 
as children older than 3 years of age to allow for some longitudinal analyses including this 
young group. See Appendix IV for a complete listing of test measures, assessed abilities, 
administration age range and estimated testing time for children.
A primary assessment goal was to select a battery that could be administered in a circumscribed 
period of time to reduce the time burden for children, their parents and the research team. The 
administration time for the test battery is 1-3 hours, dependent on the age of the child and the 
speed with which they can complete tasks. It is anticipated that the average administration time 
will be 2 hours. The maximum administration time of 3 hours is only expected for older 
children (i.e., 7-8 years of age) or children that are notably slow (e.g., unusually slow speed of 
cognitive and/or physical processing). Therefore, the administration length of the current 
battery readily lies within the standard length of typical clinical and research batteries. It is 
anticipated that parents will spend on average 45 minutes completing questionnaires while their 
child is being assessed in a separate room. 
2.1.8
Transcranial Doppler Sonography
The chronic anemia of sickle cell disease is known to result in increases in intracranial arterial 
blood flow velocities relative to the normal population.61 The use of transcranial Doppler 
sonography (TCD) is pivotal in the management of children with sickle cell anemia who are 
screened for abnormally high cerebral artery velocities that place them at high risk of stroke.61-65 
In such children, a first stroke can be prevented with initiation of prophylactic red blood cell 
transfusion.66 TCD has also proven to be valuable in the assessment of a wide variety of other 
intracranial pathologies, including: the detection and follow-up of vasoconstriction caused by 
subarachnoid hemorrhage; demonstration of major intracranial vessel stenoses or occlusion; 
determination of brain death; demonstration of the nidus of arteriovascular malformations, and; 
assessment of cerebral hemodynamics following trauma, stroke, or migraine.67-70 Components 
of the treatment are known to be associated with neurotoxicity (methotrexate, cytarabine, 
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ITMHA). In addition, the anemia induced by the lymphoma itself or by chemotherapy may 
result in abnormally elevated cerebral blood flow velocites, such is seen in children with sickle 
cell anemia. The value of TCD to monitor cerebral blood flow in cancer patients has not been 
investigated. In this study, we will obtain preliminary TCD data to gain cursory insight into 
cerebral blood flow velocities before, during and after therapy for childhood B-cell lymphoma. 
These abnormalities will be compared with neuropsychological abnormalities and with findings 
on MRI studies.
2.1.9
Neuroimaging
Based on our preliminary clinical observations, we hypothesize that conventional imaging 
findings may be quite unremarkable in the early acute stage of methotrexate-induced 
neurotoxicity. Therefore, diagnosis may heavily rely on diffusion-based MRI techniques.
In a patient with clinically symptomatic acute methotrexate neurotoxicity71 we found that ADC 
and FA changes in affected white matter were probably decoupled on the front end. This 
suggests that distinct patho-mechanisms may be responsible for the initial, very massive, 
restriction of water diffusion within lesion areas and the subsequent, somewhat delayed FA 
changes.72 Consequently, ADC changes may be an earlier and more sensitive indicator of 
methotrexate-induced neurotoxicity than FA.
Based on our observations we further speculate that methotrexate-induced leukoencephalopathy 
may be either partly or fully reversible, or irreversible; and that the magnitude, extent and 
temporal profile of ADC and FA changes may allow some prediction of outcome during the 
early phase of toxicity. Methotrexate-induced leukoencephalopathy may be clinically 
asymptomatic and therefore more frequent than believed in the past.73,74 However, even if 
clinically asymptomatic during the course of treatment, sub-clinical but not fully reversible 
changes in structural and functional integrity of white matter (and fiber tract systems) may be 
responsible for late adverse effects on neurocognitive functions in patients cured from the 
primary disease.75 Currently available diffusion-based MRI tools (DWI and DTI) provide robust 
and reliable surrogate biomarkers (ADC and FA) of tissue, in particular myelin quality and 
integrity,76,77 hence have the potential to allow detection and quantitative characterization of 
clinically asymptomatic acute methotrexate-induced toxic leukoencephalopathy, and such data 
could be ultimately correlated with late adverse neurocognitive sequelae in cured patients.78
Abnormalities in MRI neuroimaging will be compared with neuropsychological abnormalities.
2.1.10 Bone Mineral Density (BMD)
Deficits in bone mineral density have been well-documented among childhood ALL survivors 
and arise from the interaction of multiple factors.79-83 Approximately 10% of pediatric patients 
have BMD deficits at the time of diagnosis of ALL and over 67% develop such deficits during 
therapy.84 After completion of therapy, BMD deficits have been reported in up to 63% of 
survivors of pediatric ALL,80,85,86 associated with a 5-year cumulative incidence of fracture of 
28% +/- 3%. Risk factors include male sex, age at diagnosis >9 years, and treatment with 
corticosteroids.82 However, bone mineral density changes in B cell NHL patients have not been 
extensively studied. Therefore, we will prospectively study bone mineral density changes in 
this patient population.
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Corticosteroids and cytostatic therapy are associated with diminished BMD.87-90 Children, like 
postmenopausal women, are particularly at risk for steroid-induced osteoporosis because of 
their more rapid bone turnover.91,92 The mechanisms of glucocorticoid-induced osteoporosis are 
multiple. The greatest impact is on the vertebral trabecular bone.93,94 Glucocorticoids decrease 
osteoblastic replication, differentiation, and lifespan through inhibition of genes for type 1 
collagen, osteocalcin, insulin-like growth factors, bone morphogenetic proteins, RANKL, and 
transforming growth factor .95 Glucocorticoids inhibit calcium absorption and prevent 1 -
hydroxylation of vitamin D in the kidney to form the active metabolite 1,25-dihydroxy Vitamin 
D, further impairing calcium metabolism. Glucocorticoids directly inhibit many hormones and 
factors important in calcium accretion of bone.87-90,92,94,96-98 They inhibit the expression of the 
vitamin D receptor in bone and the production of osteocalcin (the major bone matrix protein) 
and decrease local production of cytokines (which normally inhibit bone resorption).92,96,98 
Glucocorticoids inhibit osteoblastic activity and reduce the renal resorption of calcium. This 
effect can result in hypercalciuria and, in turn, increase parathyroid hormone which further 
stimulates calcium loss from bone.94
In summary, to further understand the pathophysiology of BMD loss in children undergoing 
SJBC3 therapy, we will obtain serum calcium, serum magnesium, plasma 1, 25-
dihydroxyvitamin D, spot urine collection for magnesium, Creatinine, calcium, and 
calcium/creatinine ratio. If the calcium:creatinine ratio exceeds 0.2, then a 24-hour urine 
collection will be obtained for creatinine, calcium, and calcium/creatinine ratio.
2.1.11 Anthracycline Cardiotoxicity
While the anthracyclines (e.g., doxorubicin, daunorubicin) have contributed significantly to 
childhood cancer survival, they confer an excess risk of asymptomatic left ventricular 
dysfunction, cardiomyopathy, congestive heart failure and death.99-103 Anthracycline 
cardiotoxicity results in loss of cardiac myocytes, which impedes myocardial development.104 
Unfortunately, myocardial effects can be harmful, asymptomatic,101-105 and progressive.106 
Cardiotoxicity has been reported at all dose levels, but the risk increases with higher 
cumulative doses, younger age at first exposure, time from exposure, and female sex.100-103 As 
many as 5% of at-risk survivors will develop congestive heart failure 15 years after 
treatment.100 The effects may be asymptomatic in up to 57% of survivors, becoming apparent 
only with other physiologic stressors such as infection or pregnancy.101,103
Children exposed to anthracycline agents are at increased risk for cardiac toxicity that may 
become more clinically significant as they age. Late-onset anthracycline toxicity (occurring 
more than 1 year after therapy) generally manifests as left ventricular dysfunction. Although 
subclinical cardiac changes after anthracycline therapy are commonly reported, symptomatic 
disease is seen only occasionally, particularly in children treated on protocols that proactively 
restrict anthracycline cumulative doses.100,101 Because most at-risk patients are asymptomatic, 
ongoing monitoring for late cardiac complications is critical. At a minimum, a baseline 
electrocardiogram and echocardiogram is recommended after the completion of therapy, with 
serial follow-up and cardiac referral as indicated based on age at treatment and cumulative 
cardiotoxic exposures.107 
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Recommended Off Therapy Follow-Up for Late Cardiac Toxicity 
Age at treatment
Frequency of echocardiographic screening by doxorubicin dose
Group A (120 mg/m2)
Group B (120 mg/m2)
Group C (240 mg/m2)
< 1 year old
Every 2 years
Every 2 years
Every year
1 to 4 years
Every 5 years
Every 5 years
Every 2 years
> 5 years
Every 5 years
Every 5 years
Every 5 years
2.1.12 Gonadal and Reproductive Function
Among agents used for treatment of children with non-Hodgkin lymphoma, alkylating agents 
confer a dose-related risk of gonadal toxicity. Because of their faster cell division, male germ 
cells (and their supporting Sertoli cells) are more vulnerable to toxicity from cancer therapy 
than the testosterone-producing Leydig cells.108 Leydig cell failure after radiotherapy is dose-
dependent and inversely related to age at treatment. Thus, infertility is more common than 
androgen insufficiency after treatment for cancer. Treatment with combinations of alkylating 
agents or cumulative doses of cyclophosphamide exceeding 7.5 grams/m2 increases the risk of 
germ cell dysfunction.109-111 Contemporary treatment protocols for non-Hodgkin lymphoma 
attempt to restrict these exposures in an effort to preserve reproductive function. Androgen 
insufficiency as a result of chemotherapy related damage to Leydig cells occurs only 
occasionally. In fact, most males experience normal puberty and are capable of producing 
normal adult levels of testosterone. Appropriate screening for signs and symptoms of androgen 
insufficiency should be performed annually. The Children’s Oncology Group (COG) Long-
Term Follow-Up Guidelines also recommend baseline serum LH, FSH, and testosterone levels 
at age 14 and as clinically indicated in those patients with evidence of delayed puberty or 
androgen insufficiency.107 In young men at risk for infertility, semen analysis remains the most 
accurate noninvasive method for evaluating germ cell function.
In contrast to male physiology, injury to the female germ cells results in both infertility and 
endocrine dysfunction.108 Because females have a fixed number of primordial follicles at birth 
that steadily decline with age, the risk for therapy-related ovarian failure and infertility is 
directly related to age. Thus, pre-pubertal girls, having a greater reserve of follicles, are at 
lowest risk for these complications.108 Female children and adolescents treated with the 
cumulative doses of alkylators prescribed in regimens for non-Hodgkin lymphoma would be at 
relatively low risk for both gonadal or germ cell dysfunction. Evaluation for gonadal failure 
and infertility centers on a good history (to identify irregular or absent menses, history of 
pregnancy, or difficulty conceiving) and an annual physical examination with specific attention 
to Tanner staging. The COG currently recommends baseline serum LH, FSH, and estradiol 
levels at age 13 and when clinically indicated for menstrual irregularity, delayed puberty, or 
symptoms of estrogen insufficiency.
2.2 
Rationale for this Study
Treatment outcomes for children with B-cell NHL are excellent. Further improvements in 
outcome will likely be achieved through more focused study of the biology of the tumors and 
prospective studies of the late effects of treatment. Toward this end, this study features a 
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spectrum of prospective biologic and late effect studies performed in patients treated with a 
modified regimen derived from the very successful LMB-96 regimen.
3.0 
ELIGIBILITY CRITERIA AND STUDY ENROLLMENT
3.1
Inclusion Criteria
3.1.1
Participants from St. Jude and collaborating sites participating in therapeutic and 
biological objectives

Participant must have a histologic diagnosis of a mature B cell lymphoma (e.g., 
Burkitt lymphoma/leukemia, atypical Burkitt lymphoma, diffuse large B-cell 
lymphoma, mediastinal large B-cell lymphoma, mature B-cell lymphoma NOS) as 
defined in the WHO classification.

Participant must be previously untreated, (no more than 72 hours of steroids, one 
intrathecal chemotherapy treatment, and/or emergency radiation).

Participant must be < 22 years of age at the time of diagnosis.

For selected higher-risk CD20+ Group B and all CD20+ Group C participants 
receiving rituximab only (e.g., those with MLBLC, Stage III with LDH > 2 x 
ULN, and/or bone marrow/CNS involvement):
All participants who will receive rituximab must have hepatitis screening prior to 
enrollment. Participants whose results indicate that they are carrier of hepatitis B 
can still be treated as per Group B or C but will NOT receive rituximab. This 
screening must be done for eligibility for participants who will receive rituximab, 
BUT the results are not needed prior to enrollment:
o
Hepatitis B immunization status (vaccination Yes or No)
o
HBsAg
o
Anti-HBs antibody
o
Anti-HBc antibody

All participants must have screening prior to enrollment. Participants whose results 
indicate that they are carrier of hepatitis B can still be treated per Group B and C but 
will NOT receive rituximab.

HIV test has been obtained within 42 days. Participants who test positive for HIV 
cannot be enrolled on therapeutic part of study but are still eligible for biology 
studies.

Informed consent must be obtained according to St. Jude guidelines before 
enrollment into study.
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3.1.2
Participants from collaborating sites participating in biological objectives only (see also 
Appendix VII)

Participant must have a histologic diagnosis of a mature B cell lymphoma (e.g., 
Burkitt lymphoma/leukemia, atypical Burkitt lymphoma, diffuse large B-cell 
lymphoma, mediastinal large B-cell lymphoma, mature B-cell lymphoma NOS) as 
defined in the WHO classification.

Participant must be < 22 years of age at the time of diagnosis.

Participant must be previously untreated, (no more than 72 hours of steroids, one 
intrathecal chemotherapy treatment, and/or emergency radiation) at the time of the 
diagnostic biopsy.

Informed consent must be obtained by local PI or his/her designee according to 
ICH/Good Clinical Practice and local guidelines before enrollment into study.
3.2
Exclusion Criteria
3.2.1
Participants from collaborating sites participating in therapeutic and biological objectives

Participants known to be HIV positive (for therapeutic part of protocol, HIV 
participants are eligible for biology studies).

Participants who are pregnant or lactating.

Inability or unwillingness of research participant or legal guardian to consent.
3.2.2
Participants from collaborating sites participating in biological objectives only

Inability or unwillingness of research participant or legal guardian to consent.

Histologic diagnosis other than a mature B-cell lymphoma as defined in the WHO 
classification.
3.3
Research Participant Recruitment and Screening
3.3.1
St. Jude Participants
Research participants at St. Jude will be recruited by study investigators through regular 
clinical practice.
3.3.2
Collaborating sites participating in therapeutic and biological objectives
Rady Children’s Hospital San Diego will collaborate in both the therapeutic and biological 
objectives of this study. Study investigators will recruit research participants through regular 
clinical practice.  
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3.3.3
Collaborating sites participating in biological objectives only
When physicians consider enrolling a patient on the SJBC3 study (biology studies only), they 
will be asked whether frozen tumor and paraffin blocks are available.  If available, investigators 
will recruit and consent participants in accordance with local policy.  See Appendix VII for 
further details.
3.4
Enrollment on Study
3.4.1
St. Jude Participants
The Study Team or Clinical Trials Operations (CTO) will enter the Participant Eligibility 
checklist information into the central enrollment system and release the informed consent 
document. The signed consent form must be faxed to 
 or scanned and emailed the 
CTO at 
 in order to complete enrolment.
To assist with enrollments and consent release, the CTO staff is available Monday through 
Friday. After hours, weekends, and holidays, the study team is referred to the CTO webpage for 
additional resources and instructions, Link:  
3.4.2
Collaborating Sites Participants
Before the collaborating or enrolling affiliate sites screen or enroll a study participant, the site 
completes a RIN request form and submits it to CTO by e-mail: 
 or fax 
 (follow fax by a phone call to 
 to ensure receipt). A RIN is an 
eight-digit automated number beginning with an “R”. Once the form is received, St. Jude will 
register the research participant and then email the RIN to the email address provided on your 
registration form. This will register the participant only, it will not enroll the participant on the 
study. After hours, holidays, and weekends the site RIN request will be answered by the Patient 
Registration team. 
To enroll the study participant, after the RIN is obtained, the site study team will complete an 
eligibility checklist and fax it to 
 (follow fax by a phone call to 
 to 
ensure receipt). Clinical Trials Operations (CTO) will enter the Eligibility Checklist 
information into the central enrollment system to officially enroll the participant on the trial. 
Collaborating sites will scan/fax a copy of the signed informed consent document to the study 
team at 
. 
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4.0
TREATMENT PLAN
4.1
Treatment Overview
Overview - the chemotherapy regimen used varies with grouping based on extent of disease 
(see Appendix II for definition of Group A, B, and C).
Note about administration guidelines for all treatment groups:  Minor variations in the timing, 
duration of chemotherapy infusions, and hydration guidelines are acceptable, as long as the 
treating investigator determines that there is no impact on patient safety.
4.1.1
Expanded overview
Group A (resected Stage I and resected abdominal Stage II)
CR eval
COPAD ------------------> COPAD
Group B (participants not meeting criteria for Group A or C, see Appendix II
  CR eval
COP→COPAD M3→COPAD M3→CYM→
CYM
No. 1
No. 2
No. 1
No. 2
Group C (B-ALL with >25% BM blasts; CNS involvement, Group B COP failures i.e., 
<20% reduction)
COP →
COPAD M8→
No. 1
COPAD M8→
No.2
CYVE →
No. 1
      CR eval
CYVE →
No. 2
Seq. →
No. 1
Seq. →
No. 2
Seq. → 
No. 3
Seq→
No. 4
4.2
Intrathecal Treatment
Group
Pre-Phase
Induction
X 2
Consolidation
X 2
Continuation
A
------------
COPAD
-----------
-----------
B
COP
COPAD-M3
CYM
-----------
C
COP
COPAD-M8
CYVE
Seq. No 1,2,3,4
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Page 17 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
Intrathecal chemotherapy will be administered immediately after cerebrospinal fluid is 
collected for diagnosis, and the dosage is age-dependent as following: 
Age
Methotrexate
Hydrocortisone
Ara-C
Volume (mL)
<1
8 mg
8 mg
16 mg
8
> 1 & < 2
10 mg
10 mg
20 mg
10
> 2 & < 3
12 mg
12 mg
24 mg
12
> 3
15 mg
15 mg
30 mg
15
This will apply to all treatment groups for which IT is required, except participants > 3 years of 
age with Ommaya reservoir – see below.
4.2.1 Ommaya reservoir treatment (CNS+ only) – this is optional for CNS+ participants
For participants with an Ommaya reservoir, CNS-directed therapy will be administered as 
follows:
Age
Methotrexate
Hydrocortisone
Ara-C (mg/m2)
Volume (ml)
3 yr
6
15
50 mg/m2
3 
(50 mg max)
Algorithm for methotrexate dose adjustment via Ommaya reservoir
Methotrexate CSF concentration (μM*)
  Methotrexate boost dose
At 24 hours
> 40
None
15-40
None
8-15
3 mg 
4-8
4 mg
< 4
6 mg
At 48 hours
>4
None
<4
2 mg
Dose adjustment targeted a CNS methotrexate exposure ≥1.0 μM for 72 hours.
*microMolar
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Page 18 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
4.3
Chemotherapy Details
4.3.1
GROUP A Treatment Details
Participants in Group A (resected Stage I and Abdominal Stage 2) will receive two 
courses of COPAD.  The second course can be given as soon as count recovery allows; 
however, it should usually be no more than 21 days after the first course.
Vincristine
2 mg/m2 (max single dose 2 mg) IV bolus on Day 1 and 6
Prednis(ol)one*
60 mg/m2/day (divided into bid doses) orally on Days 1-7 
inclusive.
Cyclophosphamide
250 mg/m2/dose every 12 hours as a 15 minute infusion on Days 
1-3 inclusive (500 mg/m2/day).  The first dose should be given on 
day 1 prior to the start of the doxorubicin infusion.  Hydration 
should be maintained at a rate of 3000 mL/m2/day (125 
mL/m2/hour).  Continue hydration until 12 hours after the last 
dose of cyclophosphamide.
Doxorubicin
60 mg/m2 as a 6 hour infusion, after the first dose of 
cyclophosphamide
G-CSF
5 mcg/kg/day subcutaneously on Day 7-21 inclusive.  G-CSF 
should be discontinued when the post nadir ANC reaches 
2000/mm3 even if prior to day 21.
Group A:  COPAD
Days
1
2
3
4
5
6
7
Prednis(ol)one*














Vincristine


Cyclophosphamide






Doxorubicin

G-CSF^
 to D21
*Oral prednisone can be substituted with methylprednisolone at 30 mg/m2/day IV 
(divided into bid doses) for patients who cannot tolerate the oral medication.
^In the event the patient cannot receive G-CSF, Peg-Filgrastim can be substituted:
 < 45 kg: 0.1 mg/kg/dose subQ Once (to be given the same day GCSF were to 
start)
≥ 45 kg: 6 mg subQ Once (to be given the same day GCSF were to start)
The second course of COPAD should start at day 21 (or earlier), providing the blood 
count has recovered with ANC > 1.0 x 109/L and platelet count > 100 x 109/L.  G-CSF 
should have been discontinued at least 24 hours previously.
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Page 19 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
4.3.2
GROUP B Treatment Details
Intravenous fluids should be given at a rate of 3000 mL/m2/day.  Use of Rasburicase 
may preclude the need for HCO3
4.3.2.1 Group B Pre-Phase:  COP
Cyclophosphamide
300 mg/m2 as an infusion over 15 minutes on Day 1
Vincristine
1 mg/m2 (max single dose 2 mg) IV bolus on Day 1
Prednis(ol)one*
60 mg/m2/day (divided into bid doses) orally on Days 1-7
IT medications
Methotrexate, hydrocortisone, and cytarabine - IT injection on 
Day 1 (dose varies with age – see Section 4.2)**
Days
1
2
3
4
5
6
7
Vincristine

Prednis(ol)one*














Cyclophosphamide

IT MTX

IT HC

IT Ara-C

*Oral prednisone can be substituted with methylprednisolone at 30 mg/m2/day IV 
(divided into bid doses) for patients who cannot tolerate the oral medication.
**If blasts identified on this LP, participant will be moved to Group C.
Tumor response evaluation should be performed on day 7 (see Section 7.0), before 
proceeding with COPAD M3
Non-responding patients (< 20% reduction in size):  Treated with Group C:  start with 
COPADM8 #1.
In some settings (e.g. critical metabolic, renal, or clinical condition) a second course of 
COP can be considered, see Section 4.4.2 for details of management of tumor lysis.
In cases where transaminases are > 10 x ULN, a delay in COPADM3 can be considered.  
These cases should be discussed with the PI and PharmD.
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Page 20 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
Group B Induction:  COPADM3 x 2 cycles
The first COPADM3 course starts on Day 8 as long as clinical condition permits.  Any 
participant who has renal impairment during induction should be discussed with 
PharmD regarding need for additional renal studies.  
NOTE:  Group B COPAD M3 cycles for 1 and 2 are identical – not to be confused 
with Group C.   Cyclophosphamide dose is 250 mg/m2/dose every 12 hours (500 
mg/m2/day) for Group B.  
Vincristine
2 mg/m2 (max dose 2 mg) as IV bolus on Day 1
Prednis(ol)one*
60 mg/m2/day (divided into bid doses) orally on Days 1-7 inclusive
Methotrexate
3000 mg/m2 in dextrose 5% as IV infusion over 3 hours on Day 1.  
See Section 4.4.1 for further details.
Leucovorin
15 mg/m2 orally or IV every 6 hours for a total of 12 doses (or as 
required depending on methotrexate levels, see Section 4.4.1 for 
further details).  This begins at 24 hours from the start of the 
methotrexate infusion.
Cyclophosphamide
250 mg/m2/dose every 12 hours (500 mg/m2/day) as an infusion 
over 15 minutes on days 2-4 (6 doses).  The first dose is given 
before start of the doxorubicin infusion.  Continue hydration at a 
rate of 3000 mls/m2/day until 12 hours after the last dose of 
cyclophosphamide.
Doxorubicin
60 mg/m2 as a 6 hour infusion, after the first dose of 
cyclophosphamide.
IT medications
Methotrexate and hydrocortisone IT injection on Days 2 and 6 
(dose varies with age – see Section 4.2).  Administer Day 2 IT 
12-24 hours after HDMTX starts and before leucovorin 
rescue begins
Rituximab
375 mg/m2 Days1 and 3.  Rituximab treatment applies to higher-
risk Group B participants [e.g., mediastinal large B-cell 
lymphoma (MLBCL), Stage III with LDH > 2 x ULN, and/or 
bone marrow positive at diagnosis only].  See Appendix II for 
rituximab administration guidelines.
G-CSF
5 mcg/kg/day by subcutaneous injection on Days 7-21 inclusive.  
G-CSF should be discontinued when the post nadir ANC reaches 
2000/mm3, even if prior to Day 21.
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Page 21 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
Group B Induction:  COPAD M3 x 2 cycles
Days
1
2
3
4
5
6
7
Vincristine

