Conditions: Neuroblastoma
Keywords: MAB 3F8 RETINOIC ACID (CIS-9 & 13) Bone Marrow 09-159
Allocation: NA | Intervention model: SINGLE_GROUP
Enrollment: 39
Healthy volunteers: no

Brief summary:
The purpose of this study is to find out what effects, good and/or bad, the combination of 3F8 and GM-CSF has on the patient and the cancer.

Antibodies are made by the body to attack tumors and to fight infections. 3F8 is the name of one kind of antibody. It is made by mice, and it can attack neuroblastoma in people. 3F8 has been used safely in many patients, and it has killed cancer cells in some patients. One way it can kill cancer cells is by causing the patient's own white blood cells to attack the cancer. Granulocytes are one kind of white blood cell. GM-CSF increases the number of granulocytes in people, and it makes the granulocytes better able to kill the cancer cells.

Detailed description:

Arms:
arm 1: This phase II, open-label, single arm trial assesses the anti-NB activity of high-dose 3F8 (80 mg/m2/day), which is used in cycles 1-2, with return to standard 3F8 dosage (20 mg/m2/day) in subsequent cycles. Clinical results will be compared to those in the predecessor trials which used only the standard 3F8 dosage. Starting with A(8), patients no longer receive high dose 3F8 but receive only standard dose 3F8 (20mg/m2/day) for all cycles.

Interventions: intervention 1: 3F8 and 13-cis-retinoic acid
Intervention descriptions: intervention 1: 3F8 is dosed at 80 mg/m2/day (cycles 1-2) or 20 mg/m2/day (cycles 3 and beyond) and infused iv over 30-90 minutes. 13-cis-retinoic acid is dosed at 160 mg/m2/day, divided into two doses, x14 days. If a dose is missed, it can be made up at the end of the cycle. It is not taken on same days as 3F8. \*High-dose 3F8 will be administered only for patients enrolled on protocol from A(0) to A(7). Starting with A(8), patients receive standard dose (20mg/m2/day) during cycles 1 and 2.

Protocol document:
Prot_SAP_000.pdf: 
Memorial Sloan-Kettering Cancer Center
IRB Protocol 
IRB#: 09-159 A(10)
Amended: 02/24/15 
 
 
 
 
 
3F8/GM-CSF Immunotherapy Plus 13-Cis-Retinoic Acid for 
Consolidation of First Remission After Non-Myeloablative Therapy in 
Patients with High-Risk Neuroblastoma: A Phase II Study 
 
MSKCC THERAPEUTIC/DIAGNOSTIC PROTOCOL 
 
 
 
Principal Investigator/Department: Brian H. Kushner, MD 
Department of Pediatrics 
 
 
Co-Principal 
Investigator(s)/Department: 
Nai-Kong V. Cheung, MD, Ph.D Department of Pediatrics 
 
 
Investigator(s)/Department: 
Ellen Basu, MD, PhD 
Kim Kramer, MD 
Shakeel Modak, MD 
Stephen Roberts, MD 
Li-Xuan Qin, Ph.D 
Department of Pediatrics 
Department of Pediatrics 
Department of Pediatrics 
Department of Pediatrics 
Department of Epidemiology 
and Biostatistics 
 
Consenting 
Professional(s)/Department: 
Ellen Basu, MD, PhD 
Brian H. Kushner, MD 
Nai-Kong V. Cheung, MD, Ph.D 
Kim Kramer, MD 
Shakeel Modak, MD 
Stephen Roberts, MD 
Department of Pediatrics 
Department of Pediatrics 
Department of Pediatrics 
Department of Pediatrics 
Department of Pediatrics 
Department of Pediatrics 
 
 
Please Note: A Consenting Professional must have completed the mandatory Human 
Subjects Education and Certification Program. 
 
 
 
 
 
 
 
 
 
 
Memorial Sloan-Kettering Cancer Center 
1275 York Ave. New 
York, NY 10021 


Amended: 02/24/15 
Memorial Sloan-Kettering Cancer Center
IRB Protocol 
IRB#: 09-159A(10)
Table of Contents
 
 
 
 
MSKCC THERAPEUTIC/DIAGNOSTIC PROTOCOL .............................................................. 0 
1.0 
PROTOCOL SUMMARY AND/OR SCHEMA................................................................ 2 
2.0 
OBJECTIVES AND SCIENTIFIC AIMS.......................................................................... 2 
3.0 
BACKGROUND AND RATIONALE............................................................................... 2 
4.0 
OVERVIEW OF STUDY DESIGN/INTERVENTION .................................................... 8 
4.1 
DESIGN .................................................................................................................. 8 
4.2 
INTERVENTION ................................................................................................... 8 
5.0 
THERAPEUTIC/DIAGNOSTIC AGENTS ....................................................................... 9 
6.0 
CRITERIA FOR SUBJECT ELIGIBILITY ..................................................................... 11 
6.1 
SUBJECT INCLUSION CRITERIA.................................................................... 11 
6.2 
SUBJECT EXCLUSION CRITERIA................................................................... 11 
7.0 
RECRUITMENT PLAN................................................................................................... 12 
8.0 
PRETREATMENT EVALUATION (TABLE 1) ............................................................ 12 
9.0 
TREATMENT/INTERVENTION PLAN ........................................................................ 13 
10.0 
EVALUATION DURING TREATMENT/INTERVENTION ........................................ 14 
11.0 
TOXICITIES/SIDE EFFECTS ......................................................................................... 16 
12.0 
CRITERIA FOR THERAPEUTIC RESPONSE/OUTCOME ASSESSMENT............... 17 
13.0 
CRITERIA FOR REMOVAL FROM STUDY ................................................................ 18 
14.0 
BIOSTATISTICS ............................................................................................................. 19 
15.0 
RESEARCH PARTICIPANT REGISTRATION AND RANDOMIZATION 
PROCEDURES............................................................................................................................. 20 
15.1 
RESEARCH PARTICIPANT REGISTRATION................................................. 20 
16.0 
DATA  MANAGEMENT ISSUES .................................................................................. 20 
16.1 
QUALITY ASSURANCE .................................................................................... 21 
16.2 
DATA AND SAFETY MONITORING ............................................................... 21 
17.0 
PROTECTION OF HUMAN SUBJECTS ....................................................................... 22 
17.1 
PRIVACY ............................................................................................................. 22 
17.2 
SERIOUS ADVERSE EVENT (SAE) REPORTING.......................................... 22 
18.0 
INFORMED CONSENT PROCEDURES ....................................................................... 24 
19.0 
REFERENCE(S)............................................................................................................... 25 


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IRB Protocol 
IRB#: 09-159A(10)
Amended: 02/24/15
- 2 - 
 
 
 
1.0 
PROTOCOL SUMMARY AND/OR SCHEMA 
 
This phase II study of the anti-GD2 murine IgG3 monoclonal antibody 3F8 combined with 
granulocyte-macrophage colony stimulating factor (GM-CSF) will assess relapse-free 
survival in patients who are in first complete/very good partial remission (CR/VGPR) but 
are at very high risk of relapse of neuroblastoma. The high-dose 3F8 usage is four fold 
greater than the standard 3F8 dosage used in prior phase II studies.  13-cis-Retinoic Acid 
will be used in this protocol after patients are evaluated for response to 3F8/GM-CSF. 
We  will  apply  real-time  quantitative  RT-PCR  methodology  targeting  GD2   synthase 
mRNA for monitoring response in bone marrow; we will test the hypothesis that early 
bone marrow response has prognostic importance. 
 
 
 
2.0 
OBJECTIVES AND SCIENTIFIC AIMS 
 
Primary Objectives 
• 
Assess the impact of high-dose 3F8/GM-CSF on relapse-free survival in patients 
in first complete or very good partial remission, but at high risk of relapse. 
 
Secondary Objectives 
• 
Apply real-time quantitative RT-PCR to test the hypothesis that the minimal 
residual disease content of bone marrow after the first treatments with 3F8/GM- 
CSF has significant prognostic impact on relapse-free survival. 
 
• 
Monitor safety of the high-dose antibody treatment, to assure no side-effects or 
noxious sequelae develop or emerge that were not seen in the prior phase I study. 
 
