A Phase I/II Study of Pyrimethamine, a STAT3 Inhibitor, for the Treatment of Relapsed Chronic Lymphocytic Leukemia/Small Lymphocytic Lymphoma
Phase: PHASE1
Conditions: Chronic Lymphocytic Leukemia, Small Lymphocytic Leukemia
Sponsor: Dana-Farber Cancer Institute

 
Status Page  
 
 
PROTOCOL 09-421 
 
 
 
BIDMC 
 closed to accrual 
 
 
DFCI 
BWH 
 Open to accrual 
 
 
                                               
 
DATE POSTED:  05/10/16 
 
No new subjects may be enrolled in the cohort/arms/site(s) as 
described above.  
Any questions should be directed to the study’s Principal 
Investigator 

 
Status Page  
 
 
PROTOCOL 09-421 
 
 
 
Phase II 
 Closed to accrual 
 
 
 
 
 
 
 
                                                            Phase I 
Open to accrual 
 
 
 
Closure Effective Date:  12/20/13 
 
No new subjects may be enrolled in the site(s) as described above.  
Any questions regarding this closure should be directed to the 
study’s Principal Investigator 
 
 
 
 

Page 1 of 1 
Date Submitted: [03/10/16] 
Date Posted: [04/25/2016] 
Alert Page  
DF/HCC Protocol #:  [09-421] 
 
Revised: 07.01.13 
 
 
Dose Escalation Table 
 
Effective Date 
Dose Level/T-LGL and 
CLL/SLL 
Dose (insert units) 
Closed 
1 
12.5 mg, Once Daily 
Closed 
2 
25 mg, Once Daily 
Closed 
3 
50 mg, Once Daily 
11/10/14 
4 
100 mg Once Daily 
Not open yet 
5 
150 mg Once Daily 
 
Note: The row for the current dose level is highlighted and bolded. 

 
 
 
A Phase I/II Study of Pyrimethamine, a STAT3 Inhibitor, for the 
Treatment of Relapsed Chronic Lymphocytic Leukemia / Small 
Lymphocytic Lymphoma and T-Large Granular Lymphocytic 
Leukemia 
 
 
 
 
 
 
DF/HCC Biomedical Protocol Template: Version 9. October 18, 2016. 
 
 
Principal Investigator:   
Jennifer R Brown MD PhD 
Dana-Farber Cancer Institute 
450 Brookline Ave. 
Boston, MA 02215 
Tel 617-632-4894 
Fax 617-582-7909 
 
 
 
Collaborating Investigators:  
DFCI:  David Frank MD; Arnold S Freedman MD; David C Fisher MD; Ann S LaCasce MD; 
Eric Jacobsen MD; Philippe Armand MD; Caron Jacobson, MD; Matthew Davids, MD; Oreofe 
Odejide, MD 
 
 
Statisticians:  Lillian Werner; Donna Neuberg; Department of Biostatistics and Computational 
Biology, DFCI 
 
 
 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
SCHEMA 
 
 
Phase I Component  
 
 
 
2 or 3 
 
 
 
1 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
No  
 
 
 
 
 
 
Yes 
 
 
 
 
 
 
Subsequent Phase II Trial 
 
 
Single daily oral 50 mg dose: The MTD established in the Phase I portion was 50 mg. 
 
One cycle = 28 days 
 
Evaluation for toxicity & response 
 
Response of SD or better may remain on study indefinitely if toxicity is acceptable 
 
Progressive Disease will come off study  
 
 
 
 
Start at 
dose level 1 
Treat cohort 
(3 participants) 
Number 
of DLTs 
No further 
escalation 
Recruit 3 more 
participants 
Further 
DLTs? 
Stop 
Escalate dose 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
 
TABLE OF CONTENTS   
Page 
SCHEMA  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 i 
 
1:  OBJECTIVES  
 
 
 
 
 
 
 
 
 1 
1.1 Study Design  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 1 
1.2 Primary Objectives 
 
 
 
 
 
 1 
1.3 Secondary Objectives 
 
 
 
 
 
 1 
 
2: BACKGROUND                                                                                                               1 
2.1    Investigational Agent: Pyrimethamine                                                                     1 
2.2  Study Disease: Chronic Lymphocytic Leukemia                                                     2 
2.3  Rationale                                                                                                                   3 
2.4  Correlative Studies Background                                                                               6 
 
3: PARTICIPANT SELECTION 
 
 
 
 
 
 
 
 
 6 
3.1  Eligibility Criteria                                                                                                     6 
3.2  Exclusion Criteria                                                                                                     7 
3.3  Inclusion of Women, Minorities and Other Underrepresented Populations             8 
 
4: REGISTRATION PROCEDURES                                                                                 9 
4.1 General Guidelines for DF/HCC and DF/PCC Institutions     
 9 
4.2 Registration Process for DF/HCC and DF/PCC Institutions                                   9 
 
5:  TREATMENT PLAN                                                                                                      10 
 
5.1   Pre-treatment Criteria                                                                                              11 
 
5.2  Agent Administration                                                                                              11 
 
5.3  Definition of Dose- Limiting Toxicity                                                                    12 
 
5.4  General Concomitant Medication and Supportive Care Guidelines                       13 
 
5.5  Duration of Therapy                                                                                                14 
 
5.6  Duration of Follow Up                                                                                            15 
 
5.7  Criteria for Removal from Study                                                                            15 
 
 
6: EXPECTED TOXICITIES AND DOSING DELAY/DOSE MODIFICATIONS     15 
 
6.1  Anticipated Toxicities                                                                                             16 
6.2  Toxicity Management                                                                                             16 
6.3  Dose Modifications/Delays                                                                                     17 
 

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7: DRUG FORMULATION AND ADMINISTRATION                                                 19 
 
7.1   Pyrimethamine                                                                                                        19 
 
 
8: CORRELATIVE/SPECIAL STUDIES                                                                         20 
 
8.1  Pharmacokinetic Studies                                                                                          20 
 
8.2  Pharmacodynamic Studies                                                                                       21 
     8.3     Determination of CLL Biologic Prognostic Features                                              23 
 
9: STUDY CALENDAR                                                                                                      23 
 
10: MEASUREMENT OF EFFECT                                                                                  25 
10.1 Progression Free Survival                                                                                        26 
10.2 Anti-tumor Effect – Hematologic Tumors                                                              26 
 
 
11: ADVERSE EVENT REPORTING REQUIREMENTS                                            27 
11.1 General                                                                                                                    27 
11.2 Definitions                                                                                                               27 
11.3 Recording Adverse Events                                                                                      29 
11.4 Reporting Adverse Events                                                                                       29 
11.5 Sponsor Notification by Investigator                                                                       29 
11.6 Institutional Review Board (IRB) Notification by Investigator                              30 
11.7 Food and Drug Administration (FDA) Notification by Sponsor-Investigator        30 
11.8 Hospital Risk Management Notification by Investigator                                        30 
 
12: DATA AND SAFETY MONITORING                                                                       31 
12.1 Data Reporting                                                                                                         31 
12.2 Safety Meetings                                                                                                       31 
12.3 Monitoring                                                                                                               32 
  
13: REGULATORY CONSIDERATION                                                                         32 
13.1 Protocol Review and Amendments                                                                         32 
13.2 Informed Consent                                                                                                    32 
13.3 Ethics and Good Clinical Practice (GCP)                                                               33 
13.4 Study Documentation                                                                                              33 
13.5 Records Retention                                                                                                   34 
 
14: STATISTICAL CONSIDERATION                                                                           34 
 
14.1  Primary Objectives                                                                                                   34 
 
14.2  Sample Size and Patient Accrual                                                                             34 
14.3  Primary Analysis                                                                                                      35 
14.4  Secondary Objectives                                                                                               35 
14.5  Secondary Analysis                                                                                                  36 
14.6   Reporting and Exclusions                                                                                        36 
 
 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
 
15: PUBLICATION PLAN                                                                                                37 
 
16. REFERENCES                                                                                                             38 
 
17: APPENDICES                                                                                                                 
 
Appendix A                                                                                                                          40 
IW-CLL Indications for Treatment 
 
Appendix B                                                                                                                          41 
 
Performance Status Criteria 
 
Appendix C                                                                                                                          42 
 
Study Diary 
 
 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
CONFIDENTIAL 
This document is confidential. Do not disclose or use except as authorized. 
 
1 
1. OBJECTIVES 
1.1    Study Design 
 
This study includes a brief Phase I dose escalation to assess the tolerability of 
pyrimethamine in relapsed Chronic Lymphocytic Leukemia/Small Lymphocytic 
Lymphoma, CLL/SLL, and T-Large Granular Lymphocytic Leukemia, T-LGL, and to 
correlate dose with pharmacodynamic efficacy.  It then moves rapidly to a Phase II study 
to assess efficacy, using a Simon two stage design.  Due to PK and PD results that 
suggest that a higher dose between 100 and 150 mg may be required for optimal efficacy, 
and the absence of clear drug-related toxicity thus far, the protocol is being amended to 
continue the phase 1 dose escalation to a higher dose.  
1.2 
Primary Objectives 
Phase I:  To determine the MTD and recommended Phase 2 dose of 
pyrimethamine in relapsed CLL/SLL and T-LGL 
 
Phase II:  To determine the ORR of pyrimethamine in relapsed CLL/SLL and T-
LGL 
1.3 
Secondary Objectives 
• To assess the toxicity profile of pyrimethamine in relapsed CLL/SLL and T-
LGL, both acutely and over prolonged daily dosing. 
 
• To determine pyrimethamine levels in vivo with prolonged dosing. 
 
• To determine the progression-free survival following pyrimethamine for the 
treatment of relapsed CLL/SLL and T-LGL. 
 
• To determine whether pyrimethamine inhibits STAT3 in vivo by assessing 
downregulation of STAT3 dependent gene expression in CLL cells, and 
T-LGL and/or peripheral blood mononuclear cells. 
 
• To determine whether known prognostic factors in CLL/SLL and T-LGL 
correlate with response to pyrimethamine.  
 
• To assess the impact of CT scans on response evaluation in relapsed CLL/SLL 
and T-LGL.  
 
 
2. BACKGROUND 
2.1  
Investigational Agent: Pyrimethamine 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
2 
 Pyrimethamine (5-(4-chlorophenyl)-6-ethyl-2,4-pyrimidinediamine, Daraprim) is an oral 
folic acid antagonist used as an antiparasitic compound.  It is a reversible weak inhibitor of 
dihydrofolate reductase. Due to a differential requirement between host and parasite for nucleic 
acid precursors involved in growth, pyrimethamine is highly selective against plasmodia and 
Toxoplasma gondii. 
Pyrimethamine is well absorbed with a plasma half-life of approximately 96 hours. 
Pyrimethamine is 87% bound to human plasma proteins.  At the maximum dose tested in this 
trial, namely 50 mg/day, mean plasma pyrimethamine concentrations were 1,893+/-1,182 ng/ml1. 
As a folate antagonist, caution is recommended in patients with possible folate deficiency, such 
as individuals with malabsorption syndrome, alcoholism, or pregnancy, and those receiving 
therapy such as phenytoin or lorazepam, which may increase the risk of hepatotoxicity.  
The 50 mg/day dose maximum proposed for this trial is the recommended dose for the 
treatment of toxoplasmosis, and is routinely used for at least 1-3 weeks depending on treatment 
response.  At this dose, anorexia and vomiting may occur. Vomiting may be minimized by 
giving the medication with meals; it usually disappears promptly upon reduction of dosage. 
Doses used in toxoplasmosis may produce bone marrow suppression, which can be reversed with 
discontinuing the drug and with addition of leucovorin. Side effects include megaloblastic 
anemia, leukopenia, thrombocytopenia, pancytopenia, atrophic glossitis, hematuria, and 
disorders of cardiac rhythm.  Hypersensitivity reactions, occasionally severe (such as Stevens-
Johnson syndrome), have been reported but primarily when pyrimethamine is administered 
concomitantly with a sulfonamide.  
Due to the possibility of myelosuppression and given the patient population in this study, we 
include a brief Phase I dose escalation to assess the tolerability of pyrimethamine in relapsed 
CLL/SLL, and to correlate dose with pharmacodynamic efficacy.  The Phase 1 starting dose of 
12.5 mg. is the lowest convenient dose available and significantly lower than the usual dose used 
for infectious diseases. We then move rapidly to a Phase II study to assess efficacy, using a 
Simon two stage design.   
2.2 
Study Diseases: Chronic Lymphocytic Leukemia and T Large Granular 
Lymphocytic Leukemia  
Chronic lymphocytic leukemia (CLL / SLL) is the most common form of leukemia in the 
United States with more than 13,000 new cases per year2.  The disease incidence increases with 
age and is more common in men than women.  Although classified as a leukemia, the clinical 
behavior and treatment of CLL is more similar to many low-grade lymphomas.  Patients are 
observed until they are deemed to require therapy based on worsening symptoms or cytopenias, 
increasing lymphocytosis, lymphadenopathy or splenomegaly, or transformation of disease3.  
Therapy has been palliative, with no established therapy shown to improve survival.  
The last ten years have resulted in significant therapeutic advances, with the advent of 
chemoimmunotherapy combinations with complete remission rates approaching 50% and 
progression-free survivals of 3-5 years4-6.  However 10-20% of patients still fail to respond or 
relapse early, and all patients still relapse eventually.  For these patients, especially those who are 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
3 
fludarabine refractory with bulky adenopathy, or those with poor risk cytogenetics, there remains 
an urgent need for new more effective therapies that are less toxic.  The field has therefore turned 
to targeted drugs, designed to directly address the mechanism of carcinogenesis, in an effort to 
improve efficacy and reduce toxicity. 
 