Prednis(ol)one*














Methotrexate

Leucovorin

Cyclophosphamide






Doxorubicin

Rituximab – higher-
risk Group B 
participants (e.g., 
MLBCL, Stage III 
with LDH > 2 x ULN, 
and/or bone marrow 
positive at diagnosis)


IT MTX


IT HC


G-CSF^
 to D21
*Oral prednisone can be substituted with methylprednisolone at 30 mg/m2/day IV (divided into 
bid doses) for patients who cannot tolerate the oral medication.
^In the event the patient cannot receive G-CSF, Peg-Filgrastim can be substituted:
 < 45 kg: 0.1 mg/kg/dose subQ Once (to be given the same day GCSF were to 
start)
≥ 45 kg: 6 mg subQ Once (to be given the same day GCSF were to start)
   
Course #2 of COPAD M3 should start when ANC >1.0 x 109/L and platelets > 100,000 
x 109/L. The course should not be given less than 14 days after the start of COPAD M3 
#1 and should be delayed for 24 hours from the last dose of G-CSF. If hematological 
recovery has not occurred by Day 25, a bone marrow examination should be considered. 
If there are residual blasts in the marrow COPADM3 #2 should be given. If the marrow 
is hypoplastic, discuss with PI.
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Page 22 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
4.3.2.3 Group B Consolidation:  CYM x 2 cycles
Each cycle of CYM should start when ANC > 1.0 x 109/L and platelets > 100,000 x 109/L.
Methotrexate
3000 mg/m2 in dextrose 5% as IV infusion over 3 hours on Day 1.
Leucovorin
15 mg/m2 orally or IV every 6 hours for a total of 12 doses (or as 
required depending on methotrexate levels, see Section 4.4.1.3 
for further details).  This begins at 24 hours from the start of the 
methotrexate infusion.
Cytarabine
100 mg/m2 in either dextrose or saline as continuous infusion over 
24 hours.  Repeat daily from Day 2-6 inclusive (total of 5 days).
IT medications
Methotrexate and hydrocortisone IT injection on Day 2.  
Cytarabine and hydrocortisone IT injection on Day 7 (dose varies 
with age – see Section 4.2).  Administer Day 2 IT methotrexate 
and hydrocortisone 12-24 hours after HDMTX starts and 
before leucovorin begins.
Rituximab*
375 mg/m2 day 1.  Rituximab treatment applies to higher-risk Group 
B participants (e.g., mediastinal large B-cell lymphoma (MLBCL) 
and Stage III with LDH > 2 x ULN, and/or bone marrow positive at 
diagnosis.  See Appendix I for rituximab administration guidelines.
G-CSF
5 mcg/kg/day by subcutaneous injection on Days 7-21 inclusive.  
G-CSF should be discontinued when the post nadir ANC reaches 
2000/mm3, even if prior to Day 21.
Days
1
2
3
4
5
6
7
8
Methotrexate

Leucovorin

Cytarabine





Rituximab (e.g., 
MLBCL, Stage III 
with LDH > 2 x 
ULN, and/or bone 
marrow positive at 
diagnosis)

IT MTX 

IT Ara-C

IT HC


G-CSF^
 to D21
^In the event the patient cannot receive G-CSF, Peg-Filgrastim can be substituted: 
 < 45 kg: 0.1 mg/kg/dose subQ Once (to be given the same day GCSF were to 
start)
≥ 45 kg: 6 mg subQ Once (to be given the same day GCSF were to start)
 
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Page 23 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
Following recovery from CYM 1, a full assessment of response should be carried out.  Any residual masses 
should be surgically excised, or biopsied if excision is not possible (see Section 7.0).
If histology negative
Continue with CYM 2
If histology positive (even if completely resected)
Change to Arm C1 starting with CYVE 1
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Page 24 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
4.3.3
Group C Treatment Details
Intravenous fluids should be given at a rate of 3000 mL/m2/day.  Use of rasburicase 
may preclude the need for HCO3
4.3.3.1 Group C Pre-Phase:  COP
Cyclophosphamide
300 mg/m2 as an infusion over 15 minutes on Day 1
Vincristine
1 mg/m2 (max single dose 2 mg) IV bolus on Day 1
Prednis(ol)one*
60 mg/m2/day (divided into bid doses) orally on Days 1-7
IT medications
Methotrexate, cytarabine, and hydrocortisone IT injection on 
Days 1, 3, and 5 (dose varies with age – see Section 4.2)
Leucovorin
5 mg/m2/dose (max 5 mg) po given at 24 and 30 hours after IT on 
Days 2 and 4
Days
1
2
3
4
5
6
7
Cyclophosphamide

Vincristine

Prednis(ol)one*














IT MTX



IT HC



IT Ara-C



Leucovorin




*Oral prednisone can be substituted with methylprednisolone at 30 mg/m2/day IV 
(divided into bid doses) for patients who cannot tolerate the oral medication.
Tumor response evaluation should be performed on day 7.
In some settings (e.g. critical metabolic, renal, or clinical condition) a second course of 
COP can be considered.  See Section 4.4.2 for details of management of tumor lysis.
In cases where transaminases are > 10 x ULN, a delay in COPADM3 can be considered.  
These cases should be discussed with the PI or PharmD.
St. Jude Children's Research Hospital
IRB NUMBER: Pro00001026
IRB APPROVAL DATE: 07/25/2022

SJBC3
Page 25 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
4.3.3.2 Group C Induction:  COPADM8 cycle 1
COPAD M8 should start on day 8 of COP pre-phase therapy, as long as clinical 
condition permits.  Any participant who has renal impairment should be discussed with 
PharmD regarding need for additional renal studies.
Vincristine
2 mg/m2 (max dose 2 mg) as IV bolus on Day 1
Prednis(ol)one*
60 mg/m2/day (divided into bid doses) orally on Days 1-7 
inclusive
Methotrexate
8000 mg/m2 in dextrose 5% as IV infusion over 4 hours on Day 
1.  NOTE:  Higher dose than for Group B.  See Section 4.4.1 
for details.
Leucovorin
15 mg/m2 orally or IV every 6 hours for a total of 12 doses (or as 
required depending on methotrexate levels, see Section 4.4.1.3 
for further details).  This begins at 24 hours from the start of the 
methotrexate infusion.
Cyclophosphamide
250 mg/m2/dose every 12 hours (500 mg/m2/day) as an infusion 
over 15 minutes on days 2-4 (6 doses).  The first dose is given 
before start of the doxorubicin infusion.  Continue hydration until 
12 hours after the last dose of cyclophosphamide.
Doxorubicin
60 mg/m2 as a 6 hour infusion, after the first dose of 
cyclophosphamide
Rituximab
375 mg/m2 Days 1 and 3. All Group C participants will receive 
rituximab with COPADM8 cycles. See Appendix I for rituximab 
administration guidelines.
IT medications**
Methotrexate, cytarabine, and hydrocortisone IT injection on 
Days 2, 4, and 6 (dose varies with age – see Section 4.2).  
Administer Day 2 IT 12-24 hours after HDMTX starts and 
before leucovorin rescue begins.
G-CSF
5 mcg/kg/day by subcutaneous injection on Days 7-21 inclusive.  
G-CSF should be discontinued when the post nadir ANC reaches 
2000/mm3, even if prior to Day 21.
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IRB NUMBER: Pro00001026
IRB APPROVAL DATE: 07/25/2022

SJBC3
Page 26 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
Group C Induction:  COPADM8 cycle 1
Days
1
2
3
4
5
6
7
Vincristine

Prednis(ol)one*














Methotrexate

Leucovorin

Cyclophosphamide






Doxorubicin

Rituximab


IT MTX



IT HC



IT Ara-C



G-CSF^
 to D21
*Oral prednisone can be substituted with methylprednisolone at 30 mg/m2/day IV (divided into bid 
doses) for patients who cannot tolerate the oral medication.  
**CNS+ participants with Ommaya reservoirs, receive Intra-Ommaya treatment on Day 2 only [plus 
MTX boost doses as needed (section 4.2.1)]
^In the event the patient cannot receive G-CSF, Peg-Filgrastim can be substituted:
   < 45 kg: 0.1 mg/kg/dose subQ Once (to be given the same day GCSF were to 
start)
≥ 45 kg: 6 mg subQ Once (to be given the same day GCSF were to start)
St. Jude Children's Research Hospital
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IRB APPROVAL DATE: 07/25/2022

SJBC3
Page 27 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
4.3.3.3 Group C Induction: COPAD M8 Cycle 2
Course #2 of COPAD M3 should start when ANC >1.0 x 109/L and platelets > 100,000 
x 109/L.  The course should not be given less than 14 days after the start of COPAD M3 
#1 and should be delayed for 24 hours from the last dose of G-CSF.  If hematological 
recovery has not occurred by Day 25, a bone marrow examination should be considered.  
If there are residual blasts in the marrow COPADM3 #2 should be given.  If the marrow 
is hypoplastic, discuss with PI. Note: Group C COPAD M8, cycle 2 
cyclophosphamide dose is higher than Group B dose.  
Vincristine
2 mg/m2 (max dose 2 mg) as IV bolus on Day 1
Prednis(ol)one*
60 mg/m2/day (divided into bid doses) orally on Days 1-7 inclusive.
Methotrexate
8000 mg/m2 in dextrose 5% as IV infusion over 4 hours on Day 1.  
NOTE:  Higher dose than for Group B.  See Section 4.4.1 for 
further details.
Leucovorin
15 mg/m2 orally or IV every 6 hours for a total of 12 doses (or as 
required depending on methotrexate levels, see Section 4.4.1.3 
for further details).  This begins at 24 hours from the start of the 
methotrexate infusion.
Cyclophosphamide
500 mg/m2/dose every 12 hours (1000 mg/m2/day) as an infusion 
over 15 minutes on days 2-4 (6 doses).  The first dose is given 
before start of the doxorubicin infusion.  Continue hydration until 
12 hours after the last dose of cyclophosphamide.
Doxorubicin
60 mg/m2 as a 6 hour infusion, after the first dose of 
cyclophosphamide
Rituximab
375 mg/m2 Days 1 and 3.  All Group C participants will receive 
rituximab with COPADM8 cycles. See Appendix I for rituximab 
administration guidelines.
IT medications**
Methotrexate, cytarabine, and hydrocortisone IT injection on 
Days 2, 4, and 6 (dose varies with age – see Section 4.2).  
Administer Day 2 IT 12-24 hours after HDMTX starts and 
before leucovorin rescue begins.
G-CSF
5 mcg/kg/day by subcutaneous injection on Days 7-21 inclusive.  
G-CSF should be discontinued when the post nadir ANC reaches 
2000/mm3, even if prior to Day 21.
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IRB NUMBER: Pro00001026
IRB APPROVAL DATE: 07/25/2022

SJBC3
Page 28 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
Group C Induction: COPADM8 Cycle 2
Days
1
2
3
4
5
6
7
Vincristine

Prednis(ol)one*














Methotrexate

Leucovorin

Cyclophosphamide






Doxorubicin

Rituximab


IT MTX



IT HC



IT Ara-C



G-CSF^
 to D21
*Oral prednisone can be substituted with methylprednisolone at 30 mg/m2/day IV (divided into bid 
doses) for patients who cannot tolerate the oral medication.
**CNS+ participants with Ommaya reservoirs, receive Intra-Ommaya treatment on Day 2 only 
[plus MTX boost doses as needed (section 4.2.1)]
^In the event the patient cannot receive G-CSF, Peg-Filgrastim can be substituted: 
< 45 kg: 0.1 mg/kg/dose subQ Once (to be given the same day GCSF were to 
start)
≥ 45 kg: 6 mg subQ Once (to be given the same day GCSF were to start)
  
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IRB APPROVAL DATE: 07/25/2022

SJBC3
Page 29 of 74
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
4.3.3.4 Group C Consolidation CYVE x 2 cycles
The first of these two courses should start after COPAD M8 when ANC > 1.0 x 109/L 
and platelets > 100,000 x 109/L. G-CSF should have been stopped for 24 hours before 
the start of this course.  
Variations in the start/stop times of the agents below are acceptable, as long as the 
sequence of administration of the agents and duration of infusions are as close as possible.
Cytarabine
50 mg/m2 by continuous infusion over 12 hours. This should start 
at 2000 hours and run until 0800 the following day. Repeat daily 
x 5.
High-Dose Ara-C
3000 mg/m2 as IV infusion over 3 hours, to start at the end of the 
12 hour infusion of cytarabine for 4 doses (from 0800 to 1100 
hours).
Etoposide
200 mg/m2 in saline (concentration < 0.4 mg/mL) as IV infusion 
over 2 hours daily x 4 doses.  Etoposide starts at 1400 hours, 3 
hours after end of high dose cytarabine.
Rituximab
375 mg/m2 Day 1. All Group C participants will receive 
rituximab with CYVE cycles. See Appendix I for rituximab 
administration guidelines.
G-CSF
5 mcg/kg/day by subcutaneous injection on Days 7-21 inclusive. 
G-CSF should be discontinued when the post nadir ANC reaches 
2000/mm3, even if prior to Day 21. 
Days
1
2
3
4
5
6
7
Cytarabine 
(continuous)





Runs from 2000-0800
High dose Ara-C




Runs from 0800-1100
Etoposide




Runs from 1400-1600
Rituximab

See Appendix I 
G-CSF^
 to D21
^In the event the patient cannot receive G-CSF, Peg-Filgrastim can be substituted: 
  < 45 kg: 0.1 mg/kg/dose subQ Once (to be given the same day GCSF were to 
start)
≥ 45 kg: 6 mg subQ Once (to be given the same day GCSF were to start)
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CYVE Course #2: This course is the same as CYVE #1 and starts once ANC > 1.0 x 
109/L and platelets > 100,000 x 109/L (usually by days 25 to 28).
Following recovery from CYVE2, a complete response evaluation should be performed.  Residual 
masses should be surgically excised or biopsied if excision is not possible (see Section 4.6.5)
If histology negative
Continue on protocol
If histology positive
Remove from protocol treatment; additional treatment to be 
determined by primary physician.
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4.3.3.5 Group C Maintenance
Maintenance therapy includes 4 sequences/blocks of chemotherapy. Each sequence will 
begin when ANC > 1.0 x 109/L and platelets > 100,000 x 109/L.
GROUP C MAINTENANCE SEQUENCE No. 1
Vincristine
2 mg/m2 (max dose 2 mg) as IV bolus on Day 1
Prednis(ol)one*
60 mg/m2/day (divided into bid doses) orally on Days 1-7 inclusive
Cyclophosphamide
500 mg/m2/day given daily as IV infusion over 30 minutes on 
Days 2 and 3.  First dose is given before doxorubicin.
Methotrexate
8000 mg/m2 in dextrose 5% IV infusion over 4 hours on Day 1.  
NOTE: Higher dose than for Group B.  See Section 4.4.1
Leucovorin
15 mg/m2 orally or IV every 6 hours for a total of 12 doses (or as 
modified depending on methotrexate levels, see Section 4.4.1.3 
for details). This begins 24 hours from the start of the 
methotrexate infusion.
Doxorubicin
60 mg/m2 as a 6 hour infusion, after first dose of cyclophosphamide
IT medications**
Methotrexate, cytarabine, and hydrocortisone IT injection on Day 2 
(dose varies with age – see Section 4.2). Administer Day 2 IT 12-24 
hours after HDMTX starts and before leucovorin rescue begins.
G-CSF
5 mcg/kg/day by subcutaneous injection starting 24 hours after 
completion of chemotherapy. G-CSF should be discontinued 
when the post nadir ANC reaches 2000/mm3
Days
1
2
3
4
5
6
7
Vincristine

Prednis(ol)one*














Cyclophosphamide


Methotrexate

Leucovorin

Doxorubicin

IT MTX

IT HC

IT Ara-C

G-CSF^

*Oral prednisone can be substituted with methylprednisolone at 30 mg/m2/day IV (divided into bid doses) for 
patients who cannot tolerate the oral medication.
**CNS+ participants with Ommaya reservoirs, receive Intra-Ommaya treatment on Day 2 only [plus MTX 
boost doses as needed (section 4.2.1)]
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^In the event the patient cannot receive G-CSF, Peg-Filgrastim can be substituted:
 < 45 kg: 0.1 mg/kg/dose subQ Once (to be given the same day GCSF were to 
start)
≥ 45 kg: 6 mg subQ Once (to be given the same day GCSF were to start)
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GROUP C MAINTENANCE SEQUENCE No. 2
Cytarabine
50 mg/m2/dose as subcutaneous injection every 12 hours (100 mg/m2/day) 
Days 1-5
Etoposide
150 mg/m2 IV infusion over 90 minutes Days 1-3
G-CSF
5 mcg/kg/day by subcutaneous injection starting 24 hours after 
completion of chemotherapy. G-CSF should be discontinued when the 
post nadir ANC reaches 2000/mm3
IT Medications
For CNS positive participants only (see chart)
Days
1
2
3
4
5
6
Cytarabine










Etoposide



G-CSF^

IT MTX*

IT HC*

IT Ara-C*

*CNS positive participants only, (either ITMHA or Intra-Ommaya MHA).  
See Section 4.2 for boost.
^In the event the patient cannot receive G-CSF, Peg-Filgrastim can be substituted:
   < 45 kg: 0.1 mg/kg/dose subQ Once (to be given the same day GCSF were to 
start)
≥ 45 kg: 6 mg subQ Once (to be given the same day GCSF were to start)
GROUP C MAINTENANCE SEQUENCE No. 3
Vincristine
2 mg/m2 (max dose 2 mg) as IV bolus on Day 1
Prednis(ol)one
60 mg/m2/day (divided into bid doses) orally on Days 1-7 inclusive
Cyclophosphamide 500 mg/m2/day given as IV infusion over 30 minutes on Days 1 & 2. First 
dose is given before doxorubicin.  Maintain hydration at 3000 mL/m2/day 
until 12 hours after 2nd dose.
Doxorubicin
60 mg/m2 as a 6 hour infusion, after the first dose of cyclophosphamide.
G-CSF
5 mcg/kg/day by subcutaneous injection starting 24 hours after 
completion of chemotherapy. G-CSF should be discontinued when the 
post nadir ANC reaches 2000/mm3, even if prior to Day 21
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IT Medications
For CNS positive participants only (see chart)
Days
1
2
3
4
5
6
7
Vincristine

Prednis(ol)one*














Cyclophosphamide


Doxorubicin

G-CSF^

IT MTX**

IT HC**

IT Ara-C**

*Oral prednisone can be substituted with methylprednisolone at 30 mg/m2/day IV (divided into 
bid doses) for patients who cannot tolerate the oral medication.
**CNS positive patients only (either ITMHA or Intra-Ommaya MHA). See Section 4.2 for boost
^In the event the patient cannot receive G-CSF, Peg-Filgrastim can be substituted: 
  < 45 kg: 0.1 mg/kg/dose subQ Once (to be given the same day GCSF were to 
start)
≥ 45 kg: 6 mg subQ Once (to be given the same day GCSF were to start)
GROUP C MAINTENANCE SEQUENCE No. 4
Cytarabine
50 mg/m2/dose as subcutaneous injection every 12 hours (100 mg/m2/day) 
Days 1-5
Etoposide
150 mg/m2 IV infusion over 90 minutes Days 1-3
G-CSF
5 mcg/kg/day by subcutaneous injection starting 24 hours after 
completion of chemotherapy. G-CSF should be discontinued when the 
post nadir ANC reaches 2000/mm3
IT Medications
For CNS positive participants only (see chart)
Days
1
2
3
4
5
6
Cytarabine










Etoposide



G-CSF^

IT MTX*

IT HC*

IT Ara-C*

*CNS positive patients only (either ITMHA or Intra-Ommaya MHA).  See Section 4.2 for boost
^In the event the patient cannot receive G-CSF, Peg-Filgrastim can be substituted:
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  < 45 kg: 0.1 mg/kg/dose subQ Once (to be given the same day GCSF were to 
start)
≥ 45 kg: 6 mg subQ Once (to be given the same day GCSF were to start)
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4.4
Supportive Care Guidelines
These guidelines are provided to help physicians caring for participants treated on this protocol.  
They are guidelines and not protocol requirements. Nothing in these guidelines is intended to 
supplant the judgment of the treating physician regarding patient management. Current 
institutional practice may dictate other approaches to the management of the areas discussed in 
this section.
4.4.1
High-Dose Methotrexate Administration
Hydration
Pre-hydrate with 250 ml/m2/hr with alkalinization for a minimum of 2 hours in order to achieve 
a pH of > 6.5.
Methotrexate is administered in dextrose (piggyback into IVF with bicarb) at a dose of 3000 
mg/m2 over 3 hours in Group B and a dose of 8000 mg/m2 over 4 hours in Group C 
participants.
After methotrexate infusion, continue hydration at a rate of 3000 mLs/m2/day with dextrose 5% 
with added NaHCO3 (40 mEq/l) and KCL (20 mEq/l) until MTX level is < 0.5µM.  When 
MTX level < 0.5µM added NaHC03 may be removed and urine pH’s do not need to be 
checked.  Increased rate of hydration must continue until 12 hours after the last dose of 
cyclophosphamide.
Note:  doxorubicin is unstable in alkaline solution, so must give via a separate lumen from 
alkaline hydration fluids.  If no alternate lumen is available, then oral alkalinization is an 
alternative.  Give 1 mEq/kg NaHCO3 every 6 hours and increase as necessary to maintain the 
urine pH at > 7 as an alternative.  It is important to check that the urine pH is maintained > 7.
Drug interactions
Drugs which compromise renal function (e.g. aminoglycosides) can decrease clearance of 
methotrexate and lead to systemic toxicity. Avoid concurrent use of NSAIDs, omeprazole, 
azole antifungals, salicylates and sulphonamides. Large doses of penicillin may interfere with 
the active renal tubular section of methotrexate.  
Leucovorin
Leucovorin (folinic acid) 15 mg/m2 should be given orally or IV every 6 hours.  The dose 
should be rounded up to the nearest 5 mg.  The rescue begins 24 hours from the start of the 
methotrexate infusion.  Intrathecal drugs should be given before rescue starts.  If vomiting 
occurs within 30 minutes, repeat the dose.  If persistent vomiting or diarrhea occurs, give 
leucovorin by IV injection.  Methotrexate levels, urea and electrolytes should be measured 
daily for 3 days after methotrexate infusion.  Strict attention should be paid to fluid balance.  
The leucovorin dose should be modified as required based in the methotrexate level.  
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Leucovorin dosage adjustments: Patients will have their leucovorin rescue increased if they 
meet any of the following criteria:
3 g/m2
8 g/m2
MTX level
24 hour
>5 µM
>10 µM
48 hour*
>1 µM
>1 µM
72 hour
>0.1 µM
>0.1 µM
If the patient meets any of these criteria, the leucovorin dosage will be individualized and 
plasma concentrations will be monitored until the plasma level is <0.1 µM.  Concomitant 
administration of vincristine and HD MTX may delay the MTX clearance.
*If the 48 hour MTX level is < 0.15 µM/l, discontinue leucovorin rescue.
Group C:  Patients who experience Grade IV GI toxicity after COPAD M8 #1 will receive 
COPAD M3 for their subsequent courses.  Patients who experience Grade IV GI toxicity after 
COPAM M3 #1 will begin leucovorin at 18 instead of 24 hours with subsequent HDMTXs.
A further check on the MTX level should be done 24 hours after the day 6 IT in Group C 
COPADM8 courses to ensure that the level does not increase again after IT MTX and to alter 
leucovorin rescue as required.  Hydration should continue beyond 72 hours in the following 
situations:

If there is still evidence of tumor lysis

If cyclophosphamide infusion is still in process

If the MTX level is still > 0.15 mol/L (1.5 x 10-7M)
4.4.2
Management of Tumor Lysis Syndrome (TLS)
Patients with mature B cell lymphomas are at risk for Tumor Lysis Syndrome (hyperkalemia, 
hyperuricemia, hyperphosphatemia, hypocalcemia, and secondary renal dysfunction).
It is suggested that baseline serum chemistries (urate, urea, creatinine, electrolytes, Ca, Mg, and 
PO4) be obtained pre-treatment and frequently after therapy is started, and a central venous line 
(Double Lumen Hickman Catheter or DLHC) be inserted for fluid administration and 
monitoring.
Preventative measures
1. Give allopurinol 300 mg/m2 daily divided TID.  Alternatively, rasburicase may be given 
daily until the lysis period is over.  
2. Intravenous fluids should be started at least 12 hours before chemotherapy, 3000 
mL/m2/day.  NO ADDED POTASSIUM.
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Suggested monitoring during Induction chemotherapy
1. Strict monitoring of fluid balance is essential.  
2. Check blood pressure frequently.
3. Daily weights.
4. Measurement of serum chemistries at least every 6 hours.
4.4.4
Prophylactic antibiotics
Co-trimoxazole prophylaxis is strongly recommended for all participants, starting with 
COPADM #1 and continuing for 6 months after completion of treatment.
4.4.5
Fever and neutropenia
Fever with neutropenia should be evaluated immediately.  After blood cultures are obtained, 
broad spectrum antibiotics should be started as soon as possible.
4.4.6
Blood product support
Blood product support should be provided as indicated clinically.  It is suggested to keep 
platelet count >20,000 x 109/L.
4.4.7
Growth factors (G-CSF)
G-CSF should be stopped for at least 24 hours prior to next course of chemotherapy.
4.5
Dosage Modifications for Toxicity
4.5.1
Seizures
Discussion with PharmD and PI regarding anticonvulsants and potential need for modification 
of chemotherapy should be considered in the event of seizures.
4.5.2
Etoposide reactions
Cardiovascular effects
Transient hypotension has occurred in about 1 to 2 % of patients following rapid IV 
administration of etoposide during clinical trials. However, hypotension has not been 
associated with cardiac toxicity or electrocardiogram changes. Blood pressure usually 
normalizes within a few hours after discontinuation of the infusion. To avoid this complication, 
etoposide should be infused over 30 – 60 minutes. If hypotension should occur, stop the 
infusion, and if necessary, give 10 mL/kg NS bolus over 15 minutes. Repeat as necessary. Once 
symptoms resolve, resume infusion at ½ previous infusion rate until full dose administered. If 
hypotension recurs, stop infusion and administer 10 mL/kg NS bolus as indicated. Once 
hypotension resolves, resume infusion at ½ previous infusion rate until complete. Consider 
infusing NS at 1-1.5 x maintenance during remainder of infusion. For all subsequent doses, 
further dilute and infuse over 2 hours.
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Sensitivity reactions
Anaphylactoid reactions consisting principally of chills, rigors, diaphoresis, pruritis, loss of 
consciousness, nausea, vomiting, fever, bronchospasm, dyspnea, tachycardia, hypertension, 
and/or hypotension have occurred in 0.7 – 3% of patients receiving etoposide. Other 
manifestations include flushing, rash, substernal chest pain, lacrimations, sneezing, coryza, 
throat pain, back pain, abdominal cramps, and auditory impairment. Facial/lingual swelling, 
coughing, diaphoresis, cyanosis, tightness in the throat, and laryngospasm have also occurred.
If an anaphylactoid reaction should occur:
1. Stop the infusion immediately and notify H/O Fellow or Attending MD
2. Administer the following as indicated:
a) diphenhydramine 1mg/kg IV  (max dose 50 mg)
b) hydrocortisone 50 – 100 mg/m2 IV
c) epinephrine 0.01 mg/kg of a 1:1000 concentration for SQ administration
d) fluid bolus 10 mL/kg NS infused over 15 minutes
3. Once symptoms have resolved, resume infusion at ½ previous rate until infusion 
complete. Consider infusing NS at 1-1.5 x maintenance during remainder of infusion.
4. If anaphylaxis recurs, stop the infusion and re-treat as above.  Do not administer 
remainder of dose.  Consider substituting etoposide with etoposide phosphate 
(Etopophos®) for all subsequent doses.
5. If anaphylaxis does not recur, pre-medicate all subsequent doses with diphenhydramine 
1mg/kg (max 50 mg) and hydrocortisone 50 – 100 mg/m2. Consider slowing the loading 
dose to be administered over 1 hour. 
6. Have at bedside all of the following for all subsequent infusions:
a) Diphenhydramine 1mg/kg IV (max 50 mg)
b) Hydrocortisone 50 – 100 mg/m2 IV
c) Epinephrine 0.01 mg/kg of a 1:1000 concentration for SQ administration
Note: anaphylactoid reactions are still possible with etoposide phosphate.  If the 
patient cannot tolerate the substitution, drug is contraindicated and must be 
discontinued.
4.5.3
Nephrotoxicity
Consultation with PharmD and PI is suggested for severe renal dysfunction. In the event of 
toxicity secondary to high dose methotrexate, consider carboxypeptidase (glucarpidase).
4.5.4
Cardiotoxicity
Hold doxorubicin until fractional shortening is > 28%.  Consultation with cardiologist 
suggested if clinically indicated.
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4.5.5
Hepatic toxicity
Prior to starting IV methotrexate
If SGOT or SGPT is > 20 x upper limit of normal, consult with PI for further recommendation.  
If transaminases are between 10 and 20 x upper limit of normal, wait 48 hours and re-check to 
ensure that they are decreasing.
Anthracyclines and vincristine
Anthracyclines and vincristine dosages should be modified in patients with elevated direct 
bilirubin concentrations or other evidence of biliary obstruction.  

