Clinical trial NCT00490529
Conditions: Lymphoma, Mantle-Cell
Sponsor: Ronald Levy
Start date: 2009-08-01; Completion date: 2017-12-14
Enrollment: 59 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
Mantle cell lymphoma (MCL) is a sub-type of non-Hodgkin's lymphoma (NHL) which is generally considered incurable with current therapy. Participants will receive an autologous vaccine against their individual lymphoma after undergoing stem cell transplantation. This vaccination may prolong the time which patients will stay in remission from their disease.

Detailed description:
Study treatment is a complex set of steps of research procedures and regular medical care. By using a participant's cancer cells as an immungen, the study hopes to improve freedom from molecular residual disease (MRD).

PRIMARY OBJECTIVE Freedom from molecular residual disease at 1-year post-autologous transplant.

SECONDARY OBJECTIVE Time To Clinical Progression (TTP)

This study has 2 research agents, PF-03152676 and CpG-MCL Vaccine.

PF-03152676 is a synthetic DNA molecule, 24 nucleotides in length with a nuclease-resistant phosphorothioate backbone. It is an immunostimulatory, single-stranded oligodeoxynucleotide (oligo-DNA) containing unmethylated cytosine and guanine (CpG) motifs and synthesized with a nuclease-resistant phosphorothioate backbone. PF-03512676 acts as an agonist of human Toll-like receptor 9, leading to activation of antigen-presenting cells and a cascade of anti-tumor immune reactions.

CpG-MCL Vaccine is the primary study agent. It is prepared by dissociating a participant's harvested tumor cells into a single-cell suspension, and culturing them with PF-03152676 for 72 hours at 37 degrees C, 5% CO2 to allow for up-regulation of antigen-presenting and co-stimulatory molecules, then irradiated to 200 Gy to destroy any remaining cancer propagating ability.

The study procedure is summarized as 12 steps, listed below.

* Step 1. Undergo excisional tumor biopsy or apheresis to obtain tumor cells, which will be used to generate the CpG-MCL vaccine .
* Step 2. Receive standard induction chemotherapy (regular medical care).
* Step 3. Once in remission, receive 3 vaccinations of CpG-MCL Vaccine over 3 weeks. With each CpG-MCL vaccination, a concurrent subcutaneous injection of PF-3512676 is administered as an adjuvant.
* Step 4. About 4 weeks later, receive rituximab 375 mg/m² to minimize any residual tumor.
* Step 5. Apheresis procedure to harvest the CpG-MCL Vaccine-primed T-cells. Each collection is \~1 x 10e10 CD3+ T-cells.
* Step 6. High-dose cytoxan and filgrastim to mobilize peripheral blood progenitor cell (PBPC).
* Step 7. Undergo separate apheresis procedure to harvest PBPC).
* Step 8. Receive myeloablative chemotherapy (regular medical care).
* Step 9. Receive PBPC infusion (also known as autologous hematopoietic cell transplant, AHCT).
* Step 10. Within 3 days of AHCT (but typically 1 day), receive infusion of CpG-MCL Vaccine-primed T-cells, followed within 1 hour by a with 4th vaccination with CpG-MCL Vaccine (1st booster vaccination).
* Step 11. After hematopoietic recovery, receive 5th vaccination with CpG-MCl (2nd booster vaccination).
* Step 12. Monitor participants for general health and disease status through at least 3 years.

Arms: arm 1: An autologous anti-tumor vaccine.
Interventions: intervention 1: CpG-MCL vaccine intervention 2: PF-3512676 intervention 3: Vaccine-primed T-cells intervention 4: Autologous hematopoietic stem cell transplant (HSCT) intervention 5: Rituximab intervention 6: Standard induction chemotherapy intervention 7: Cyclophosphamide intervention 8: Filgrastim
Intervention descriptions: intervention 1: CpG-MCL vaccine is a vaccine prepared by co-culturing cells from the participant's mantle cell lymphoma suspension with 3 mcg/mL PF-3512676, then irradiated to 200 Gy. 1 x 10e8 CpG-MCL cells will be given as a subcutaneous injection. intervention 2: PF-03152676 is a synthetic immunostimulatory, single-stranded oligodeoxynucleotide (oligo-DNA) moledule containing unmethylated cytosine and guanine (CpG) motifs. PF-03512676 acts as an agonist of human Toll-like receptor 9, leading to activation of antigen-presenting cells and a cascade of antitumor immune reactions. intervention 3: Vaccine primed T-cells are the post-vaccination leukapheresis harvest of peripheral blood mononuclear cells. Each collection is approx 1 x 10e10 CD3+ T-cells. intervention 4: Regular medical procedure intervention 5: 375 mg/m² by infusion intervention 6: Patient-specific, regular medical care treatment as determined by treating oncologist intervention 7: Regular medical care treatment to mobilize peripheral blood progenitor cell (PBPC) intervention 8: Regular medical care treatment to mobilize peripheral blood progenitor cell (PBPC)