T-LGL is an uncommon low-grade lymphoproliferative disorder that can occur as an isolated 
entity or in the context of rheumatologic or neoplastic disease.  Many patients remain 
asymptomatic and do not require treatment.  However, some patients develop anemia (either 
autoimmune or hypoproliferative), neutropenia, infection, or disease related symptoms such as 
fatigue.  Oral methotrexate and oral cyclophosphamide have been the most commonly utilized 
agents with response rates ranging from 38-64%7.  These agents are not curative and can have 
significant toxicity including myelosuppression, mucositis, and liver function abnormalities.  
Other agents that have been investigated with varying success include cyclosporine and 
alemtuzumab8,9 However, cyclosporine can cause renal insufficiency and is immunosuppressive 
while alemtuzumab is profoundly myelosuppressive and immunosuppressive and can be 
challenging to obtain commercially due to marketing restrictions. 
 
A large body of literature has emerged demonstrating the importance of STAT3 and STAT5b 
mutations in T-LGL. Most of these mutations are activating somatic mutations in the SH2 
domain of STAT3 though other STAT3 mutations have been described10,11. Therefore we 
propose amending the existing protocol to include patients with T-LGL.  We are not requiring a 
demonstrable STAT3 mutation since the presence of these mutations is not clearly known to be 
predictive of response to STAT3 inhibitors. 
 
2.3 
Rationale 
2.3.1 Biology of the STAT family of Transcription Factors 
The function of lymphoid cells is regulated by cytokines which control processes such as 
survival, self-renewal, proliferation, and differentiation.  Although cytokines activate a number 
of signaling pathways, increasing evidence suggests that STATs, for signal transducers and 
activators of transcription, play a key role in this process.  STATs are present in the cytoplasm of 
cells under basal conditions.  When stimulated by a growth factor or cytokine, STATs become 
phosphorylated on a unique tyrosine residue which triggers their dimerization.  STAT dimers 
then translocate to the nucleus where they bind to specific nine base pair sequences in the 
regulatory region of target genes, thereby activating transcription (Figure 1).  Given that STATs 
control genes that regulate critical biological functions including cell proliferation and survival, 
physiological STAT activation is rapid and transient.  Roughly one hour after STAT activation, 
the STATs have been dephosphorylated and have returned to the cytoplasm.  Although 
classically STATs are activated by tyrosine phosphorylation, certain STAT family members, 
including STAT3, have a highly conserved serine residue towards the carboxyl terminus that is 
also phosphorylated in a highly regulated manner.  Increasing evidence indicates that 
phosphorylation of this residue, serine 727, can have activating effects independent of tyrosine 
phosphorylation12,13. 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
4 
 
Given the importance of STATs in 
mediating the physiological regulation of 
these genes, it is not surprising that 
inappropriate activation of STATs occurs 
commonly in hematologic cancers.  In 
fact some abnormality in STAT signaling 
has been reported in every form of 
leukemia and lymphoma.  We had found 
that STAT3 is phosphorylated on serine 
727 in CLL cells isolated from essentially 
all patients with this disease14.  Serine 
phosphorylated STAT3 can be found in 
the nucleus, and can drive expression of 
genes regulating survival and 
proliferation15.  Furthermore, natural 
products with therapeutic activity toward 
CLL can decrease the serine 
phosphorylation of STAT316.  Genes whose expression are activated by STAT3 control critical 
cellular functions such as cell survival (e.g., Mcl-1 and  Bcl-xl), which is a major factor in both 
the pathogenesis of CLL, and its resistance to curative cytotoxic therapy17.  While continued 
STAT3 function is critical for tumor cell survival, STAT3 can be inhibited in normal cells with 
little consequence18.  Taken together, these findings suggest that STAT3 is an excellent target for 
the treatment of CLL and T-LGL. 
 
2.3.2  Identification of STAT3 inhibitors.  Given that inappropriate phosphorylation of STAT3 
is observed nearly universally in CLL and appears to contribute directly to the pathobiology of 
CLL cells, there would be great utility in the development of small molecules that could 
specifically and effectively inhibit this pathway.  Since the goal was to identify drugs that can be 
translated rapidly into human clinical trials, the Prestwick Collection, a library containing 1120 
compounds most of which have already been tested in humans and are either FDA-approved or 
known to be safe, was screened with a STAT3 inhibition assay in the laboratory of Dr. David 
Frank, Director of the Frank Laboratory, a division of the Louis B. Mayer Research Laboratories 
at DFCI.  See Section 8, Correlative/Special Studies. 
The compound that led to the greatest specific inhibition of STAT3 activity in the initial screen 
was pyrimethamine, a compound that is FDA approved for treating parasitic infections such as 
toxoplasmosis and malaria.  It is orally bioavailable and has relatively few significant side 
effects.  Given its slow elimination, it is generally dosed once daily.  Pyrimethamine shows half-
maximal activity (EC50) at approximately 1.5 µM, well within the plasma concentrations found 
with clinical use.  Pyrimethamine has no effect on STAT3 phosphorylation or nuclear 
translocation, but it appears to inhibit STAT3 localization to its cognate genomic DNA 
sequences, thereby inhibiting its function. Although pyrimethamine is known as an inhibitor of 
dihyrofolate reductase (DHFR), albeit a weak one, the potent DHFR inhibitor methotrexate did 
not inhibit STAT3 activity in this assay.  Therefore, STAT3 inhibition by pyrimethamine is 
probably distinct from its effect on DHFR.  The effect of pyrimethamine was examined in MDA-
MB-231 cells, which are characterized by constitutive STAT3 activation. Using quantitative RT-
TTCNNNAA
cytokine
Jak
Jak
STAT
ADP
ATP
STAT
P
STAT
P
STAT
P
STAT
P
STAT
P
Figure 1.  Activation of STATs in response to cytokines or 
other signals leads to the regulation of genes which play a 
key role in cellular proliferation and survival.

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
5 
PCR normalized to GAPDH, we found that 1 µM pyrimethamine, a concentration achievable in 
vivo, decreased expression of Bcl-xl and cyclin D1 by approximately 60%.  Pyrimethamine 
showed no toxicity in non-transformed hematopoietic cells, epithelial cells, or fibroblasts.   
 
2.3.3  Pyrimethamine shows therapeutic effects in CLL cells in vitro.  In model systems, 
pyrimethamine effectively inhibits STAT3-dependent gene transcription at low micromolar 
concentrations that can be achieved clinically in humans (Figure 2).  By contrast, pyrimethamine 
has no effect on gene expression driven by NF-κB or the closely related transcription factor 
STAT5. Pyrimethamine (5 µM) significantly decreased expression of a panel of STAT3-
dependent target genes, including BCL3, BCL6, cyclin D1, and SMAD7, while showing no 
effect on expression of “housekeeping” genes including tubulin and GAPDH, or NF-κB-
dependent genes including A20 and TLR4.  Dr. Frank’s lab next assessed the effect of 
pyrimethamine on the viability of primary CLL cells in culture.  As early as 48 hours after 
treatment, pyrimethamine caused a significant dose-dependent decrease in survival of CLL cells, 
beginning at concentrations as low as 1 µM, and it does so through induction of apoptosis as 
measured by annexin V staining.  Thus, pyrimethamine, a well tolerated FDA approved drug, 
inhibits STAT3 and induces apoptosis of primary CLL cells in vitro. 
 
 
2.3.4 Study Rationale 
Based on these data, we propose a clinical trial of the STAT3 inhibitor pyrimethamine in 
relapsed CLL/SLL and T-LGL.  The drug is FDA approved and has been well-tolerated in a 
wide variety of patients, suggesting that its toxicity profile is favorable.  The in vitro data suggest 
that pyrimethamine may be highly effective against CLL/SLL in vivo, warranting a clinical test.    
 
2.3.5  Results from Initial Phase 1 Study 
 
             In the first phase of this study, patients were accrued to the three planned dose levels, 
sixteen in all.  Three patients each were enrolled on cohorts 1 and 2 with no DLTs and no 
significant drug-related toxicities.  Cohort 3 enrolled 10 patients, also with no DLTs observed.  
The reason 10 patients were enrolled is that 4 were removed from study prior to one month of 
therapy and were therefore not evaluable for DLT.  Patients 7, 11, and 13 were all removed for 
early progressive disease and not evaluable for DLT.  Patient 8 was removed due to pneumonia 
that was not considered drug-related as this patient had had many prior serious pneumonias and 
the pneumonia began at the same time or even prior to study initiation.  Thus, only patients 9, 10, 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
6 
and 12 were all evaluable and had no evidence of DLT.  The cohort was then expanded to six, 
where no patients experienced DLTs. The few significant toxicities at least possibly related to 
study drug in these six patients all occurred after cycle 1, which was the DLT evaluation period. 
These toxicities did include one instance of asymptomatic rapidly reversible grade 
3 transaminitis (which occurred also in the setting of initiation of doxycycline and a change in 
type of hormone replacement therapy); transient grade 4 thrombocytopenia in a patient with 
baseline grade 3 thrombocytopenia and a history of prior stem cell transplant; and transient grade 
4 thrombocytopenia concomitant with infection.  None of these would have met DLT criteria 
even had they occurred in cycle 1.  We therefore moved ahead and opened the phase 2 portion at 
what we thought was the recommended phase 2 dose of 50 mg daily.   
 
 
Subsequent PK and PD data suggest that the 50 mg dose is just reaching the lower 
end of the efficacy range in blood in a subset of the patients.  Given that the drug was well 
tolerated and patients with toxoplasmosis normally receive significantly higher doses, we 
have decided to go back to phase 1 and do further dose escalation, in hopes of achieving 
higher drug levels which may then enhance our PD results and hopefully by extension 
efficacy.  Best response at the lower doses was prolonged stable disease (7 months in a very 
heavily pretreated patient)  
2.4 
Correlative Studies Background 
To optimally utilize pyrimethamine as a therapy for CLL, it will be essential to determine 
if pyrimethamine is, in fact, inhibiting STAT3 function in CLL cells.  If pyrimethamine is 
clinically effective, then it will be important to determine the lowest dose necessary to achieve 
STAT3 inhibition, to minimize the chance for non-specific toxicities.  If pyrimethamine does not 
show activity in these patients, then it will be essential to know whether STAT3 was successfully 
inhibited. 
 
In order to address these questions, at the time of each clinic visit, heparinized blood 
samples will be drawn for several assays.  First, pharmacokinetics will be performed on plasma 
to assess steady-state levels of pyrimethamine over time, and drug accumulation over time.  
Second, two pharmacodynamic assays will be performed.  The first will employ quantitative 
flow cytometry to assess total STAT3 levels as well as phosphorylation of STAT3 in vivo in 
CLL cells over time.  The second will use PCR to assess the expression of a panel of ten STAT3 
target genes that Dr. Frank’s lab has found to be most informative of pyrimethamine-induced 
STAT3 inhibition in CLL cells treated in vitro, as well as four genes they have found to be 
invariant which will be used for normalization.  Taken together these assays should enable us to 
determine whether adequate pyrimethamine levels were present and whether pyrimethamine 
successfully inhibited STAT3 phosphorylation and activation of target genes.    
 
Preliminary results of PK and PD analyses from the first three dose cohorts, up to 50 mg 
daily, suggest that the achieved blood levels of pyrimethamine thus far may be lower than 
required for most effective STAT3 inhibition.  For that reason we are adding dose cohorts to the 
Phase 1 study. 
 
3.  PARTICIPANT SELECTION 

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Protocol v. 8 22JAN16    
 
 
7 
 
3.1  
Eligibility Criteria 
 
Participants must meet the following criteria on screening examination to be eligible to 
participate in the study: 
 
Subjects with CLL/SLL: 
 
3.1.1  Subjects must be diagnosed with CLL / SLL (chronic lymphocytic leukemia / small 
lymphocytic lymphoma) based on the standard histologic and immunophenotypic criteria 
described in the WHO classification of lymphoid malignancies, including immunophenotypic 
confirmation that the tumor cells co-express B cell antigens CD19 / 20 and CD5.  Mantle cell 
lymphoma should be excluded based on positive staining of the tumor cells for CD23, or the 
absence of staining of the tumor cells for cyclin D1 or the absence of t(11;14).  This diagnosis 
should be confirmed at a Dana-Farber Harvard Cancer Center institution (DFCI, BWH, MGH, 
BIDMC) within approximately one month after the subject is registered. Any question on 
histology confirmation should be brought to the attention of the Principal Investigator. 
3.1.2  Participants must have measurable disease, defined as lymphocytosis ≥ 
5,000 / ul, or at least one palpable or CT measurable lesion > approximately 1.5 
cm, or bone marrow involvement > approximately 30%. 
 
3.1.3 Subjects must have relapsed after at least one prior purine analogue-containing 
regimen (fludarabine, cladribine or pentostatin), OR at least two non-purine 
analogue containing regimens.  
 
Subjects with T-LGL: 
 T-LGL judged by the investigator to require therapy based upon:   
1) Severe neutropenia (absolute neutrophil count <500/microL) 
2) Moderate neutropenia (absolute neutrophil count <1000/microL) with recurrent infections 
3) Symptomatic or transfusion dependent anemia 
4) Severe thrombocytopenia (<50,000/microL) 
5) Hepatic infiltration resulting in abnormal liver function tests 
6) Symptomatic splenomegaly 
 
Patients with previously untreated or relapsed/refractory disease will be eligible 
All Subjects: 
3.1.4  Age ≥18 years. 
 