Direct bilirubin 2-4 mg/dl:
50% dosage decrease

Direct bilirubin 4-6 mg/dl:
75% dosage decrease

Direct bilirubin >6 mg/dl:
withhold dose
4.5.7
Modifications for age
For participants < 1 year of age, a 1/3 dosage reduction should be made; if this course is well 
tolerated, an increase to full dosage may be considered.   Intrathecal doses are age specific, see 
Section 4.2.
4.6 
Surgical Guidelines
4.6.1
Surgical intervention at diagnosis
Obtaining tissue for diagnosis should be done with the least invasive procedure possible.  
Surgical resection of Stage I or localized abdominal disease (i.e., ileo-caecal area with 
involvement of regional mesenteric lymph nodes only) should be considered in consultation 
with surgery.
4.6.2
Abdominal disease
Debulking procedures are not indicated (see Section 4.6.1).
4.6.3
Mediastinal mass
Consultation with Anesthesia, Surgery, and Interventional Radiology is suggested to determine 
the safest procedure for obtaining tissue for diagnosis.
4.6.4
Bone primary
Consultation with Surgery is suggested for those presenting with primary involvement of bone.
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4.6.5
Secondary surgery
For residual masses prior to CYM #2 in Group B patients, after CYVE #2 in Group C patients
Histological evaluation of residual masses should be done in consultation with Surgery.  
Complete resection is advisable; however, biopsy (with interventional radiology) may be 
considered depending on site of disease and potential risk.
Group B patients
If there is histologic confirmation of residual tumor (even if this is resected with the surgery) the 
patient should be transferred to Group C (starting with CYVE #1).
Group C patients
If there is histologic confirmation of residual tumor (even if this is resected with the surgery) 
the participant is considered a treatment failure and should be taken off treatment.
Residual bone lesions
Residual diagnostic imaging abnormalities are common.  Consultation with Orthopedics and 
Radiology is suggested as clinically indicated.
4.7
Radiation Therapy Guidelines
There are no specific indications for RT in this study.
5.0
DRUG INFORMATION
5.1
Cytarabine (Ara-C) (Cytosar-U)
Source and pharmacology: Cytarabine is a deoxycytidine analogue.  It must be tri-
phosphorylated to its active form, ARA-CTP, by deoxycytidine kinase and other nucleotide 
kinases.  Ara-CTP inhibits DNA polymerase. In addition, ara-CTP is incorporated into DNA as 
a false base, causing inhibition of DNA synthesis. It is cell cycle, S phase specific.  Cytarabine 
does penetrate the blood brain barrier. It is converted to its inactive form, uracil arabinoside, by 
pyrimidine nucleoside deaminase. Approximately 80% of the dose is recovered in the urine, 
mostly as uracil arabinoside (ara-U).
Formulation and stability: Cytarabine is available in multi-dose vials containing 100, 500, 1000 
and 2000mg of lyophilized drug. Intact vials can be stored at room temperature. For IV use, 
either sterile water for injection or bacteriostatic water for injection can be used to reconstitute 
the lyophilized drug. For intrathecal use, only sterile water for injection should be used for 
reconstitution. The 100 and 500 mg vials are reconstituted with 2 and 10 ml respectively 
resulting in a final concentration of 50mg/ml.  he 1000 and 2000mg vials are reconstituted with 
20ml and 40 ml respectively resulting in a final concentration of 50mg/ml. After reconstitution, 
the drug is stable for 8 days at room temperature.
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Supplier:  Commercially available
Toxicity: Myelosuppression is the dose limiting adverse effect, with leukopenia and 
thrombocytopenia being predominant. Other adverse effects reported commonly include nausea 
and vomiting (may be severe at high doses), diarrhea, mucositis, anorexia, alopecia, skin rash 
and liver dysfunction. A flu-like syndrome characterized by fever, muscle and bone aches is 
common. Less common side effects include allergic reactions and cellulitis at the injection site. 
High doses of cytarabine can cause conjunctivitis, hepatitis, and a group of CNS symptoms 
including somnolence, peripheral neuropathy, ataxia, and personality changes. CNS symptoms 
are usually reversible and are more common in patients who have received previous cranial 
irradiation. In addition, a syndrome of sudden respiratory distress progressing to pulmonary 
edema has occurred.
5.2
Cytarabine (High Dose Ara-C)
Source and pharmacology: Cytarabine is a deoxycytidine analogue. It must be tri-
phosphorylated to its active form, ARA-CTP, by deoxycytidine kinase and other nucleotide 
kinases. Ara-CTP inhibits DNA polymerase. In addition, ara-CTP is incorporated into DNA as 
a false base, causing inhibition of DNA synthesis. It is cell cycle, S phase specific. Cytarabine 
does penetrate the blood brain barrier. It is converted to its inactive form, uracil arabinoside, by 
pyrimidine nucleoside deaminase. Approximately 80% of the dose is recovered in the urine, 
mostly as uracil arabinoside (ara-U).
Formulation and stability: Cytarabine is available in multi-dose vials containing 100, 500, 1000 
and 2000mg of lyophilized drug. Intact vials can be stored at room temperature. For IV use, 
either sterile water for injection or bacteriostatic water for injection can be used to reconstitute 
the lyophilized drug. For intrathecal use, only sterile water for injection should be used for 
reconstitution. The 100 and 500 mg vials are reconstituted with 2 and 10 ml respectively 
resulting in a final concentration of 50mg/ml. The 1000 and 2000mg vials are reconstituted 
with 20ml and 40 ml respectively resulting in a final concentration of 50mg/ml. After 
reconstitution, the drug is stable for 8 days at room temperature.
Supplier: Commercially available.
Toxicity: Myelosuppression is the dose limiting adverse effect, with leukopenia and 
thrombocytopenia being predominant. Other adverse effects reported commonly include nausea 
and vomiting (may be severe at high doses), diarrhea, mucositis, anorexia, alopecia, skin rash 
and liver dysfunction. A flu-like syndrome characterized by fever, muscle and bone aches is 
common. Less common side effects include allergic reactions and cellulitis at the injection site.   
High doses of cytarabine can cause conjunctivitis, hepatitis, and a group of CNS symptoms 
including somnolence, peripheral neuropathy, ataxia, and personality changes. CNS symptoms 
are usually reversible and are more common in patients who have received previous cranial 
irradiation. In addition, a syndrome of sudden respiratory distress progressing to pulmonary 
edema has occurred.
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5.3
Cyclophosphamide
Source and pharmacology: Cyclophosphamide is a nitrogen mustard derivative. It acts as an 
alkylating agent that causes cross-linking of DNA strands by binding with nucleic acids and 
other intracellular structures, thus interfering with the normal function of DNA. 
Cyclophosphamide is cell-cycle, phase non-specific. Cyclophosphamide is well absorbed from 
the GI tract with a bioavailability of > 75%. Cyclophosphamide is a prodrug that requires 
activation.  It is metabolized by mixed-function oxidases in the liver to 4-
hydroxycyclophosphamide, which is in equilibrium with aldofosfamide. Aldofosfamide 
spontaneously splits into cyclophosphamide mustard, which is considered to be the major 
active metabolite, and acrolein. In addition, 4-hydroxycyclophosphamide may be enzymatically 
metabolized to 4-ketocyclophosphamide and aldofosfamide may be enzymatically metabolized 
to carboxyphosphamide which are generally considered to be inactive. Cyclophosphamide and 
its metabolites are excreted mainly in the urine. Dosage adjustments should be made in patients 
with a creatinine clearance of < 50 ml/min.
Formulation and stability: Cyclophosphamide is available in 25 and 50 mg tablets.  
Cyclophosphamide is also available in vials containing 100, 200, 500, 1000 and 2000mg of 
lyophilized drug and 75 mg mannitol per 100 mg of cyclophosphamide. Both forms of the drug 
can be stored at room temperature. The vials are reconstituted with 5, 10, 25, 50 or 100 ml of 
sterile water for injection respectively to yield a final concentration of 20 mg/ml. Reconstituted 
solutions may be further diluted in either 5% dextrose or 0.9% NaCl containing solutions. 
Diluted solutions are physically stable for 24 hours at room temperature and 6 days if 
refrigerated, but contain no preservative, so it is recommended that they be used within 24 
hours of preparation.
Supplier: Commercially available
Toxicity: Dose limiting toxicities of cyclophosphamide are bone marrow suppression and 
cardiac toxicity. Cardiac toxicity is typically manifested as congestive heart failure, cardiac 
necrosis or hemorrhagic myocarditis and can be fatal. Hemorrhagic cystitis may occur and 
necessitates withholding therapy. The incidence of hemorrhagic cystitis is related to 
cyclophosphamide dose and duration of therapy. Forced fluid intake and/or the administration 
of mesna decrease the incidence and severity of hemorrhagic cystitis. Other toxicities reported 
commonly include nausea and vomiting (may be mild to severe depending on dosage), 
diarrhea, anorexia, alopecia, immunosuppression and sterility. Pulmonary fibrosis, SIADH, 
anaphylaxis and secondary neoplasms have been reported rarely.
5.4
Doxorubicin (Adriamycin)
Source and pharmacology: Doxorubicin is an anthracycline antibiotic produced by 
Streptomyces peucetius. Doxorubicin exerts its anti-tumor effects in several different ways. 
Doxorubicin intercalates between base pairs of DNA causing steric obstruction, disruption of 
DNA function and inhibition of RNA synthesis. In addition, doxorubicin inhibits 
topoisomerase II, an enzyme responsible for allowing strands of DNA to pass through one 
another as they unwind. Lastly, doxorubicin undergoes enzymatic electron reduction to 
generate highly reactive species, including the hydroxyl free radical, which is thought to be 
responsible for the drug’s cardiac toxicity, but may play a role in its anti-tumor activity as well. 
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Doxorubicin is cell-cycle, phase non-specific. Doxorubicin is widely distributed in the tissues 
and plasma, but does not cross the blood brain barrier to an appreciable extent. It is metabolized 
to doxorubicinol, which is thought to be the major active metabolite, and aglycones. 
Doxorubicin and its metabolites are excreted mainly in the bile and feces (80%). The 
remainder is excreted in the urine. Dosage should be reduced in patients with liver dysfunction 
(bilirubin > 1.2 mg/dl) or renal dysfunction (creatinine > 3 mg/dl).
Formulation and stability: Doxorubicin is available in vials containing 10 mg, 20 mg, 50 mg 
and 200 mg as a 2mg/ml red-orange solution. It is also available in vials containing 10 mg, 20 
mg, 50 mg, 100 mg and 150 mg of doxorubicin as a red-orange lyophilized powder. Intact vials 
of doxorubicin solution should be stored under refrigeration while the lyophilized product 
should be stored at room temperature. Both products should be protected from light. 
Lyophilized doxorubicin can be reconstituted by adding 5, 10, 25, 50 or 75 ml of 0.9% NaCl 
respectively to the 10, 20, 50, 100 and 150 mg vials to produce a final concentration of 2 
mg/ml. Bacteriostatic diluents are not recommended. After reconstitution, the resultant solution 
should be protected from light and is stable for 7 days at room temperature and 15 days if 
refrigerated.
Supplier: Commercially available
Toxicity: Dose-limiting toxicities include myelosuppression and cardiotoxicity. Two forms of 
cardiac toxicity can occur. Acute toxicity may take the form of arrhythmias, heart block or 
pericarditis and may be fatal. The chronic form of cardiotoxicity is related to total cumulative 
dose and is characterized by heart failure. Mediastinal radiotherapy and/or other cardiotoxic 
drugs may increase the risk of cardiotoxicity. In general, total lifetime dosages of 450-
550mg/m2 should not be exceeded. Other toxicities include nausea and vomiting, mucositis, 
alopecia, diarrhea and red discoloration of the urine and other body fluids. Severe tissue 
damage and necrosis can occur upon extravasation. Radiation recall reactions can occur and 
can be severe. Rarely, allergic reactions have occurred. Typhlitis can occur when combined 
with cytarabine.
5.5
Etoposide (VP16) (Vepesid)
Source and pharmacology: Etoposide is an epipodophyllotoxin derived from 
Podophyllumpelatatum. It is thought to act mainly by inhibiting topoisomerase II, causing 
double and single strand DNA breaks. Etoposide is cell cycle, phase-specific, with activity in 
the G2 and S phases. Absorption of etoposide is approximately 30-40% and is highly variable 
and somewhat dose-dependent. It is extensively bound to serum proteins and is metabolized in 
the liver, including cytochrome P450 3A metabolism to several moieties that include a reactive 
oxidized species. Etoposide and its metabolites are excreted mainly in the urine with a smaller 
amount excreted in the feces. Dosage adjustments should be considered in patients with liver 
dysfunction, kidney dysfunction or hypoalbuminemia.
Formulation and stability: Etoposide is available in multi-dose vials containing 100mg, 150mg, 
500mg and 1000mg of etoposide as a 20mg/ml solution and 30% alcohol. Etoposide is also 
available as a 50 mg capsule. The intact vials of etoposide solution should be stored at room 
temperature. The capsules should be stored under refrigeration. Etoposide solution should be 
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diluted in D5W or 0.9% NaCl prior to administration. Solutions with a final concentration of 
0.2 and 0.4 mg/ml are stable at room temperature for 96 hours and 24 hours respectively.
Supplier: Commercially available
Toxicity: Dose limiting toxicity is myelosuppression. Nausea and vomiting (usually of low to 
moderate severity), diarrhea, mucositis (particularly with high doses), alopecia and anorexia are 
fairly common. Hypotension can occur with rapid infusions. Other side effects reported less 
commonly include hepatitis, fever and chills, anaphylaxis and peripheral neuropathy. 
Secondary leukemia has been reported.
5.6
Etoposide phosphate (Etopophos®)
To be used in case of etoposide reactions (see Section 4.4.3).
Source and pharmacology: Etoposide is an epipodophylotoxin derived from 
Podophyllumpelatatum. It is thought to act mainly by inhibiting topoisomerase II, causing 
double and single strand DNA breaks. Etoposide is cell cycle, phase-specific, with activity in 
the G2 and S phases. Absorption of etoposide is approximately 30-40% and is highly variable 
and somewhat dose-dependent. It is extensively bound to serum proteins and is metabolized in 
the liver, including cytochrome P450 3A metabolism to several moieties that include a reactive 
oxidized species. Etoposide and its metabolites are excreted mainly in the urine with a smaller 
amount excreted in the feces. Dosage adjustments should be considered in patients with liver 
dysfunction, kidney dysfunction or hypoalbuminemia.
Formulation and stability: Etoposide phosphate is a water-soluble ester of etoposide. The 
higher water solubility of etoposide phosphate than that of etoposide lessens the potential for 
precipitation following dilution and during administration. Etoposide phosphate is available in 
single-dose vials containing etoposide phosphate equivalent to 100mg etoposide. The intact 
vials of etoposide solution should be stored at 2 to 8 degrees Celsius. Etoposide phosphate 
solution should be diluted in D5W or 0.9% NaCl prior to administration. Solution is stable at 
room temperature for 24 hours.
Supplier: Commercially available.
Toxicity: Dose limiting toxicity is myelosuppression. Nausea and vomiting (usually of low to 
moderate severity), diarrhea, mucositis (particularly with high doses), alopecia and anorexia are 
fairly common. Hypotension can occur with rapid infusions. Other side effects reported less 
commonly include hepatitis, fever and chills, anaphylaxis and peripheral neuropathy. 
Secondary leukemia has been reported.
5.7
Filgrastim (G-CSF, Neupogen)
Source and pharmacology: G-CSF (granulocytic colony stimulating factor), is a biosynthetic 
hematopoietic agent that is made using recombinant DNA technology in cultures of 
Escherichia coli. G-CSF stimulates production, maturation and activation of neutrophils. In 
addition, endogenous G-CSF enhances certain functions of mature neutrophils, including 
phagocytosis, chemotaxis and antibody--dependent cellular cytotoxicity.
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Formulation and stability: G-CSF is supplied in vials containing 300 mcg and 480 mcg of G-
CSF at a concentration of 300 mcg/ml. The intact vials should be stored under refrigeration. 
The vials can be left out of refrigeration for 24 hours but should be discarded if left at room 
temperature for longer periods of time. G-CSF can be drawn up into tuberculin syringes for 
administration and stored under refrigeration for up to 7 days prior to usage. G-CSF can be 
further diluted for IV infusion in 5% dextrose. Do not dilute in saline---precipitate may form. If 
the final concentration of this product is < 15 mcg/ml, it is recommended that albumin be added 
to a final concentration of 2mg/ml (0.2%) to minimize adsorption of the drug to infusion 
containers and equipment.
Supplier: Commercially available
Toxicity: G-CSF causes marked leukocytosis. Adverse reactions reported commonly include 
bone pain, thrombocytopenia, diarrhea, nausea, rash, alopecia, fever, anorexia and pain or 
bruising at the injection site. Allergic reactions, MI, atrial fibrillation, and splenomegaly have 
been reported rarely. G-CSF is contraindicated in patients with allergy to E. coli derived 
products.
5.8
Peg-filgrastim (pegylated filgrastim, PEG-filgrastim, Neulasta®)
Source and pharmacology: Pegfilgrastim is the pegylated form of recombinant methionyl human 
G-CSF (filgrastim). Pegfilgrastim is produced by covalently binding a 20-kilodalton (kD) 
monomethoxypolyethylene glycol molecule to the N-terminal methionyl residue of filgrastim. 
The molecular weight of pegfilgrastim is 39 kD. G-CSF is a lineage specific colony-stimulating 
factor which regulates the production of neutrophils within the bone marrow and affects 
neutrophil progenitor proliferation, differentiation, and selected end-cell functional activation 
(including enhanced phagocytic ability, priming of the cellular metabolism associated with 
respiratory burst, antibody dependent killing, and the increased expression of some functions 
associated with cell surface antigens).
After subcutaneous injection the elimination half-life of pegfilgrastim ranges from 15 to 80 hours 
and the time to peak concentration ranges from 24 to 72 hours. Serum levels are sustained in 
most patients during the neutropenic period post-chemotherapy and begin to decline after the 
start of neutrophil recovery, consistent with neutrophil-dependent elimination.
Toxicity: mild to moderate medullary bone pain, local irritation at the injection site, headache, 
elevations in alkaline phosphatase, LDH, uric acid, thrombocytopenia, low grade fever, allergic 
reactions, splenomegaly, splenic rupture, exacerbation of pre-existing skin rashes, sickle cell 
crisis in patients with SCD, excessive leukocytosis, ARDS. Fetal toxicities and teratogenic 
effects in humans are unknown. There is conflicting data in animals. It is unknown whether the 
drug is excreted in breast milk.
Formulation and stability: Supplied as a preservative-free solution containing 6 mg (0.6 mL) of 
pegfilgrastim (10 mg/mL) in a single dose syringe with 27 g, ½ inch needle with an UltraSafe® 
Needle Guard. Store refrigerated at 2º-8ºC (36º-46ºF) and in the carton to protect from light. Prior 
to injection, pegfilgrastim may be allowed to reach room temperature protected from light for a 
maximum of 48 hours. Avoid freezing.
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Guidelines for administration: Pegfilgrastim should not be administered in the period between 2 
weeks before and 24 hours after chemotherapy. Do not shake. Dose and administration as 
clinically indicated, per institutional guidelines. See Treatment and Dose Modifications sections 
of the protocol.
Supplier: Commercially available.
5.9
Hydrocortisone, Intrathecal (Cortef, Solu-Cortef)
Source and pharmacology: Hydrocortisone is a synthetic steroid akin to the natural adrenal 
hormone cortisol. Hydrocortisone has phase-specific cytotoxicity, killing lymphoblasts 
primarily during S phase. It has catabolic effect on proteins and alters the kinetics of peripheral 
blood leukocytes. It is excreted in the urine and catabolized in the liver.
Formulation and stability: Solu-Cortef sterile powder is supplied in the following package: 100 
mg plain, and 100 mg, 250 mg, 500 mg, and 1000 mg ACT-O-VIAL (MIX-O-VIAL). Store 
unreconstituted product at controlled room temperature 15-30C (59-86F). Store reconstituted 
solution in the refrigerator and protect from light. Unused solution should be discarded after 3 
days. Use Solu-Cortef (plain vial) for intrathecal use, and reconstitute with 0.9% sodium 
chloride, USP for injection.
Supplier: Commercially available
Toxicity: If given intrathecally, sterile arachnoiditis may occur. Headache, seizures, unusual 
feelings or sensations, loss of feeling or ability to move arms or legs, and difficulty with 
urination or bowel movements may also occur.
5.10
Leucovorin (folinic acid)
Source and pharmacology: Leucovorin is a racemic mixture of tetrahydrofolic acid, which is 
involved as a cofactor for 1-carbon transfer reactions in the synthesis of purine and 
pyrimidines. Leucovorin is a potent antidote for both the hematopoietic and reticuloendothelial 
toxic effects of folic acid antagonists by replenishing reduced folate pools. It is postulated that 
in some cancers, leucovorin enters and “rescues” normal cells from the toxic effects of folic 
acid antagonists, in preference to tumor cells, because of differences in membrane transport and 
affinity for polyglutamylation. Leucovorin is converted in the intestinal mucosa and the liver to 
5-methyl-tetrahydrofolate, which is also active as a reduced folate. It is excreted primarily in 
the urine with minor excretion occurring in the feces.
Formulation and stability: Leucovorin is supplied in 5, 15 and 25 mg tablets and vials 
containing 50, 100 or 350 mg of leucovorin as a lyophilized powder. The tablets and the 
lyophilized powder can be stored at room temperature. The 50 mg and 100 vials can be 
reconstituted by adding 5 or 10 ml of sterile water or bacteriostatic water for injection 
respectively to yield a final concentration of 10 mg/ml. The 350 mg vials can be reconstituted 
with 17 ml of sterile water or bacteriostatic water for injection to yield a final concentration of 
20 mg/ml. The reconstituted solution is stable for at least 7 days at room temperature. 
Leucovorin may be further diluted in 5% dextrose or 0.9% NaCl containing solutions.
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Supplier: Commercially available
Toxicity: Leucovorin is generally well tolerated. Toxicities that have been reported 
uncommonly include rash, mild nausea, headache, and wheezing (possible allergic reaction). 
Intrathecal leucovorin is contraindicated and has caused neurotoxic deaths. There have been 
rare reports of leucovorin promoting seizures.
5.11
Methotrexate
Source and pharmacology: Methotrexate is a folate analogue that acts by inhibiting 
dihydrofolatereductase. Dihydrofolatereductase is an enzyme important in the conversion of folic 
acid to tetrahydrofolic acid, which is necessary in the synthesis of purine nucleotides and 
thymidylate. By inhibiting the production of tetrahydrofolic acid, methotrexate interferes with 
DNA, RNA and protein synthesis. Methotrexate is poorly and variably absorbed orally, with an 
average of  40% for doses of < 30 mg/m2. At higher dosages, the extent of absorption decreases.  
Methotrexate is approximately 50% protein bound. It distributes widely into body tissues and 
fluids with sustained concentrations in the kidney and the liver. Methotrexate undergoes 
metabolism by cytosolic aldehyde oxidase to hydroxy methotrexate. It is excreted mainly in the 
urine as unchanged drug with small amounts being excreted in the bile and feces. The percent 
recovered as unchanged drug in the urine is higher with short infusions than with prolonged 
infusions. Methotrexate has a biphasic elimination with an initial half-life of  2-3 hours and a 
terminal half-life of 10-12 hours. Methotrexate may be “sequestered” in body fluid collections 
and eliminated slowly from these areas. Patients with effusions or GI obstruction should have 
plasma levels monitored closely for delayed excretion following high-dose methotrexate.
Formulation and stability: Methotrexate is supplied in single-dose vials containing 50mg, 
100mg, 200mg, and 250 mg of methotrexate as a 25 mg/ml preservative-free solution and in 
vials containing 20mg, 50 mg, 100mg, 250 mg and 1000mg of lyophilized drug. It is also 
available in 2.5 mg tablets. Methotrexate preservative-free solution and lyophilized drug should 
be stored at room temperature and protected from light.  Methotrexate tablets can also be stored 
at room temperature. The vials containing 20, 50, 100 and 250 mg of lyophilized product can 
be reconstituted by adding sterile water, 0.9% NaCl or D5W to a final concentration not 
exceeding 25 mg/ml. The 1000mg vials containing lyophilized product are reconstituted to a 
final concentration of 50 mg/ml.
Supplier: Commercially available.
Toxicity: The dose limiting toxicities of methotrexate are generally bone marrow suppression, 
ulcerative stomatitis, severe diarrhea or acute nephrotoxicity. Toxicities reported frequently 
include nausea and vomiting, diarrhea, anorexia, alopecia, hepatic toxicity and alopecia. Less 
common side effects include blurred vision, photosensitivity, anaphylaxis, headache, 
pneumonitis, skin depigmentation or hyperpigmentation, rash, vasculitis and encephalopathy. 
During high-dose methotrexate therapy, most patients experience a transient decrease in GFR, 
but renal failure can occur, particularly if the patient does not receive urinary alkalinization and 
aggressive hydration before, during and after receiving high dose methotrexate. Leucovorin 
rescue should be initiated within 48 hours of starting high-dose methotrexate and adjusted 
based on MTX levels to prevent bone marrow toxicity and mucositis. Leucovorin may also be 
necessary after IT administration, especially if IT methotrexate therapy is given to patients with 
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renal dysfunction. Patients with Down syndrome have a tendency to have delayed methotrexate 
clearance and a greater risk of toxicity, despite increased leucovorin rescue.
5.12
Prednisone (prednisolone)
Source and pharmacology: Prednisone is a synthetic congener of hydrocortisone, the natural 
adrenal hormone. Prednisone is a white or yellowish crystalline powder. It binds with steroid 
receptors on nuclear membranes, impairs cellular mitosis and inhibits protein synthesis. 
Prednisone also has potent anti-inflammatory effects and suppresses the immune system. 
Prednisone is well absorbed orally. It is converted to prednisolone, the pharmacologically 
active metabolite, in the liver. Prednisolone is further metabolized to inactive compounds in the 
liver. The metabolites are excreted mainly in the urine.
Formulation and stability: Prednisone is available as various strength tablets and oral solution 
from multiple manufacturers. All dosage forms can be stored at room temperature. At St. Jude 
Children’s Research Hospital, prednisolone oral solution may be substituted for prednisone 
liquid at equal doses due to its superior palatability.
Supplier: commercially available.
Toxicity: Side effects of prednisone vary depending on the duration of its use. Side effects that 
can occur with short term use include sodium and water retention with associated hypertension, 
peptic ulcer with possible perforation and hemorrhage, increased susceptibility to infections, 
emotional instability, insomnia, increased appetite, weight gain, acne and hyperglycemia. Side 
effects more commonly associated with prolonged use include cataracts, increased intraocular 
pressure and associated glaucoma, development of a “cushingoid” state, compression fractures, 
menstrual irregularities, suppression of growth in children, secondary adrenocortical and 
pituitary unresponsiveness particularly in times of stress as in trauma, surgery or illness, 
osteoporosis and muscle wasting.
5.13
Rituximab (Rituxan)
Source and pharmacology: Rituximab is a murine /human chimeric monoclonal antibody. It is 
specific for the CD20 antigen located on B-cells. Rituximab has been shown to mediate 
complement-dependent tumor cell lysis and antibody-dependent cellular cytotoxicity. Direct 
binding to the CD20 antigen is thought to play a role in inhibition of cell growth. Rituximab is 
administered intravenously. The mean serum half-life after a single IV dose of 375 mg/m2 is 
59.8 hours (range 11.1-104.6 hours). 
Formulation and stability: Rituximab is available as 100 mg/10 ml single-use and 500 mg/50 
ml single-use vials. Each vial also contains sodium chloride 9mg/ml, sodium citrate 7.35 
mg/ml, polysorbate 80 0.7 mg/ml and water for injection. Rituximab for injection concentration 
must be diluted with 5% Dextrose or 0.9% NaCl prior to administration. After dilution, unused 
drug is stable for 24 hours when refrigerated (2-8 degrees Celsius) and 12 hours at room 
temperature. Vials should be protected from direct sunlight. 
Supplier: Commercially available.
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Toxicity: Hypersensitivity reactions may occur; therefore, premedication with acetaminophen 
and diphenhydramine should be considered before each infusion. The most common toxicities 
are infusion related and may include chills, fever, headache, nausea, vomiting, angioedema 
(13%), hypotension (10%), bronchospasm (8%), and arrhythmia. Other possible adverse 
reactions include thrombocytopenia, myalgias, arthralgias, asthenia, and throat irritation.
Dosage and route of administration: Do not administer as an intravenous push or bolus.  
Infusions should be initiated at 50 mg/hour for 30 minutes. If hypersensitivity or infusion-
related events do not occur, subsequent infusions can be administered at an initial rate of 100 
mg/hour and increased by 100 mg/hour every 30 minutes as tolerated. Infuse at a maximum 
rate of 400 mg/hour.
5.14
Vincristine
Source and pharmacology: Vincristine is an alkaloid obtained from the periwinkle (Vinca 
rosea) plant. It reversibly binds to microtubule and spindle proteins causing metaphase arrest. 
Vincristine has poor penetration into the CSF. It is approximately 75% protein bound.  
Extensive metabolism occurs in the liver. Excretion is primarily in the bile. A dosage decrease 
is recommended in patients with a bilirubin > 3 mg/dl.
Formulation and stability: Vincristine is supplied in multiple-dose 1 mg/ml vials containing 1 ml, 
2 ml and 5 ml. The intact vials should be stored under refrigeration and protected from light.
Supplier: Commercially available.
Toxicity: Dose limiting toxicity is neurotoxicity. This can be characterized by constipation 
and/or paralytic ileus, ptosis, vocal cord paralysis, weakness, jaw pain, abdominal pain, 
peripheral neuropathies, loss of deep tendon reflexes and “foot drop”. Peripheral neuropathy is 
often the first sign of neurotoxicity and is initially reversible. Other toxicities reported include 
alopecia, mild nausea and vomiting, SIADH, myelosuppression, orthostatic hypotension, optic 
atrophy, transient cortical blindness, and auditory damage. Acute shortness of breath and severe 
bronchospasm have been reported following the administration of vinca alkaloids.  
Myelosuppression is rare at usual doses. Vincristine is a vesicant and may cause severe tissue 
damage if extravasation occurs. NOTE: dose reduction may be necessary in patients < 1 year of 
age. Dosing on a per kg (rather than per m2) basis has been advocated for infants in order to 
decrease toxicity.
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6.0 
EVALUATIONS, TESTS, AND OBSERVATIONS
6.1
Suggested Evaluations Before and During Therapy – for Good Medical Care**
All entry/eligibility studies should be performed within 2 weeks prior to entry onto the trial (unless otherwise specified).  Imaging 
studies are required within 4 weeks prior to study entry.
Pre-
Treatmen
t
COP
Induction
Consolidation
Continuation
Off 
Therapy
Follow-Up
H & P
X
Weekly
Weekly
Prior to each Rx
X
See Section 
6.4
CBC & Diff
X
Weekly
Weekly or as 
indicated
Weekly or as indicated
X
See Section 
6.4
CMP, Mg++, PO4, LDH, uric 
acid
X
Daily
Weekly
Weekly
Weekly or as indicated
X
Hepatitis Screening (Section 
3.1.1) – required for patients 
receiving rituximab
X
PT/PTT, INR, fibrinogen, 
ESR
X
U/A
X
Daily
Prior to Rx
Prior to Rx
Prior to 
Cyclophosphamide
X
Bone marrow aspiration & 
biopsy (bilateral)
X
See below
CR Evaluation
See below
CR Evaluation
X
1 year from 
enrollment
CSF evaluation*
X
X
1 year from 
enrollment
CT – Neck, chest, abdomen, 
pelvis
X
CT of tumor on day 7 
COP
See below
CR Evaluation
Group A
See below
CR Evaluation
Groups B and C
As 
clinically 
indicated
As 
clinically 
indicated
FDG-PET 
X
Day 7 (research – See 
Table 6.2)
(to be done only in patients 
who do not require sedation)
See below
CR Evaluation
Group A
See below
CR Evaluation
Groups B and C
As 
clinically 
indicated
As 
clinically 
indicated
EKG, 2D-echocardiogram
X
Prior to each dose of 
Adriamycin, if 
clinically indicated
X
2 and 5 
years from 
enrollment
CMP-comprehensive metabolic panel (Na+, K+, Cl, CO2, Ca++, BUN, Cr, Blood sugar, total protein, albumin, Alk Phos),
Obtain other studies as needed for good patient care.
* Also with all IT Treatments; first CSF examination done with first intrathecal treatment after enrollment
**Participants of childbearing potential require a negative pregnancy test prior to starting treatment.
CR Evaluation:  Includes CSF exam (not required if no CNS involvement at diagnosis), bone marrow exam (not required if bone marrow negative at diagnosis) and imaging studies of primary tumor sites or as 
clinically indicated.  The time of evaluation varies with group:
Group A - after completion of COPAD #2, Group B - after completion of CYM #1, Group C - after completion of CYVE #2
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6.2
Suggested Evaluations for Research
The following research tests are optional, but strongly encouraged
Pre-
Treatment***
COP
Induction
Consolidation
Continuation
Off Therapy
Follow-Up
Plasma 1,25-dihydroxyvitamin D
X
X
25-hydroxyvitamin D, osteocalcin
X
X
Spot urine for Mg, creatinine, calcium, and 
calcium:creatinine ratio (obtain 24 hour if 
calcium:creatinine exceeds 0.2).
X
X
Fertility Studies (See Section 6.4.2.1)
X
X
2 and 5 years 
from 
enrollment
Serum Troponin T, Natriuretic hormone 
levels (NT-PRO BNP) (see Section 6.4.2.2)
X
X
2 and 5 years 
from 
enrollment
Serum for XLP studies**
X
Transcranial Doppler studies (TCD) ¥
X
X (Day 15 
COPADM)
X
1 year from 
enrollment
QCT- BMD¥
X
X
1 and 5 years 
from 
enrollment
PET-CT¥
(to be done only in patients who do not 
require sedation)
Day 7 - 
research
X
MRI Brain*¥
(to be done only in patients who do not 
require sedation)
X
X (Day 15 
COPADM #1)
See below
CR Evaluation
X
1 year from 
enrollment
Psychological Testing¥
X
1, 3 and 5 
years from 
enrollment
PET-CT on day 7 is research and should only be done in patients who do not require sedation. Other PET-CT scans performed as part of good medical care per table 6.1
CR Evaluation: Includes CSF exam, bone marrow exam and imaging studies of primary tumor sites or as clinically indicated. The time of evaluation varies with group:
Group A - after completion of COPAD #2; Group B - after completion of CYM #1; Group C - after completion of CYVE #2
*For Group B and C only
**XLP studies may be obtained after chemotherapy has started. The results of XLP studies will be reported to the treating physician for those patients enrolled outside of St. Jude. For those 
treated at St. Jude, the primary MD will discuss the results with the family and the patient will be referred for genetic counseling, if clinically indicated. 
***If research imaging and psych testing cannot be scheduled pre-treatment, can obtain as soon as possible after therapy starts.
¥Day 7 PET, psychological testing, Brain MRIs, QCT-BMD, transcranial Doppler studies, Troponin T levels, natriuretic hormone levels will not be done at Rady Children’s 
Hospital/UCSD.
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6.3
Advanced MR Imaging Methods
6.3.1
MR diffusion and perfusion imaging
In addition to the conventional MR neuroimaging prescribed for clinical evaluation, diffusion 
tensor imaging (DTI) will be acquired as a set of diffusion images in six or more directions. 
DTI will be acquired using bipolar diffusion-encoding gradients to reduce gradient-induced 
eddy currents that cause image distortion and degradation.112,113 All images will be acquired at 
either 1.5 or 3 T using a double spin echo EPI pulse sequence (TR/TE = 10/100 ms, b=1000 or 
700 ms). Imaging sets will be acquired as forty 3 mm thick contiguous sections with whole-
head coverage and a 128 square matrix. These data sets will be used to fully define a diffusion 
tensor for each point in the image114,115 Once the tensors have been calculated, Eigen values can 
be derived and used to calculate apparent diffusion coefficient (ADC) and fractional anisotropy 
(FA) maps for the whole brain. A commercial workstation and software will be used for post 
processing of this data as well as other custom analysis software developed at St. Jude. These 
measures are related to the integrity of the myelin sheath which restricts the diffusion of water 
perpendicular to the axons within the white matter fiber tracts.
Perfusion imaging will be evaluated with a dynamic-susceptibility contrast imaging. Imaging is 
acquired as fifteen 4 mm thick contiguous sections centered to ensure coverage across the 
entire corpus callosum and with a 256 square matrix to match the resolution of the conventional 
imaging. T2*-weighted images will be acquired with an axial echo-planer gradient-echo 
sequence (TR/TE = 2000/50 ms, 1 average). Fifty imaging sets with a temporal sampling of 
every 2 seconds will be acquired after the contrast injection. Standard bolus tracking of 0.1 
mmol/kg gadolinium followed by a 20 ml saline flush, both delivered at 1 ml/s will be 
performed. A commercial workstation and software will be used for post processing of this data 
as well as other custom analysis software developed at St. Jude.
6.3.2
Proton MR Spectroscopy
Proton MR spectroscopy will be performed using FDA approved commercially available data 
acquisition technology, provided by vendor (Siemens Medical Systems). Typically, 2D CSI 
(chemical shift imaging) technique will be used, depending on lesion extension within cerebral 
white matter, either single slice, or multislice. Occasionally, single voxel technique may be 
employed. Post processing of raw data will be done off line, using a multimodality workstation, 
provided by the same vendor using proprietary software. Individual, voxel-based MR spectra as 
well as metabolic maps of the entire region of interest for NAA (N-acetylaspartate), Cho 
(choline compounds), Cr (creatine) will be generated. Metabolic ratios (typically Cho/Naa) will 
be calculated based on relative metabolic concentrations.
6.4
Suggested Long-Term Follow-up Evaluations
6.4.1
Investigations at completion of therapy
At completion of therapy all initially positive sites should be investigated to confirm remission 
(radiological, bone marrow, and cerebrospinal fluid examinations).
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6.4.2
Suggested Follow-up and Late Effects Monitoring
Upon completion of therapy, it is suggested that participants be seen on a monthly basis (PE, 
CBC, chem panel including LDH) until the one year anniversary date from diagnosis. After one 
year from diagnosis, it is suggested that participants be seen at 3 monthly intervals (PE, CBC, 
chem panel including LDH) for the next year, and then annually (PE, CBC, chem panel 
including LDH). The late effects that may arise following this treatment include cardiotoxicity 
and infertility.
Suggested Fertility Studies
The following tests/evaluations are recommended to monitor for late effects on fertility at the 
following time points: Pre-treatment, off therapy, two years from enrollment, five years from 
enrollment, at Tanner III and V (if not already studied):
Males

Pubertal development evaluation (annually)

FSH, LH, and testosterone (annually)

Semen analysis (through Fertility Associates of Memphis)

Inhibin B level
Females

Pubertal development evaluation (annually)

FSH, LH, and Estradiol (annually)

Anti-mullerian hormone (AMH)
Suggested Cardiac Studies
The following tests/evaluations are recommended to monitor for cardiac late effects at the 
following time points:  pre-treatment, off therapy, two year from enrollment, and five years 
from enrollment:

2D-Echocardiogram

Troponin T levels (will not be done at UCSD)

Natriuretic hormone levels (NT-PRO BNP) (will not be done at UCSD).
6.5
Modifications for Collaborating Sites Participating in Therapeutic & Biological 
Objectives
The evaluations listed in Table 6.2 as indicated by “¥” will not be performed at Rady 
Children’s Hospital, San Diego.  This includes the PET scan on Day 7, Psychological testing 
(all evaluations), Brain MRI (all evaluations), pre-treatment bone density/BMD-QCT, 
transcranial Doppler studies, Troponin T levels, natriuretic hormone levels.
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7.0 
EVALUATION CRITERIA
7.1
Definition of Evaluable Disease
All abnormal sites at diagnosis should be evaluated using whatever modality is most appropriate. 
If bone marrow or CNS is involved, marrow and CSF should be examined.  The timing of 
investigations is described in Section 6.0.
7.2
Definition of Response
Complete response:  Complete disappearance of all measurable or evaluable lesions (except 
bone), no L3 blasts in the bone marrow or in the CSF.
Incomplete response:  20-99% reduction in the product of the two largest diameters 
(perpendicular) of measurable lesions and/or in the case of leukemia 20-99% reduction in the 
number of L3 blasts in the bone marrow and/or in the CSF.
Non-response:  < 20% tumor reduction of the product of the two largest diameters 
(perpendicular) of measurable lesions, or tumor progression, or tumor re-growth after initial 
shrinkage.
Relapse:  Defined as recurrence of disease at any site after achieving a CR.
Failure:  defined as relapses, deaths and failures to achieve a CR within the time frame 
described in the protocol.
7.3
Response Evaluations According to Treatment Group
7.3.1
Group A:  participants who are not in remission after two courses of COPAD 
(confirmed histologically) will advance to induction in Group C (COPADM8 Course 2). 
7.3.2
Group B:  Participants with < 20% tumor reduction after COP will move to Group C, 
starting at COPADM8 Course #1. Those remaining in Group B should be in complete remission 
after completing CYM Course #1. Residual masses are to be biopsied (surgical consultation is 
suggested). If there is histologic confirmation of viable tumor, the participant will be moved to 
Group C, starting with CYVE #1 as per Section 4.6.6. If the mass contains no histological 
evidence of viable tumor, the participant is considered to be in complete remission and will stay 
on treatment.
7.3.3
Group C: Participants should be in complete remission after completing CYVE Course 
#2. Residual masses are to be biopsied (surgical consultation is suggested). If the mass contains 
histologic evidence of viable tumor, the participant will be removed from protocol treatment. If 
the mass contains no viable tumor, the participant is considered to be in complete remission and 
stays on treatment.
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7.4
Toxicity Evaluation Criteria
Common Terminology Criteria for Adverse Events v4.0 (CTCAE): This study will utilize the 
CTCAE of the National Cancer Institute (NCI) for toxicity and performance reporting. A copy 
of the current version of the CTCAE can be downloaded from the Cancer Therapy Evaluation 
Program (CTEP) home page (http://ctep.info.nih.gov). Additionally, toxicities are to be 
reported on the appropriate data collection screens/forms.
7.5
Acceptable Percentage of Missed Doses for Commercially Available Drugs
It is understood that deviations from the protocol should be avoided, except when necessary to 
eliminate an immediate hazard to a research participant. In such case, the deviation must be 
reported to the IRB. However, it is expected that participants will occasionally miss some doses 
or receive the wrong dose of oral chemotherapy. Compliance with oral medication will be 
captured in the CRIS database and appropriately documented in the participants’ medical 
records. Appropriately documented doses of missed or wrong doses of chemotherapy will not 
constitute a deviation unless the amount in question is over 10% of the expected total dose due 
in the respective protocol cycles (these are specified in the CRIS SJBC3 database). Missed 
doses do not include doses held or reduced for medical reasons (toxicity, illness) and will not 
be considered protocol deviations or violations.
8.0
OFF PROTOCOL TREATMENT AND OFF-STUDY CRITERIA
8.1
Off Treatment Criteria
A patient will be taken off SJBC3 treatment if any of the following occurs:

Failure to achieve a complete response to induction therapy:
-
Group A:  failure to achieve CR after CYVE #2
-
Group C:  failure to achieve CR after CYVE #2

Bone marrow or extramedullary relapse

Second malignancy (e.g., therapy-induced AML or brain tumor)

Development of unacceptable toxicity during treatment (with concurrence of the PI or 
co-PI)

Patients with severe congenital immunodeficiency (e.g. ataxia telangiectasia) or HIV 
infection will be taken off SJBC3 therapy and will receive alternate therapy (to prevent 
excessive toxicities) with curative intent.