 
 
3.0 
BACKGROUND AND RATIONALE 
 
3.1       Disease background:  Neuroblastoma  (NB)  is  the  most  common  extracranial 
solid tumor of childhood; 50-60% of patients present with an unresectable primary tumor 
and metastases in bone marrow (BM).1   Intensive induction chemotherapy and aggressive 
surgery have improved remission rates in young patients2-4; results have been less 
impressive  in  adolescents  and  adults  in  whom  NB  is  especially  chemoresistant.5,6 
Realization of an effective strategy for eradicating minimal residual disease (MRD) has 
remained a formidable challenge.  Post-surgical use of local radiotherapy helps control 
MRD in the primary site.7  Myeloablative therapy (with stem-cell support) has been the 
most common approach for eradicating MRD in distant sites.  The long-term relapse-free 
survival rate in a recent national study was only ~20%, although the vitamin A derivative 
13-cis-retinoic  acid  helped  to  improve  overall  survival.8       These  results,  plus  the 
potentially severe toxicities of chemotherapy and radiotherapy, are compelling reasons 
for pursuing novel therapeutic approaches, including immunotherapy mediated by anti- 
GD2 monoclonal antibodies (MoAbs)9-22 which interact with granulocytes.23 


Memorial Sloan-Kettering Cancer Center
IRB Protocol 
IRB#: 09-159A(10)
Amended: 02/24/15
- 3 - 
 
 
 
3.2       Rationale for 3F8: 3F8 is a murine IgG3 MoAb that is well suited for targeted 
immunotherapy.  The reasons are several.  First, the intensive chemotherapy required to 
produce in NB patients the minimal disease state optimal for immunotherapy results in 
prolonged lymphopenia. This setting is unfavorable for active immunotherapy but allows 
passive immunotherapy since the patient is unable to reject allogeneic, xenogeneic, or 
genetically-engineered MoAbs.   Second, 3F8 recognizes the ganglioside GD2.24     This 
target antigen is highly expressed on NB (but has restricted distribution in normal human 
tissues), is not modulated from the cell surface when bound by MoAbs, and is genetically 
stable (unlike tumor-associated antigens such as immunoglobulin idiotypes on lymphoma 
cells).25-27    Third, scintigraphy using 131I-3F8 confirms that 3F8 localizes selectively to 
GD2(+) tumor deposits in patients.28    The excellent targeting potential of 3F8 was 
evidenced by its high tumor to non-tumor ratio, the high percent injected dose per gram 
uptake, and the limited, if any, nonspecific liver and spleen uptake.28,29    Fourth, 3F8 
mediates destruction in vitro, in a dose-dependent manner, of GD2(+) human tumor cells 
by human complement30 and by human lymphocytes, cultured monocytes, and 
neutrophils.31-33  Finally, the capacity of 3F8 to activate complement (C’) on NB cells 
(which lack decay accelerating factor34) raises the possibility of C’-mediated lysis in 
patients and the release of C’ fragments that may elicit an inflammatory influx of 
granulocytes capable of lysing 3F8-labeled tumor cells.  Furthermore, the deposition of 
C’ fragments C3b and iC3b on NB cells may enhance ADCC because the receptor for 
iC3b – variously called Mac-1, CR3, CD11b/CD18, or άMβ2-integrin – is a key element 
in anti-GD2 MoAb-mediated tumor-cell kill by neutrophils,35,36 which are the most 
abundant circulating class of leukocyte. 
 
3.3       Rationale for GM-CSF:  GM-CSF can amplify 3F8 anti-NB activity in patients 
via effects on granulocytes and macrophages.  Reasons for combining GM-CSF with 3F8 
include the following.  First, granulocyte production is only transiently suppressed by 
chemotherapy, and GM-CSF increases neutrophil and eosinophil production and is well 
tolerated compared to other cytokines such as interleukin-2.37  Second, granulocytes from 
patients receiving chemotherapy and normal volunteers mount ADCC against NB cells 
via non-oxidative mechanisms, and GM-CSF enhances the ADCC.36,38-42 Third, 
eosinophilic infiltration of some cancers has favorable prognostic significance, and 
eosinophils exhibit potent antitumor activity in animal models.43,44   Fourth, activated 
monocytes-macrophages efficiently phagocytose NB cells, and exposure in vitro or in 
vivo 
to 
GM-CSF 
primes 
monocytes-macrophages 
for 
greater 
antineoplastic 
cytotoxicity.33,39,45-50  Fifth, GM-CSF enhances the proliferation, maturation, and function 
of  antigen-presenting  cells,  including  antigen  processing  and  presentation  by 
macrophages  and  dendritic  cells37,51,52   -  effects  that  might  promote  induction,  or 
antitumor activity, of an idiotypic network.53-56   Finally, GM-CSF is not a growth factor 
for NB cells in vitro.57 


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IRB Protocol 
IRB#: 09-159A(10)
Amended: 02/24/15
- 4 - 
 
 
 
3.4       Rationale  for  quantitating  BM  disease  by  real-time  RT-PCR:    Curative 
strategies for stage 4 NB must encompass control of both the primary mass and distant 
disease.  Dose-intensive chemotherapy has had a favorable impact on resectability of 
primary tumors and surgical removal of all visible tumor, followed by local radiotherapy, 
reduces the risk of relapse in the primary site to <10%.4,7   Dose-intensive chemotherapy 
eliminates histologically evident BM involvement in 90% of high-risk cases.3    These 
encouraging results have shifted the focus of curative strategies to the problem of MRD – 
which is readily assessed in BM. 
 
Adjuvant treatments, such as immunotherapy with MoAbs, may make gradual and not 
quantum changes in MRD, which by definition is beyond the sensitivity of conventional 
histologic58 or radiologic techniques. The availability of more sensitive methods for 
quantitating occult tumor cells in sequential BM specimens can allow determination of 
the efficacy and optimal duration of adjuvant treatment.  The absence of detectable tumor 
cells in BM can serve as a surrogate end point for the adjuvant treatment strategy, while 
quantitative data on BM disease will help detect relapse earlier and indicate a need for a 
change in treatment. 
 
We58-65a and others66-70 have reported a series of studies aimed at improving the utility of 
BM testing, focusing on immunological and molecular methods for identifying MRD. 
The limit of detection for immunocytology is 1/105  to 1/106  cells; RT-PCR has slightly 
superior sensitivity with a range of 1/105 to 1/107 cells.  The results suggest that GD2 
synthase might serve as a sensitive marker of MRD in BM and that its level might reflect 
clinical disease status as well as correlate with histologic response to immunotherapy. 
 
Histologic examination of aspirates and biopsies from bilateral posterior iliac crests has 
been the standard method for detecting NB in BM.  We have routinely also obtained 
specimens from bilateral anterior iliac crests since sampling from multiple sites increases 
detection rates.65a    Histologic studies alone, however, yield an underestimate of BM 
involvement, as shown by immunocytology with MoAbs.68.69    For >15 years, we have 
used anti-GD2 immunocytology to enumerate NB cells in BM of patients.92     Molecular 
markers have also been tested for detecting NB.  Thus, the gene transcript for tyrosine 
hydroxylase, which is a key enzyme in the catecholamine biosynthesis pathway, is useful 
for identifying NB in BM but its expression is down-regulated in some cells.64,67   We 
found cancer-testis antigen GAGE to be comparable in sensitivity to immunocytology in 
detecting NB in BM,59 and results with GAGE proved to have prognostic significance.61 
 
Ease of use, superior sensitivity, and greater specificity prompted us to study the utility of 
real-time quantitative RT-PCR of GD2 synthase mRNA for the detection of rare NB cells 
in BM.  The marker GD2 synthase is particularly pertinent because NB cells express 
ganglioside  GD2  at  high  density,  with  little  heterogeneity  within  tumors  or  among 
patients, and this surface antigen has limited expression in normal human tissues.25-27 
The 
key 
enzyme 
required 
for 
GD2 
synthesis 
is 
GD2 
synthase 
(β1,4-N- 
acetylgalactosamineyltransferase, EC 2.4.1.92).62 


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IRB Protocol 
IRB#: 09-159A(10)
Amended: 02/24/15
- 5 - 
 
 
Peak serum 3F8 (ug/ml) 
 
In a study of 155 BM samples (100 patients), the transcript levels correlated well with the 
number of NB cells in BM as measured by immunocytology.63   Further, in a study of BM 
findings in 86 NB patients treated with 3F8 and GM-CSF, molecular responders had a 
significantly lower risk of disease progression.65    The data provide compelling support 
for the utility of this molecular approach for assessing MRD. 
 