3.1.5   Life expectancy of greater than 3 months.  
3.1.6  ECOG performance status <2 (Karnofsky >60%, see Appendix B).  

Pyrimethamine for CLL 
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8 
3.1.7 Participants must have normal organ function as defined below: 
 total bilirubin < 1.5X institutional ULN unless due to indirect 
hyperbilirubinemia related to Gilbert’s syndrome or hemolysis, or to 
disease infiltration 
 AST (SGOT)/ALT (SGPT) < 5 X institutional upper limit of normal 
 creatinine <2X institutional ULN  
3.1.8   Subjects must require treatment based on IWCLL 2008 criteria (Appendix A). 
   3.1.9   The effects of pyrimethamine on the developing human fetus are unknown. 
For this reason and because pyrimethamine is teratogenic in animals, women of child-
bearing potential and men must agree to use adequate contraception (hormonal or 
barrier method of birth control; abstinence) prior to study entry and for the duration of 
study participation. Should a woman become pregnant or suspect she is pregnant 
while participating in this study, she should inform her treating physician immediately. 
 
3.1.10  Ability to understand and the willingness to sign a written informed consent  
          document.  
3.2       Exclusion Criteria 
 
Participants who exhibit any of the following conditions at screening will not be eligible 
for admission into the study. 
3.2.1  Participants who have had chemotherapy or radiotherapy within 3 weeks 
prior to entering the study or those who have not recovered from clinically 
significant adverse events due to agents administered more than 3 weeks 
earlier. The Principal Investigator or treating investigator determine if any 
abnormal laboratory values or toxicities are due to prior agents administered vs. 
disease and if they are clinically significant. 
3.2.2  Participants may not be receiving any other study agents. 
3.2.3  Known CNS involvement with CLL or T-LGL.  
3.2.4  History of allergic reactions or sensitivity to pyrimethamine.  
3.2.5  Patients taking folic acid are eligible if the folic acid is discontinued prior to 
pyrimethamine administration and not taken for the duration of time enrolled on 
this study. 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
9 
3.2.6 
Prior allogeneic SCT is an exclusion only if the subject has active graft vs host 
disease or requires immunosuppression other than a constant stable dose of 
glucocorticoids (the latter is permitted).  
3.2.7 
Uncontrolled intercurrent illness including, but not limited to, ongoing or 
active infection not controlled by appropriate antibacterial, antiviral or 
antifungal therapy, symptomatic congestive heart failure, unstable angina 
pectoris, clinically significant cardiac arrhythmia, or psychiatric illness/social 
situations that would limit compliance with study requirements. 
3.2.8  Pregnant women are excluded from this study because pyrimethamine is a/an 
class C agent with the potential for teratogenic or abortifacient effects. Because 
there is an unknown but potential risk of adverse events in nursing infants 
secondary to treatment of the mother with pyrimethamine, breastfeeding should 
be discontinued if the mother is treated with pyrimethamine. These potential 
risks may also apply to other agents used in this study.  
3.2.9  HIV-positive individuals on combination antiretroviral therapy are ineligible 
because of the potential for pharmacokinetic interactions with pyrimethamine. 
In addition, these individuals are at increased risk of lethal infections when 
treated with marrow-suppressive therapy. Appropriate studies will be 
undertaken in participants receiving combination antiretroviral therapy when 
indicated. 
3.3    Inclusion of Women, Minorities and Other Underrepresented Populations 
The investigators will work hard to enroll women and minorities on this clinical trial.  
CLL is approximately two-fold more common in men than women but we will aim for 
representation of women at least equivalent to the balance among those diagnosed with 
the disease.  The Inclusion / Exclusion criteria are broad and should not have a negative 
impact on accrual of underrepresented populations.  
 
 
4. REGISTRATION PROCEDURES 
4.1  
General Guidelines for DF/HCC and DF/PCC Institutions 
Institutions will register eligible participants with the DF/HCC Quality Assurance 
Office for Clinical Trials (QACT) central registration system. Registration must 
occur prior to the initiation of therapy. Any participant not registered to the protocol 
before treatment begins will be considered ineligible and registration will be denied.  
 
A member of the study team will confirm eligibility criteria and complete the 
protocol-specific eligibility checklist.  

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Following registration, participants may begin protocol treatment within two weeks. 
Issues that would cause treatment delays should be discussed with the Principal 
Investigator. Minor delays due to physician clinic schedule, holidays, vacations, etc. are 
permitted.  If a participant does not receive protocol therapy following registration as 
described herein, the participant’s protocol status should be changed. Notify the QACT 
Registrar of participant status changes as soon as possible. 
 
4.2  
Registration Process for DF/HCC and DF/PCC Institutions 
The QACT registration staff is accessible on Monday through Friday, from 8:00 AM to 
5:00 PM Eastern Standard Time. In emergency situations when a participant must 
begin treatment during off-hours or holidays, call the QACT registration line at 617-
632-3761 and follow the instructions for registering participants after hours.   
The registration procedures are as follows: 
1. 
Obtain written informed consent from the participant prior to the performance 
of any study related procedures or assessments. 
2. 
Complete the protocol-specific eligibility checklist using the eligibility 
assessment documented in the participant’s medical/research record. To be 
eligible for registration to the study, the participant must meet each 
inclusion and exclusion criteria listed on the eligibility checklist unless 
granted an eligibility deviation from the Regulatory Sponsor and the IRB.  
Reminder: Confirm eligibility for ancillary studies at the same time as 
eligibility for the treatment study. Registration to both treatment and ancillary 
studies will not be completed if eligibility requirements are not met for all 
studies.  
3. 
Fax the eligibility checklist(s) and all pages of the consent form(s) to the 
QACT at 617-632-2295.  
Exception: DF/PCC Affiliate sites must fax the entire signed consent form 
including HIPAA Privacy Authorization and the eligibility checklist to the 
Network Affiliate Office. The Network Affiliate Office will register the 
participant with the QACT.  
4. 
The QACT Registrar will (a) validate eligibility, (b) register the participant on 
the study, and (c) randomize the participant when applicable.  
5. 
The QACT Registrar will send an email confirmation of the registration 
and/or randomization to the person initiating the registration immediately 
following the registration and/or randomization.  
 
 
5. TREATMENT PLAN  

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11 
 
Treatment will be administered on an outpatient basis. Expected toxicities and potential 
risks as well as dose modifications for pyrimethamine are described in Section 6 (Expected 
Toxicities and Dosing Delays/Dose Modifications). No investigational or commercial 
agents or therapies other than those described below may be administered with the intent to 
treat the participant's malignancy. 
 
Subjects will self-administer this oral drug once daily, in the morning, with food.  Dose is to be 
held until lab work results are reported on days that subjects are scheduled for clinic (i.e, Cycle 1 
Day 1, 8 and 15, Cycle 2 Day 1, etc.). Therapy will be continuous, with one cycle arbitrarily 
defined as 28 days (+/- 3 days).  All subjects will initially be treated for one 28 day cycle, 
followed by response and toxicity evaluation.  On the day of a clinic visit, subjects will delay 
taking their dose until after their laboratory tests have been drawn at clinic.  The subject may 
delay eating or eat again after the dose. This will be noted on the diary. 
 
At screening all subjects will be assessed by blood draw for folate deficiency prior to initiating 
study therapy.  Those found to be deficient will be repleted with folic acid, 1 mg. per day for at 
least 7 days, until level reaches normal range. If deficient at baseline, folate level should be 
redrawn approximately every month while on study drug, or at required visits, whichever is less.  
Once within normal range, subjects should discontinue supplemental folate administration.  
Baseline assessments will not need to be repeated if they fall out of screening window due to 
folate repletion. 
 
The initial component of this study will be a Phase I dose escalation to assess the safety of 
pyrimethamine in this patient population.  The first three dose levels below were completed with 
expansion, but the PK and PD data suggested inadequate blood drug levels.  For that reason and 
because the safety data thus far look good (see below), cohorts 4 and 5 have been added as per 
the table. 
 
 
 
 
 
 
Table 1: Dose-Escalation Schedule 
 
Dose-Escalation Schedule 
1 cycle = 28 days (+/- 3 days) 
 
Dose Level 
Dose of 
Pyrimethamine 
Starting Dose → 
1 
12.5 mg, Once Daily* 
 
2 
25 mg, Once Daily 
 
3  
50 mg, Once Daily 
 
4 
100 mg Once Daily 
 
5 
150 mg Once Daily 
 

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Following completion of the additional cohorts of the Phase 1 study, the PK, PD and safety 
data will be assessed to determine the recommended Phase 2 dose.  Any subjects who are 
continuing on study from the Phase I portion, who are deriving clinical benefit without 
significant toxicity and who are currently on a lower dose than the recommended Phase 2 
dose, may be considered for dose escalation to the newly determined Phase 2 dose, at the 
discretion of the treating physician and the Principal Investigator.  
 
*The 25 mg tablet is scored, and will be split in half either by the research nurse or 
pharmacist.   
 
5.1 
Pre-treatment Criteria 
 
5.1.1 Cycle 1, Day 1 
 
There should be no clinically significant change in the status of the patient between 
screening and C1D1. If there is, the laboratory values of the eligibility criteria 
should be rerun, and the patient approved for study start by the treating physicians 
or PI.  As per Section 9, Study Calendar, CBCD and chemistries should be drawn on 
Day 1 of each cycle. 
 
5.1.2 Subsequent Cycles 
 
All toxicities for which pyrimethamine was held must have resolved to grade 1 or 
less prior to re-initiation of study therapy.  For a detailed discussion of the 
management of hematologic toxicity please see section 6.3.1 
  
5.2 
Agent Administration  
 
  5.2.1    Pyrimethamine 
 
• Administration – Subjects will self-administer Pyrimethamine once 
daily, in the morning, with food.   
 
• Dosing – The daily dose will be determined during the Phase I study by the cohort 
to which the study is enrolling. (See Table 1: Dose-Escalation Table, Section 5.0)  
Following the determination of the Phase 2 dose, all subjects will be enrolled to 
that dose level.  
 
• Hydration – No hydration is required. *  
 
• Oral Doses – The daily dose should be administered with food.  Any doses 
vomited within one hour of administration should be retaken.  Any doses missed 
can be taken later on the same day they were missed, with food.  If a missed dose 
is not discovered until the following day, the dose should not be made up.  There 
are no foods or liquids that are contraindicated.  
 

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* Investigators should monitor patients for tumor lysis with labs twice 
per week in the first cycle and at least weekly thereafter (in the first 
cycle), and subjects will be encouraged to maintain oral hydration.  
Intravenous hydration or additional treatment for TLS should commence if 
any laboratory evidence of such is demonstrated. At a minimum, 
hyperuricemia, hydration, and electrolyte disturbances will be monitored 
closely and addressed promptly. 
 
 
5.3 Definition of Dose-Limiting Toxicity  
 
Dose limiting toxicity (DLT) will be defined as any non-hematologic toxicity of grade 3 or 
greater severity (excluding asymptomatic grade 3 laboratory abnormalities that are not life-
threatening and respond to treatment; grade 3 fatigue; grade 3 nausea, vomiting or diarrhea 
occurring without optimal prophylaxis).  Infectious toxicities will only be considered dose-
limiting if grade 4 or greater.  Any grade 4 non-hematological toxicity, as well as any 
irreversible grade 2 cardiac, renal or neurologic toxicities, will be considered dose-limiting.  
Grading of non-hematologic toxicities will be according to CTEP Active Version of the 
NCI Common Terminology Criteria for Adverse Events (CTCAE). DLT refers to toxicities 
experienced during the first cycle of treatment only.  
 
Hematologic toxicity will be graded according to the 2008 IWCLL criteria (see table in 
section 6.3.1 under dose modifications)19.  Hematologic toxicity will only be considered 
dose-limiting as follows: 
 
 
 
 
Grade 4 neutropenia persisting for at least 7 days after discontinuation of study drug 
in Cycle 1 only. Neutropenia will not considered a DLT for subjects with T-LGL. 
 
 
 
 
Grade 4 thrombocytopenia persisting for at least 7 days after discontinuation of drug 
in Cycle 1 only. 
 
Abnormal hematologic values are not noted as inclusion or exclusion criteria. Subjects 
with starting ANC  ≤ 1000 (or greater than 1000 but requiring growth factor support) or 
starting platelet count ≤20,000 will not be evaluable for hematologic toxicity or 
hematologic DLT, as per consensus recommendations for clinical trials in CLL and T-
LGL.  Determination of whether to hold pyrimethamine for hematologic toxicities will be 
determined in these patients on a case by case basis by the treating physician and the 
Principal Investigator.  As in Section 5.4 following, growth factor support and/or platelet 
transfusion are allowed at the investigator’s discretion.    
 
  Dose-limiting toxicities will be determined during the 1st 28-day cycle only. There are no 
DLT parameters post Cycle 1. 
Management and dose modifications associated with the above adverse events are 
outlined in Section 6 (Expected Toxicities and Dosing Delays/Dose Modifications). 

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Dose escalation will proceed within each cohort according to the following scheme.  
 
Number of Participants with DLT  
at a Given Dose Level 
 
Escalation Decision Rule 
 
0 out of 3 
Enter 3 participants at the next dose level. 
                                  
                                 
                                  > 2 
Dose escalation will be stopped. This dose 
level will be declared the maximally 
administered dose (highest dose 
administered). Three (3) additional 
participants will be entered at the next lowest 
dose level if only 3 participants were treated 
previously at that dose. 
 