Completion of all protocol-prescribed treatment

Participant/family decision to withdraw from protocol treatment at any time for any 
reason

Investigator decides continued protocol treatment is no longer in patient’s best interest
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8.2
Off Study Criteria
A patient is taken off SJBC3 study if any of the following occurs:

Death

Lost to follow-up

Withdrawal of consent for continued follow-up

Found to be ineligible (e.g., incorrect diagnosis)
Follow up will stop at the time the patient is off study. The follow up time of an off-study 
patient is censored at this point, and no outcome data beyond the off-study time will be used in 
analyses.
9.0
SAFETY AND ADVERSE EVENT REPORTING REQUIREMENTS
9.1
Reporting Adverse Experiences (AEs) and Deaths to the St. Jude IRB 
Only “unanticipated problems involving risks to participants or others” referred to hereafter as 
“unanticipated problems” are required to be reported to the St. Jude IRB promptly, but in no 
event later than 10 working days after the investigator first learns of the unanticipated problem. 
Regardless of whether the event is internal or external (for example, an IND safety report by 
the sponsor pursuant to 21 CFR 312.32), only adverse events that constitute unanticipated 
problems are reportable to the St. Jude IRB.  
As further described in the definition of unanticipated problem, this includes any event that in 
the PI’s opinion was:

Unexpected (in terms of nature, severity, or frequency) given (1) the research 
procedures that are described in the protocol-related documents, such as the IRB-
approved research protocol and informed consent document, as well as other relevant 
information available about the research; (2) the observed rate of occurrence (compared 
to a credible baseline for comparison); and (3) the characteristics of the subject 
population being studied; and

Related or possibly related to participation in the research; and

Serious; or if not serious suggests that the research places subjects or others at a greater 
risk of harm (including physical, psychological, economic, or social harm) than was 
previously known or recognized. 
Deaths that occur > 30 days after protocol treatment that expected and unrelated to protocol 
therapy do not require reporting to the IRB. Though death is “serious”, the event must meet the 
other two requirements of “related or possibly related” and “unexpected/unanticipated” to be 
considered reportable.
Deaths meeting reporting requirements are to be reported immediately to the St. Jude IRB, but 
in no event later than 48 hours after the investigator first learns of the death; this includes any 
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death that occurs while the participant is on treatment or that occurs < 30 days of last protocol 
treatment, regardless of cause.
The following definitions apply with respect to reporting adverse experiences: 
Serious Adverse Event: Any adverse event temporally associated with the subject’s 
participation in research that meets any of the following criteria:

results in death;

is life-threatening (places the subject at immediate risk of death from the event as it 
occurred);

requires inpatient hospitalization or prolongation of existing hospitalization;

results in a persistent or significant disability/incapacity;

results in a congenital anomaly/birth defect; or

any other adverse event that, based upon appropriate medical judgment, may jeopardize 
the subject’s health and may require medical or surgical intervention to prevent one of 
the other outcomes listed in this definition (examples of such events include:  any 
substantial disruption of the ability to conduct normal life functions, allergic 
bronchospasm requiring intensive treatment in the emergency room or at home, blood 
dyscrasias or convulsions that do not result in inpatient hospitalization, or the 
development of drug dependency or drug abuse), a congenital anomaly/birth defect, 
secondary or concurrent cancer, medication overdose, or is any medical event which 
requires treatment to prevent any of the medical outcomes previously listed.  
Unexpected Adverse Event: 
Any adverse event for which the specificity or severity is not consistent with the 
protocol-related documents, including the applicable investigator brochure, IRB 
approved consent form, Investigational New Drug (IND) or Investigational Device 
Exemption (IDE) application, or other relevant sources of information, such as product 
labeling and package inserts; or if it does appear in such documents, an event in which 
the specificity, severity or duration is not consistent with the risk information included 
therein; or 
The observed rate of occurrence is a clinically significant increase in the expected rate 
(based on a credible baseline rate for comparison); or 
The occurrence is not consistent with the expected natural progression of any 
underlying disease, disorder, or condition of the subject(s) experiencing the adverse 
event and the subject’s predisposing risk factor profile for the adverse event. 
Internal Events: Events experienced by a research participant enrolled at a site under the 
jurisdiction of St. Jude IRB for either multicenter or single-center research projects.
External Events: Events experienced by participants enrolled at a site external to the 
jurisdiction of the St. Jude Institutional Review Board (IRB) or in a study for which St. Jude is 
not the coordinating center or the IRB of record.
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Unanticipated Problem Involving Risks to Subjects or Others: An unanticipated problem 
involving risks to subjects or others is an event which was not expected to occur, and which 
increases the degree of risk posed to research participants.  
Such events, in general, meet all of the following criteria: 

unexpected; 

related or possibly related to participation in the research; and 

suggests that the research places subjects or others at a greater risk of harm (including 
physical, psychological, economic, or social harm) than was previously known or 
recognized.  An unanticipated problem involving risk to subjects or others may exist 
even when actual harm does not occur to any participant. 
Consistent with FDA and OHRP guidance on reporting unanticipated problems and adverse 
events to IRBs, the St. Jude IRB does not require the submission of external events, for 
example IND safety reports, nor is a summary of such events/reports required; however, if an 
event giving rise to an IND safety or other external event report constitutes an “unanticipated 
problem involving risks to subjects or others” it must be reported in accordance with this 
policy.  In general, to be reportable external events need to have implications for the conduct of 
the study (for example, requiring a significant and usually safety-related change in the protocol 
and/or informed consent form).
Although some adverse events will qualify as unanticipated problems involving risks to 
subjects or others, some will not; and there may be other unanticipated problems that go 
beyond the definitions of serious and/or unexpected adverse events.  Examples of unanticipated 
problems involving risks to subjects or others include:

Improperly staging a participant’s tumor resulting in the participant being assigned to an 
incorrect arm of the research study;

The theft of a research computer containing confidential subject information (breach of 
confidentiality); and 

The contamination of a study drug.  Unanticipated problems generally will warrant 
consideration of substantive changes in the research protocol or informed consent 
process/document or other corrective actions in order to protect the safety, welfare, or 
rights of subjects or others. 
9.2
Recording AEs and SAEs
Adverse events (AEs) will be evaluated and documented by the clinical staff and investigators 
throughout inpatient hospitalizations and each outpatient visit.  CRAs are responsible for 
reviewing documentation related to AEs and entering directly into CRIS protocol-specific 
database.  The data to be recorded are 1) the event description, 2) the NCI CTCAE v4.0 code 
and grade, 3) the onset date, 4) the resolution date (or ongoing), 4) action taken for event, 5) 
patient outcome 6) relationship of AE to protocol treatment/interventions, 7) if AE was 
expected or unexpected, and 8) comments, if applicable.  AEs that are classified as serious, 
unexpected, and at least possibly related will be notated as such in the database as “SAEs”.  
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These events will be reported expeditiously to the St. Jude IRB within the timeframes as 
described above.  
Cumulative summary of Grade 3-5 events will be reported as part of the progress reports to 
IRB at the time of continuing review.  Specific data entry instructions for AEs and other 
protocol-related data will be documented in protocol-specific data entry guidelines, which will 
be developed and maintained by study team and clinical research informatics.
The study team will meet regularly to discuss AEs (and other study progress as required by 
institutional DSMP).  The PI will review Adverse Event reports generated from the research 
database, and corrections will be made if applicable.  Once the information is final the PI will 
sign and date reports, to acknowledge his/her review and approval of the AE as entered in the 
research database. 
9.3
Reporting AEs to and from St. Jude and Collaborating Sites/Affiliates
Adverse events from collaborating sites will also be reviewed by the PI and discussed in study 
team meetings as described above.  SAE report from collaborating sites for AEs that are 
serious, unexpected, and at least possibly related to protocol treatment or interventions will be 
reported to site IRB and the St. Jude IRB within the reporting requirements described above.  
The PI will determine if this is an event that will need to be reported expeditiously to all 
participating sites, considering the following criteria:

Is the AE serious, unexpected, and related or possibly related to participation in the 
research?

Is the AE expected, but occurring at a significantly higher frequency or severity than 
expected?

Is this an AE that is unexpected (regardless of severity that may alter the IRB’s analysis 
of the risk versus potential benefit of the research and, as a result, warrant consideration 
of substantive changes in the research protocol or informed consent process/document?
With the submission of the “Reportable Event” in St. Jude TRACKS application, the PI will 
indicate if all sites should be notified to report to their IRBs, and if the protocol and/or consent 
should be amended (consent will be amended if event is information that should be 
communicated to currently enrolled subjects). Generally, only events that warrant an 
amendment to the protocol and/or consent will be reported expeditiously to all sites. However, 
any event may be reported expeditiously to all sites at the discretion of the PI. 
A cumulative summary of Grade 3-5 AEs and expected/unrelated deaths that occur more than 
30 days off last protocol treatment will be reported to all sites with study progress report at the 
time of continuing review. 
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10.0
DATA COLLECTION, STUDY MONITORING, AND CONFIDENTIALITY
10.1
Data Collection
Electronic case report forms (e-CRFs) will be completed by the St. Jude Leukemia/Lymphoma 
CRAs. Data will be entered from record directly into a secure CRIS database, developed and 
maintained by St. Jude Clinical Research Informatics. 
Data Management will be supervised by the Director of Clinical Trials Management, and 
Manager of Clinical Research Operations for the Leukemia/Lymphoma Division, working with 
Dr. Ribeiro or his designee. All protocol-specific data and all grade 3-5 adverse events will be 
recorded by the clinical research associates into the CRIS database, ideally within 2-4 weeks of 
completion of study phase. All questions will be directed to the attending physician and/or PI 
and reviewed at regularly-scheduled working meetings. The attending physicians (or their 
designees) are responsible for keeping up-to-date roadmaps in the patient’s primary St. Jude 
medical chart.
Regular (at least monthly) summaries of toxicity and protocol events will be generated for the PI 
and the department of Biostatistics to review.
10.2
Data Collection Instructions for Collaborating Sites
Collaborating sites will collect abstract data from medical records and submit by using e-CRFs 
via remote electronic data entry. All protocol-specific data and all grade 3-5 adverse events will 
be recorded by the clinical research associates into the CRIS database, ideally within 2-4 weeks 
of completion of study phase.
10.3
Study Monitoring
Monitoring of this protocol is considered to be in the moderate risk category, as per the St. 
Jude Data Safety and Monitoring Plan (DSMP).
The study team will hold monthly meetings and review case histories or quality summaries on 
participants.
Source document verification of eligibility and informed consent for 50% of St. Jude 
participants will be performed by the Eligibility Coordinators within 5 working days of 
completion of enrollment.
The Clinical Research Monitor will perform monitoring of applicable essential regulatory 
documentation. Also, reviewing for the timeliness of serious adverse event reporting (type, 
grade, attribution, duration, timeliness and appropriateness) for selected study participants 
semi-annually and track accrual continuously. The monitor will verify those data points relating 
to the primary study objective for a certain number of study enrollees as specified in the 
Moderate Risk monitoring plan checklist for this study. Protocol compliance monitoring will 
include participant status, safety assessments, eligibility, the informed consent process, 
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participant protocol status, off-study, and off-therapy criteria. The Monitor will generate a 
formal report which is shared with the Principal Investigator (PI), study team and the Internal 
Monitoring Committee (IMC).
Monitoring may be conducted more frequently if deemed necessary by the CTO or IMC. 
Continuing reviews by the IRB and CT-SRC will occur at least annually. In addition, SAE 
reports in TRACKS (Total Research and Knowledge System) are reviewed in a timely manner 
by the IRB/OHSP.
This will include verification of appropriate documentation of consent. Monitoring of 
timeliness of serious adverse event reporting will be done as events are reported in TRACKS. 
St. Jude affiliates and domestic collaborating study sites will be monitored on-site by a 
representative of St. Jude at intervals specified in the DSMP. 
10.4
Confidentiality
Study numbers will be used in place of an identifier such as a medical record number. No 
research participant names will be recorded on the data collection forms. The list containing the 
study number and the medical record number will be maintained in a locked file and will be 
destroyed after all data have been analyzed. The medical records of study participants may be 
reviewed by the St. Jude IRB, FDA, and St. Jude clinical research monitors.
11.0
STATISTICAL CONSIDERATIONS
11.1
Biological Aims
The SJBC3 trial has several exploratory biological studies as its primary objectives. Statistical 
considerations on accrual, sample size, and levels of statistical errors for the first aim are given 
below.
To perform transcriptional profiling and genome-wide analysis of DNA copy number 
abnormalities and loss-of-heterozygosity using DNA microarrays in children with newly 
diagnosed mature B cell lymphomas (e.g. Burkitt lymphoma/leukemia, DLBCL, and MLBCL) 
from different parts of the world.
Accrual. It is difficult to get a precise projection of accrual rate at this point although we have 
some expectation based on prior experience. Below we give a conservative projection for the 
biological study in terms of number of evaluable tumor samples after 5 year accrual although 
we anticipate the trial may run beyond 5 years. Experience from the SJBCII trial indicates that 
the annually 6 patients can be accrued at St. Jude, giving approximately 30 patients after 5 
years of accrual at this institution. Because there is not another childhood B-cell lymphoma 
trial currently in US, we anticipate the accrual may increase to 8 to 12 patients per year. The 
genomic analysis requires tumor biopsy samples from participating international institutions 
along with St. Jude. The international institutions will participate in the biological studies and 
provide biopsy samples but will not participate in the SJBC3 therapy. We also note that it is 
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rather difficult to obtain tumor biopsy samples from patients enrolled here for various logistical 
reasons. We expect that there will be about 10, 1 to 2, and 5 per year from Latin America, 
North America (St. Jude), and other geographic regions respectively. Five year accrual can 
provide about 50, 5 to 10, and 25 tumor samples respectively. Approximately 80% of the 
tumors are expected to be Burkitt’s lymphoma (BL). Moreover, we conservatively assume that 
80% of the samples from each region are eventually evaluable for genomic analyses, resulting 
in 40, 8 and 20 evaluable tumor samples from Latin America, North America, and other regions 
respectively. 
Five year accrual of evaluable tumor samples from each region for this aim is summarized 
below. 
Number of evaluable tumor samples after 5-year accrual
BL
Non-BL
Total
Latin America
32
8
40
North America
6
2
8
Other
16
4
20
Total
54
14
68
At this point we set the accrual target for the biological aims as 68 evaluable tumor samples and 
anticipate completing accrual in 5 to 6 years for the biological aims. We set the accrual target 
for other aims to 60 patients treated at St. Jude and anticipate completion in 5 to 6 years. We 
will amend the protocol to extend the enrollment time if the targets cannot be met in 5 years.
Amendment 3.0 update:
We recognize that we need an extension in duration of study to complete the biology 
(genomics) objectives. We anticipate that the study will need to remain open for an additional 5 
years. We estimate that we will accrue an additional 40 participants on the therapeutic and 
biology part of the study during that time, for a total enrollment of 168 patients (100 
therapeutic and biology, and 68 biology only).
Increasing treatment enrollment/accrual for the “Therapeutic and Biology” component is 
justified by (1) providing potentially more tumor samples from North America, and (2) 
strengthening the analyses of the exploratory objectives, especially the very promising 
preliminary results of the neuroimaging studies (abstract included in earlier continuing review 
report).
Analyses. There will be three major components in the analyses for this objective. Each is 
discussed below, with very limited preliminary genomic data in pediatric lymphomas. 
Gene differential profiling of BL vs. non-BL. Identification of gene differential expression 
signatures requires inference (comparison) of gene expression levels in BL vs. non-BL. It is 
important to realize that the tumor samples come from dramatically different geographical 
regions where ethnic and environmental factors (e.g., diet) can greatly affect gene expression. 
Additionally, the extent of differential expression between BL and non-BL may also be 
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affected by environmental, endemic and related biological factors (e.g., involvement of EBV in 
BL). Thus, it is important to adjust the gene expression comparison by geographic region, 
which serves as an aggregate of all the possible region-related confounding and stratification 
factors, in the absence of the data on such factors. Each participating institute defines a 
geographic region; from example, St. Jude defines the “North America” region. At this point 
we expect in addition to St. Jude, there will be one institute in Brazil (Latin America) and one 
in Singapore (Asia) participating the biological studies. Modeling, testing, and interpretation 
for a single gene (probeset) is given below, followed by methods of handling massive multiple 
tests. For samples with suitable RNA quality, gene expression profiling will be performed 
using transcriptome sequencing in preference to microarray analysis, in view of the more 
complete data provided by this platform.
The following two-factor Poisson regression model will be used for each gene, where 
E[Y(bgi)] denotes the mean expression of a gene in the i-th tumor sample from region g 
(g=Latin America, North America, etc.) and disease b (b=BL, non-BL):
Log{E[Y(bgi)]}=μ+α(b)+β(g)+αβ(g,b).
From a completely biological stand point the primary interest here is the main effect α(b). For 
each mRNA expression we will test the null hypothesis α(BL)= α(non-BL)=0 against the two-
sided alternative at least one of them is non-zero. Statistical significance of this effect indicates 
that it is likely the gene is differentially expressed between BL and non-BL. From a molecular 
epidemiology (and to a lesser extent biological) stand point the disease-region interaction 
effect αβ(g,b) is important, as it signifies that the extent of differential expression between BL 
and non-BL may be affected by geographical region. Statistical significance of this interaction 
effect indicates that it is likely the differential expression between BL and non-BL is different 
from one region to another; this would be an epidemiologically and biologically interesting 
finding as it helps to define the RNA-level differences between endemic and non-endemic 
Burkitt lymphoma. 
Hence, we will be testing the main effect α(b) and the interaction effect for each RNA  
expression measurement. Because EBV positivity in BL may have an important effect on gene 
differential expression, in the final analysis we will also consider defining the disease main 
effect α(b) as a three-level factor: b = EBV+ BL, EBV- BL, non-BL.
It is imperative to assess the levels of statistical errors, especially false positives, made in these 
massive multiple tests. An assessment of the level of false negative errors with the available 
sample size will be given in the next paragraph. Following the published microarray studies 
many of which from St. Jude, we will first assess the ensemble distribution of the P values and 
use the profile information criterion116  to obtain a data-suggested level of statistical 
significance and the corresponding estimated false discovery rate (FDR),116,117 and then 
estimate the false discovery rate corresponding to a set of statistical significance levels 
specified around the data-suggested level. Estimates of FDR will be computed using methods 
in the literature.116,118,119 We will analyze further the “significant genes” so identified by 
modeling there expressions with disease subtype as well as potential confounding factors as 
explanatory variables.
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Recognizing the fast changes in biotechnology, we’ll use the most effective technology and 
appropriate statistical methods at the time of final analyses.
Levels of statistical errors – an assessment by published data. All of the published gene 
expression profiling studies of mature B-cell lymphomas focused on adults, but Hummel et al. 
(2006)120 included a number of pediatric samples (age≤18 years). None of the published 
studies considered geographic region as a factor and thus there is no preliminary data available. 
Therefore the preliminary microarray gene expression data on pediatric mature B-cell 
lymphomas are very limited. Nonetheless we provide a rough assessment of the levels of false 
positive and false negative errors that we may encounter in the analysis based on the pediatric 
data120 followed by a discussion of caveats and limitations of this assessment.
Here we consider the tests of the main effect α(b) for the m (≈ 22000) Affymetrix GeneChip 
probesets.  Adapting the concept of anticipated FDR (aFDR)121 to the fixed sample size 
situation and the planned analysis outlined above, we define the aFDR at a given significance 
level α as
.