 
3.5       Clinical experience with 3F8/GM-CSF:   Prior clinical studies of 3F8/GM-CSF 
established 1) this combination’s excellent activity against refractory BM disease (IRB 
protocols 94-018 and 03-077); 2) the safety of markedly augmented 3F8 dosing (IRB 
protocol 05-015: “3F8 antibody dose escalation plus granulocyte-macrophage colony- 
stimulating factor in high-risk neuroblastoma: A phase I trial”) 71a  compared to dosages 
derived from pilot studies in the 1980s, an important finding since in vitro studies show a 
3F8 dose-dependent lysis of NB cells (see section 3.2); and 3) the superiority of 
subcutaneous  over  intravenous  (iv)  use  of  GM-CSF,  for  both  anti-NB  effect  and 
treatment simplification (which helps patients and medical staff - and promotes wide 
availability of this treatment)71-73 
 
The long-standing standard daily dosage of 3F8 has been 20 mg/m2.  In the IRB protocol 
05-015 dose escalation phase I study, 3F8 dosages up to 160 mg/m2  were possible 
without dose-limiting toxicity, as reported71a  (That protocol also used a small dose of 
heat-modified  3F8  –  2  mg/m2,  iv  over  ~5  minutes  –  administered  just  before  the 
treatment dose, as a way to modify the major acute toxicity of pain.) For the current 
protocol, we chose daily 3F8 dosage 80 mg/m2  because in the dose escalation phase I 
study, serum levels were not significantly different at a 3F8 daily dosage of >80 mg/m2 
(see Figure 1), and excellent anti-NB activity was seen at 80 mg/m2. 
 
 
 
Figure 1: 
 
 
 
 
60 
 
50 
 
40 
 
30 
 
20 
 
10 
 
0 
0 
20 
40 
60 
80 
100 
120 
140 
160 
Dose of 3F8 (mg/m2) 


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IRB Protocol 
IRB#: 09-159A(10)
Amended: 02/24/15
- 6 - 
 
 
 
3.6    HAMA response/idiotypic network:   In a study of 34 patients with MRD treated 
with 3F8 alone, patients with a self-limited (transient) HAMA response had significantly 
better  PFS  than  patients  who  had  persistently  high  HAMA  titers  or  no  HAMA 
formation.15   This  association  of  a  transient  low  level  HAMA/anti-idiotype  (Ab2) 
antibody   response   with   better   PFS   was   a   seemingly   paradoxical   finding   since 
HAMA/Ab2 blocks 3F8 binding to tumor cells.  The unexpected observation regarding 
HAMA/Ab2 and survival led to the hypothesis of a host idiotypic network induced by 
3F8 (Ab1) and responsible for long-term tumor control.  The idiotypic network was first 
postulated by Jerne75 for immune regulation and later demonstrated in human disease 
models, including cancer patients.76-79 Anti-idiotypic antibodies (Ab2) in the HAMA pool 
may mimic antigen GD2 and stimulate an Ab3 antibody response.  We previously reported 
the association of anti-anti-idiotype (Ab3) with prolonged survival among patients who 
underwent myeloablative therapy with autologous stem-cell rescue.53 Other investigators 
have postulated the development of an idiotypic network in patients treated with 17-1A 
antibodies, where both Ab3 antibodies as well as anti-idiotypic and anti-anti-idiotypic T 
cells have been demonstrated.54,55 
 
In our study of 34 patients, consistent with the emergence of an idiotypic network, Ab3 
(anti-anti-idiotype) and Ab3' (anti-GD2) response at 6-14 months after treatment with 3F8 
alone correlated with improved PFS, and higher Ab3 titers showed a stronger correlation 
with  improved  outcome,  while  non-idiotype  antibody  responses  (anti-mouse  or  anti- 
tumor nuclear antigen) had no apparent impact on outcome.56    Since GD2 is an auto- 
antigen, induced Ab3' response represented a breakdown in self-tolerance.   We 
hypothesized  that  significant  rises  in  Ab3  or  Ab3'  would  not  be  possible  unless 
suppressor pathways were removed and naive T or B cells were allowed to repopulate. 
Following intensive chemotherapy that eliminates a large part of the lymphoid system, 
exposure to tumor-selective Ab1s (3F8) might induce unique Ab2s that could bias the 
recovering repertoire towards the GD2 antigen network.  The potential for biasing the 
immune system towards specific antigens has been well documented in murine models80 
and human disease states.81,82   If true, one would expect such an idiotypic network to be 
most successful following intensive immunosuppressive therapy, and not easily induced 
in immunocompetent patients.  In fact, we found that myeloablative therapy suppressed 
HAMA and Ab2 but was followed by the appearance of Ab3 which in turn correlated 
with prolonged survival.53,56    The prerequisite of strongly immunosuppressive therapy 
may explain why in most clinical studies of MoAbs, such an idiotypic network is not 
commonly observed.   However, when found in patients with colorectal or ovarian 
carcinoma treated with specific MoAbs, the idiotypic network gave patients superior 
survival advantage.54,55,76,77,83-87 


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IRB Protocol 
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Amended: 02/24/15
- 7 - 
 
 
 
3.7       Isotretinoin (13-cis-retinoic acid) was shown in a randomized national study to 
significantly decrease the risk of early relapse in patients treated in complete remission 
and to significantly improve overall survival.8 This agent subsequently became standard 
of care for NB patients in remission.  It will be used in this protocol after patients are 
evaluated for response to 3F8/GM-CSF – same dosage, same timing as in the predecessor 
3F8/GM-CSF (IRB 03-077) protocol. 
 
3.8       Interpretations and implications:   Various strategies have undergone clinical 
testing to promote immune-mediated attack against cancer.  However, few clinical trials 
have used MoAb-targeted immunotherapy against solid tumors in children or adults, 
especially with ADCC as the principal underlying immune cytotoxic mechanism.  Also, 
an  antineoplastic  role  for  granulocytes  has  received  scant  attention.23      A  treatment 
program combining 3F8 and GM-CSF thus represents a step towards filling a gap in the 
emerging field of immunotherapy. 
 
This phase II trial exploits recent findings that promise to optimize the anti-NB activity of 
3F8/GM-CSF – namely, high dosing of 3F8 and subcutaneous use of GM-CSF.  Further, 
RT-PCR analysis of response in BM, which has proved informative in past studies, will 
be applied to monitor anti-NB activity of the immunotherapy, with an emphasis on early 
recognition of treatment efficacy.   The studies focus on BM; findings may not reflect 
total body tumor burden.  Metastatic NB is, however, a systemic disease, and, as with 
leukemia,88 BM status appears to approximate the overall disease state.  If the RT-PCR 
method proves to be sensitive, it will provide a useful tool for measuring NB-cell content 
in BM as a patient goes through treatment.  The ability to identify patients not likely to 
benefit from further use of a given adjuvant therapy will allow a more timely switch to a 
possibly better treatment.  We hypothesize that quantifying GD2 synthase transcript levels 
will help stratify patients by tumor load, define tumor thresholds that correlate with 
favorable clinical outcome, and provide prognostic classification for patients receiving 
immunotherapy.    Risk-related  molecular  guidelines  can  be  rapidly  incorporated  into 
future treatment programs. 
 
Our  RT-PCR  method  provides  a  quantitative  measure,  such  that  the  percentage  of 
patients who achieve CR in BM, and the degree of response with 3F8/GM-CSF, can be 
calculated and compared to other adjuvant treatments.  With NB, where randomized trials 
are difficult to carry out because of the small number of affected patients, such 
quantitative analyses acquire even greater utility for rapidly identifying both the best 
agent and its optimal use in a curative treatment program. 
 
The findings regarding RT-PCR utility in the adjuvant setting, granulocyte activation, 
and idiotypic network will have general implications for MoAb-based therapeutic 
strategies against solid tumors in humans. 


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IRB Protocol 
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Amended: 02/24/15
- 8 - 
 
 
 
The ease of administration of the treatments (a subcutaneous injection of GM-CSF, 
which is given at home, and a 30-minute iv infusion of 3F8 in the outpatient clinic), plus 
their transient acute side-effects, are compatible with usage of this regimen beyond our 
hospital – a strong positive factor in furthering product development, which in fact is 
underway in a multi-institutional study of 3F8/GM-CSF using the previous 3F8 dosage. 
 