 
 
 
1 out of 3 
Enter at least 3 more participants at this dose 
level. 
• If 0 of these 3 participants experience 
DLT, proceed to the next dose level. 
• If 1 or more of this group suffer DLT, then 
dose escalation is stopped, and this dose is 
declared the maximally administered 
dose. Three (3) additional participants 
will be entered at the next lowest dose level 
if only 3 participants were treated 
previously at that dose. 
<1 out of 6 at highest dose level below the 
maximally administered dose 
This is generally the recommended Phase 
II dose. At least 6 participants must be 
entered at the recommended Phase II 
dose. 
 
 5.4 
 General Concomitant Medication and Supportive Care Guidelines 
 
Patients will receive the following additional supportive care medications: 
 
• Allopurinol 300 mg po QD day 1-10 after initiation of study therapy.  This would 
be considered standard of care, unless the subject has an allergy. Subjects with T-
LGL are not required to receive allopurinol. 
 
• Anti-emetics may be prescribed as needed; no specific premedication is required 
or suggested.   
 
• Subjects who have been taking trimethoprim-sulfamethoxazole for PCP 
prophylaxis should be assessed for their real risk of PCP.  Due to an increased risk 
of hypersensitivity reactions and/or myelosuppression when given together with 

Pyrimethamine for CLL 
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15 
pyrimethamine, this medication should be discontinued if possible or changed to 
atovaquone or other equivalent PCP prophylaxis during the time on study therapy. 
 
• Subjects receiving acyclovir or equivalent prophylaxis should continue these 
medications.  Consideration should be given to initiating acyclovir prophylaxis at 
400 mg po TID (or equivalent) for those patients not yet receiving it. 
 
• Lorazepam given concurrently with pyrimethamine has been associated with an 
increased risk of transaminitis.  If substitution with another benzodiazepine or 
other medication is possible, then that is preferred; otherwise close monitoring is 
appropriate. 
 
• All patients requiring transfusions should receive irradiated, leukopore filtered 
blood products. 
 
• Subjects may receive neupogen, pegfilgrastim, erythropoietin or related growth 
factors at the discretion of the treating physician.  Subjects in whom study therapy 
has been held due to myelosuppression may be given a short course of leucovorin 
10 mg po QD, at the discretion of the treating physician in consultation with the 
Principal Investigator. 
• Because there is a potential for interaction of pyrimethamine with other 
concomitantly administered drugs, especially sulfonamides or 
myelosuppressive drugs, the case report form must capture the concurrent use 
of all other drugs, over-the-counter medications, or alternative therapies. 
 5.5 
Duration of Therapy 
 
 
Initial response assessment will occur after 28 days of study therapy.  Those subjects with 
stable or responding disease may continue on study indefinitely if their treating physician and  
the Principal Investigator feel that they are deriving clinical benefit.  Subjects with significant 
adverse events or progressive disease should be removed from study. 
Duration of therapy will depend on individual response, evidence of disease progression 
and tolerance. In the absence of significant adverse events, treatment may continue 
indefinitely or until one of the following criteria applies: 
• Disease progression, 
• Intercurrent illness that prevents further administration of treatment, 
• Unacceptable adverse event(s), 
• Participant decides to withdraw from the study, or 

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16 
• General or specific changes in the participant's condition or compliance 
render the participant unacceptable for further treatment in the opinion 
of the treating investigator. 
 
5.6 
Duration of Follow Up  
Participants will be followed until initiation of their next therapy or until death, 
whichever occurs first.  Their general medical condition and survival status will be 
followed for the duration of their life, even after additional therapies.  Subjects not 
presenting in clinic on a regular basis may be called by the study staff when possible. 
Participants removed from study for unacceptable adverse events will be followed until 
resolution or stabilization of the adverse event.  
 
5.7 
Criteria for Removal from Study  
Participants will be removed from study when any of the criteria listed in Section 
5.5 apply.  The reason for study removal and the date the participant was 
removed must be documented in the study-specific case report form (CRF). 
Alternative care options will be discussed with the participant. 
In the event of unusual or life-threatening complications, participating 
investigators must immediately notify the Principal Investigator (or Protocol 
Chair), Jennifer R Brown MD PhD at 617-632-4894. 
 
6. EXPECTED TOXICITIES AND DOSING DELAYS/DOSE MODIFICATIONS  
 
Dose delays and modifications will be made using the following recommendations, and as per 
Table 2, Section 6.3. Toxicity assessments for non-hematologic toxicities will be done using 
CTEP Active Version of the NCI Common Terminology Criteria for Adverse Events (CTCAE) 
which is identified and located on the CTEP website at: http://ctep.cancer.gov/protocol 
Development/electronic_applications/ctc.htm.   Please see below for appropriate evaluation of 
hematologic toxicities, which will be according to the IW-CLL 2008 criteria.   
 
If possible, symptoms should be managed symptomatically. In the case of toxicity, appropriate 
medical treatment should be used (including anti-emetics, anti-diarrheals, etc.). 
 
All adverse events experienced by participants will be collected from the time of the first dose of 
study treatment, through the study and until the final study visit. Participants continuing to 
experience toxicity at the off study visit may be contacted for additional assessments until the 
toxicity has resolved or is deemed irreversible.  
 
 

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6.1 Anticipated Toxicities  
A list of the adverse events and potential risks associated with the agents administered 
in this study appear below and will determine whether dose delays and modifications 
will be made or whether the event requires expedited reporting in addition to routine 
reporting.  
 
6.1.1  
Adverse Event Lists(s) for  Pyrimethamine  
Hypersensitivity reactions, occasionally severe (such as Stevens-Johnson syndrome, toxic 
epidermal necrolysis, erythema multiforme, and anaphylaxis), and hyperphenylalaninemia, 
can occur particularly when pyrimethamine is administered concomitantly with a 
sulfonamide. Anorexia and vomiting may occur. Vomiting may be minimized by giving the 
medication with meals; it usually disappears promptly upon reduction of dosage. Higher 
doses may produce megaloblastic anemia, leukopenia, thrombocytopenia, pancytopenia, 
atrophic glossitis, hematuria, and disorders of cardiac rhythm. Hematologic effects, 
however, may also occur at low doses in certain individuals.  Pulmonary eosinophilia has 
been reported rarely. 
6.2 Toxicity Management 
 
 
Following the ingestion of 300 mg or more of pyrimethamine, gastrointestinal and/or 
central nervous system signs may be present, including convulsions. The initial 
symptoms are usually gastrointestinal and may include abdominal pain, nausea, severe 
and repeated vomiting, possibly including hematemesis. Central nervous system toxicity 
may be manifest by initial excitability, generalized and prolonged convulsions which may 
be followed by respiratory depression, circulatory collapse, and death within a few hours. 
Neurological symptoms appear rapidly (30 minutes to 2 hours after drug ingestion), 
suggesting that in gross overdosage pyrimethamine has a direct toxic effect on the central 
nervous system.  The fatal dose is variable, with the smallest reported fatal single dose 
being 375 mg. There are, however, reports of pediatric patients who have recovered after 
taking 375 to 625 mg. 
 
There is no specific antidote to acute pyrimethamine poisoning. In the event of 
overdosage, symptomatic and supportive measures should be employed. Gastric lavage is 
recommended and is effective if carried out very soon after drug ingestion. Parenteral 
diazepam may be used to control convulsions. Folinic acid should also be administered 
within 2 hours of drug ingestion to be most effective in counteracting the effects on the 
hematopoietic system. Due to the long half-life of pyrimethamine, daily monitoring of 
peripheral blood counts is recommended for up to several weeks after the overdose until 
normal hematologic values are restored. 
6.3 Dose Modifications/Delays 
As noted above, an individual patient who experiences DLT will have therapy held until 
resolution to grade 1 or less, and may then resume therapy at one dose level below their 

Pyrimethamine for CLL 
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18 
previous level at the discretion of the investigator and treating physician.  If the toxicity 
persists at >grade 1 for more than 4 weeks, the subject must be removed from the study.    
 
In the Phase 2 study, all grade 3 (excluding asymptomatic laboratory abnormalities, or 
grade 3 nausea, vomiting or diarrhea readily controlled with appropriate supportive care) 
or grade 4 non-hematologic toxicities will result in at least temporary discontinuation 
from study drug.  When the toxicity resolves to grade 1 or less, the treating physician and 
the Principal Investigator may make a decision about the appropriateness of continuing 
the patient on study.  Toxicity grading is based on the CTEP Active Version of the NCI 
Common Terminology Criteria for Adverse Events (CTCAE)  non-hematologic 
toxicities.  Any non-hematologic toxicity (excluding asymptomatic laboratory 
abnormalities, or grade 3 nausea, vomiting or diarrhea readily controlled with appropriate 
supportive care) that persists at > grade 1 for more than four weeks will result in 
permanent discontinuation from the study.  
 
 
Table 2: Dose-Modification  
 
 
Dose-Modification 
 
 
Dose Level 
Reduce Dose To:  
Starting Dose 
150 mg 
100 mg 
 
100 mg 
50 mg 
 
50 mg 
25 mg 
 
25 mg 
12.5 
 
12.5 
Hold 
 
 
Subjects removed from the study for toxicity will be followed until initiation of next 
therapy or death.   
 
 
6.3.1 Hematologic Toxicity 
 
 
 
Hematologic toxicity will be graded according to IWCLL 2008 criteria19, as enumerated 
in the table below.   
 
ANC / µl  
 
Toxicity Grade 
Decrease in Plts from Pre-Tx 
Value 
(Use this correction if starting plt 
count < 100,000) 
>= 2,000 
0 
0-10% 
>= 1,500 - 2000 
1 
11-24% 
>= 1,000 – 1,500 
2 
25-49% 

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19 
>= 500 – 1,000 
3 
50-74% 
< 500 
4 
>=75% 
 
 
Subjects will not be evaluable for hematologic toxicity, hematologic dose-limiting 
toxicity or holding of therapy based on low ANC if the starting ANC < 1000 or the 
starting ANC is greater than 1000 with the assistance of growth factors.  The ANC may 
be supported by growth factors throughout study therapy for this group of patients.  In 
particular, LGL patients with pre-existing neutropenia will not be evaluable for toxicity 
related to neutropenia and may continue study drug at their assigned dose regardless of 
neutrophil count. 
 
Subjects with baseline platelets < 20,000 will not be evaluable for platelet toxicity. 
Subjects with baseline platelets ≥100,000 will be evaluated per CTEP Active Version of 
the NCI Common Terminology Criteria for Adverse Events (CTCAE); i.e, do not use the 
correction in the table above for assessing platelet toxicity when baseline platelets 
≥100,000. When baseline and Cycle 1 Day 1 ANC and/or platelets differ, the lower of the 
two values should be used for assessing DLT, dose modifications and hematologic 
toxicity. 
For subjects with initial ANC =/>1000, neutropenia grading will follow the table above 
and development of grade 4 neutropenia will result in temporary discontinuation of study 
drug.  Consideration may be given to administration of folic acid, low-dose leucovorin or 
growth factors to these patients.  Once recovery occurs subjects may be restarted at their 
previous dose.  Those whose grade 4 ANC persists for at least 7 days despite these 
supportive care measures will be considered to have suffered dose limiting toxicity if this 
happens in the first cycle, only.   If these subjects experiencing DLT are continued on 
study after resolution, they should undergo a one dose level reduction in dose.  Although 
NOT considered a DLT in cycle 2 and beyond, if a subject experiences grade 4 ANC that 
persists for at least 7 days despite the supportive care measures listed above, the subject 
may  undergo a one dose level reduction in dose at investigator discretion  
 
Thrombocytopenia will be graded based on the table above, using adjusted parameters for all 
subjects with a starting platelet count < 100,000.  For those subjects with a starting platelet 
count =/> 100,000, development of thrombocytopenia will be graded based on CTEP Active 
Version of the NCI Common Terminology Criteria for Adverse Events (CTCAE).  Any 
subject with a starting platelet count < 20,000 will not be evaluable for hematologic toxicity, 
hematologic dose-limiting toxicity or holding of therapy based on thrombocytopenia.  For 
any subject with a starting platelet count > 20,000, development of a platelet count < 20,000 
is considered grade 4 thrombocytopenia and will result in holding of study drug.  
Consideration may be given to administration of folic acid or low-dose leucovorin.  
Persistence of grade 4 thrombocytopenia for > 7 days despite these supportive care measures 
will be considered a DLT if this happens in the first cycle.  If these subjects experiencing 
DLT are continued on study after resolution within one month, a dose reduction by one dose 

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20 
level per the dose escalation table is required. If grade 4 thrombocytopenia persists for > 7 
days in cycle 2 or beyond, consideration may be given to a one dose level reduction, at the 
discretion of the Principal Investigator and treating investigator.  
 
 
6.3.2 Rash 
 
For grade 3 or greater skin rash, study drug should be discontinued.  If the rash resolves 
(back to grade 1 or less, within 4 weeks) with supportive care and is not IgE mediated or 
accompanied by other signs of allergic hypersensitivity, patients may resume study 
therapy at one lower dose level, or if already at the lowest dose level, may resume at that 
dose with addition of prophylactic antihistamine.  Consideration may be given to use of a 
prophylactic antihistamine, particularly diphenhydramine, even in patients resuming at a 
reduced dose.   
 