AP
aFDR
)
1(
0
0
0









Here π0 is the proportion of true null hypotheses (non-differentially expressed genes), and AP is 
the average power which depends on the effect size, sample size, and significance level. The 
quantity (1- π0)(1-AP) is the expected proportion of true alternative hypotheses the multiple 
tests fail to capture, hence a measure of the level of false negative errors; we call this the false 
negative proportion.116
Hummel et al. (2006) included 32 pediatric samples run on Affymetrix U133A GeneChip.120 In 
these pediatric lymphoma samples 6 are traditional BL, 10 are atypical BL, 12 are DLBCL, and 
4 are aggressive B-NHL. The 32 samples are put into two groups: BL (traditional and atypical 
BL) and non-BL (DLBCL and aggressive B-NHL). To get some preliminary information about 
π0, per-probeset comparisons of BL vs. non-BL by the two-sample t test yield 22,215 P values, 
one for each probeset. The profile information criterion (Ip)116 produces a significance 
threshold (P value cut-off) to declare the significant tests in a way to balance the levels of false 
positive and false negative errors, and a conservative (biased up) estimate of the corresponding 
FDR; it also provides an estimate of π0 as part of the procedure. Applying this procedure to the 
22,215 P values yields an estimate of π0 0.6525 (65.25%), the significance threshold 0.004547, 
and the corresponding FDR 0.1377 (13.77%) among the 478 probesets declared differentially 
expressed at the 0.004547 significance level. For each probeset, the mean in the BL and non-
BL group along with the within-group variance are computed to provide an estimate of the non-
centrality parameter of the t test, and then is shrunk toward zero by the q-value weighting as 
described.121 The probesets are then sorted by the magnitude of the shrunken non-centrality 
parameter in descending order. To be on the conservative side, for each postulated null 
proportion (π0) in the table below, the non-centrality parameters of the top m(1-π0) probesets 
are all set to the same as the [m(1-π0)]th largest one, and the remaining non-centrality 
parameters are set to zero. Next, average power (AP) and π0 is estimated after this additional 
shrinkage, giving enlarged π0.
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Based on the π0 estimate and the significance threshold suggested by Ip, we provide the final 
non-centrality after shrinkage, estimates of aFDR, the false negative proportion FNP, and 
average power (AP)  for a range of postulated and enlarged π0 values and significance levels (P 
value cutoffs) in the following table.
Anticipated False Discovery Rate and False Negative Proportion
π0
Postulated
Enlarged
Non-centrality
0.60
0.7969
3.39
0.65
0.7994
4.17
0.80
0.8469
7.57
0.90
0.9117
11.82
0.95
0.9532
15.81
0.99
0.9902
24.38
0.0010   aFDR
              FNP
              AP
0.1107
0.3874
0.0315
0.0823
0.3344
0.0443
0.0388
0.1726
0.1371
0.0306
0.0673
0.3268
0.0372
0.0236
0.5274
0.1116
0.0019
0.8079
0.0025
0.1471
0.3773
0.0568
0.1141
0.3229
0.0773
0.0621
0.1583
0.2086
0.0599
0.0564
0.4358
0.0737
0.0180
0.6405
0.2241
0.0012
0.8783
0.0045
0.1775
0.3673
0.0818
0.1417
0.3120
0.1086
0.0850
0.1464
0.2680
0.0830
0.0487
0.5134
0.1142
0.0144
0.7114
0.3330
0.0009
0.9144
0.0050
0.1836
0.3651
0.0872
0.1473
0.3096
0.1153
0.0899
0.1440
0.2798
0.0891
0.0472
0.5278
0.1235
0.0138
0.7237
0.3554
0.0008
0.9201
0.0075
0.2093
0.3555
0.1111
0.1715
0.2994
0.1444
0.1120
0.1342
0.3287
0.1171
0.0416
0.5838
0.1658
0.0115
0.7696
0.4474
0.0006
0.9396
α
0.0100
0.2298
0.3474
0.1315
0.1911
0.2909
0.1687
0.1311
0.1267
0.3667
0.1419
0.0376
0.6239
0.2030
0.0100
0.8003
0.5160
0.0005
0.9514
For example, if the proportion of non-differentially expressed probesets π0=0.65 (65%) and all 
probesets with P value ≤ α=0.0045 are declared as significantly differentially expressed 
between BL and non-BL, we anticipate that the false discovery rate aDFR=0.1417, i.e., 14.17% 
of these “significant” probesets are false positives; and that the false negative proportion 
FNP=0.3120, i.e., we expect to miss 31.2% of the truly differentially expressed probesets. 
It should be noted that the non-centrality parameters estimated above in fact correspond to one-
factor ANOVA. Unfortunately however, there is no preliminary data on the second factor 
(geographic region). The numerator of the non-centrality parameter determined from the one-
factor model could be too optimistic for the two-factor mode, but the denominator (the within-
group/residual standard deviation) determined by the one-factor model may be larger than that 
from the two-factor model.
The total number of tests performed for this analysis will be over 44,000 although the above 
error assessment was done for 22,000+ tests. Note however, aFDR and FNR do not depend 
directly on the total number of tests; rather they depend directly on the proportion of true null 
hypothesis π0.
It should be noted that there will be very limited statistical power for gene expression profiling 
for BL vs. non-BL in a single geographic region, except perhaps in Latin America where the 
sample size is the largest among all regions.
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Additionally, unsupervised analyses such as hierarchical clustering will be performed to 
explore novel subtypes in childhood lymphomas. Also, identified gene differential signatures 
will be compared with published results (in adult lymphomas) using methods such as gene set 
enrichment analysis.
Catalog and estimate frequencies of copy number variations in childhood lymphomas. 
There is no preliminary data on copy number variations (CNVs) in childhood lymphomas. A 
study of DLBCL in adults10  reported the prevalence of CNVs in certain subsets ranged roughly 
from 4% to 40%. So, conservatively, with 14 samples in the non-BL group, if a CNV is 
observed in 1 sample (sample proportion 0.07), then the exact 95% confidence interval is 
[0.0037, 0.3122]; if a CNV is observed in 6 samples (sample proportion 0.43), the exact 95% 
confidence interval is [0.2061, 0.6878]. Additionally, we will compare the prevalence of CNVs 
between different subtypes of childhood lymphomas and geographic regions with exact chi-
square or Fisher’s test, taking population structures into account if necessary. DNA extracted 
from peripheral blood will be used as matched control tissue. Loss of heterozygosity (LOH) 
will be analyzed by comparison of SNP genotype data of the tumor sample to matched 
germline samples, or, in the case of samples lacking a matched germline sample, pools of 
reference samples already accrued at St Jude. LOH data will be analyzed using publicly 
available algorithms (e.g. dChip using the Hidden Markov Model) and alternative algorithms 
that will be tested and developed at St Jude during the course of the study. These analyses will 
complement analyses identifying copy number alteration, and will permit identification of both 
deletional and copy-neutral loss-of-heterozygosity.
Integrated analysis of CNVs and gene expressions. For CNVs with a reasonable prevalence, 
we will explore the association between the identified CNVs and gene expressions in the study 
cohort.  We will identify individual gene expression correlated with a CNV, and a gene 
expression profile (signature) associated with a CNV. General linear models will be used to 
perform statistical tests, and multiple tests will be addressed using FDR, as described before.
To describe the types and frequency of mutations in the ARF-HDM2-TP53 pathway, in “non-
endemic” B-cell lymphomas (United States) and those found in selected geographic regions of 
the world (Brazil, Chile, Guatemala, Honduras, El-Salvador, Singapore, and others) 
This exploratory aim will be analyzed by enumerating the types of mutations and estimating the 
frequency of each type of mutation in each geographical region as well as in all regions pooled. 
Point estimates and 95% confidence intervals (CI) will be provided.
To describe the expression of ARF-HDM2-TP53 and PUMA-associated pathways in non-
endemic B-cell lymphomas (United States) and that found in B-cell lymphomas of other 
selected geographic regions of the world (see above)
Protein as well as mRNA expression levels of important genes on these pathways from various 
bioassays will be summarized using descriptive statistics including mean, standard deviation, 
and the five-number summary in each geographical region as well as in all regions pooled.
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To describe the pattern and frequency of XLP gene mutations presenting with B-cell 
lymphomas in the United States and selected geographic regions.
Point estimates and 95% CIs of the frequency of XLP mutation among boys will be calculated 
in each geographical region as well as in all regions pooled. Relationships of XLP mutations 
with clinical phenotypes will be explored by appropriate statistical models including 
contingency table analysis and/or logistic regression modeling (for categorical/binary 
phenotypes), general linear models for continuous phenotypes, and hazard regression models 
for right-censored failure times.
To describe the frequency of EBV-positive Burkitt lymphomas between “non-endemic” and 
endemic B-cell lymphomas;
a. To describe the pattern of EBV protein and gene expression (e.g., EBNA 3) in EBV-
positive lymphomas.
b. To relate EBV protein and gene expression with clinical, laboratory and outcome data.
It is expected that among BL, about 80% are EBV positive in endemic BL (Latin America, 
Africa) and 45% are EBV positive in non-endemic BL (US, Mideast), and about 80% of the 
available tumor samples are BL. Point estimates and 95% CIs of the frequency of EBV-positive 
BL will be calculated for each geographical region. EBV protein expression will be described 
by summary statistics including mean, standard deviation, and the five-number summary in 
each geographical region as well as in all regions pooled. Relationship of EBV protein 
expression with clinical, laboratory, and outcome features will be explored using appropriate 
statistical methods such as general linear models.
11.2
Exploratory Aims
Acute and Late Effects Objectives
These objectives are completely exploratory. Only patients treated at St. Jude will be asked to 
participate in these studies. As stated earlier, we anticipate the accrual rate of 8-12 patients per 
year and expect to accrual approximately 100 patients in 12 years. We also note that although 
we planned 6 year accrual at St. Jude for this and the outcome aim, similar to SJBCII, the 
SJBC3 trial will run beyond five years in this institution. SJBCII opened in September 1994 
with the first and last patient enrolled on 12/6/1994 and 9/23/2005 respectively. Descriptive 
statistics and appropriate statistical models (regression, logistic regression, etc.) will be applied 
to analyze the data obtained for these aims.
To estimate the complete response, event-free survival, and overall survival rates in patients 
with Burkitt lymphoma (BL), Burkitt leukemia/B-cell acute lymphoblastic leukemia (B-ALL) 
and diffuse large B-cell lymphoma (DLBCL) treated with a stage-adapted regimen based on 
the SJBCII protocol.
All eligible patients treated at St. Jude will be included in the analysis. SJBCII experience 
shows the accrual rate at St. Jude is about 6 per year; thus approximately 30 patients after 5 
years. It should be noted that although we plan 6 year accrual at St. Jude, similar to SJBCII, the 
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SJBC3 trial may run beyond 6 years in this institution. SJBCII opened in September 1994 with 
the first and last patient enrolled on 12/6/1994 and 9/23/2005 respectively. 
Complete response (CR), event-free survival (EFS), and overall survival (OS) will be analyzed 
only for eligible patients treated at St. Jude. Point estimate and exact 95% CI based on the 
binomial distribution will be calculated for the rate (probability) of CR. For EFS, failures will 
include (1) death while in continuous CR, (2) relapse, (3) secondary malignancy, and (4) failure 
to achieve CR. Failure time is defined to be the time between on-study and the time of failure 
or the last follow-up time for patients who had not failed at the last follow-up time. The failure 
time of patients failing to achieve CR is set to zero. The survival distribution of EFS and OS 
will be estimated by the Kaplan-Meier estimator, and the standard deviation will be calculated 
by Peto’s method. CR rate, EFS and OS of patients treated on SJBC3 at St. Jude will be 
compared historically with those treated on SJBCII. This comparison defines the continued 
improvement in the treatment of childhood mature B-cell lymphomas and leukemia in this 
institution. 
Of the 64 patients on the SJBCII trial there are 3 patients in Group A, 37 in Group B, and 24 in 
Group C. Two (2/64=3.1%) patients failed to achieve CR. As of May 2007 the median follow-
up time was 6.07 years (range 1.05 to 12.07 years). Ninety five percent (95%) of the patients 
were contacted within a year. 
The overall five-year EFS and OS were 90.1% ± 4.9% and 93.6% ± 3.7% respectively (see also 
figure below).
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Five-year EFS and OS in each treatment group in the SJBCII trial are summarized below.
Five-year EFS and OS by Group on SJBCII
Group
5-Y EFS
5-Y OS
A
100%
100%
B
96.4%±3.7%
100%
C
79.2%±1.2%
82.9%±9.9%
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To determine the feasibility of administration and toxicity profile of the SJBC3 protocol, which 
features the use of rituximab for selected higher-risk Group B and all Group C participants.
Frequencies of possible rituximab-related toxicities will be described in terms of the estimates 
of the number of episodes and the cumulative incidence throughout the treatment period for 
each type of toxicity.
12.0 
OBTAINING INFORMED CONSENT
12.1
Consent/Assent at Enrollment
The process of informed consent for SJBC3 will follow institutional policy. The informed 
consent process is an ongoing one that begins at the time of diagnosis and ends after the 
completion of therapy. Informed consent should be obtained by the attending physician or 
his/her designee, in the presence of at least one non-physician witness. Initially, informed 
consent will be sought for the institutional banking protocol (research study), blood transfusion 
and other procedures as necessary. After the diagnosis of mature B-cell NHL or leukemia is 
established, we will invite the patient to participate in the SJBC3 protocol.  
Throughout the entire treatment period, participants and their parents receive constant 
education from health professionals at St. Jude and collaborating sites, and are encouraged to 
ask questions regarding alternatives and therapy. All families have ready access to chaplains, 
psychologists, social workers, and the St. Jude ombudsperson for support, in addition to that 
provided by the primary physician and other clinicians involved in their care.
We will also obtain verbal assent from children 7 to 14 years old and written assent for all 
participants >14 years of age. Participants who reach the age of majority while on study will be 
re-consented for continued participation on SJBC3, according to Cancer Center and 
institutional policy.
12.2
Consent at Age of Majority
The age of majority in the state of Tennessee is 18 years old. Research participants must be 
consented at the next clinic visit after their 18th birthday. If an affiliate or collaborating site is 
located in a country or state where a different age of majority applies, that location must 
consent the participants according to their local laws.  
12.3
Consent When English is Not the Primary Language
When English is not the patient, parent, or legally authorized representative’s primary 
language, the Social Work department will determine the need for an interpreter. This 
information documented in the participant’s medical record. Either a certified interpreter or the 
telephone interpreter’s service will be used to translate the consent information. The process for 
obtaining an interpreter and for the appropriate use of an interpreter is outlined on the 
Interpreter Services, OHSP, and CTO websites.
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Collaborating Sites will follow institutional policy for consenting non-English speaking 
participants (and will provide institutional policy to St. Jude).
12.4
Collection of Collaborating Institution Consent Forms
Signed collaborating institution’s consent forms will be faxed/scanned to 
 
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13.0
REFERENCES
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composition, and height in long-term survivors of acute lymphoblastic leukemia in childhood. 
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leukemia. J Clin Oncol 19:3066-72, 2001
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survivors of childhood acute lymphoblastic leukemia. J Pediatr Hematol Oncol 20:234-40, 1998
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Arikoski P, Voutilainen R, Kroger H: Bone mineral density in long-term survivors of childhood 
cancer. J Pediatr Endocrinol Metab 16 Suppl 2:343-53, 2003
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Lukert BP, Raisz LG: Glucocorticoid-induced osteoporosis. Rheum Dis Clin North Am 20:629-
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Joseph JC: Corticosteroid-induced osteoporosis. Am J Hosp Pharm 51:188-97; quiz 229-30, 1994
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98.
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lymphoblastic leukemia. J Pediatr 117:238-44, 1990
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Adams MJ, Lipshultz SE: Pathophysiology of anthracycline- and radiation-associated 
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induced clinical heart failure in children: a systematic review. Ann Oncol 13:503-12, 2002
101. Kremer LC, van der Pal HJ, Offringa M, et al: Frequency and risk factors of subclinical 
cardiotoxicity after anthracycline therapy in children: a systematic review. Ann Oncol 13:819-29, 
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102. Sorensen K, Levitt GA, Bull C, et al: Late anthracycline cardiotoxicity after childhood cancer: a 
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119. Pounds S, Cheng C: Robust estimation of the false discovery rate. Bioinformatics 22:1979-87, 
2006
120. Hummel M, Bentink S, Berger H, et al: A biologic definition of Burkitt's lymphoma from 
transcriptional and genomic profiling. N Engl J Med 354:2419-30, 2006
121. Pounds S, Cheng C: Sample size determination for the false discovery rate. Bioinformatics 
21:4263-71, 2005
122. West AN, Ribeiro RC, Jenkins J, et al: Identification of a novel germ line variant hotspot mutant 
p53-R175L in pediatric adrenal cortical carcinoma. Cancer Res 66:5056-62, 2006
123. Broniscer A, Baker SJ, West AN, et al: Clinical and molecular characteristics of malignant 
transformation of low-grade glioma in children. J Clin Oncol 25:682-9, 2007
124. West AN, Neale GA, Pounds S, et al: Gene expression profiling of childhood adrenocortical 
tumors. Cancer Res 67:600-8, 2007
125. Livak KJ, Schmittgen TD: Analysis of relative gene expression data using real-time quantitative 
PCR and the 2(-Delta Delta C(T)) Method. Methods 25:402-8, 2001
126. Wensing B, Farrell PJ: Regulation of cell growth and death by Epstein-Barr virus. Microbes 
Infect 2:77-84, 2000
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Appendix I
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
APPENDIX I:  PEDIATRIC RITUXIMAB INFUSION GUIDELINES
Dose = 375 mg/m2

Dilute in NS to final concentration 1 mg/ml for ease of administration.

Administer intravenously through a dedicated line
Pre-medicate with:

Acetaminophen 15 mg/kg PO (max 650 mg)

Benadryl 1 mg/kg IV or PO (max 50 mg)
FIRST INFUSION:
See current St. Jude formulary from Intranet. 
SUBSEQUENT INFUSIONS:
See current St. Jude formulary from Intranet.
GUIDELINES:
1. Monitor blood pressure, pulse, respiration and temperature every 15 minutes for the first hour or 
until the participant is stable, then hourly until the infusion is complete.
2. Have epinephrine and diphenhydramine available along with resuscitation equipment for the 
emergency management of anaphylactic reactions.
3. If infusion related events occur, slow the infusion or stop the infusion until resolution, treating the 
participant if necessary.  If it is determined that the participant can be safely re-challenged, begin 
the infusion at 50% of the rate at which the infusion was running when stopped.
4. Verify that the participant has not taken antihypertensive medications within 12 hours of beginning 
the rituximab infusion. Hypotension may occur during the infusion.
5. Participants with a high number of circulating cancer cells (> 25,000/mm3) may be at a higher risk 
for tumor lysis syndrome. These patients should receive allopurinol and intravenous hydration and 
should be monitored.
6. Participants who experience severe infusional symptoms may need to be hospitalized for 
observation. Participants and families should be counseled to be seen in the emergency room if 
infusion-related symptoms occur again after the infusion is complete (i.e. at home at a later time).
7. Monitor participants during and for a few weeks after receiving rituximab for the development of 
mucositis with a sore throat or mouth ulcers followed by a diffuse skin rash which can worsen 
rapidly and result in total body skin sloughing and can become life threatening.
8. Carriers of hepatitis B should be closely monitored for clinical and laboratory signs of active HBV 
infection and for signs of hepatitis throughout their study participation. 
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SJBC3
Appendix II
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
APPENDIX II: GROUP CLASSIFICATION/MURPHY STAGE
Group Classification
Group A - Completely resected stage I or completely resected abdominal stage II lesions.
Group B - All cases not eligible for Group A or Group C. (Murphy Stage III and non-CNS Stage IV)
Group C - Any CNS involvement and/or bone marrow involvement ≥ 25% blasts.  For CNS 
involvement one or more of the following applies:
(1)
Any L3 blasts in CSF
(2)
Cranial nerve palsy (if not explained by extracranial tumor)
(3)
Clinical spinal cord compression
(4)
Isolated intracerebral mass
(5)
Parameningeal extension: cranial and/or spinal
Murphy’s Staging
Stage 1: A single tumor (extranodal) or a single anatomical site (nodal) with exclusion of the 
mediastinum or abdomen.
Stage 2: A single tumor (extranodal) with regional involvement:
-
Two or more nodal areas on the same side of the diaphragm
-
Two single (extranodal) tumors with or without regional node involvement on the same 
side of the diaphragm.  A primary gastrointestinal tract tumor, usually in the ileocecal 
area, with or without involvement of associated mesenteric nodes only, grossly 
completely resected
Stage 3: Two single tumors (extranodal) on opposite sides of the diaphragm
-
Two or more nodal areas above and below the diaphragm
-
All primary intrathoracic tumors (mediastinal, pleural, thymic)
-
All extensive primary intra-abdominal disease, unresectable
-
All paraspinal or epidural tumors, regardless of other tumor sites
Stage 4: Any of the above with initial CNS and/or bone marrow involvement
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SJBC3
Appendix III
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
APPENDIX III:  BIOLOGICAL SPECIMENS
To ensure high quality data from the genomic profiling studies, it is important to immediately process 
tissue obtained at the time of tumor sampling to maximize DNA and RNA integrity. Tissue samples 
should be placed in liquid nitrogen as soon as possible (or after an intermediate freezing step, such as 
in a dry ice and ethanol slurry). If access to very low temperature freezing is not available, a portion of 
the tissue sample should be placed immediately in a solution that prevent RNA degradation, and permit 
subsequent RNA extraction, e.g. RNAlater (Ambion).
DNA studies:
DNA will be isolated from peripheral blood and tumors using commercially available reagents, such as 
the DNA blood and tissue mini kit (Qiagen, Valencia, CA) or by phenol-chloroform extraction. DNA 
purity and integrity will be assessed by spectrophotometry and by agarose gel electrophoresis. Analysis 
of DNA copy number alterations and LOH will be performed using Affymetrix SNP 6.0 arrays 
(Affymetrix, Santa Clara, CA) using a bioinformatic analysis pipeline established and developed at St. 
Jude. Validation of CNA will be performed using genomic quantitative PCR and/or fluorescence in 
situ hybridization on paraffin embedded tissue. Mutational analysis of genes including TP53, c-MYC 
and ARF¸ and other genes found to be targeted by recurring genetic alterations on SNP array analysis, 
will be performed by PCR and direct sequencing on ABI 3730 analyzers at St. Jude25,122,123. 
Methylation analysis of individual genes will be performed using mass spectrometry of bisulfate 
treated DNA using the MassARRAY platform (Sequenom, San Diego, CA).
RNA studies:
RNA will be extracted from frozen tissue samples or samples placed in RNAlater using TriZOL 
reagent, or alternative commercially available reagents, such as the QiagenRNeasy RNA Midi-Prep kit 
(Valencia, CA) according to the Manufacturer’s recommendations122. The RNA will be resuspended in 
diethylpyrocarbonate-treated water, quantified by UV absorbance at 260/280 nm and stored at -80oC. 
a) Quantitative Real-Time PCR: cDNA will be generated from 1 g total RNA using 
commercially available kits (e.g., iScriptcDNA amplification kit, Bio-Rad Laboratories, 
Hercules, CA). Specific gene transcripts will be identified and quantified by amplifying 
cDNA using DNA oligonucleotide primers and intercalating fluorescent dyes  (e.g., 
SybrGreen) or custom specific fluorescent probes (e.g. 6FAM and MGB labeled probes 
designed using Primer Express, Applied Biosystems, Foster City, CA).122,124 GAPDH, 
RNAseP or other housekeeping genes will also be analyzed as an internal loading control. 
Expression levels will be determined by the comparison of standard curves orCt 
methods125. 
b) Affymetrix Gene Expression: Intact total cellular RNA (~5-10 g) will be processed 
according to the Affymetrix one-cycle labeling protocol 
(http:www.affymetrix.com/support/technical/manual/expression_manual.affx) and analyzed 
using the Affymetrix U133 Plus 2.0GeneChip through the assistance of the Hartwell 
Center. Data will be analyzed using the AffymetrixMAS 5.0, Bioconductor (e.g. 
limmamodule) software or other appropriate programs. 
St. Jude Children's Research Hospital
IRB NUMBER: Pro00001026
IRB APPROVAL DATE: 07/25/2022