4.0 
OVERVIEW OF STUDY DESIGN/INTERVENTION 
4.1       Design 
This phase II, open-label, single arm trial assesses the anti-NB activity of high-dose 3F8 
(80 mg/m2/day), which is used in cycles 1-2, with return to standard 3F8 dosage (20 
mg/m2/day) in subsequent cycles. Clinical results will be compared to those in the 
predecessor trials which used only the standard 3F8 dosage. 
 
Starting with A(8), patients no longer receive high dose 3F8 but receive only standard 
dose 3F8 (20mg/m2/day) for all cycles. 
 
The patients are in 1st complete/very good partial remission (CR/VGPR)89 after 
conventional therapy, with no evidence of NB by standard studies, but are at high risk for 
relapse. 
 
Real-time quantitative RT-PCR63-65  will be used to assess MRD in BM.  13-cis-retinoic 
acid is started after cycle 2. 
 
4.2       Intervention 
Road Map/Schema (1st CR/VGPR after conventional therapy): A cycle consists of treatment 
with 3F8 for 5 days. GM-CSF is started 5 days in advance of each 3F8 cycle.  The break 
between end of a cycle of 3F8/GM-CSF and start of next cycle is approximately 2-to-4-weeks 
through 4 cycles; subsequent breaks are ~6-8 weeks.  13-cis-retinoic acid is started after cycle 
2. 
Cycle 1 
high-dose 3F8* + GM-CSF 
2-4 weeks off 
Cycle 2 
high-dose 3F8* + GM-CSF 
2-4 weeks off– 13-cis-retinoic acid x14 days on, x14 days off 
Cycle 3 
3F8 + GM-CSF 
2-4 weeks off– 13-cis-retinoic acid x14 days on, x14 days off 
Cycle 4 
3F8 + GM-CSF 
6-8 weeks off – 13-cis-retinoic acid x14 days on, x14 days off, x14 days on 
Cycle 5 
3F8 + GM-CSF 
6-8 weeks off – 13-cis-retinoic acid x14 days on, x14 days off, x14 days on 
Cycle 6 
3F8 + GM-CSF 
Continue with ~6-8-week breaks through 24 months from 1st dose of 3F8.  The treatment 
schedule may require minor adjustment as clinically indicated (e.g., due to PDH closure for 
holidays).  Patients can complete the missed day of 3F8 the following week, so they receive 


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a full 5-day cycle.  GM-CSF will be continued through the 3F8 make-up day at the 500 
mcg/m2 dose. 
 
*High-dose 3F8 will be administered only for patients enrolled on protocol from A(0) to 
A(7).   Starting with A(8), patients receive standard dose (20mg/m2/day) during cycles 1 
and 2. 
 
 
5.0 
THERAPEUTIC/DIAGNOSTIC AGENTS 
 
5.1       3F8 monoclonal antibody (IND number of BB-IND-8449) 
 
5.1.1    Source  and  Pharmacology:    Monoclonal  antibody  3F8  is  purified  by 
column chromatography. The final product is tested to assure that it is free of nucleic 
acid, murine viruses, bacteria, fungi, mycoplasma and pyrogens. Supplier: 3F8 is 
manufactured as an investigational agent. 
 
5.1.2    Formulation and Stability: The purified antibody is frozen at –80oC, at 2 
mg/mL 0.1 M citrate-phosphate pH 4.2 buffer in glass vials.  At –80oC it is stable for at 
least 3 years.  For iv administration, 3F8 should be diluted into 10 ml 5% human serum 
albumin and millipore (0.2 µm) filtered before use.  Route of Administration: iv infusion. 
 
5.2 
Yeast-derived human recombinant granulocyte-macrophage colony stimulating f 
actor GM-CSF; LEUKINE (sargramostim)) is produced by Berlex Laboratories, Seattle, 
WA. 
 
5.2.1    LEUKINE  is  a  sterile,  white,  preservative-free,  lyophilized  powder 
suitable for iv infusion upon reconstitution of 250 mcg vials; these vials should be 
reconstituted aseptically with 1.0 ml Sterile Water for Injection, USP (without 
preservative). The reconstituted LEUKINE solutions are clear, colorless, isotonic with a 
pH of 7.4 ± 0.3, and contain 250 mcg/ml of Sargramostim.  The single-use vial should 
not be re-entered or reused.   Do not save any unused portion for later administration. 
Vials containing 500 mcg of LEUKINE are already in solution and are multiple dose 
vials. 
 
5.2.2    During  reconstitution  the  Sterile  Water  for  Injection,  USP  should  be 
directed at the side of the vial and the contents gently swirled to avoid foaming during 
dissolution. Avoid excessive or rigorous agitation; do not shake. 
 
5.2.3   Dilution for iv infusion should be performed in 0.9% Sodium Chloride 
Injection. USP. If the final concentration of LEUKINE is below 10 mcg/ml, Albumin 
(Human), add 1 mg Albumin (Human) per l ml 0.9% Sodium Chloride Injection, USP 
(e.g. use 1 ml 5% Albumin [Human] in 50 ml 0.9% Sodium Chloride Injection, USP). 


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IRB Protocol 
IRB#: 09-159A(10)
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5.2.4 
An in-line membrane filter is not to be used for iv infusion of LEUKINE. 
 
5.2.5    LEUKINE contains no antibacterial preservative and therefore should be 
administered as soon as possible, and within 6 hours following reconstitution and/or 
dilution for iv infusion. Store LEUKINE powder, reconstituted solutions, or diluted 
solutions under refrigeration at 2-8°C (36-46°F); do not freeze or shake. LEUKINE vials 
are intended for single use only; discard any unused solution after 6 hours.  Do not use 
beyond the expiration date printed on the vial. 
5.2.6  In the absence of compatibility and stability information, no other 
medication should be added to infusion solutions containing LEUKINE.  Use only 0.9% 
Sodium Chloride Injection, USP to prepare iv infusion solutions. 
 
5.2.7    Aseptic technique should he employed in the preparation of all LEUKINE 
Solutions. To assure correct concentration following reconstitution, care should be 
exercised to eliminate any air bubbles from the needle hub of the syringe used to prepare 
the diluent. Parenteral products should be inspected visually for particulate matter and 
discoloration prior to administration whenever solution and container permit. 
 
5.3 
 
13-cis-Retinoic 
Acid 
(NDC 
#0004-0155-01,0004-0169, 
0004-0156-01- 
ISOTRETINOIN, ACCUTANE) 
 
5.3.1    Source and Pharmacology:    The exact mechanism of retinoic acid (RA)- 
induced maturation of tumor cells is not known. In NB cell lines it has been shown to 
down regulate MYCN RNA and protein expression, and such down regulation correlates 
with the ability of isotretinoin to induce tumor cell growth arrest.  Recent studies using 
gene transfection directly implicate down regulation of MYCN expression by isotretinoin 
as a key event in achieving sustained arrest of NB cell proliferation. RA also appears to 
enhance  normal  hematopoietic  differentiation  by  increasing  the  responsiveness  of 
myeloid  and  erythroid  progenitor  cells  to  the  action  of  myeloid  colony  stimulating 
activity and erythropoietin, respectively. Metabolism: RA is 99.9% bound in plasma 
(almost entirely to albumin) and has a half-life of 10-20 hours. The major metabolite is 4- 
oxoisotretinoin, and excretion is in the urine and feces. A single oral dose of 100mg/m2 
isotretinoin will produce peak plasma levels of 1-2mM. The mean peaktime as 3.2 hours 
after 80mg orally, with a terminal t½ of 10 to 20 hours. Administering 160 mg/m2/day to 
children after autologous bone marrow transplantation has been shown to achieve 13-cis- 
RA levels of 5 to 7 micromolar. 
 
5.3.2    Formulation  and  Stability:  Isotretinoin,  which  is  the  13-cis  isomer  of 
retinoic acid, will be used. This is a yellow--orange crystalline powder with a molecular 
weight of 300.44.  Isotretinoin is sensitive to light and oxygen, and so it should not be 
removed from the capsule for longer than one hour prior to administration to the patient 
and it should be kept in subdued light as much as possible. 