6.3.3   Anorexia/Vomiting: 
 
For grades 1-4 anorexia/vomiting, study drug should be continued with initiation of 
antiemetic therapy.  If drug needs to be held until patient can tolerate drug with 
supportive care, patient may resume at same dose level.  If antiemetics do not bring relief, 
then the dose should be reduced one level. Patients with grade 3 or 4 anorexia/vomiting 
that do not respond to supportive care should be discontinued from study drug, but will 
remain on study for follow-up procedures until initiation of another therapy. 
 
7. DRUG FORMULATION AND ADMINISTRATION  
7.1 Pyrimethamine 
7.1.1 Description 
 
The chemical name of pyrimethamine is 5-(4-chlorophenyl)- 6-ethyl-2,4-
pyrimidinediamine.  It acts as a folate antagonist and a STAT3 inhibitor.   
Pyrimethamine is well absorbed orally with peak levels occurring between 2 to 6 hours 
following administration. It is eliminated slowly and has a plasma half-life of 
approximately 96 hours. Pyrimethamine is 87% bound to human plasma proteins. 
7.1.2 Form 
 
Pyrimethamine is supplied as 25 mg tablets.  Depending on dose level, 0.5, 1 or 2 tablets 
will be taken by mouth once per day while on study.  Do not crush or chew.  *The 25 mg 
tablet is scored, and will be split in half either by the research nurse or pharmacist.   
 
7.1.3 Storage and Stability 

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21 
Pyrimethamine is stored at room temperature and protected from light. 
7.1.4 Availability 
The agent is commercially available, but will be provided free of charge to patients 
enrolled on this protocol. 
7.1.5  Ordering 
The agent will be ordered by the Investigational Pharmacy who will maintain adequate 
stocks for this protocol. 
7.1.6 Accountability 
The investigator, or a responsible party designated by the investigator, must maintain a 
careful record of the inventory and disposition of the agent using the NCI Drug 
Accountability Record Form (DARF) or another comparable drug accountability form. 
(See the CTEP home page at http://ctep.cancer.gov for the Procedures for Drug 
Accountability and Storage or to obtain a copy of the DARF.) 
7.1.7   Destruction and Return 
 
Study drug for daily oral administration will be supplied by the DFCI pharmacy as 25 mg 
tablets in bottles.  Drug accountability will be performed as per current DF/HCC SOP All 
unused study drug should be returned by the patient to the Investigator or research 
nurse/PA. 
At the end of the study, unused supplies of pyrimethamine should be destroyed 
according to institutional policies. Destruction will be documented in the Drug 
Accountability Record Form.  
 
8. CORRELATIVE/SPECIAL STUDIES  
8.1  
Pharmacokinetic Studies 
 
 
Pharmacokinetics will be performed to assess pyrimethamine accumulation over time, 
given the long half-life of the drug.  1 heparin-anticoagulated green top tube will be 
drawn prior to therapy, prior to dose on one of days 2-5 (optional) and prior to dose on 
each subsequent clinic visit (weekly during cycle 1, every other week during cycle 2, 
and monthly thereafter until discontinuation from study drug.  Pharmacokinetics will 
also be drawn at time of off study visit and once again 2-4 weeks later if the patient has 

Pyrimethamine for CLL 
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22 
not started additional therapy. These samples will be collected at all of the above 
timepoints and transferred for processing and freezing. 
 
PK samples are drawn, processed and stored as follows: 
 
The Frank Laboratory – a division of  
Louis B. Mayer Research Laboratories 
David Frank, MD, Laboratory Director 
Mayer 5-522B 
Dana-Farber Cancer Institute 
44 Binney St. 
Boston, MA  02115 
 
 
The frozen samples will be processed in batches by the DF/HCC Core Pharmacokinetics 
laboratory run by Dr. Jeff Supko: 
 
Clinical Pharmacology Lab 
Jeffrey Supko, MD, Director 
Gray / Jackson 
Massachusetts General Hospital 
55 Fruit St. 
Boston, MA  02114 
8.2 
Pharmacodynamic Studies 
Pharmacodynamic studies will be run in Dr. Frank's lab.  
 
Two 6 cc heparin-anticoagulated green top tubes (or equivalent) will be collected and 
transferred to Dr. David Frank’s lab on Mayer 5 on the following days/times: 
-Screening  
-Pre and two hours post drug on day 1 of cycle 1 
-One day among Days 2-5 therapy (optional) 
-Day 8, 15, 22 cycle 1  
-Day 1 and 15 cycle 2    
-Day 1 of subsequent cycles up to cycle 6  
-Off study visit 
-2-4 weeks after off study visit (if subsequent therapy has not begun) 
-3 month follow up appointment (if subsequent therapy has not begun) 
 
As outlined above, prior to initiating therapy, and at the time of each subsequent clinic visit 
while on study, patients will have approximately 10 ml of blood drawn into a heparinized 
tube.  Mononuclear cells will be isolated by Ficoll-based density separation, and the 
proportion of CLL cells will be determined by flow cytometry.  If greater than 90% of the 
isolated cells are CLL cells (CD5+CD19+CD20+), these will be used for subsequent 
analyses.  If less than 90% display a CLL phenotype, then we will use antibody-based 

Pyrimethamine for CLL 
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23 
magnetic cell separation to increase the purity to this level.  It will be necessary to isolate a 
minimum of 5 x 106 cells for the required analyses. 
 
Cells will be analyzed in two ways in parallel, as follows.   
 
8.2.1  Laboratory Correlative Studies:  Determination of STAT3 Inhibition In Vivo 
 
8.2.1.1  STAT3 Phosphorylation 
 
An aliquot of 1 x 106 cells will be used for a quantitative flow cytometric assay to 
measure total STAT3 and serine phosphorylation of STAT3.  Cells will be fixed, 
permeabilized, and stained with the specific antibodies, employing a technique we 
have used successfully on clinical CLL specimens in the past 20.  This analysis will 
indicate the concordance of normalized expression of STAT3 target genes (as 
described below) with the level and phosphorylation of STAT3.  In addition, it will 
allow a longitudinal analysis of changes in the quantity and phosphorylation state of 
STAT3 in CLL cells of patients on pyrimethamine.  If pyrimethamine is decreasing 
the survival of CLL cells in these patients, then it will be expected that the total 
number of CLL cells will decline over time.  Furthermore, if CLL cells persist, it will 
allow a determination of whether a change in the proportion of CLL cells with 
activated STAT3 is occurring over time. 
 
8.2.2.1  STAT3 Target Gene Expression 
 
We will also analyze expression of a panel of 10 STAT3 target genes that we have 
found to be most informative of pyrimethamine-induced STAT3 inhibition in CLL 
cells treated in vitro, as well as four genes we have found to be invariant which will 
be used for normalization.  STAT3 target gene expression will be quantitated for 
individual genes, and a weighted normalized score will also be generated 12.  This 
analysis will provide a correlation of STAT3-dependent gene expression with 
STAT3 phosphorylation by the flow cytometric assay, a determination of the effect 
of pyrimethamine dosing on STAT3-dependent gene expression, and a longitudinal 
analysis of STAT3 dependent gene expression with pyrimethamine treatment in 
individual patients.  Furthermore, in conjunction with the pharmacokinetic analyses 
described above, these studies will provide correlations between plasma 
concentrations of pyrimethamine and effects on STAT3 phosphorylation and 
STAT3-dependent gene expression. 
 
 
 
8.3 
Determination of CLL and T-LGL Biologic Prognostic Features 
 
For CLL patients, peripheral blood will be collected in two 6 cc green top tubes and 
one 6 cc red top tube prior to therapy on all DFCI patients, along with saliva under the 
auspices of protocols 01-206 or 99-224 (if the patient has a low white blood cell 
count, then four 6 cc green top tubes will be collected instead). These samples can be 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
24 
drawn at screening or Day 1 (prior to therapy). The CLL Research Consortium Tissue 
Bank, our collaborator on Protocol 99-224, will determine IgVH mutational status 
and ZAP70 expression on all samples from DFCI subjects. This will be determined 
from three 6 cc green top tubes.  *BIDMC subjects are unable to be registered to 99-
224, but will have three 6 cc green top tubes drawn as part of this protocol, and they 
will be banked here for prognostic factor measurements later.   
 
In addition, and for all T-LGL patients, two 6 cc green tops & one 6 cc red top will be 
used for banking of peripheral blood mononuclear cells in the JFB 426 Brown lab, at 
screening and at day 15 of therapy (if the patient has a low white blood cell count, 
then four 6 cc green top tubes will be collected instead).  For those patients seen at 
BIDMC, these green-topped tubes and red-topped tube will be collected as part of this 
protocol and transferred to  JFB426 Brown lab for banking.   
 
The clinical flow cytometry laboratory will determine CD38 expression.  The 
DFPCC/BWH clinical cytogenetics laboratory will perform interphase FISH 
cytogenetics.   
 
 
9. STUDY CALENDAR 
 
Baseline evaluations are to be conducted within 1 month prior to start of protocol 
therapy.  CBCD and chemistry blood will be redrawn on Cycle 1 Day 1. Scans must be 
done <4 weeks prior to the start of therapy. In the event that the participant's condition 
is significantly deteriorating, laboratory evaluations should be repeated within 48 hours 
prior to initiation of the next cycle of therapy. 
All assessments must be performed prior to administration of any study medication. 
Failure to perform any assessment prior to administration of study drug will constitute a 
protocol violation. The Principal Investigator will determine if the event is a minor or 
major violation, per DF/HCC SOPs.  Informed Consent must take place prior to any 
study specific procedures. All study assessments and medications should be 
administered within + 3 days of the protocol-specified date, unless otherwise noted. 
 
 
 
  
Study Calendar Follows 
 
Test/Procedures 
Pre-
Study 
Weekly in 
1st 28 day 
Every 2 wks 
in 2nd cycle 
Every 
3-6 
months 
at 
investigator discretion and 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
25 
cycle (TLS 
blood 
X2 
1st week of  
C1) 
and 
then 
monthly 
at discontinuation of study 
treatment 
Informed Consent 
X 
 
 
 
History/PE/AE 
evaluation/ Perf. status 
X 
X 
X 
X 
Weight/VS 
X 
X 
X 
X 
Screen & Replete Folate 
(replete as necessary) 
X 
 
X10 
 
Concomitant Meds 
X 
X 
X 
 
CBC with Diff* 
X 
X 
X 
X 
CMP1,* 
X 
X 
X 
X 
Tumor Lysis 
Monitoring1A 
X 
X 
 
 
CAP CT**  
X 
 
X2 
X2 
Τumor Measurement by 
physical exam 9 
X 
 
X9 
X9 
Bone Marrow Biopsy 
X 
 
 
X7 
FISH8  
X 
 
 
 
Patient Diary 
X4 
X4 
X4 
X4 
Pharmacokinetic Studies 
X5a 
X5a 
X5a 
X5a   
Pharmacodynamic 
Studies 
X5b 
X5b 
X5b 
X5b 
CD4 
count 
(T 
cell 
subsets) 
X 
 
X6 
X6 
IgG, IgA, IgM 
X 
 
X6 
X6 
Beta-2-microglobulin 
X 
 
X6 
X6 
Flow 
cytometry 
for 
CD388 
X 
 
 
 
Samples 
for 
CLL 
prognostic factors/tissue 
banking3 
X3 
X3 
 
 
IgVH Testing 
X 
 
 
 
 
1  Comprehensive Metabolic Panel: Creatinine, glucose, total bilirubin, alkaline phosphatase, 
and SGOT/SGPT, LDH.  Serum beta-HCG within seven days prior to initiation of therapy for 
women of child-bearing potential. 
1A Investigators should monitor patients for tumor lysis with calcium, magnesium, phosphate, 
uric acid and LDH, in addition to a full comprehensive metabolic panel, twice during the 
first week, weekly for the rest of cycle 1, and then subsequently at times of other lab 
draws for the rest of the study. Treatment for TLS should commence should evidence of such 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
26 
be demonstrated. At a minimum, hyperuricemia, hydration, and electrolyte disturbances 
should be monitored closely and addressed promptly. 
2  CT scans (chest/abdomen/pelvis) to be performed at screening in all patients and again at 
the end of the 1st, 3rd and 6th months of therapy; subsequent scans shall be at the 
discretion of the treating physician if needed for adequate response evaluation. 
3  Please see Section 8.3 "Determination of CLL Biologic Prognostic Factors" for details on 
sample collection.   
4  
Diary dispensed with study drug and retrieved at scheduled visits.  
5a   One heparin-anticoagulated green top tube  (pharmacokinetic studies) to be collected during screening, on cycle 
1 day 1 prior to drug; prior to drug on one day among days 2, 3, 4 or 5, (optional); and prior to dose (trough)  at 
each subsequent clinic visit as per Section 8.1, above. A heparin-anticoagulated green top tube will also be 
collected at time of off study visit and once again 2-4 weeks later if the patient has not started additional 
therapy. All samples are to be delivered to the David Frank lab on Mayer 5. This includes BIDMC patients.  
5b   Two 6 cc heparinized blood for cell isolation (pharmacodynamic studies)  is to be collected during screening,  
on cycle 1 day 1 prior to drug and approximately two hours after drug administration, prior to dose (trough)  on  
days 2, 3 4 or 5 of therapy, pre drug on cycle 1 days 8, 15,and 22, cycle 2 days 1 and 15, and day 1 of 
subsequent cycles monthly up to cycle 6. Pharmacodynamic studies will also be collected at time of off study 
visit, 2-4 weeks later if the patient has not started additional therapy, and lastly at the 3 month follow up 
appointment (if subsequent therapy has not started).  These are to be delivered to the David Frank lab on Mayer 
5.  This includes BIDMC patients. 
6 
These laboratory tests will be done at baseline, at the end of cycles 1, 3, 6, end of treatment, and every 3-6 
months in follow-up.   
 7     Bone marrow biopsy will be done at screening (unless done within previous 6 months and the Principal 
Investigator feels it does not need to be repeated, or if prior marrow was positive and the patient has not had 
CLL treatment since) and thereafter only to confirm CR.  Confirmation of complete response by bone marrow 
biopsy should occur approximately 3 months after the clinical criteria for CR are first met.  Bone marrow 
biopsy is not required at off study.  
8 
To be performed on bone marrow if aspirate obtained; otherwise on peripheral blood prior to therapy initiation. 
9 
Up to 6 palpable nodes should be followed as targets, with bi-dimensions noted by the clinician whenever 
possible.  To be performed at screening in all patients and again at the end of the 1st, 3rd 
and 6th months of therapy, and in follow up if needed for adequate response evaluation. 
10 If deficient at baseline, folate level should be redrawn approximately every month while on study drug, or at 
required visits, whichever is less.   
* 
 Hematology and chemistry blood tests should be resulted and patient assessed for toxicity prior to taking dose 
of pyrimethamine.  
** Subjects with T-LGL do not require radiology 
 