SJBC3
Appendix III
Amendment 6.0, dated: 04-22-2022 
Protocol document date: 04-22-2022
Protein studies:
Protein will be isolated from whole cell extracts in NuPAGE (Invitrogen), T-PER (Pierce Chemical, 
Rockford, IL) or other suitable lysis buffers  containing a protease and or phosphatase inhibitor 
cocktail (Roche Diagnostics Corporation, Indianapolis, IN) or by other appropriate methods. Total 
protein will be separated by SDS-PAGE and transferred to nylon or nitrocellulose membranes. The 
filters will be blocked in buffer containing milk or bovine serum albumin and probed with primary 
antibodies that are specific for p53, HDM2, p14ARF, c-MYC and other constituents of the TP53 
signaling pathway, and other genes found to be targets of recurring genetic alterations. Horseradish 
peroxidase-linked secondary antibodies and chemiluminescent-based reagents (e.g., Supersignal West 
Femto kit, Pierce Chemical) will be used to detect the level of expression of each protein.
Return of results:
As in other tumor profiling projects, germline material is being used to serve as a comparator to tumor 
material to facilitate the identification of tumor-specific (i.e. somatic) mutations. We will not perform 
interrogative germline analyses.   These studies are being done in a non-CLIA certified research 
laboratory, and therefore the results will not be returned to the patient/parent.  This has been made 
clear in the consent form.  In the unlikely event that a genetic abnormality was discovered that is 
thought to have clinical significance (after consultation with experts such as members in the cancer 
predisposition team), we would approach the IRB for a decision as to how to best handle the situation.  
Patients will all be enrolled on the TBANK protocol, which permits contact with participant/family if 
something is discovered that could potentially affect the health and wellbeing of a child under the age 
of 18.
The non-CLIA certified laboratory results for the participants from international sites will not be 
returned to the primary physician or family, as indicated in the consent form.
EBV METHODS
Initial identification of EBV-positive BLs will be determined by in situ hybridization with EBV EBER 
RNA. Tumors will be analyzed for the presence of a variant latency program where EBNA-3 proteins 
are expressed or for the latency I program found in the majority of BL.
Classification of EBV genomes:  To identify the latency program expressed in these tumors, the EBV 
genome will be examined by PCR. In African tumors, this pattern of latency is associated with a 
deletion in the region of the genome encoding the EBNA-2 latency protein; the known deletions fall 
within a defined region and can be identified by long range PCR analysis using a common set of 
primers. These tumors also carry wild-type EBV genomes that are silenced by an as yet undefined 
mechanism. Therefore, tumors will be analyzed for the presence of both wild-type and deleted 
genomes, allowing the detection of wild-type genomes. Detection of wild-type genomes will verify the 
presence of EBV in the tumor and serve as a positive control for the PCR reaction. DNA will be 
amplified using the CG Rich PCR System (Roche) according to the manufacturer’s instructions. Each 
50l reaction will contain 1 x GC Rich PCR buffer, 200 M each dNTP, 400 nM forward and reverse 
primer, 100ng DNA and 2units of GC Rich enzyme mix. Primers to amplify the deleted EBV genome 
will be 5’-TCCTCTCCAACCTTCGCTCC-3’ (forward primer) and 5’-
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GCCTTCGCTGGCTTCTAACATC-3’ (reverse primer); primers for amplification of wild type EBV 
will be 5’-CTCTGCCACCTGCAACACTA-3’ (forward primer) and 5’-
GAGGGTGCATTGATTGGTCT-3’ (reverse primer). Conditions for PCR have been previously 
established in the laboratory and these primers have been found to amplify EBV DNA from 4 distinct 
tumors. Products will be examined by agarose gel electrophoresis.  
EBV gene expression:  The presence or absence of the EBNA-3 proteins will be determined using RT-
PCR analysis of RNA isolated from the tumors. If sufficient sample is available (106 or fewer cells are 
required), these results will be confirmed by immunoblot analysis using monospecific antibodies to 
each individual EBNA-3 protein and the EBNA-1 protein. Ongoing studies in the Sample laboratory 
are directed at identifying cellular changes in the subset of BL associated with the expression of the 
EBNA-3 proteins, and these will be incorporated into the analysis as appropriate. 
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APPENDIX IV:  PSYCHOLOGICAL TESTING
Cognitive 
Domain
Measure
Broad Skill
Task Characteristics
Age  Range
Admin.
Time
IQ Estimate
SB-V Routing 
Subtests 
Global Cognitive Function
Object Series/Matrices and Descriptive Vocabulary 
subtests
3+
20 min.
Attention & 
Executive 
Function
Digit Span- 
WISCIV Integrated
Forward- Brief Auditory 
Attention
Backward- Auditory Working 
Memory
Forward- repeat random digit strands of increasing 
length
Backward- repeat random digit strands in reverse 
order
*6-17
(WAIS- 
17+)
5 min.
Conners’ CPT
Sustained Visual Attention
Selectively respond to target stimuli (all letters 
except X) and not a distractor (X); computerized
4+
20 min.
Auditory Working  
Memory- WJIII
Auditory Working Memory
Repeat words and numbers after organizing based 
on rule
4+
5 min.
Retrieval Fluency- 
WJIII
Verbal Fluency
Name foods, people names & animals, each in 1 
min. trial
3+
5 min.
(Parent 
Measure)
BRIEF 
Questionnaire
Executive Function
Questionnaire assessing executive dysfunction in 
community
2-5/6-18
15 min.
Memory
CVLT-C/CVLT-II
Verbal List Learning
Learn 15 item list with multiple presentations, 
interference, delay & recognition
5-17
(CVLTII- 
17+)
15 min.
Bead Memory- 
SBIV
Visual Learning & Memory
Recreate bead patterns (color & shape) of 
increasing length
3+
5 min.
Rey-O
Complex Visual Memory Task
Copy a complex figure followed by drawing from 
memory after short and long delays; recognition 
trial included
6+
20 min.
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Processing 
Speed
Visual Matching- 
WJIII
Processing Speed
Find matching pairs; timed
3+
3 min.
Decision Speed- 
WJIII
Processing Speed
Pick two conceptually similar pairs; timed
3+
3 min.
Visual Spatial/ 
Visual Motor
Beery VMI 
Visual Perception &
Visual-Motor Integration
Blue- Match shapes of decreasing size
Green- Draw shapes of increasing complexity
3-19
15 min.
Achievement**
Letter- Spelling, 
Letter Word 
Identification & 
Applied Problems- 
WJIII 
Academic Achievement
Single word reading, spelling to dictations and 
solving math word problems
3+
20 min.
Motor
Purdue Pegboard
Fine Motor Speed & Dexterity
Place pegs using dominant, nondominant & both 
hands; timed
3+
3 min.
Adaptive
ABAS-II
Adaptive Functioning
Inquires about adaptive skills across functional 
domains
birth-89
15 min.
Social-
Emotional
BASC-II
Psycho-social Adjustment
Inquires about 9 behavioral domains- aggression, 
anxiety, etc.
2.0- 21.11
15 min.
* Tests that will be tried at ages below normative population for longitudinal assessment of individual change 
Table Abbreviations: - SB-V- Stanford-Binet, Fifth Edition; WISC-IV- Wechsler Intelligence Scale for Children, Fourth Edition; WJIII- Woodcock 
Johnson, Third Edition; TOVA- Test of Variables of Attention; BRIEF- Brief Rating Inventory of Executive Function; Beery VMI- Beery-Buktenica 
Test of Visual-Motor Integration; CVLT-C- California Verbal Learning Test- Children’s Version; CVLT-II- California Verbal Learning Test, Second 
Edition Rey-O- Rey-Osterrieth Complex Figure; ABAS-II- Adaptive Behavior Assessment System, Second Edition; BASC-II- Behavior Assessment 
System for Children, Second Edition;
**Only administered at end of therapy in lieu of the Rey-O
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APPENDIX V:  COLLECTION AND SUBMISSION OF SPECIMENS FOR BIOLOGY STUDIES
Collection of Tissues:  Tumor tissue obtained at diagnosis must be immediately processed to ensure 
high quality, non-degraded nucleic acid material (DNA and RNA) for genomic analysis.
 One of two methods can be used:
1. Snap freezing in liquid nitrogen
Small tissue pieces (less than 0.5 cm in any one direction) should be wrapped in foil and snap 
frozen either in liquid nitrogen, or in dry ice/ethanol slurry, and then subsequently stored in 
liquid nitrogen. This material can then be shipped to St Jude on dry ice, and used for subsequent 
DNA and RNA extraction. 
OR
2. Storage in RNA later
If liquid nitrogen storage is not available, small tissue pieces (less than 0.5 cm in any one 
direction) should be placed in approximately 5 volumes of RNAlater (Ambion). e.g. 0.5g 
sample requires approximately 2.5ml of RNA later. This solution permeates the cells, 
stabilizing the RNA for subsequent extraction. The solution may then be stored for 1 day at 
37°C, 1 week at 25°C, 1 month at 4°C or indefinitely at -20°C. This tissue will then be 
processed for RNA and DNA.
Germline DNA will be obtained from peripheral blood samples, and may be drawn after chemotherapy 
has started (primary tumor for genomics still must be obtained pre chemo apart from listed exceptions 
as specified in the protocol eligibility). 5ml of EDTA-anti-coagulated blood should be collected and 
buffy coat leukocyte preparations made. These cell preparations may then be used for DNA extraction 
using either phenol-chloroform or column based methods. For sample from non-St Jude sites, the blood 
sample may be centrifuged, the buffy coat aspirated, washed in phosphate buffered saline, and then the 
leukocytes pelleted, and the cell pellet frozen on dry ice. The pellet is then stored on liquid nitrogen. If 
liquid nitrogen is not available, ship the EDTA tube to St. Jude within 24 hours of collection at 
room temperature.
Mail all specimens to:
Charles Mullighan / Audrey Phillips
St Jude Children's Research Hospital
262 Danny Thomas Place
Pathology, Mail Stop 342, Room D4036
Memphis, TN, 38105
For questions, e-mail or call Dr. Mullighan, or call Assistant: Carolyn Reed: T: 
or Lab 
Manager: Audrey Phillips: T: 
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APPENDIX VI:  CLINICAL DATA FOR PARTICIPANTS AT COLLABORATING SITES 
PARTICIPATING IN BIOLOGIC OBJECTIVES ONLY
Age
Race
Gender
Diagnosis Date
LDH
CBC with diff
Diagnosis
Sites of Disease
Stage
Treatment Regimen(s)
Treatment Start Date
Treatment Best Response
Date of Response
Date Completed Therapy
Significant Events (e.g. relapse, induction failure, death, cause of death, second malignancy)
Date Event
Last Contact Date
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APPENDIX VII: SUB-STUDY FOR COLLABORATING INSTITUTIONS 
PARTICIPATING IN BIOLOGICAL OBJECTIVES ONLY
1.0
OBJECTIVES
1.1
Biological Primary Objectives
1.1.1
To perform genomic analysis of childhood newly diagnosed mature B cell 
lymphomas (e.g. Burkitt lymphoma/leukemia, DLBCL, and MLBCL) obtained 
from different parts of the world.
1.1.2
To describe the types and frequency of mutations in the ARF-HDM2-TP53 
pathway, in B-cell lymphomas in the United States and those found in selected 
geographic regions of the world.
1.1.3
To describe the expression of ARF-HDM2-TP53 and PUMA-associated pathways 
in B-cell lymphomas in the United States and that found in B-cell lymphomas of 
other selected geographic regions of the world.
1.1.4
To describe the pattern and frequency of XLP gene mutations presenting with B-cell 
lymphomas in the United States and selected geographic regions.
1.1.5
To describe the frequency of EBV-positive B-cell lymphomas in the United States 
and selected geographical regions of the world.

To describe the pattern of EBV protein and gene expression (e.g., EBNA 3) in 
EBV-positive lymphomas.

To compare patterns of EBV protein and gene expression with clinical, 
laboratory and outcome data.
1.2
Exploratory Aim
1.2.1
Tumor Bank
To establish a tumor bank of mature B-cell lymphoma samples for future molecular 
studies aimed to clarify the role of c-MYC oncogene and its associated pathways, and 
other pathways found to harbor genetic alterations from the genomic profiling studies 
described in 1.1.1.
2.0 
BACKGROUND AND RATIONALE
2.1
Background
2.1.1
Genome-Wide Profiling of Genetic Alterations in Lymphoma
Genome-wide profiling of the gene expression and genetic alterations, including DNA 
copy number abnormalities (CNA) and loss of heterozygosity (LOH), and genome 
sequencing has provided valuable insights into the pathogenesis of lymphoid 
neoplasms.1,2 Gene expression profiling has been found to be an effective tool in 
differentiating BL from other mature B-cell lymphomas like DLBCL in adults.3,4 These 
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studies have shown that some cases that cannot be classified unequivocally by classical 
morphological or cytogenetic modalities express a clear genetic signature that allows for 
their classification.  Three major DNA microarray studies have looked into the biological 
and clinical heterogeneity of adult DLBCL.5-7 The initial DLBCL gene expression 
profiling study used genes that define the germinal center stage of B-cell differentiation 
to identify 2 prominent DLBCL subgroups. The “germinal center B-cell-like” DLBCL 
(GCB DLBCL) expressed genes characteristic of normal germinal center B cells (e.g. CD 
10, BCL6, A-myb), whereas the “activated B cells “ DLBCL subgroup (ABC DLBCL) 
expressed genes that are induced during mitogenic activation of peripheral blood B cells 
(e.g. BCL2, IRF-4, cyclin D2)5. These two different patterns of differentiation were 
associated with significantly different outcomes. Another larger gene expression profiling 
study of DLBCL cases confirmed the existence of these two subgroups as well as a third 
one called “type 3” that did not resemble either and may represent an additional 
subgroup.6 Genome-wide profiling of CNA in DLBCL has identified multiple recurring 
regions of DNA gain and loss, many of which are associated with the DLBCL subtypes 
identified by gene expression profiling. These include amplification of a region of 
chromosome 19 harboring the SPIB transcription factor, deletion of CDKN2A/B and 
trisomy 3 in ABC DLBCL; and amplification of the mir-17-92 microRNA cluster and 
deletion of PTEN and GCB DLBCL. To date, there are limited genome-wide profiling 
data of genetic alterations pediatric B-NHL. Our aim is to perform integrated analysis of 
genomic data to identify genetic pathways that are altered in this disease, to identify 
tumor subtypes, to examine variation in genetic features according to geographic region, 
and to perform a comparative analysis with existing data from adult cases.5,9
Techniques for studying the genomic profile of patient tumors (somatic and germline) 
continue to advance.  These approaches to be used, depending on sample availability and 
quality include RNA (transcriptome) sequencing, exome sequencing, single nucleotide 
polymorphism array analysis and whole genome sequencing. These will be done using 
standard established approaches in the Mullighan laboratory and the Genomic 
Sequencing Facility of the Hartwell Center at St. Jude.
2.1.2
ARF-HDM2-TP53 and PUMA
Burkitt lymphoma (BL) and mature B-cell leukemia (B-ALL) are characterized by one of 
three reciprocal chromosomal translocations [t(8;14);t(2;8);t(8;22)].  The unifying feature 
of these translocations is the juxtaposition of the c-MYC proto-oncogene and one of the 
immunoglobulin genes.  These translocations result in the constitutive over-expression of 
c-MYC, a transcription factor which is necessary and sufficient for promoting cell 
proliferation and for provoking angiogenesis.  In normal B-cells, c-MYC expression is 
tightly regulated and rapidly responds to mitogenic or growth inhibitory signals.  These 
controls are lost in BL/B-ALL, resulting in aberrantly high levels of c-MYC protein.  The 
c-MYC translocations that occur in BL/B-ALL have been extensively studied with respect 
to their breakpoint patterns and their mechanism of de-regulation of c-MYC expression.  
However, recent insights into the molecular pathogenesis of BL/B-ALL have come from 
a detailed analysis of E-myc transgenic mice, a model which accurately mimics the 
chromosomal abnormality and pathology of BL/B-ALL12.  In normal cells, including B 
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lymphocytes, c-Myc expression activates the Arf-Mdm2-p53 pathway resulting in 
apoptosis.13-15 Arf (mouse p19Arf /human p14ARF) is encoded by an alternative reading 
frame of the Ink4a locus and limits lymphomagenesis by antagonizing Mdm2 (murine 
double minute-2; HDM2 in man), a negative regulator of the tumor suppressor p53.16,17  
The c-Myc transgenic animal studies have shown that 80% of the lymphomas that arise in 
these mice carry mutations affecting apoptotic checkpoints, from either deletions of 
Ink4a/Arf (25%), mutation of p53 (30%) or overexpression of Mdm2 (50%).15,18-22  
Although data suggest that one-third of human BL/B-ALL may have mutations of 
TP5323,24 many of the studies performed have been anecdotal and/or limited to the study 
of BL/B-ALL cell lines25,26. Therefore, a detailed study of alterations in this pathway is 
currently lacking in human mature B cell malignancies.
Although these studies established that c-Myc activates Arf and p53, and that functional 
loss of this pathway impairs Myc-induced apoptosis, Myc activation can still trigger cell 
death in the absence of Arf and/or p53.15 Activation of c-Myc in normal B cells 
suppresses the expression of Bcl-2 or Bcl-XL, which are anti-apoptotic proteins that 
protect the cells from a variety of insults, including chemotherapeutic drugs. Importantly, 
an analysis of lymphomas arising in E-Myc transgenic mice has established that this 
pathway is also bypassed during lymphoma development and that this also occurs at an 
equal frequency in tumors having alterations in the ARF-Mdm2-p53 pathway.
Recent studies have shown that about one third of BL in humans harbor c-MYC 
mutations, and two recurrent mutant c-MYC alleles derived from BL have been shown to 
be able to evade the p53 tumor surveillance network and promote rapid lymphoma 
development in mice.25 Another possible mediator in the altered apoptotic pathways of 
BL is PUMA (p53-up-regulated modulator of apoptosis). PUMA, a pro-apoptotic BH3-
only Bcl-2 family protein, is an essential mediator of p53- dependent and independent 
cell death,26 and recent observations have indicated that 40% of primary BLs (n=19) fail 
to express detectable levels of PUMA, and that in some cases PUMA is inactivated by 
epigenetic silencing.27
Understanding the exact mechanism by which the apoptotic pathways are affected in 
these lymphomas will provide an opportunity for the development of novel therapies. The 
development of more rational therapies with a selective effect in specific tumor pathways 
while sparing normal tissue can have a great impact in the survival of cancer especially in 
the developing world, where children are at high risk of dying of the toxic effects of 
aggressive chemotherapy due to suboptimal hospital infrastructure.
2.1.3
Epstein-Barr Virus
Epstein–Barr virus (EBV) is a B lymphotropic herpes virus linked to several human 
diseases including B-cell malignancies.126 Greater than 90% of the world’s population is 
persistently infected with EBV. It is a ubiquitous γ-herpes virus that, upon primary 
infection, causes a brief and confined replication of the virus at the site of infection in the 
immunocompetent host.29 In industrialized societies, infection with EBV generally occurs 
later in life, compared to early childhood infection common in countries with limited 
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resources. EBV is transmitted orally from oropharyngeal secretions of asymptomatic 
carriers and patients with infectious mononucleosis.28 While EBV infects B lymphocytes 
early in the course of primary infection, it normally does not replicate in them, but rather 
establishes a latent infection characterized by the limited expression of a subset of 
latency-associated genes.30  The latency genes consist of six nuclear antigens (EBNAs 1, 
2, 3A, 3B, 3C and LP) and three membrane proteins (LMPs 1, 2A and 2B).31 There have 
been three different latency-gene expression programs described after EBV infection. In 
BL/B-ALL only a very limited repertoire of the latency-associated genes (EBNA-1, 
EBV-encoded RNA (EBER)-1 and EBER-2 and the BamHI-A rightwards transcripts 
[BARTs]) are routinely expressed (Latency I) contributing to immune evasion.32 The 
Latency II program includes additional expression of LMP1 and LMP2A/B, and is 
commonly observed in EBV-positive Hodgkin lymphoma, some types of T and NK-cell 
lymphomas and some cases of B-cell lymphoma.33 Latency III is expressed in 
lymphoblastoid cell lines and newly infected B cells in vivo, and includes the entire array 
of EBV latency-associated genes. These cells would normally be very susceptible to 
killing by EBV-specific T-cells, and, therefore, this pattern of expression is only seen in 
the EBV-associated lymphoproliferative disease (EBV-LPD) associated with primary or 
secondary immunodeficiency.33 The EBER RNAs are present in all previously described 
programs of EBV latency, and their presence within tumor samples is evidence of a latent 
EBV infection.34
The EBNA3 proteins (3A, 3B and 3C) are not normally expressed in classic endemic BL 
(eBL); however, recent studies have revealed a subset of eBLs (~ 15%) that express these 
EBV proteins as a consequence of a deletion in the EBNA2 gene.35,36 These tumors are 
particularly resistant to apoptotic stimuli. Dr. Clare Sample’s laboratory has determined 
that, in a SCID mouse model of tumorigenicity, these variant EBNA-3+ eBLs are 
particularly aggressive relative to the eBLs that do not carry the EBNA2 deletion (C. 
Sample, personal communication) and additional unpublished data suggests that the 
EBNA-3 proteins may contribute directly to proliferation of the tumor cells. Thus, it is 
important to determine whether the pattern of EBV gene expression is associated with 
biologic and clinical and outcome features. 
EBV has not been shown to be directly involved in BL pathogenesis, but has rather been 
suggested to increase the target pool of cells susceptible to malignant transformation. 
However, an interesting study by Ruf, et. al has demonstrated that the malignant 
phenotype of a group I BL line requires the EBV genome, and that this is associated with 
EBV’s ability to impair the c-MYC apoptotic program.37 Therefore, at least in this 
scenario, malignant transformation requires EBV gene expression altering apoptotic 
pathways. These observations suggest that EBV may influence c-MYC’s ability to trigger 
apoptosis in BL and possibly other pathways as well. Hence, we propose to evaluate and 
correlate EBV gene expression with alterations in the ARF-Mdm-p53 and Bcl-2/Bcl-X 
apoptotic pathways.
There are two main clinical presentations of BL clustering in different geographic 
regions. The eBL, which affects younger children and often involves the jaw, is found 
mainly in equatorial Africa and New Guinea.38-40 The majority of patients with eBL have 
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been infected with malaria. By contrast, non-endemic or sporadic BL (sBL) occurs 
worldwide and mainly affects young adults. Both forms of BL differ in their association 
with EBV: eBL is almost always associated with EBV, whereas sBL has a more irregular 
association, ranging from 10% to 30%.41 However, our knowledge of other clinical and 
biological forms of BL, particularly in developing countries outside of Africa, is very 
limited. A study examining EBV association with BL in temperate countries of South 
America, found a 50% EBV association with predominantly abdominal presentation,42 
suggesting an intermediate form. The pattern of EBV gene expression has not been 
determined in these BLs. The rate of South American patients with BL infected with 
malaria is extremely low. Studies in Northeastern Brazil and Northern Africa have shown 
about 85-90% positivity for EBV (like eBL), but have a predominant abdominal 
presentation of the tumors (90%) similar to sBL, suggesting that other still unknown 
genetic or environmental factors might modify the clinical presentation, and possibly 
response to therapy of BL in these regions.43,44
2.1.4
X-Linked Lymphoproliferative syndrome
X-linked lymphoproliferative (XLP) syndrome is a sex-linked immunodeficiency 
syndrome. Boys with this syndrome are at increased risk for the development of B-cell 
lymphomas and fatal infectious mononucleosis. Most are diagnosed by the time they are 
5 years of age, although some have gone undiagnosed until much later in life (i.e. 5th or 
6th decade). There may be an associated hypogammaglobulinemia; however, there is 
generally not a significant history of infections. As a result, patients are usually 
undiagnosed until they develop a B-cell lymphoma or die from infectious mononucleosis. 
Currently, boys with newly diagnosed high grade B-cell NHL are not screened for the 
XLP gene mutation. Hence, the actual frequency of these mutations in boys with primary 
high grade B-cell lymphoma is not known.
We have identified five boys with B-cell lymphoma who have subsequently been found 
to have an XLP gene mutation. These cases were only identified after their clinical course 
suggested the possibility of XLP. Now that we have an accurate and efficient assay to 
screen for the XLP gene mutations, we believe that boys with BL and other B-cell 
lymphomas should routinely be screened for XLP mutations. It is not known whether 
different XLP mutations result in the same clinical phenotype or have prognostic 
importance.
3.0 
ELIGIBILITY CRITERIA AND STUDY ENROLLMENT 
3.1
Inclusion criteria for subjects participating in biological objectives only

Participant must have a histologic diagnosis of a mature B cell lymphoma 
(e.g., Burkitt lymphoma/leukemia, atypical Burkitt lymphoma, diffuse large 
B-cell lymphoma, mediastinal large B-cell lymphoma, mature B-cell 
lymphoma NOS) as defined in the WHO classification.

Participant must be < 22 years of age at the time of diagnosis.
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
Participant must be previously untreated, (no more than 72 hours of steroids, 
one intrathecal chemotherapy treatment, and/or emergency radiation) at the 
time of the diagnostic biopsy.

Informed consent must be obtained by local PI or his/her designee according to 
ICH/Good Clinical Practice and local guidelines before enrollment into study.
3.2
Exclusion criteria for subjects participating in biological objectives only

Inability or unwillingness of research participant or legal guardian to consent.