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5.3.3   Guidelines for Administration: PO with fat-containing food or milk to 
enhance absorption.  Administration of the entire capsule is to be encouraged and small 
children can be trained to swallow them using similar sized candy. 
 
5.3.4  Supplier: Isotretinoin is available commercially under the trade name 
ACCUTANE (Roche Laboratories) in 10mg, 20mg, and 40mg soft gelatin capsules.  See 
package insert for further information. 
 
6.0 
CRITERIA FOR SUBJECT ELIGIBILITY 
 
6.1 Subject Inclusion Criteria 
 
6.1.1 
Diagnosis  of  NB  as  defined  by  a)  histopathology  (confirmed  by  the 
MSKCC Department of Pathology), or b) BM metastases or MIBG-avid 
lesion(s) plus high urine catecholamine levels. 
 
6.1.2 
High-risk  NB  as  defined  by  risk-related treatment  guidelines1   and  the 
International NB Staging System,89 i.e., stage 4 with (any age) or without 
(>18 months of age) MYCN amplification, MYCN-amplified stage 2 or 
stage 3 (any age), or MYCN-amplified stage 4S. 
 
6.1.3 
The patients are in first CR/VGPR after conventional therapy.  They have 
no measurable MIBG-avid soft tissue tumor assessable for response. 
 
6.1.4 
Signed  informed  consent  indicating  awareness  of  the  investigational 
nature of this program. 
 
6.2 Subject Exclusion Criteria 
 
6.2.1 
Creatinine > 3.0 mg/dL 
 
6.2.2 
ALT, AST and Alkaline Phosphatase > 5.0 times the upper limit of normal 
 
6.2.3 
Bilirubin > 3.0 mg/dL 
 
6.2.4 
Patients with grade 3 or higher toxicities (using the CTCAE v4.0) related 
to cardiac, neurological, pulmonary or gastrointestinal function as 
determined by physical exam. Patients must have normal blood pressure 
for age. 
 
6.2.5 
Progressive disease 
 
6.2.6 
History of allergy to mouse proteins. 
 
6.2.7 
Active life-threatening infection. 


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6.2.8 
Human anti-mouse antibody (HAMA) titer >1000 Elisa units/ml. 
 
6.2.9 
Inability to comply with protocol requirements. 
 
 
 
7.0 
RECRUITMENT PLAN 
 
In each of the past 2 years, 46 patients were treated on the predecessor of this protocol. 
We anticipate at least maintaining this accrual rate, which will allow rapid realization of 
preliminary conclusions, with completion of the trial in 3-4 years.  Given the favorable 
prior  efficacy-toxicity  profile  with  3F8/GM-CSF,  we  expect  that  all  patients  will 
complete the trial.  Patients will be offered participation in this study by their attending 
physician in the Department of Pediatrics, Memorial Sloan-Kettering Cancer Center.  No 
patient will be identified by chart review or direct advertising.  The attending physician 
will be responsible for explaining the study, obtaining written informed consent, and 
registering the patient on study.  Patients will mainly be children and adolescents because 
of the nature of NB (90% of patients are <6 years old at diagnosis).   Patients of both 
sexes and all ethnic backgrounds are eligible for this study. 
 
 
8.0 
PRETREATMENT EVALUATION 
 
Pretreatment evaluations should be completed within 30 days of start of treatment. Refer to 
Table 1. 
 
8.1       Complete history and physical examination. 
 
8.2 
Complete blood count, serum creatinine, blood urea nitrogen, serum aspartate 
aminotransferase, serum alanine aminotransferase, and serum total bilirubin 
 
8.3       Echocardiogram. 
 
8.4       Complete extent-of-disease evaluation. 
 
8.4.1 
Urine catecholamine levels - optional. 
 
8.4.2 
BM aspirates from bilateral anterior and bilateral posterior iliac crests, and 
biopsies from any two separate sites.  The specimens are studied by: 
 
▪ 
Standard histochemical methods for the presence of tumor cells. 
 
▪ 
Immunocytology using a panel of monoclonal antibodies that react with 
antigens associated with NB (to Dr. Cheung’s lab). 


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▪ 
RT-PCR quantitation of tumor cells (to Dr. Cheung’s lab) 
 
8.4.3 
CT or MRI of primary- tumor site, plus other suspected sites of tumor. 
 
8.4.4 
Scintigraphic studies (MIBG scan90 or PET scan91). 
 
8.5 
Biologic (blood) studies for phenotypes of leukocytes, and for DNA to do HLA 
and KIR typing, FcR2A and FcR3A polymorphism (to be drawn prior to 
treatment). 
 
9.0 
TREATMENT/INTERVENTION PLAN 
 
9.1 
Schedule: For patients consented on A(0) to A(7), 3F8 is dosed at 80 mg/m2/day 
(cycles 1-2) or 20 mg/m2/day (cycles 3 and beyond) and infused iv over 30-90 minutes. 
Starting with A(8), patients receive 3F8 at 20 mg/m2/day for all cycles. To modulate pain 
from 3F8, patients are premedicated with analgesics (e.g., hydromorphone) and 
antihistamines (e.g., diphenhydramine or hydroxyzine). 3F8 is started ~1 hr after GM- 
CSF. GM-CSF is dosed at 250 mcg/m2/day from day –5 to day +1 (i.e. Wednesday to 
Tuesday is customary), and is 500 mcg/m2/day on days +2 to +4 (i.e. Wednesday to 
Friday), as in the predecessor protocols.18,74     Patients come off study if progressive 
disease occurs or if there is life-threatening grade 4 toxicity from 3F8. 
 
The break from end of a cycle to start of next cycle is approximately 2-to-4 weeks 
through 4 cycles; subsequent breaks are ~6-8 weeks.  13-cis-retinoic acid is started after 
cycle 2.  Road map/schema is in section 4.2. 
 
Patients who develop HAMA which precludes timely treatments with 3F8/GM-CSF are 
eligible to receive low-dose maintenance regimens such as irinotecan alone,92 
temozolomide  alone,93    irinotecan-temozolomide,94    or  cyclophosphamide-topotecan.95 
They can also receive anti-HAMA agents such as rituximab and cyclophosphamide. 
They resume treatment with 3F8/GM-CSF if HAMA becomes negative. 
 
9.2       3F8/GM-CSF Treatment Schedule 
 
9.2.1   Treatment Schedule Cycles 1-2*: 
 
Days -5 to -1: 
GM-CSF 250 mcg/m2/day, subcutaneously. 
Days 0 and +1:   GM-CSF 250 mcg/m2/day, subcutaneously. 
3F8 80 mg/m2/day by iv infusion over 30-90 minutes. 
Days +2 to +4:   GM-CSF 500 mcg/m2/day, subcutaneously. 


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3F8 80 mg/m2/day by iv infusion over 30-90 minutes. 
 
*For patients enrolled on protocol from A(0) to A(7) 
 
 
 
9.2.2   Treatment Schedule Cycle 3 and onwards for patients enrolled on A(0) to 
A(7) and Treatment Schedule for all Cycles for patients enrolled on A(8) and 
onwards: 
Days -5 to -1: 
GM-CSF 250 mcg/m2/day, subcutaneously. 
Days 0 and +1:   GM-CSF 250 mcg/m2/day, subcutaneously. 
3F8 20 mg/m2/day by iv infusion over 30-90 minutes. 
 
Days +2 to +4:   GM-CSF 500 mcg/m2/day, subcutaneously. 
3F8 20 mg/m2/day by iv infusion over 30-90 minutes. 
 
Note:   GM-CSF is not administered if the ANC is >20,000/ul. If HAMA develops, 
cycles are deferred until HAMA titer decreases to <1000 Elisa units/ml. 
 
9.3       13-cis-retinoic acid is dosed at 160 mg/m2/day, divided into two doses, x14 days. 
If a dose is missed, it can be made up at the end of the cycle. It is not taken on same days 
as 3F8.  The treatment can be repeated after a minimum rest period of 14 days. 13-cis- 
retinoic acid is started after cycle 2 of 3F8/GM-CSF, but it can be started sooner if early 
HAMA develops. All patients take a total of 6 cycles of 13-cis-retinoic acid. Dose 
reductions due to expected side effects of 13-cis-retinoic acid (e.g., headaches, dry skin, 
etc.) are allowed. 
 