10. MEASUREMENT OF EFFECT  
Responses will be formally assessed initially at the conclusion of the 1st 28 day cycle, then at the 
conclusion of three cycles (three months), and then every three months thereafter, until disease 
progression, start of another treatment, or death.  Response will be assessed in two ways:  (1) By 
NCI-WG 1996 criteria3 (as updated in the IWCLL 2008 criteria Section 10.2)19 without the use 
of CT scans.  In this case lymph nodes will be assessed based on physical exam measurements; 
(2) With the incorporation of CT findings as suggested for clinical trials in IWCLL 2008.  Thus 
CT scans will be performed on all patients at the end of the 1st month, 3rd month and 6th month, if 
still on study we will evaluate the impact of the CT scans (1) if used only to confirm CR; (2) if 
used to assess lymphadenopathy regardless of PE findings.  After the 6 month evaluation, further 
CT scans will generally not be done, but may be done at the discretion of the treating physician.  

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
27 
Following the initial screening bone marrow biopsy, additional bone marrow biopsies will be 
performed only to confirm complete remission.    
T-LGL Response Criteria21: 
Response to treatment should be evaluated by history, physical examination, and complete blood 
count with differential: 
●Complete response — A hematologic complete response is defined as the complete 
normalization of blood counts (ie, hemoglobin >12 g/dL; platelets >150,000/microL; 
absolute neutrophil count >1500/microL; absolute lymphocyte count <4000/microL) in 
the setting of a circulating LGL count of less than 500/microL.  
●Partial response — A hematologic partial response is defined as an improvement in 
blood counts that does not meet criteria for complete remission (eg, an absolute 
neutrophil count >500/microL or decreasing transfusion requirements). 
●Treatment failure — Treatment failure is defined as an inability to achieve an at least 
partial response after four months of therapy. 
●Progressive disease — Progressive disease is defined as a worsening of cytopenias, 
hepatomegaly, or splenomegaly 
 
10.1  Progression-Free Survival 
Progression-Free Survival (PFS) is defined as the duration of time from start of treatment to 
time of objective disease progression or death, whichever occurs earlier.  
10.2 
Antitumor Effect – Hematologic Tumors 
 
Parameter  
 CR*  
 PR*  
 PD*  
 
 
 
 
 Group A  
   
   
   
     Lymphadenopathy^ 
 None > 1.5 cm  
 Decrease ≥50%  
 Increase ≥50%  
     Hepatomegaly  
 None  
 Decrease ≥50%  
 Increase ≥50%  
     Splenomegaly  
 None  
 Decrease ≥50%  
 Increase ≥50%  
     Blood lymphocytes  
 <4000/µL  
 Decrease ≥50% from 
baseline  
 Increase ≥50% over 
baseline  
     Marrow# 
 Normocellular, < 30% 
lymphocytes, no B-
lymphoid nodules.  
≥ 30% lymphocytes, 
or B-lymphoid 
nodules, or not done 
  Increase of 
lymphocytes to more 
than 30% from 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
28 
normal. 
 Hypocellular marrow  
defines CRi  
 
Constitutional 
Symptoms@ 
None 
Any 
Any 
  
Group B  
   
   
   
     Platelet count  
 > 100 000/µL  
 > 100 000/µL or 
increase ≥50% over 
baseline  
 Decrease of ≥50% 
from baseline 
secondary to CLL  
     Hemoglobin  
 > 11.0 g/dL  
(untransfused and w/o 
erythropoietin) 
 > 11 g/dL or 
increase ≥50% over 
baseline  
 Decrease of > 2 
g/dL from baseline 
secondary to CLL  
     Neutrophils# 
 > 1500/µL  
 > 1500/µL or > 50% 
improvement over 
baseline  
 Any  
 
 
 
 
* CR (complete remission): all of the criteria have to be met, and patients have to lack disease-
related constitutional symptoms 
* PR (partial remission): at least two of the criteria of group A plus one of the criteria of group B 
have to be met. PR to be held at least two months and confirmed by restaging. 
* SD is absence of progressive disease (PD) and failure to achieve at least a PR; PD: at least one of 
the above criteria of group A or group B has to be met. 
 
 
 
 
^Sum of the products of multiple lymph nodes (as evaluated by CT scans in clinical trials, or by 
physical examination in general practice). 
#These parameters are irrelevant for some response categories. 
 
@Constitutional symptoms:  Unintentional weight loss ≥10% within the 
previous 6 months, significant fatigue (ECOG PS 2 or worse, Fevers 
>100.5F or 38.0 C for 2 or more weeks without other evidence of infection, 
night sweats >1 month without evidence of infection 
 
11. ADVERSE EVENT REPORTING REQUIREMENTS  
 
11.1General  
 
Adverse event collection and reporting is a routine part of every clinical trial. This study 
will use the descriptions and grading scales found in the CTEP Active Version of the NCI 
Common Terminology Criteria for Adverse Events (CTCAE)  that is available at 
http://ctep.cancer.gov/reporting//ctc.html and the IWCLL criteria for hematologic 
malignancies as outlined in Section 6.3.1.   
 
Information on all adverse events, whether reported by the participant, directly observed, 
or detected by physical examination, laboratory test or other means, will be collected, 
recorded, followed and reported as described in the following sections.  
 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
29 
Adverse events experienced by participants will be collected and reported from initiation 
of study medication, throughout the study, and within 30 days of the last dose of study 
medication. Participants who experience an ongoing adverse event related to a study 
procedure and/or study medication beyond 30 days will continue to be contacted by a 
member of the study team until the event is resolved, stabilized, or determined to be 
irreversible by the participating investigator.  
 
Participants should be instructed to report any serious post-study event(s) that might 
reasonably be related to participation in this study. The investigator should notify the IRB 
and any other applicable regulatory agency of any unanticipated death or adverse event 
occurring after a participant has discontinued or terminated study participation that may 
reasonably be related to the study.  
 
11.2 Definitions  
 
11.2.1  Adverse Event (AE) 
 
 
An adverse event is any undesirable sign, symptom or medical condition or 
experience that develops or worsens in severity after starting the first dose of 
study treatment or any procedure specified in the protocol, even if the event is not 
considered to be related to the study.  
 
11.2.2 Serious Adverse Event (SAE) 
 
A serious adverse event is an undesirable sign, symptom, or medical condition 
which: 
• is fatal or life-threatening; 
• requires or prolongs inpatient hospitalization; 
• results in persistent or significant disability/incapacity; 
• constitutes a congenital anomaly or birth defect; or 
• jeopardizes the participant and requires medical or surgical intervention to 
prevent one of the outcomes listed above.  
 
Events not considered to be serious adverse events are hospitalizations for: 
• routine treatment or monitoring of the studied indication, not associated 
with any deterioration in condition, or for elective procedures 
• elective or pre-planned treatment for a pre-existing condition that did not 
worsen 
• emergency outpatient treatment for an event not fulfilling the serious 
criteria outlined above and not resulting in inpatient admission 
• respite care 
 
11.2.3 Expectedness  
 
Adverse events can be 'Expected' or 'Unexpected.'  
 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
30 
11.2.3.1 Expected adverse event 
 
 
Expected adverse events are those that have been previously identified as 
resulting from administration of the agent. For the purposes of this study, 
an adverse event is considered expected when it appears in the current 
adverse event list, the Investigator’s Brochure, the package insert or is 
included in the informed consent document as a potential risk.   
 
Refer to Section 6.1 for a listing of common expected adverse events 
associated with the study agent(s).   
 
11.2.3.2. Unexpected adverse event 
 
 
For the purposes of this study, an adverse event is considered unexpected 
when it varies in nature, intensity or frequency from information 
provided in the current adverse event list, the Investigator’s Brochure, 
the package insert or when it is not included in the informed consent 
document as a potential risk.   
 
 
11.2.4 Attribution  
 
Attribution is the relationship between an adverse event or serious adverse event 
and the study treatment. Attribution will be assigned as follows: 
 
• Definite – The AE is clearly related to the study treatment. 
• Probable – The AE is likely related to the study treatment. 
• Possible – The AE may be related to the study treatment.  
• Unlikely - The AE is doubtfully related to the study treatment. 
• Unrelated - The AE is clearly NOT related to the study treatment. 
 
 
11.3  Recording Adverse Events 
 
Adverse event information will be obtained at each contact with the participant. All 
adverse events will be recorded on the appropriate study-specific case report forms 
(CRFs).  
 
11.4 
Reporting Adverse Events  
 
Each adverse event will be assessed to determine if it meets the criteria for serious 
adverse event. If a serious adverse event occurs, expedited reporting will follow local 
policies, and federal guidelines and regulations as appropriate.  
 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
31 
It is the responsibility of the participating investigator to notify the Principal 
Investigator (or Protocol Chair), IRB, and others of all serious adverse events as 
required in the protocol.  
 
The Principal Investigator (or Protocol Chair) will provide information with respect to 
adverse events and safe use of the study treatment (e.g., safety reports, Action Letters) 
to all participating investigators as soon as the information becomes available.  
 
11.5    Sponsor Notification by Investigator  
 
 
 11.5.1 
Serious Adverse Event Reporting Requirements 
 
All events meeting the criteria for Serious Adverse Event (see Section 11.2.2) 
that occur after the initial dose of study treatment, during treatment, or within 
30 days of the last dose of treatment must be reported as serious adverse 
events. Grade 4 hematogic toxicites are excluded from SAE reporting after the 
first cycle.  Grade 4 ALC reporting is excluded from SAE reporting during all 
cycles. 
 
The participating investigator must report each serious adverse event, 
regardless of attribution, to the Principal Investigator (or Protocol Chair) 
within 24 hours of learning of the occurrence. In the event that the 
participating investigator does not become aware of the serious adverse event 
immediately (e.g., participant sought treatment elsewhere), the participating 
investigator is to report the event within 24 hours after learning of it and 
document the time of his or her first awareness of the adverse event. Report 
serious adverse events by telephone and facsimile to: 
 
 
  
 
 
Jennifer R Brown MD PhD  
Tel 617-632-4894 
 
Fax 617-582-7909 
 
Within the following 24-48 hours, the participating investigator must provide 
follow-up information on the serious adverse event. Follow-up information 
should describe whether the event has resolved or continues, if and how the 
event was treated, and whether the participant will continue or discontinue 
study participation.  
 
11.5.2 Non-Serious Adverse Event Reporting Requirements 
 
Non-serious adverse events will be reported to the Principal Investigator (or 
Protocol Chair) on the toxicity Case Report Forms.   
 
 
11.6   Institutional Review Board (IRB) Notification by Investigator 
 
The participating investigator will report all adverse events and serious adverse events 
to the Principal Investigator (or Protocol Chair) and to the IRB according to the local 
IRB’s policies and procedures in reporting adverse events.  

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
32 
 
In the event of a multi-center study, the Principal Investigator (or Protocol Chair) will 
report adverse events and serious adverse events from all participating sites to the DFCI 
IRB according to the IRB’s policies and procedures in reporting adverse events.  
 
11.7   Food and Drug Administration (FDA) Notification by Sponsor-Investigator  
 
The Sponsor-Investigator will report to the FDA any adverse event that is serious, 
unexpected and reasonably related (i.e., possible, probable, definite) to the study 
treatment. 
 
Unexpected fatal or life-threatening experiences associated with the use of the study 
treatment will be reported to FDA as soon as possible but in no event later than 7 
calendar days after initial receipt of the information.  
 
All other serious unexpected experiences associated with the use of the study treatment 
will be reported to FDA as soon as possible but in no event later than 15 calendar days 
after initial receipt of the information. 
 
  
Events will be reported to the FDA by telephone (1-800-FDA-1088) or by fax (1-800-
FDA-0178) using Form FDA 3500A (Mandatory Reporting Form for investigational 
agents) or FDA Form 3500 (Voluntary Reporting Form for commercial agents). Forms 
are available at http://www.fda.gov/medwatch/getforms.htm.  
 