Histologic diagnosis other than a mature B-cell lymphoma as defined in the 
WHO classification.
4.0
DESIGN AND METHODS
4.1
Biological Specimens
To ensure high quality data from the genomic profiling studies, it is important to 
immediately process tissue obtained at the time of tumor sampling to maximize DNA and 
RNA integrity. Tissue samples should be placed in liquid nitrogen as soon as possible (or 
after an intermediate freezing step, such as in a dry ice and ethanol slurry). If access to 
very low temperature freezing is not available, a portion of the tissue sample should be 
placed immediately in a solution that prevent RNA degradation, and permit subsequent 
RNA extraction, e.g. RNAlater (Ambion). See Section 6.2 of main protocol for further 
details.
4.1.1
DNA studies:
DNA will be isolated from peripheral blood and tumors using commercially available 
reagents, such as the DNA blood and tissue mini kit (Qiagen, Valencia, CA) or by 
phenol-chloroform extraction. DNA purity and integrity will be assessed by 
spectrophotometry and by agarose gel electrophoresis. Analysis of DNA copy number 
alterations and LOH will be performed using Affymetrix SNP 6.0 arrays (Affymetrix, 
Santa Clara, CA) using a bioinformatic analysis pipeline established and developed at St. 
Jude. Validation of CNA will be performed using genomic quantitative PCR and/or 
fluorescence in situ hybridization on paraffin embedded tissue. Mutational analysis of 
genes including TP53, c-MYC and ARF¸ and other genes found to be targeted by 
recurring genetic alterations on SNP array analysis, will be performed by PCR and direct 
sequencing on ABI 3730 analyzers at St. Jude24,122,123. Methylation analysis of individual 
genes will be performed using mass spectrometry of bisulfate treated DNA using the 
MassARRAY platform (Sequenom, San Diego, CA).
4.1.2
RNA studies:
RNA will be extracted from frozen tissue samples or samples placed in RNA later using 
TriZOL reagent, or alternative commercially available reagents, such as the 
QiagenRNeasy RNA Midi-Prep kit (Valencia, CA) according to the Manufacturer’s 
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recommendations.122 The RNA will be resuspended in diethylpyrocarbonate-treated 
water, quantified by UV absorbance at 260/280 nm and stored at -80oC. 
a) Quantitative Real-Time PCR: cDNA will be generated from 1 g total RNA using 
commercially available kits (e.g., iScriptcDNA amplification kit, Bio-Rad 
Laboratories, Hercules, CA). Specific gene transcripts will be identified and 
quantified by amplifying cDNA using DNA oligonucleotide primers and 
intercalating fluorescent dyes  (e.g., SybrGreen) or custom specific fluorescent 
probes (e.g. 6FAM and MGB labeled probes designed using Primer Express, 
Applied Biosystems, Foster City, CA).122,124 GAPDH, RNAseP or other 
housekeeping genes will also be analyzed as an internal loading control. 
Expression levels will be determined by the comparison of standard curves 
orCt methods.125
b) Affymetrix Gene Expression: Intact total cellular RNA (~5-10 g) will be 
processed according to the Affymetrix one-cycle labeling protocol 
(http:www.affymetrix.com/support/technical/manual/expression_manual.affx) 
and analyzed using the Affymetrix U133 Plus 2.0 GeneChip through the 
assistance of the Hartwell Center. Data will be analyzed using the 
AffymetrixMAS 5.0, Bioconductor (e.g. limmamodule) software or other 
appropriate programs. 
The non-CLIA certified laboratory results for the participants from international sites will 
not be returned to the primary physician or family, as indicated in the consent form
4.1.3
Protein studies:
Protein will be isolated from whole cell extracts in NuPAGE (Invitrogen), T-PER (Pierce 
Chemical, Rockford, IL) or other suitable lysis buffers  containing a protease and or 
phosphatase inhibitor cocktail (Roche Diagnostics Corporation, Indianapolis, IN) or by 
other appropriate methods. Total protein will be separated by SDS-PAGE and transferred 
to nylon or nitrocellulose membranes. The filters will be blocked in buffer containing 
milk or bovine serum albumin and probed with primary antibodies that are specific for 
p53, HDM2, p14ARF, c-MYC and other constituents of the TP53 signaling pathway, and 
other genes found to be targets of recurring genetic alterations. Horseradish peroxidase-
linked secondary antibodies and chemiluminescent-based reagents (e.g., Supersignal 
West Femto kit, Pierce Chemical) will be used to detect the level of expression of each 
protein.
4.1.4
EBV METHODS
Initial identification of EBV-positive BLs will be determined by in situ hybridization 
with EBV EBER RNA. Tumors will be analyzed for the presence of a variant latency 
program where EBNA-3 proteins are expressed or for the latency I program found in the 
majority of BL.
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Classification of EBV genomes:  To identify the latency program expressed in these 
tumors, the EBV genome will be examined by PCR. In African tumors, this pattern of 
latency is associated with a deletion in the region of the genome encoding the EBNA-2 
latency protein; the known deletions fall within a defined region and can be identified by 
long range PCR analysis using a common set of primers. These tumors also carry wild-
type EBV genomes that are silenced by an as yet undefined mechanism. Therefore, 
tumors will be analyzed for the presence of both wild-type and deleted genomes, 
allowing the detection of wild-type genomes. Detection of wild-type genomes will verify 
the presence of EBV in the tumor and serve as a positive control for the PCR reaction. 
DNA will be amplified using the CG Rich PCR System (Roche) according to the 
manufacturer’s instructions. Each 50l reaction will contain 1 x GC Rich PCR buffer, 
200 M each dNTP, 400 nM forward and reverse primer, 100ng DNA and 2units of GC 
Rich enzyme mix. Primers to amplify the deleted EBV genome will be 5’-
TCCTCTCCAACCTTCGCTCC-3’ (forward primer) and 5’-
GCCTTCGCTGGCTTCTAACATC-3’ (reverse primer); primers for amplification of 
wildtype EBV will be 5’-CTCTGCCACCTGCAACACTA-3’ (forward primer) and 5’-
GAGGGTGCATTGATTGGTCT-3’ (reverse primer). Conditions for PCR have been 
previously established in the laboratory and these primers have been found to amplify 
EBV DNA from 4 distinct tumors. Products will be examined by agarose gel 
electrophoresis.
EBV gene expression:  The presence or absence of the EBNA-3 proteins will be 
determined using RT-PCR analysis of RNA isolated from the tumors. If sufficient sample 
is available (106 or fewer cells are required), these results will be confirmed by 
immunoblot analysis using monospecific antibodies to each individual EBNA-3 protein 
and the EBNA-1 protein. Ongoing studies in the Sample laboratory are directed at 
identifying cellular changes in the subset of BL associated with the expression of the 
EBNA-3 proteins, and these will be incorporated into the analysis as appropriate. 
4.2
Collection and Submission of Specimens for Biology Studies
4.2.1
Collection of tissues:
Tumor tissue obtained at diagnosis must be immediately processed to ensure high quality, 
non-degraded nucleic acid material (DNA and RNA) for genomic analysis.
One of two methods:
1. Snap freezing in liquid nitrogen
Small tissue pieces (less than 0.5 cm in any one direction) should be wrapped 
in foil and snap frozen either in liquid nitrogen, or in dry ice/ethanol slurry, 
and then subsequently stored in liquid nitrogen. This material can then be 
shipped to St Jude on dry ice and used for subsequent DNA and RNA 
extraction. 
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OR
2. Storage in RNA later
If liquid nitrogen storage is not available, small tissue pieces (less than 0.5 cm 
in any one direction) should be placed in approximately 5 volumes of 
RNAlater (Ambion), e.g. 0.5g sample requires approximately 2.5ml of RNA 
later. This solution permeates the cells, stabilizing the RNA for subsequent 
extraction. The solution may then be stored for 1 day at 37°C, 1 week at 25°C, 
1 month at 4°C or indefinitely at -20°C. This tissue will be processed for RNA 
and DNA.
4.2.2
Collection of Blood for Germline DNA
5ml of EDTA-anti-coagulated blood should be collected and buffy coat leukocyte 
preparations made. These cell preparations may then be used for DNA extraction using 
either phenol-chloroform or column based methods. For sample from non-St Jude sites, 
the blood sample may be centrifuged, the buffy coat aspirated, washed in phosphate 
buffered saline, and then the leukocytes pelleted, and the cell pellet frozen on dry ice. The 
pellet is then stored on liquid nitrogen. If liquid nitrogen is not available, ship the EDTA 
tube to St. Jude within 24 hours of collection at room temperature.
This sample may be collected any time before or after chemotherapy. 
4.2.3
Mail all specimens to:
Charles Mullighan / Audrey Phillips
St Jude Children's Research Hospital
262 Danny Thomas Place
Pathology, Mail Stop 342, Room D4036
Memphis, TN, 38105
Email: 
. For questions, e-mail or call Dr. Mullighan, or call 
his assistant: Carolyn Reed:  T: 
, or Lab Manager: Audrey Phillips: T: 
.
5.0
DATA COLLECTION AND PROCEDURES FOR SITES PARTICIPATING IN 
BIOLOGICAL OBJECTIVES ONLY
5.1
Pre-enrollment:
Before the collaborating or enrolling affiliate sites screen or enroll a study participant, the 
site completes a RIN request form and submits it to CTO by e-mail: 
or fax 
(follow fax by a phone call to 
 
 to ensure receipt). A RIN is an eight-digit automated number beginning with an 
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“R”. Once the form is received, St. Jude will register the research participant and then 
email the RIN to the email address provided on your registration form. This will register 
the participant only, it will not enroll the participant on the study. After hours, holidays, 
and weekends the site RIN request will be answered by the Patient Registration team. 
To enroll the study participant, after the RIN is obtained, the site study team will 
complete an eligibility checklist and fax it to 
 (follow fax by a phone call to 
to ensure receipt). Clinical Trials Operations (CTO) will enter the 
Eligibility Checklist information into the central enrollment system to officially enroll the 
participant on the trial. 
Collaborating sites will scan/fax a copy of the signed informed consent document to the 
study team at 
. 
5.2
Post-enrollment:
1) The St. Jude PI and the study coordinator receive a notification of the 
enrollment completion from the St. Jude Data Warehouse Service
2) The PI completes the Registration Form and the study coordinator enters the 
information into a secured study database.
3) The research subject and the referring physician receive a copy of the signed 
consent form.
4) The referring physician sends St. Jude paraffin blocks, slides, frozen tumor, 
and blood samples if they are available and patient/parents/guardians have 
signed the consent form (Specimen Guidelines – Appendix III).
5) If the diagnosis is not confirmed, the biologic studies will not be performed, 
and the samples will be returned or discarded.
5.3
Yearly follow-up:
The St. Jude PI or designee contacts the treating physician for yearly follow up 
information.
5.4
Clinical Data
Age
Treatment regimen(s)
Race
Treatment start date
Gender
Treatment best response
Diagnosis date
Date of response
LDH
Date completed therapy
CBC with diff
Significant events (e.g., relapse, induction failure,
Diagnosis
death, cause of death, second malignancy)
Sites of disease
Date of event
Stage
Last contact date and survival status
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6.0
STATISTICAL CONSIDERATIONS
6.1
Biological Aims
The SJBC3 trial has several exploratory biological studies as its primary objectives. 
Statistical considerations on accrual, sample size, and levels of statistical errors for the 
first aim are given below. 
To perform transcriptional profiling and genome-wide analysis of DNA copy number 
abnormalities and loss-of-heterozygosity using DNA microarrays in children with newly 
diagnosed mature B cell lymphomas (e.g. Burkitt lymphoma/leukemia, DLBCL, and 
MLBCL) from different parts of the world.
Accrual. It is difficult to get a precise projection of accrual rate at this point although we 
have some expectation based on prior experience. Below we give a conservative 
projection for the biological study in terms of number of evaluable tumor samples after 5 
year accrual although we anticipate the trial may run beyond 5 years. Experience from 
the SJBCII trial indicates that the annually 6 patients can be accrued at St. Jude, giving 
approximately 30 patients after 5 years of accrual at this institution. Because there is not 
another childhood B-cell lymphoma trial currently in US, we anticipate the accrual may 
increase to 8 to 12 patients per year.  The genomic analysis requires tumor biopsy 
samples from participating international institutions along with St. Jude. The international 
institutions will participate in the biological studies and provide biopsy samples but will 
not participate in the SJBC3 therapy. We also note that it is rather difficult to obtain 
tumor biopsy samples from patients enrolled here for various logistical reasons. We 
expect that there will be about 10, 1 to 2, and 5 per year from Latin America, North 
America (St. Jude), and other geographic regions respectively. Five-year accrual can 
provide about 50, 5 to 10, and 25 tumor samples respectively. Approximately 80% of the 
tumors are expected to be Burkitts Lymphoma (BL). Moreover, we conservatively 
assume that 80% of the samples from each region are eventually evaluable for genomic 
analyses, resulting in 40, 8 and 20 evaluable tumor samples from Latin America, North 
America, and other regions respectively. Five-year accrual of evaluable tumor samples 
from each region for this aim is summarized below. 
Number of evaluable tumor samples after 5-year accrual
BL
Non-BL
Total
Latin America
32
8
40
North America (St. Jude)
6
2
8
Other
16
4
20
Total
54
14
68
At this point we set the accrual target for the biological aims as 68 evaluable tumor 
samples and anticipate completing accrual in 12 years for the biological aims. We set the 
accrual target for other aims to 100 patients treated at St. Jude and anticipate completing 
in 12 years. 
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Analyses. There will be three major components in the analyses for this objective. Each 
is discussed below, with very limited preliminary genomic data in pediatric lymphomas. 
Gene differential profiling of BL vs. non-BL. Identification of gene differential 
expression signatures requires inference (comparison) of gene expression levels in BL vs. 
non-BL. It is important to realize that the tumor samples come from dramatically 
different geographical regions where ethnic and environmental factors (e.g., diet) can 
greatly affect gene expression. Additionally, the extent of differential expression between 
BL and non-BL may also be affected by environmental, endemic and related biological 
factors (e.g., involvement of EBV in BL). Thus, it is important to adjust the gene 
expression comparison by geographic region, which serves as an aggregate of all the 
possible region-related confounding and stratification factors, in the absence of the data 
on such factors. Each participating institute defines a geographic region; from example, 
St. Jude defines the “North America” region. At this point we expect in addition to St. 
Jude, there will be one institute in Brazil (Latin America) and one in Singapore (Asia) 
participating the biological studies. Modeling, testing, and interpretation for a single gene 
(probeset) is given below, followed by methods of handling massive multiple tests. For 
samples with suitable RNA quality, gene expression profiling will be performed using 
transcriptome sequencing in preference to microarray analysis, in view of the more 
complete data provided by this platform.
The following two-factor Poisson regression model will be used for each gene, where 
E[Y(bgi)] denotes the mean expression of a gene in the i-th tumor sample from region g 
(g=Latin America, North America, etc.) and disease b (b=BL, non-BL):
Log{E[Y(bgi)]}=μ+α(b)+β(g)+αβ(g,b).
From a completely biological stand point the primary interest here is the main effect α(b). 
For each mRNA expression we will test the null hypothesis α(BL)= α(non-BL)=0 against 
the two-sided alternative at least one of them is non-zero. Statistical significance of this 
effect indicates that it is likely the gene is differentially expressed between BL and non-
BL. From a molecular epidemiology (and to a lesser extent biological) stand point the 
disease-region interaction effect αβ(g,b) is important, as it signifies that the extent of 
differential expression between BL and non-BL may be affected by geographical region. 
Statistical significance of this interaction effect indicates that it is likely the differential 
expression between BL and non-BL is different from one region to another; this would be 
an epidemiologically and biologically interesting finding as it helps to define the RNA-
level differences between endemic and non-endemic Burkitt lymphoma. 
Hence, we will be testing the main effect α(b) and the interaction effect for each RNA  
expression measurement. Because EBV positivity in BL may have an important effect on 
gene differential expression, in the final analysis we will also consider defining the 
disease main effect α(b) as a three-level factor: b = EBV+ BL, EBV- BL, non-BL.
It is imperative to assess the levels of statistical errors, especially false positives, made in 
these massive multiple tests. An assessment of the level of false negative errors with the 
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available sample size will be given in the next paragraph. Following the published 
microarray studies many of which from St. Jude, we will first assess the ensemble 
distribution of the P values and use the profile information criterion116  to obtain a data-
suggested level of statistical significance and the corresponding estimated false discovery 
rate (FDR),116,117 and then estimate the false discovery rate corresponding to a set of 
statistical significance levels specified around the data-suggested level. Estimates of FDR 
will be computed using methods in the literature.116,118,119 We will analyze further the 
“significant genes” so identified by modeling there expressions with disease subtype as 
well as potential confounding factors as explanatory variables.
Recognizing the fast changes in biotechnology, we’ll use the most effective technology 
and appropriate statistical methods at the time of final analyses.
Levels of statistical errors – an assessment by published data. All of the published 
gene expression profiling studies of mature B-cell lymphomas focused on adults, but 
Hummel et al.120 included a number of pediatric samples (age≤18 years). None of the 
published studies considered geographic region as a factor and thus there is no 
preliminary data available. Therefore, the preliminary microarray gene expression data on 
pediatric mature B-cell lymphomas are very limited. Nonetheless we provide a rough 
assessment of the levels of false positive and false negative errors that we may encounter 
in the analysis based on the pediatric data120 followed by a discussion of caveats and 
limitations of this assessment.
Here we consider the tests of the main effect α(b) for the m (≈ 22000) Affymetrix 
GeneChip probesets.  Adapting the concept of anticipated FDR (aFDR)121 to the fixed 
sample size situation and the planned analysis outlined above, we define the aFDR at a 
given significance level α as

AP
aFDR
)
1(
0
0
0









Here π0 is the proportion of true null hypotheses (non-differentially expressed genes), and 
AP is the average power which depends on the effect size, sample size, and significance 
level. The quantity (1- π0)(1-AP) is the expected proportion of true alternative hypotheses 
the multiple tests fail to capture, hence a measure of the level of false negative errors; we 
call this the false negative proportion.116 
Hummel et al. (2006) included 32 pediatric samples run on Affymetrix U133A 
GeneChip120. In these pediatric lymphoma samples 6 are traditional BL, 10 are atypical 
BL, 12 are DLBCL, and 4 are aggressive B-NHL. The 32 samples are put into two 
groups: BL (traditional and atypical BL) and non-BL (DLBCL and aggressive B-NHL). 
To get some preliminary information about π0, per-probeset comparisons of BL vs. non-
BL by the two-sample t test yield  22,215 P values, one for each probeset. The profile 
information criterion (Ip)116 produces a significance threshold (P value cut-off) to declare 
the significant tests in a way to balance the levels of false positive and false negative 
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errors, and a conservative (biased up) estimate of the corresponding FDR; it also provides 
an estimate of π0 as part of the procedure. Applying this procedure to the 22,215 P values 
yields an estimate of π0 0.6525 (65.25%), the significance threshold 0.004547, and the 
corresponding FDR 0.1377 (13.77%) among the 478 probesets declared differentially 
expressed at the 0.004547 significance level. For each probeset, the mean in the BL and 
non-BL group along with the within-group variance are computed to provide an estimate 
of the non-centrality parameter of the t test, and then is shrunk toward zero by the q-value 
weighting as described.121 The probesets are then sorted by the magnitude of the 
shrunken non-centrality parameter in descending order. To be on the conservative side, 
for each postulated null proportion (π0) in the table below, the non-centrality parameters 
of the top m(1-π0) probesets are all set to the same as the [m(1-π0)]th largest one, and the 
remaining non-centrality parameters are set to zero. Next, average power (AP) and π0 is 
estimated after this additional shrinkage, giving enlarged π0.
Based on the π0 estimate and the significance threshold suggested by Ip, we provide the 
final non-centrality after shrinkage, estimates of aFDR, the false negative proportion FNP, 
and average power (AP)  for a range of postulated and enlarged π0 values and significance 
levels (P value cutoffs) in the following table.
Anticipated False Discovery Rate and False Negative Proportion
π0
Postulated
Enlarged
Non-centrality
0.60
0.7969
3.39
0.65
0.7994
4.17
0.80
0.8469
7.57
0.90
0.9117
11.82
0.95
0.9532
15.81
0.99
0.9902
24.38
0.0010   aFDR
              FNP
              AP
0.1107
0.3874
0.0315
0.0823
0.3344
0.0443
0.0388
0.1726
0.1371
0.0306
0.0673
0.3268
0.0372
0.0236
0.5274
0.1116
0.0019
0.8079
0.0025
0.1471
0.3773
0.0568
0.1141
0.3229
0.0773
0.0621
0.1583
0.2086
0.0599
0.0564
0.4358
0.0737
0.0180
0.6405
0.2241
0.0012
0.8783
0.0045
0.1775
0.3673
0.0818
0.1417
0.3120
0.1086
0.0850
0.1464
0.2680
0.0830
0.0487
0.5134
0.1142
0.0144
0.7114
0.3330
0.0009
0.9144
0.0050
0.1836
0.3651
0.0872
0.1473
0.3096
0.1153
0.0899
0.1440
0.2798
0.0891
0.0472
0.5278
0.1235
0.0138
0.7237
0.3554
0.0008
0.9201
0.0075
0.2093
0.3555
0.1111
0.1715
0.2994
0.1444
0.1120
0.1342
0.3287
0.1171
0.0416
0.5838
0.1658
0.0115
0.7696
0.4474
0.0006
0.9396
α
0.0100
0.2298
0.3474
0.1315
0.1911
0.2909
0.1687
0.1311
0.1267
0.3667
0.1419
0.0376
0.6239
0.2030
0.0100
0.8003
0.5160
0.0005
0.9514
For example, if the proportion of non-differentially expressed probesets π0=0.65 (65%) 
and all probesets with P value ≤ α=0.0045 are declared as significantly differentially 
expressed between BL and non-BL, we anticipate that the false discovery rate 
aDFR=0.1417, i.e., 14.17% of these “significant” probesets are false positives; and that 
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the false negative proportion FNP=0.3120, i.e., we expect to miss 31.2% of the truly 
differentially expressed probesets. 
It should be noted that the non-centrality parameters estimated above in fact correspond 
to one-factor ANOVA. Unfortunately, however, there is no preliminary data on the 
second factor (geographic region). The numerator of the non-centrality parameter 
determined from the one-factor model could be too optimistic for the two-factor mode, 
but the denominator (the within-group/residual standard deviation) determined by the 
one-factor model may be larger than that from the two-factor model.
The total number of tests performed for this analysis will be over 44,000 although the 
above error assessment was done for 22,000+ tests. Note however, aFDR and FNR do not 
depend directly on the total number of tests; rather they depend directly on the proportion 
of true null hypothesis π0.
It should be noted that there will be very limited statistical power for gene expression 
profiling for BL vs. non-BL in a single geographic region, except perhaps in Latin 
America where the sample size is the largest among all regions.
Additionally, unsupervised analyses such as hierarchical clustering will be performed to 
explore novel subtypes in childhood lymphomas. Also, identified gene differential 
signatures will be compared with published results (in adult lymphomas) using methods 
such as gene set enrichment analysis.
Catalog and estimate frequencies of copy number variations in childhood 
lymphomas. There is no preliminary data on copy number variations (CNVs) in 
childhood lymphomas. A study of DLBCL in adults8 reported the prevalence of CNVs in 
certain subsets ranged roughly from 4% to 40%. So, conservatively, with 14 samples in 
the non-BL group, if a CNV is observed in 1 sample (sample proportion 0.07), then the 
exact 95% confidence interval is [0.0037, 0.3122]; if a CNV is observed in 6 samples 
(sample proportion 0.43), the exact 95% confidence interval is [0.2061, 0.6878]. 
Additionally, we will compare the prevalence of CNVs between different subtypes of 
childhood lymphomas and geographic regions with exact chi-square or Fisher’s test, 
taking population structures into account if necessary. DNA extracted from peripheral 
blood will be used as matched control tissue. Loss of heterozygosity (LOH) will be 
analyzed by comparison of SNP genotype data of the tumor sample to matched germline 
samples, or, in the case of samples lacking a matched germline sample, pools of reference 
samples already accrued at St Jude. LOH data will be analyzed using publicly available 
algorithms (e.g. dChip using the Hidden Markov Model) and alternative algorithms that 
will be tested and developed at St Jude during the course of the study. These analyses 
will complement analyses identifying copy number alteration and will permit 
identification of both deletional and copy-neutral loss-of-heterozygosity.
Integrated analysis of CNVs and gene expressions. For CNVs with a reasonable 
prevalence, we will explore the association between the identified CNVs and gene 
expressions in the study cohort. We will identify individual gene expression correlated 
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with a CNV, and a gene expression profile (signature) associated with a CNV. General 
linear models will be used to perform statistical tests, and multiple tests will be addressed 
using FDR, as described before.
To describe the types and frequency of mutations in the ARF-HDM2-TP53 pathway, in 
“non-endemic” B-cell lymphomas (United States) and those found in selected geographic 
regions of the world (Brazil, Chile, Guatemala, Honduras, El-Salvador, Singapore, and 
others).
This exploratory aim will be analyzed by enumerating the types of mutations and 
estimating the frequency of each type of mutation in each geographical region as well as 
in all regions pooled. Point estimates and 95% confidence intervals (CI) will be provided.
To describe the expression of ARF-HDM2-TP53 and PUMA-associated pathways in non-
endemic B-cell lymphomas (United States) and that found in B-cell lymphomas of other 
selected geographic regions of the world (see above)
Protein as well as mRNA expression levels of important genes on these pathways from 
various bioassays will be summarized using descriptive statistics including mean, 
standard deviation, and the five-number summary in each geographical region as well as 
in all regions pooled.
To describe the pattern and frequency of XLP gene mutations presenting with B-cell 
lymphomas in the United States and selected geographic regions.
Point estimates and 95% CIs of the frequency of XLP mutation among boys will be 
calculated in each geographical region as well as in all regions pooled. Relationships of 
XLP mutations with clinical phenotypes will be explored by appropriate statistical 
models including contingency table analysis and/or logistic regression modeling (for 
categorical/binary phenotypes), general linear models for continuous phenotypes, and 
hazard regression models for right-censored failure times.
To describe the frequency of EBV-positive Burkitt lymphomas between “non-endemic” 
and endemic B-cell lymphomas;
a. To describe the pattern of EBV protein and gene expression (e.g., EBNA 3) in 
EBV-positive lymphomas.
b. To relate EBV protein and gene expression with clinical, laboratory and outcome 
data.
It is expected that among BL, about 80% are EBV positive in endemic BL (Latin 
America, Africa) and 45% are EBV positive in non-endemic BL (US, Mideast), and 
about 80% of the available tumor samples are BL. Point estimates and 95% CIs of the 
frequency of EBV-positive BL will be calculated for each geographical region. EBV 
protein expression will be described by summary statistics including mean, standard 
deviation, and the five-number summary in each geographical region as well as in all 
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regions pooled. Relationship of EBV protein expression with clinical, laboratory, and 
outcome features will be explored using appropriate statistical methods such as general 
linear models.
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