10.0  EVALUATION DURING TREATMENT/INTERVENTION 
 
Note: If for any reason tests cannot be completed on day 0, they will be completed on day 1 
 
10.1 
CBC on day 0 and +3 of each cycle. 
 
10.2 
C3 and CH50, on day 0 and +4 (i.e. Monday and Friday) of cycles 1 and 2. 
 
10.3 
Phenotype of leukocytes, on day 0 prior to treatment and day +4 (i.e. Monday and 
Friday) of cycles 1 and 2. 
 
10.4 
Serum (2-5 ml) for 3F8 levels pre 3F8 infusion on Day 0 (i.e. Monday) and post- 
3F8 infusion on Day 4 (i.e. Friday) for cycle 1 and 2. 
 
10.5 
Blood for HAMA is obtained approximately every 1-2 months while on study (to 
Dr. Cheung’s lab). 


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10.6 
Liver and kidney function blood tests on day 0 and +4 of each cycle. 
 
10.7 
Assessment of response is the same as in prior studies,96 including the predecessor 
3F8/GM-CSF (IRB 03-077) protocol: BM studies (8.4.2) after cycle 2 and 
complete extent-of-disease evaluation (section 8.4) after cycle 4 of 3F8/GM-CSF 
and at least every 3 months through 2 years.   BM studies are repeated in 
conjunction with MIBG or PET scan (section 8.4.4) through 2 years in patients 
with history of BM or cortical bone involvement, but are repeated ~every 6 
months in other patients (e.g., patients who were stage 4 by virtue of metastases in 
distant lymph nodes, patients with MYCN-amplified stage 4S lacking BM 
involvement, or patients with MYCN-amplified stage 2 or stage 3). 
 
10.8 
Pregnancy screen (females of child-bearing age) before starting cis-retinoic acid. 
 
10.9 
Blood to study minimal residual disease (to Dr. Cheung’s lab)98 
 
10.10   Patients receiving 80 mg/m2 of 3F8 will be monitored for their blood pressure for 
at least 2 days after 3F8 until a normal blood pressure has been assured. Not 
applicable starting with A(8) 
 
Table 1. PRETREATMENT AND DURING THE STUDY EVALUATION 
WHAT? 
WHEN? 
Complete history and physical examination. 
Pretreatment evaluation 
Complete blood count 
1. Pretreatment evaluation 
2. On day 0 (i.e. Monday) and +3 (i.e. Thursday). 
Liver and kidney function blood tests 
1. Pretreatment 
2. On day 0 (i.e. Monday) and +4 (i.e. Friday) 
Blood for studies of DNA to do HLA and KIR 
typing, FcR2A and FcR3A polymorphism (to Dr. 
Cheung’s lab) 
Pretreatment or on Cycle 1 Day 0 prior to 3F8. 
Blood for studies of phenotypes of leukocytes (to Dr. 
Cheung’s lab) 
Day 0 (i.e. Monday) prior to treatment and Day 4 (i.e. Friday), 
cycles 1 and 2 
Blood (2-5 ml) for 3F8 levels (to Dr. Cheung’s lab) 
1. Pre 3F8 infusion day 0 (i.e. Monday), cycles 1  and 2 
2. Post 3F8 infusion day 4 (i.e. Friday), cycles 1 and 2 
Blood for minimal residual disease studies (to Dr. 
Cheung’s lab) 
1. Drawn with post cycle 2 bone marrow studies 
2. Drawn with bone marrows during first work-up (~3 
months from start of study) 
C3, CH50 
On day 0 and +4 (i.e. Monday and Friday), cycles 1 and 2. 
HAMA (Red Top Tube) (to Dr. Cheung’s lab) 
~Every 1-2 months while on study. 


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Bone marrow studies (including 10 ml heparinized 
for immunocytology and/or RT-PCR, to Dr. 
Cheung’s lab) 
1. Pretreatment 
2. At end of cycle 2 
3. Subsequently, 
• 
BM studies are repeated with MIBG or PET 
through 2 years for patients with history of 
BM or cortical bone involvement. 
• 
But are repeated ~every 6 months in other 
patients (e.g., patients who were stage 4 by 
virtue of metastases in distant lymph nodes) 
while on study. 
CT or MRI 
1. Pretreatment: of primary tumor site, plus other specific 
or suspected sites of tumor. 
2. Followup: of primary site ~every 3 months through 1 
year. 
MIBG scan90 or PET scan91 
Pretreatment and ~every 3 months through 2 years 
Echocardiogram 
Pretreatment 
Urine catecholamine levels (optional) 
1. Pretreatment 
2. ~Every 3 months through 2 years. 
Pregnancy screen (females of child-bearing age) 
Before starting 13-cis-retinoic acid. 
Blood pressure monitoring with 80 mg/m2  of 3F8 – 
not applicable starting with A(8) 
Measured  for  at  least  2  days  after  3F8  until  normal  blood 
pressure is assured. 
Note: If for any reason tests cannot be completed on day 0, they will be completed on day 1 
* Pretreatment evaluation should be completed within 30 days of treatment 
 
11.0 
TOXICITIES/SIDE EFFECTS 
 
11.1     Toxicities are graded by the Common Toxicity Criteria (Version 3.0) developed 
by the National Cancer Institute (NCI) of the USA. All toxicities during cycle 1 will be 
recorded. After cycle 1, only toxicities ≥ grade 3 will be recorded. 
 
11.2 
3F8: Reversible side-effects include pain, paresthesia, hypertension, hypotension, 
tachycardia, urticaria, fever, nausea, emesis, and rarely, diarrhea, serum sickness, 
hyponatremia, somnolence and posterior reversible encephalopathy syndrome (PRES). 
Other potential side-effects that may occur are bronchospasm, anaphylaxis, peripheral 
neuropathy, impaired accommodation of the eye and poor reactivity of pupils to light. 
 
11.2.1 The definition of unacceptable pain is the same as in the IRB 05-015 phase 
I 3F8 dose-escalation study, as reported71a: >7 doses of opioids administered within two 
hours. One dose of analgesic is defined as hydromorphone 0.015 mg/kg or equianalgesic 
dose of morphine sulfate. 
 
11.2.2  The definition of unacceptable hypertension is the same as in the IRB 05- 
015 phase I 3F8 dose-escalation study, as reported71a: hypertension necessitating 
medicinal intervention for >24 hours. 


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11.3     GM-CSF: Common side-effects include bone pain, flushing, local reaction at site 
of injection, leukopenia shortly after injection, and decrease in platelet count.  Rare side- 
effects (predominantly in adults) include allergic reactions, weight gain, pleural or 
pericardial effusion, pericardial embolism, thrombosis, and difficulty breathing after first 
injection. 
 
11.4     Isotretinoin: Dry skin, dry mouth, cheilitis, dry eyes, hypercalcemia, pseudotumor 
cerebri, rash, muscle or bone pain, fatigue, headache, cholesterol, nausea and vomiting, 
changes in skin color, upset stomach, dizziness, low red and white blood cells, retinoic 
acid syndrome, hepatotoxicity, teratogenic effect on fetus, depression, suicidal ideation. 
 
 
 
12.0 
CRITERIA FOR THERAPEUTIC RESPONSE/OUTCOME ASSESSMENT 
 
12.1     Response duration is calculated from first day of treatment with 3F8. 
 
12.2     Patients are considered a treatment failure if progressive disease develops. 
 
12.3     Disease status is defined by the International NB Response Criteria.89 
 
▪ 
Complete response/remission (CR): no evidence of disease by imaging 
studies and standard BM histology (section 8.4). 
 
▪ 
Very  good  partial  response/remission  (VGPR):  >90%  decrease  in  all 
disease markers, except bone scan unchanged or improved; BM must be 
free of disease by histology and by scintigraphic studies. 
 
▪ 
Partial response/remission: >50% decrease in all disease markers, except 
bone scan unchanged or improved; <1 positive BM site; MIBG scan must 
be negative or improved in all abnormal sites. 
 
▪ 
Minor response: >50% decrease in >1 but not all disease markers; MIBG 
scan must be negative or improved in at least one site. 
 
▪ 
No response: <50% decrease in all tumor markers, no change in MIBG 
scan. 
 
▪ 
Progressive disease (PD): new lesion, or >25 % increase in any disease 
marker. 
 
12.4 
All patients who fulfill the eligibility requirements and receive a first dose of 3F8 
will be included in the analysis. 