11.8  Hospital Risk Management Notification by Investigator 
 
The participating investigator will report to the Principal Investigator (or Protocol Chair) 
and to local Risk Management any subject safety reports or sentinel events that require 
reporting according to institutional policy. 
12. DATA AND SAFETY MONITORING 
 
12.1  Data Reporting  
12.1.1 Method  
The QACT will collect, manage, and monitor data for this study.  
12.1.2 Data Submission  
The schedule for completion and submission of case report forms (paper or electronic) to the 
QACT is as follows: 
Form 
Submission Timeline 
Eligibility Checklist 
Complete prior to registration with QACT 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
33 
On Study Form 
Within 14 days of registration 
Baseline Assessment Form 
Within 14 days of registration 
Treatment Form 
Within 10 days of the last day of the cycle 
Adverse Event Report Form 
Within 10 days of the last day of the cycle 
Response Assessment Form 
Within 10 days of the completion of the cycle 
required for response evaluation 
Off Treatment/Off Study Form Within 14 days of completing treatment or 
being taken off study for any reason 
Follow up/Survival Form 
Within 14 days of the protocol defined follow 
up visit date or call 
 
12.2  Safety Meetings 
 
The DF/HCC Data and Safety Monitoring Committee (DSMC) will review and monitor 
toxicity and accrual data from this trial. The committee is composed of clinical specialists 
with experience in oncology and who have no direct relationship with the study. 
Information that raises any questions about participant safety will be addressed with the 
Principal Investigator and study team. 
 
The DSMC will meet quarterly and/or more often if required to review toxicity and 
accrual data.  Information to be provided to the committee may include:  up-to-date 
participant accrual; current dose level information; DLT information; all grade 2 or higher 
unexpected adverse events that have been reported; summary of all deaths occurring 
within 30 days for Phase I or II protocols; for gene transfer protocols, summary of all 
deaths while being treated and during active follow-up; any response information; audit 
results, and a summary provided by the study team. Other information (e.g. scans, 
laboratory values) will be provided upon request.  
 
12.3  Monitoring 
 
Involvement in this study as a participating investigator implies acceptance of potential 
audits or inspections, including source data verification, by representatives designated 
by the Principal Investigator (or Protocol Chair) or DF/HCC. The purpose of these 
audits or inspections is to examine study-related activities and documents to determine 
whether these activities were conducted and data were recorded, analyzed, and 
accurately reported in accordance with the protocol, institutional policy, Good Clinical 
Practice (GCP), and any applicable regulatory requirements.  
 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
34 
All data will be monitored for timeliness of submission, completeness, and adherence to 
protocol requirements. Monitoring will begin at the time of participant registration and 
will continue during protocol performance and completion.  
 
 
13. REGULATORY CONSIDERATIONS 
 
13.1  Protocol Review and Amendments 
This protocol, the proposed informed consent and all forms of participant information 
related to the study (e.g., advertisements used to recruit participants) and any other 
necessary documents must be submitted, reviewed and approved by a properly 
constituted IRB governing each study location.  
Any changes made to the protocol must be submitted as amendments and must be 
approved by the IRB prior to implementation. Any changes in study conduct must be 
reported to the IRB. The Principal Investigator (or Protocol Chair) will disseminate 
protocol amendment information to all participating investigators.  
All decisions of the IRB concerning the conduct of the study must be made in 
writing. 
 
13.2   Informed Consent 
 
All participants must be provided a consent form describing this study and 
providing sufficient information for participants to make an informed decision 
about their participation in this study. The formal consent of a participant, using the 
IRB approved consent form, must be obtained before the participant is involved in 
any study-related procedure. The consent form must be signed and dated by the 
participant or the participant’s legally authorized representative, and by the person 
obtaining the consent. The participant must be given a copy of the signed and dated 
consent document. The original signed copy of the consent document must be 
retained in the medical record or research file.  
 
13.3  Ethics and Good Clinical Practice (GCP) 
This study is to be conducted according to the following considerations, which 
represent good and sound research practice: 
• ICH Consolidated Good Clinical Practice: Guidelines (E6) 
www.fda.gov/cder/guidance/iche6.htm  
• US Code of Federal Regulations (CFR) governing clinical study conduct 
and ethical principles that have their origin in the Declaration of Helsinki 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
35 
 Title 21 Part 11 – Electronic Records; Electronic Signatures 
www.access.gpo.gov/nara/cfr/waisidx_02/21cfr11_02.html 
 Title 21 Part 50 – Protection of Human Subjects 
www.access.gpo.gov/nara/cfr/waisidx_02/21cfr50_02.html  
 Title 21 Part 54 – Financial Disclosure by Clinical Investigators 
www.access.gpo.gov/nara/cfr/waisidx_02/21cfr54_02.html  
 Title 21 Part 56 – Institutional Review Boards 
www.access.gpo.gov/nara/cfr/waisidx_02/21cfr56_02.html  
 Title 21 Part 312 – Investigational New Drug Application 
www.access.gpo.gov/nara/cfr/waisidx_02/21cfr312_02.html  
• State laws 
• Institutional research policies and procedures 
www.dfhcc.harvard.edu/clinical-research-support/clinical-research-
operations-cro/policies-and-procedures 
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 according to the local reporting policy.  
 
13.4  Study Documentation 
 
The investigator must prepare and maintain adequate and accurate case histories designed 
to record all observations and other data pertinent to the study for each research 
participant. This information enables the study to be fully documented and the study data 
to be subsequently verified. 
 
Original source documents supporting entries in the case report forms include but are not 
limited to hospital records, clinical charts, laboratory and pharmacy records, recorded 
data from automated instruments, microfiches, photographic negatives, microfilm or 
magnetic media, and/or x-rays.  
 
 
13.5  Records Retention 
 
All study-related documents must be retained for the maximum period required by 
applicable federal regulations and guidelines or institutional policies.  
 
 
14. STATISTICAL CONSIDERATIONS  
 
14.1 Primary Objectives 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
36 
 
14.1.1 Primary Objectives for Phase I Trial 
 
A standard 3+3 dose-escalation design is used in the Phase I portion for assessing the 
safety and determining the MTD and recommended dose for the Phase II portion of this 
study of pyrimethamine in relapsed CLL/SLL and T-LGL patients.   
 
14.1.2 Primary Objectives for Phase II Trial 
 
After MTD is established in Phase I, 20 additional patients will be enrolled in the Phase 
II study and treated with pyrimethamine at the MTD.  The primary objective for this 
portion of the study is to determine the overall response rate of pyrimethamine in these 
patients, including both CLL/SLL and T-LGL.   
 
14.2 Sample Size and Patient Accrual 
 
14.2.1 Phase I 
 
To assess the MTD of pyrimethamine, a standard 3+3 dose-escalation design is used, 
with 5 planned pyrimethamine dose levels (Section 5); the observation period is one 
cycle (28 days).  3 patients are enrolled in the study initially, if no DLT is observed in the 
first 3 patients at a given dose level of pyrimethamine, escalation will proceed.  If 2 or 3 
DLTs are observed in the first 3 patients, escalation stops.  If 1 DLT is observed, an 
additional 3 patients will be enrolled at the same dose level.  If no DLT is observed in the 
additional 3 patients, escalation proceeds.  If any DLTs are observed among the 3 
additional patients, escalation stops.   
 
Based on this dose escalation scheme, the probability of escalating to the next dose level 
for various true underlying rates of DLT is given in Table 1. 
 
Table 1 Probability of Escalation for Various True Underlying DLT Rates 
for Pyrimethamine 
 
True DLT Rate  1.0%   10.0%  20.0%   30.0%  40.0%  50.0%   
 
Probability of 
Escalation          0.99     0.91     0.71      0.49     0.31     0.17      
 
If the true underlying proportion of DLT is 30% at the current dose during the treatment 
period, there is a 49% chance of escalating to the next dose.  We anticipate that a 
minimum of 2 to a maximum of 12 additional patients will be enrolled in the Phase I 
portion of the study.   
 
 
14.2.2 Phase II 
 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
37 
The Phase II portion is a single arm study which consists of determining the overall 
response rate of pyrimethamine in relapsed CLL/SLL and T-LGL patients.  The overall 
response rate is not expected to differ between these two diseases and therefore one 
overall response rate will be reported.  A 35% overall response rate will be considered 
promising.  The first response evaluation will be assessed after the first cycle of treatment 
then at the end of three cycles.   
 
Six Phase I patients treated at the MTD of pyrimethamine will be enrolled in the Phase II 
part of the study.  A Simon’s two stage optimal design is used to compute the sample 
size.  A total number of 26 patients is needed in order to detect a 35% response rate, 
assuming the ORR for the null hypothesis is 10%, with 90% power and 10% type I error.  
The study will stop early if only 1 or 0 responses out of the first 10 patients is observed, 
and the drug will not be declared promising if less than 5 responses are seen in the total 
26 patients.  The probability of stopping at the first stage if the true ORR is less than 35% 
is 0.736.   
 
14.3 Primary Analysis 
 
The overall response rate and a 90% confidence interval will also be calculated, using the 
method of Atkinson and Brown for a two stage study.  Descriptive statistics for patients’ 
baseline characteristics will be summarized using descriptive statistics (median, 
interquartile range, proportions).   
 
14.4 Secondary Objectives 
 
• To assess the toxicity profile of pyrimethamine in relapsed CLL/SLL and T-LGL, 
both acutely and over prolonged daily dosing. 
• To determine pyrimethamine levels in vivo with prolonged dosing. 
• To determine the progression free and overall survival following pyrimethamine 
for the treatment of relapsed CLL/SLL and T-LGL. 
• To determine whether pyrimethamine inhibits STAT3 in vivo by assessing 
downregulation of STAT2 dependent gene expression in CLL cells and/or 
peripheral blood. 
• To determine whether known prognostic factors in CLL/SLL correlate with 
reponse to pyrimethamine. 
• To assess the impact of CT scans on response evaluation in relapsed CLL/SLL 
and T-LGL. 
 
14.5 Secondary Analysis 
 
Kaplan Meier method will be used to estimate the median of overall and progression free 
survival.  The number and percent of patients who experienced toxicities on 
pyrimethamine after at least one dose of treatment will also be summarized. The paired t-
test or Wilcoxon’s signed rank test, depending on the symmetry of the data, will be used 
in univariate analyses for continuous variables of gene expression data, based on the ratio 
of post-therapy to baseline expression.  Spearman’s correlation coefficients will be used 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
38 
for assessing relationships between continuous variables for some known prognostic 
factors and Fisher’s exact test will be used for finding associations between categorical 
variables.   
 
14.6  Reporting and Exclusions 
 
14.6.1  Evaluation of toxicity. All participants who take at least one dose of 
pyrimethamine on study will be evaluable for toxicity from the time of their 
first treatment. 
14.6.2  Evaluation of response.  All participants included in the study must be assessed 
for response to treatment, even if there are major protocol treatment deviations 
or if they are ineligible. Each participant should be assigned one of the following 
categories: 1) complete response, 2) partial response, 3) stable disease, 4) 
progressive disease, 5) early death from malignant disease, 6) early death from 
toxicity, 7) early death because of other cause, 8) CRi or 9) unknown (not 
assessable, insufficient data). By arbitrary convention, category 9 usually 
designates the "unknown" status of any type of data in a clinical database. 
              
All of the participants who met the eligibility criteria should be included in the 
main analysis of the response rate. Participants in response categories 4-9 should be 
considered to have a treatment failure (disease progression). Thus, an 
incorrect treatment schedule or drug administration does not result in 
exclusion from the analysis of the response rate.  If a bone marrow is not 
assessed at restaging it will be noted on the appropriate CRF, and an otherwise 
complete response would become a PR. 
 
 
 
15. PUBLICATION PLAN 
The results will be made public within approximately 24 months of the end of data collection. 
The initial report may be a meeting abstract followed by a manuscript for a peer-reviewed 
journal.  This study will be registered at clinicaltrials.gov after IRB approval and activation. 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
39 
16. REFERENCES  
 
1. Klinker H, Langmann P, Richter E. Plasma pyrimethamine concentrations during long-term 
treatment for cerebral toxoplasmosis in patients with AIDS. Antimicrob Agents Chemother. 
1996;40:1623-1627. 
2. Jemal A, Murray T, Ward E, et al. Cancer Statistics, 2005. CA A Cancer Journal for 
Clinicians. 2005;55:10-30. 
3. Cheson BD, Bennett JM, Grever M, et al. National Cancer Institute-sponsored Working 
Group guidelines for chronic lymphocytic leukemia: revised guidelines for diagnosis and 
treatment. Blood. 1996;87:4990-4997. 
4. Byrd JC, Peterson BL, Morrison VA, et al. Randomized phase 2 study of fludarabine with 
concurrent versus sequential treatment with rituximab in symptomatic, untreated patients with B-
cell chronic lymphocytic leukemia: results from Cancer and Leukemia Group B 9712 (CALGB 
9712). Blood. 2003;101:6-14. 
5. Keating MJ, O'Brien S, Albitar M, et al. Early results of a chemoimmunotherapy regimen of 
fludarabine, cyclophosphamide, and rituximab as initial therapy for chronic lymphocytic 
leukemia. J Clin Oncol. 2005;23:4079-4088. 
6. Tsimberidou AM, Wen S, O'Brien S, et al. Assessment of chronic lymphocytic leukemia and 
small lymphocytic lymphoma by absolute lymphocyte counts in 2,126 patients: 20 years of 
experience at the University of Texas M.D. Anderson Cancer Center. J Clin Oncol. 
2007;25:4648-4656. 
7. Loughran TP, Zickl L, Olson TL, et al, Immunosuppressive therapy of LGL leukemia: 
prospective multicenter phase II study by the Eastern Cooperative Oncology Group (E5998), 
Leukemia 2015; 29:886-894 
8. Osuji N, Matutes E, Tjonnfjord G, et al, T-cell large granular lymphocyte leukemia: A report 
on the treatment of 29 patients and a review of the literature, Cancer 2006; 107:570-578 
9. Osuji N, Del Giudice I, Matutes E, et al, CD52 expression in T-cell large granular lymphocyte 
leukemia--implications for treatment with alemtuzumab, Leuk Lymphoma. 2005 May;46:723-
727 
10. Andersson E, Kuusanmäki H, Bortoluzzi S, et al, Activating somatic mutations outside the 
SH2-domain of STAT3 in LGL leukemia, Leukemia. 2015   
11. Rajala HL, Eldfors S, Kuusanmäki H, et al, Discovery of somatic STAT5b mutations in large 
granular lymphocytic leukemia, Blood 2013; 121:4541-4550 
12. 
Kumar A, Commane M, Flickinger TW, Horvath CM, Stark GR. Defective TNF-α-
induced apoptosis in STAT1-null cells due to low constitutive levels of caspases. Science. 
1997;278:1630-1632. 
13. 
Wegrzyn J, Potla R, Chwae Y-J, et al. Function of Mitochondrial Stat3 in Cellular 
Respiration. Science. 2009;323:793-797. 
14. 
Frank DA, Mahajan S, Ritz J. B lymphocytes from patients with chronic lymphocytic 
leukemia contain signal transducer and activator of transcription (STAT) 1 and STAT3 
constitutively phosphorylated on serine residues. J Clin Invest. 1997;100:3140-3148. 
15. 
Hazan-Halevy I, Ferrajoli A, Harris D, et al. Constitutive Serine Phosphorylation of 
STAT3 Induces Proliferation of Chronic Lymphocytic Leukemia Cells. Cytokine. 2007;39:15- 

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Protocol v. 8 22JAN16    
 
 
40 
16. 
Ghosh AK, Kay NE, Secreto CR, Shanafelt TD. Curcumin Inhibits Prosurvival Pathways 
in Chronic Lymphocytic Leukemia B Cells and May Overcome Their Stromal Protection in 
Combination with EGCG. Clinical Cancer Research. 2009;15:1250-1258. 
17. 
Alvarez JV, Febbo PG, Ramaswamy S, Loda M, Richardson A, Frank DA. Identification 
of a genetic signature of activated signal transducer and activator of transcription 3 in human 
tumors. Cancer Res. 2005;65:5054-5062. 
18. 
Frank DA. STAT inhibition in the treatment of cancer:  Transcription factors as targets for 
molecular therapy. Curr Cancer Therapy Reviews. 2006;2:57-65. 
19. 
Hallek M, Cheson BD, Catovsky D, et al. Guidelines for the diagnosis and treatment of 
chronic lymphocytic leukemia: a report from the International Workshop on Chronic 
Lymphocytic Leukemia updating the National Cancer Institute-Working Group 1996 guidelines. 
Blood. 2008;111:5446-5456.20. Friedberg JW, Dong DA, Li S, et al. Oral fludarabine has 
significant activity in patients with previously untreated chronic lymphocytic leukemia, and leads 
to increased STAT1 levels in vivo. Leukemia Res. 2004;28:139-147. 
21. Lamy T, Loughran TP Jr, How I treat LGL leukemia, Blood 2011; 117:2764 
 
 
 
 

Pyrimethamine for CLL 
Protocol v. 8 22JAN16    
 
 
41 
Appendix A   IWCLL 2008 Indications for Therapy for CLL 
 
1. Evidence of progressive marrow failure as manifested by the development of, or 
worsening of, anemia and/or thrombocytopenia  
2. Massive (ie, at least 6 cm below the left costal margin) or progressive or symptomatic 
splenomegaly  
3. Massive nodes (ie, at least 10 cm in longest diameter) or progressive or symptomatic 
lymphadenopathy  
4. Progressive lymphocytosis with an increase of more than 50% over a 2-month period or 
lymphocyte doubling time (LDT) of less than 6 months. LDT can be obtained by linear 
regression extrapolation of absolute lymphocyte counts obtained at intervals of 2 weeks 
over an observation period of 2 to 3 months. In patients with initial blood lymphocyte 
counts of less than 30 x 109/L (30 000/µL), LDT should not be used as a single parameter 
to define a treatment indication. In addition, factors contributing to lymphocytosis or 
lymphadenopathy other than CLL (eg, infections) should be excluded.  
5. Autoimmune anemia and/or thrombocytopenia that is poorly responsive to corticosteroids 
or other standard therapy (see section 10.2).  
6. Constitutional symptoms, defined as any one or more of the following disease-related 
symptoms or signs:  
a. Unintentional weight loss of 10% or more within the previous 6 months;  
b. Significant fatigue (ie, ECOG PS 2 or worse; inability to work or perform usual 
activities);  
c. Fevers higher than 100.5°F or 38.0°C for 2 or more weeks without other evidence 
of infection; or  
d. Night sweats for more than 1 month without evidence of infection.  
 
 
 
 
 

Pyrimethamine for CLL 
Protocol v. 7 24JUL14    
 
42 
Appendix B 
Performance Status Criteria 
 
 
ECOG Performance Status Scale 
 
 
Karnofsky Performance Scale 
Grade 
Description 
Percent 
Description 
0 
Normal activity. Fully active, able 
to carry on all pre-disease 
performance without restriction. 
100 
Normal, no complaints, no 
evidence of disease. 
90 
Able to carry on normal activity; 
minor signs or symptoms of 
disease. 
1 
Symptoms, but ambulatory. 
Restricted in physically strenuous 
activity, but ambulatory and able to 
carry out work of a light or 
sedentary nature (e.g., light 
housework, office work). 
80 
Normal activity with effort; 
some signs or symptoms of 
disease. 
70 
Cares for self, unable to carry on 
normal activity or to do active 
work. 
2 
In bed < 50% of the time. 
Ambulatory and capable of all self-
care, but unable to carry out any 
work activities. Up and about more 
than 50% of waking hours. 
60 
Requires occasional assistance, 
but is able to care for most of 
his/her needs. 
50 
Requires considerable assistance 
and frequent medical care. 
3 
In bed >50% of the time. Capable 
of only limited self-care, confined 
to bed or chair more than 50% of 
waking hours. 
40 
Disabled, requires special care 
and assistance. 
30 
Severely disabled, 
hospitalization indicated. Death 
not imminent. 
4 
100% bedridden. Completely 
disabled. Cannot carry on any self-
care. Totally confined to bed or 
chair. 
20 
Very sick, hospitalization 
indicated. Death not imminent. 
10 
Moribund, fatal processes 
progressing rapidly.  
5 
Dead. 
0 
Dead. 
 

 
 
 
43
Appendix C 
 
Study Diary 
  
 

 
 
 
44
 

Pyrimethamine Study Drug Diary
Page 1
OTHER MEDICATIONS TAKEN
Study Participant
Self-Administration
If you take a daily medication (prescribed or otherwise), please use one
Study Drug Diary
line per drug and indicate the start and stop dates under the "Date(s) 
Dana-Farber/Harvard Cancer Center
Taken" section (i.e., 6/2/09 - 6/5/09). 
Dose
Dates Taken
Reason Taken
Participant Identifier: ____________________
300 mg
Jan 1 to 10
Per MD
Protocol # :  
Your MD ____________________ Phone ________________
Your RN _____________________ Phone _______________
STUDY DRUG INSTRUCTIONS:
Study Drug: Pyrimethamine
How Much:  Your dose is 
How Often: You will take Pyrimethamine once daily
When: You should take your dose orally, 
in the morning, with food.
HOLD DOSE ON DAYS THAT YOU REPORT TO CLINIC  UNTIL
YOUR DOCTOR OR NURSE TELLS YOU THAT YOUR LAB
RESULTS ARE COMPLETED
SPECIAL INSTRUCTIONS:
Study Participant Initials __________
Date _________
*
Please remember to take Allopurinal as prescribed, 
 300 mg once per day, on days 1 through 10
*
Please keep track of the Allopurinol on this diary, 
FOR STUDY TEAM USE ONLY
 as in example to the left.
*
Drug must be kept in original container. 
Staff Initials:
*
There are no restricted foods or beverages
Date Dispensed:
Date Returned:
*
Any doses missed can be taken later on the same day 
# pills/caps/tabs dispensed:
# pills/caps/tabs returned:
 they were missed, with food.  
*
If a missed dose is not discovered until the following 
# pills/caps/tabs that should have been taken:
 day, the dose should not be made up.
*
Any doses vomited within one hour of administration 
Discrepancy Notes:
 should be retaken
*
Please remember to bring any unused study drug, all 
 empty containers, and diary to the next clinic visit. 
Drug Name
Example: Allopurinol

Page 2
DOSING LOG
SYMPTOMS/SIDE EFFECTS
Pyrimethamine
Cycle: ___
For each AM dose take: 
Please record any side effects experienced during this cycle.
Include the date the particular symptom started and when it
ended. Please evaluate the severity of the symptom according to
Please indicate the date, time, amount taken and any comments. 
the following scale:
Mild: Awareness of sign or symptom; easily tolerated and did 
Dose
Time
Food
not affect ability to perform normal daily activities. Symptom 
Ex:
6/1/2009
 X mg
9:00 AM
vomited PM pill
yes
did not require medication or therapeutic intervention.
Day 1
Day 2
Moderate: Significant  discomfort which interfered with ability 
Day 3
to perform normal daily activities. Symptom was easily resolved  
Day 4
with at home medication or simple therapeutic intervention.
Day 5
Day 6
Severe: Marked discomfort with an inability to carry out 
Day 7
normal daily activities. Symptom required new medication 
Day 8
and/or therapeutic intervention in order to resolve.
Day 9
Day 10
Please Note: The severity should reflect the most severe level 
Day 11
experienced during the time period. 
Day 12
Day 13
Symptom
Start Date End Date Severity
Day 14
Day 15
Day 16
Day 17
Day 18
Day 19
Day 20
Day 21
Day 22
Day 23
Day 24
Day 25
Day 26
Day 27
Day 28
Comments
Date

 
Final 2/12/10 DMM; R: 6/13/12 DMM/AM #20 KAF 4/27/16: VP 4/27/16 
Dana-Farber Cancer Institute 
Nursing Protocol Education Sheet 
 
Protocol Number: 
09-421 
Protocol Name: 
A phase I/II study of Pyrimethamine, a STAT3 inhibitor, for the 
treatment of relapsed Chronic Lymphocytic Leukemia/Small 
Lymphocytic Lymphoma and T-LGL 
DFCI Site PI: 
Jennifer Brown, MD, PhD 
DFCI ResearchNurses: 
 Karen Francoeur, RN; Victoria Patterson, RN; Kathleen McDermott, RN 
Page the DFCI research nurse or DFCI site PI if there are any questions/concerns about the protocol.   
Please also refer to ONC 15: Oncology Nursing Protocol Education Policy 
 
           SPECIAL NURSING CONSIDERATIONS UNIQUE TO THIS PROTOCOL 
Study Design 
 
Pyrimethamine is an oral folic acid antagonist used as an antiparasitic agent. See section 
2.1 for description of this agent, precautions in people with possible folate deficiency and 
for potential toxicities, like myelosuppression, vomiting, anorexia, etc… 
Current dose as per A.P. 
 Each cycle = 28 days. 
Dose 
Calcul-
ation 
Doses of Pyrimethamine are fixed in mg 
  The Recommended Phase II dose is found as noted on Alert Page 
  
Study Drug and 
Administration 
Pyrimethamine- See TREATMENT PLAN – sec. 5 
 Self-administered, once daily in the AM with food. 
 Instruct patient to not take drug at home on clinic visit days (e.g. Cycle 1 Day 1,8, and 
15, C2D1, etc…). Dose is not to be taken until labs results are reviewed on those days. 
 Instruct patients on Drug Diary use. 
 Vomited doses not made up unless vomited within 1 hr of intake. Missed, e.g. forgotten  
doses can be made up on same day but not later 
 Section 5.1 Pre treatment criteria 
 Section 5.2 Agent Administration, to include twice weekly labs in Cycle 1 for TLS 
monitoring. 
Dose 
Modifications 
and Toxicity 
Definition of DLT- sect. 5.3 
 
Anticipated toxicities – sect. 6.1; management per Setion 6.2 
 
Dose Modifications/Delays – see sect. 6.3. Note guidelines for Heme toxicity in 
sect. 5.3, for Rash in 6.3.2, and for Anorexia/Vomiting in sect. 6.3.3 
Concomitant 
Meds 
Review sect. 5.4 for all Supportive care guidelines and conc. Meds allowed, prohibited or to 
used with caution 
 Allopurinol days 1-10 daily unless pt. has allergy to it. 
 Antiemetics as needed but Lorazepam is to be used with caution due to risk of ↑LFT’s 
 Use of Bactrim discouraged; Acyclovir prophylactic is recommended.  
Required Data 
(ECG, V/S,  
etc…) 
 Note folic acid blood draws at screening and periodically – see sect. 5 and 9 (Study 
Calendar) 
 PD blood draws per Section 8.2 and Study Calendar 
 PKs per Section 8.2 and Study Calendar 
 

 
Final 2/12/10 DMM; R: 6/13/12 DMM/AM #20 KAF 4/27/16: VP 4/27/16 
Charting 
Tips 
Please be sure to DOCUMENT time of drug administration,  PD blood draws, any patient teaching.