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13.0 
CRITERIA FOR REMOVAL FROM STUDY 
 
13.1     PD any time after cycle 1. However, if a patient develops recurrent disease that 
can be completely eliminated surgically or eliminated by other means, that patient may 
remain on study at the discretion of the Principal Investigator as long as the patient does 
not receive systemic therapy and the recurrence is promptly eliminated. 
 
13.2     Life-threatening grade 4 toxicity clearly attributable to 3F8. 
 
13.3     The investigators will make every reasonable effort to keep each patient in the 
study until all planned treatments and assessments have been performed.   The 
investigators may discontinue study drug treatment for the following reasons: 
 
• 
Adverse events, including unacceptable toxicity or exacerbation of underlying disease, 
associated with study drug administration and necessitating discontinuation of treatment. 
Patients who are removed from the study due to adverse events will be treated and 
followed according to established, acceptable medical practice. All pertinent information 
concerning the outcome of such treatment will be entered in the Case Report Form or on 
the Serious Event Report, as applicable. 
 
• 
Withdrawal of consent.  The patient's desire to withdraw from the study may occur at 
any time.  The investigator should carefully consider whether the patient’s withdrawal of 
consent is due to an adverse event, and if so, record the adverse event as the reason for 
withdrawal. 
 
• 
Withdrawal by the physician for clinical reasons not related to study drug treatment, for 
example, clinical need to administer a concomitant medication that is excluded by the 
protocol, in the absence of an adverse event. 
 
• 
Violation of the study protocol, including failure to return for required treatments or 
assessments.  Patients who fail to return for treatments will be withdrawn from the study 
if more than 4 scheduled doses are missed 
 
13.4     Patients will not be removed from study in order to receive radiotherapy. 
 
13.5 
Patients removed from the study will not be replaced. Every effort will be made to 
follow  these  patients,  with  extent-of-disease  evaluations  performed  at  least  every  3 
months through >2 years from study enrollment. 


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14.0 
BIOSTATISTICS 
 
The objective of this phase II trial is to assess the anti-NB activity of high-dose 3F8 plus 
GM-CSF in patients in first CR/VGPR from high-risk NB after conventional therapy. 
The 3F8 dosing is four fold greater than the standard 3F8 dosage used in prior phase II 
studies. 
 
In A(8), treatment was changed to use standard dose 3F8 (20 mg/m2/day) for all cycles 
instead of high-dose 3F8 for cycles 1 and 2.  Patients treated as such are evaluable for 
toxicity only. 
 
14.1     Primary Endpoint 
 
The primary endpoint is two-year relapse-free survival (RFS). 
 
14.2     Sample Size 
 
We will use a Simon’s two-stage design.97 The type I error rate is set as 0.1 and the type 
II error rate as 0.1. 
 
In the predecessor trial (IRB 03-077: 3F8 at standard dosage of 100 mg/m2/cycle, 
(administered initially as 10 mg/m2/day x10 days per cycle, and subsequently as 20 
mg/m2/day x5 days per cycle) combined with GM-CSF, plus 13-cis-retinoic acid, the 56 
patients treated in CR/VGPR after conventional therapy achieved 70% two-year RFS. By 
taking into account of the small sample variation, we will consider a two-year RFS rate 
0.64 as not promising and 0.80 as promising with the new treatment. Twenty-two patients 
will be accrued initially, and if there are less than 15 RFSs, further accrual to the protocol 
will be stopped and the new treatment will be declared ineffective in this protocol.   If 
there are at least 15 RFSs, then an additional 36 patients will be accrued for a total of 58. 
If at least 42 RFSs out of 58 patients, the protocol would be considered to have a positive 
result and the new treatment would be considered worthy of further testing in this patient 
population.  The  probability  of  stopping  the  study  earlier  is  57%  under  the  null 
hypothesis. 
 
14.3     Accrual Rate 
 
We expect to accrue 10-15 patients per year.   This trial can be completed within 3-4 
years. The accrual will continue after the first stage of the study, and the two-stage design 
is used to guard against many early recurrences. 


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14.4 
Secondary Endpoint 
 
The association between histologic examination and real time RT-PCR regarding BM 
status will be assessed by the Wilcoxon statistic.  The clinical significance (relapse-free 
survival) of histologic results and real time RT-PCR findings will be assessed by log-rank 
test and Cox regression model. 
 
 
 
15.0 
RESEARCH PARTICIPANT REGISTRATION AND RANDOMIZATION 
PROCEDURES 
 
15.1 
Research Participant Registration 
 
Confirm eligibility as defined in the section entitled Criteria for Patient/Subject 
Eligibility. 
 
Obtain informed consent, by following procedures defined in section entitled Informed 
Consent Procedures. 
 
During the registration process registering individuals will be required to complete a 
protocol specific Eligibility Checklist. 
 
All participants must be registered through the Protocol Participant Registration (PPR) 
Office at Memorial Sloan-Kettering Cancer Center. PPR is available Monday through 
Friday from 8:30am – 5:30pm at 646-735-8000. Registrations must be submitted via the 
PPR Electronic Registration System (http://ppr/). The completed signature page of the 
written consent/RA or verbal script/RA, a completed Eligibility Checklist and other 
relevant documents must be uploaded via the PPR Electronic Registration System. 
 
 
16.0 
DATA  MANAGEMENT ISSUES 
 
A Research Study Assistant (RSA) will be assigned to the study.  The responsibilities of 
the RSA include project compliance, data collection, abstraction and entry, data 
reporting, regulatory monitoring, problem resolution and prioritization, and coordinate 
the activities of the protocol study team. 
 
 
 
The data collected for this study will be entered into a secure database.  Source 
documentation will be available to support the computerized patient record. 


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16.1 
Quality Assurance 
 
Weekly registration reports will be generated to monitor patient accruals and 
completeness of registration data.  Routine data quality reports will be generated to assess 
missing data and inconsistencies.  Accrual rates and extent and accuracy of evaluations 
and follow-up will be monitored periodically throughout the study period and potential 
problems will be brought to the attention of the study team for discussion and action. 
 
Random-sample data quality and protocol compliance audits will be conducted by the 
study team, at a minimum of two times per year, more frequently if indicated. 
 
16.2 
Data and Safety Monitoring 
 
The  Data  and  Safety  Monitoring  (DSM)  Plans  at  Memorial  Sloan-Kettering  Cancer 
Center were approved by the National Cancer Institute in September 2001.  The plans 
address the new policies set forth by the NCI in the document entitled “Policy of the 
National Cancer Institute for Data and Safety Monitoring of Clinical Trials” which can be 
found at:  http://cancertrials.nci.nih.gov/researchers/dsm/index.html.   The DSM Plans at 
MSKCC were established and are monitored by the Office of Clinical Research.  The 
MSKCC Data and Safety Monitoring Plans can be found on the MSKCC Intranet at: 
http://inside2/clinresearch/Documents/MSKCC%20Data%20and%20Safety%20Monitori 
ng%20Plans.pdf 
 
There are several different mechanisms by which clinical trials are monitored for data, 
safety and quality.  There are institutional processes in place for quality assurance (e.g., 
protocol monitoring, compliance and data verification audits, therapeutic response, and 
staff education on clinical research QA) and departmental procedures for quality control, 
plus there are two institutional committees that are responsible for monitoring the 
activities of our clinical trials programs.  The committees: Data and Safety Monitoring 
Committee (DSMC) for Phase I and II clinical trials, and the Data and Safety Monitoring 
Board (DSMB) for Phase III clinical trials,  report to the Center’s Research Council and 
Institutional Review Board. 
 
During the protocol development and review process, each protocol will be assessed for 
it’s level of risk and degree of monitoring required.  Every type of protocol (e.g., NIH 
sponsored, in-house sponsored, industrial sponsored, NCI cooperative group, etc.) will be 
addressed and the monitoring procedures will be established at the time of protocol 
activation. 
 
Institutional monitoring plan for phase I and phase II trials: Memorial Sloan-Kettering 
Cancer Center (MSKCC) has established standard procedures for data safety monitoring 
of clinical research (see appendix).   For Phase II trials, these procedures include 
consideration of accrual rates, toxicity, adherence to dose-escalation schedules, adverse 
event notification and data recording. 


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Therapeutic responses are logged on a central database and approximately 50% are 
reviewed by an independent committee.  An annual report of the trial’s progress is sent to 
the IRB.  All of the procedures for Phase II studies outlined in MSKCC’s policy are 
applicable to the current trial and will be followed by the investigators.  The analysis of 
safety  will  include  all  patients  who  receive  at  least  one  dose  of  study  medication. 
Adverse events, including all toxic effects of treatment, will be tabulated individually, 
and summarized by body system, according to dosage of study medication (single dose as 
well as cumulative doses of 3F8 and GM-CSF), and according to severity or toxicity 
grade.  Laboratory data will be tabulated and summarized by descriptive statistics, as well 
as on the basis of MSKCC specified normal ranges. 
 
17.0 
PROTECTION OF HUMAN SUBJECTS 
 
The investigator agrees to conduct this study in accordance with the International 
Conference on Harmonization (ICH) principles of Good Clinical Practice and with the 
Declaration of Helsinki (1989).  The investigator will conduct all aspects of this study in 
accordance with all national, state, and local laws of the applicable regulatory agencies. 
 
Most patients will be children, adolescents, and young adults because of the nature of 
these tumors.  Patients of both sexes and all ethnic backgrounds are eligible for this study. 
Alternative treatments are available and will be discussed with patient or legal guardian. 
Patients are responsible for the costs of physician visits and usual (routine) laboratory 
tests, all scans, usual (standard) bone marrow tests, hospitalizations, and outpatient care, 
GM-CSF, and isotretinoin. There is no charge for 3F8. If there is an injury as a result of 
this research study, emergency care, hospitalization, and outpatient care will be made 
available by Memorial Hospital and billed to the patient as part of the medical expenses. 
No money will be provided by Memorial Hospital as compensation for research-related 
injury. 
 
17.1         Privacy 
MSKCC’s Privacy Office may allow the use and disclosure of protected health 
information pursuant to a completed and signed Research Authorization form.  The use 
and disclosure of protected health information will be limited to the individuals 
described in the Research Authorization form.  A Research Authorization form must be 
completed by the Principal Investigator and approved by the IRB and Privacy Board. 
 
17.2     Serious Adverse Event (SAE) Reporting 
 
Any SAE must be reported to the IRB/PB as soon as possible but no later than 5 calendar 
days. The IRB/PB requires a Clinical Research Database (CRDB) SAE report be 
submitted electronically to the SAE Office at sae@mskcc.org containing the following 
information: 


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Fields populated from the CRDB: 
• 
Subject’s name (generate the report with only initials if it will be sent outside of 
MSKCC) 
• 
Medical record number 
• 
Disease/histology (if applicable) 
• 
Protocol number and title 
 
Data needing to be entered: 
• 
The date the adverse event occurred 
• 
The adverse event 
• 
Relationship of the adverse event to the treatment (drug, device, or intervention) 
• 
If the AE was expected 
• 
The severity of the AE 
• 
The intervention 
• 
Detailed text that includes the following information: 
o A explanation of how the AE was handled 
o A description of the subject's condition 
o Indication if the subject remains on the study 
o If an amendment will need to be made to the protocol and/or consent form 
The PI’s signature and the date it was signed are required on the completed report. 
For IND/IDE protocols: 
The CRDB AE report should be completed as above and the FDA assigned IND/IDE 
number written at the top of the report. The report will be forwarded to the FDA by the 
Institutional SAE Manager through the IND Office. 
 
 
17.2.1  All SAEs must be entered into the CRDB SAE form page. 
 
 
SAEs are defined as grade 4 toxicities other than fatigue, weight loss or weight gain, 
anorexia, nausea, anxiety, constipation, urinary retention from opioid analgesics, 
somnolence/hallucinations/ 
disorientation/confusion/ 
agitation/anxiety 
from 
anti- 
histamine and opioid premedications, hypomagnesemia, fever, rash, dry skin from 
Accutane, urticaria from 3F8 or GMCSF, myelosuppression from the combination of 3F8 
and  GM-CSF,  pain  and  transient  hypoxia  from  opioids,  and  breath-holding  with  or 
without 
transient 
oxygen requirement. Preexisting conditions e.g. hearing loss, 
hyperbilirubinemia or elevated LFTs from TPN, alopecia are not counted as SAEs. 
Hospitalizations that arise from complications of chemotherapy are considered part of 
standard care and therefore will not be reported. 


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18.0 
INFORMED CONSENT PROCEDURES 
 
Before protocol-specified procedures are carried out, consenting professionals will 
explain full details of the protocol and study procedures as well as the risks involved to 
participants prior to their inclusion in the study. Participants will also be informed that 
they are free to withdraw from the study at any time. All participants must sign an 
IRB/PB-approved consent form indicating their consent to participate. This consent form 
meets the requirements of the Code of Federal Regulations and the Institutional Review 
Board/Privacy Board of this Center. The consent form will include the following: 
 
1.   The nature and objectives, potential risks and benefits of the intended study. 
2.   The length of study and the likely follow-up required. 
3.   Alternatives to the proposed study. (This will include available standard and 
investigational therapies. In addition, patients will be offered an option of 
supportive care for therapeutic studies.) 
4.   The name of the investigator(s) responsible for the protocol. 
5.   The right of the participant to accept or refuse study interventions/interactions and 
to withdraw from participation at any time. 
 
Before any protocol-specific procedures can be carried out, the consenting professional 
will fully explain the aspects of patient privacy concerning research specific information. 
 
In addition to signing the IRB Informed Consent, all patients must agree to the Research 
Authorization component of the informed consent form. 
 
Each participant and consenting professional will sign the consent form. The participant 
must receive a copy of the signed informed consent form. 


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19.0     REFERENCE(S) 
 
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2.         Kushner  BH,  Kramer  K,  LaQuaglia  MP,  Modak  S,  Yataghene  K,  Cheung  N-KV: 
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4.         La Quaglia MP, Kushner BH, Heller G, Bonilla MA, Lindsley KL, Cheung N-KV:  Stage 
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7. 
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Hyperfractionated  low-dose  (21  Gy)  radiotherapy  for  high-risk  neuroblastoma  following 
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8.         Matthay KK, Reynolds CP, Seeger RC, et al:  Long-term results for children with high- 
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9. 
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anti-ganglioside GD2 antibody ch14.18 in patients with neuroblastoma.  Eur J Cancer 31A:261- 
267, 1995 


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12.       Frost JD, Hank JA, Reaman GH, et al:  A phase I/IB trial of murine monoclonal anti- 
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13.       Yu AL, Batova A, Alvarado C, et al:  Usefulness of a chimeric anti-GD2 (ch14.18) and 
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16:513a, 1997 
 
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15.       Cheung N-KV, Kushner BH, Cheung IY, Canete A, Gerald W, Liu C, Finn R, Yeh SJ, 
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more than 1 year of age. J Clin Oncol 16:3053-3060, 1998 
 
16.       Cheung N-KV, Kushner BH, Yeh SJ, Larson SM:  3F8 monoclonal antibody treatment of 
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19.       Cheung N-KV, Sowers R, Vickers AJ, Cheung IY, Kushner BH, Gorlick R: FCGR2A 
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23.       Di  Carlo  E,  Forni  G,  Lollini  PL,  et  al:    The  intriguing  role  of  polymorphonuclear 
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27.     Kramer K, Gerald WL, Kushner BH, Larson SM, Hameed M, Cheung N-KV: 
Disialoganglioside GD2  loss following monoclonal antibody therapy is rare in neuroblastoma. 
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28.     Yeh SDJ, Larson SM, Burch L, Kushner BH, Finn R, Larson S, Cheung N-KV: 
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1988 


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35.       Kushner BH, Cheung N-KV:  Absolute requirement of CD11/CD18 adhesion molecules, 
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36.       Metelitsa LS, Gillies SD, Super M, et al:  Antidisialoganglioside/granulocyte macrophage 
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47.      Chachoua A, Oratz R, Hoogmoed R, et al:  Monocyte activation following systemic 
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51.       Young JW, Szabolcs P, Moore MA:   Identification of dendritic cell colony-forming units 
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57.       Foulke RS, Marshall MH, Trotta PP, Von Hoff DD: In vitro assessment of the effects of 
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78.       Fagerberg J, Frodin JE, Wigzell H, Mellstedt H:   Induction of an immune network 
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IRB Protocol 
IRB#: 09-159A(10)
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20.0 APPENDICES 
Appendix A: Patient diaries.