Protocol documents:
Prot_SAP_000.pdf: 
TITLE:  Phase 1/2 Study of a CpG-Activated Whole Cell Vaccine Followed by 
Autologous “Immunotransplant” for Mantle Cell Lymphoma 
 
NCT00490529 
 
 
IND Number: 
BB-IND 14089 
 
Protocol Number: 
 eProtocol # 5089, LYMNHL0040 
 
Drug: 
CpG-MCL vaccine 
 
Indication: 
Mantle Cell Lymphoma 
 
Treatment Center:            
Stanford University Medical Center 
 
 
Coordinating Center:          Stanford University Medical Center 
 
*Principal Investigator:      Ronald Levy, MD 
                                                Professor, Division of Oncology 
                                                Stanford University Medical Center 
                                                 
                                                Stanford, CA.  94305 
Levy@Stanford.edu 
 
Co-investigators: 
Robert S. Negrin, MD; Ranjana H. Advani, MD; Wen Kai Weng, 
MD,PhD; David Miklos, MD, PhD; Jonathan Benjamin, MD; 
Lauren S. Maeda, MD; Everett Meyer, MD; Sally Arai, MD; 
Matthew Frank, MD, PhD; Neel Gupta, MD; Michael Khodadoust, 
MD, PhD; Arash Alizadeh, MD, PhD; Sunil Reddy, MD; Andrew 
Rezvani, MD; Irene Wapnir, MD 
Responsible Research 
Coordinators:                       Ami Okada 
Responsible Data 
Manager:  
Ami Okada 
 
Biostatistician:                     Phillip Lavori, PhD 
 
Protocol Version:                 Amendment 5  April 4, 2016  – 
 
(Supersedes Version Amendment 4 dated December 18 2012 ) 
 
 
 


 
 
 
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STUDY SYNOPSIS 
 
 
TITLE 
Phase 1/2 Study of a CpG-Activated Whole Cell Vaccine 
Followed by Autologous “Immunotransplant” for Mantle 
Cell Lymphoma 
STUDY PHASE 
Phase 1/2 
INDICATION 
Newly Diagnosed Adult Mantle Cell Lymphoma 
INVESTIGATIONAL PRODUCT 
OR PROCEDURE 
CpG-MCL Vaccine 
Vaccine-primed T cells 
PRIMARY OBJECTIVE(S) 
The primary objective of the study is to evaluate freedom 
from molecular residual disease at one year post-autologous 
transplant.  
SECONDARY OBJECTIVE(S) 
Secondary objectives are Time To Clinical Progression 
(TTP),  and evaluation of anti-tumor immune responses 
after vaccination, and after immunotransplant.   
TREATMENT SUMMARY 
Patients will undergo excisional tumor biopsy or apheresis 
(for patients with significant peripheral blood involvement), 
to obtain at least 1.5 x 109 malignant cells, which will be 
used to produce a patient-specific “CpG-MCL” vaccine.  
Patients will receive standard induction chemotherapy.  
Once in remission and after recovery of sufficient number 
of T cells, patients will receive three ‘priming’ CpG-MCL 
vaccinations at within a period of 21 days (4-7 day 
intervals).  Within approximately four weeks thereafter, 
patients will receive an infusion of rituximab as an in vivo 
“purge” as per standard institutional protocol, followed by 
leukapheresis to harvest vaccine-primed T cells.  In 
preparation for AHCT, patients will then undergo 
peripheral blood progenitor cell (PBPC) harvesting, 
myeloablative chemotherapy, and AHCT per standard 
institutional protocol.  Between day 1 and day 3 post-
AHCT, patients will receive an infusion of their primed T 
cells together with a booster vaccination of CpG-MCL. 
After hematopoietic recovery post transplant, patients will 
receive a second booster vaccination. 
 
SAMPLE SIZE  
The goal is to enroll patients sufficient to obtain a total of 
59 evaluable patients over 60 months 
STATISTICAL 
CONSIDERATIONS 
Primary Endpoint 
The primary endpoint of the trial is freedom from molecular 
residual disease at the landmark of one-year post-transplant.   
 
Based on the 2-Stage Simon Optimal Design with both 
error rates below 0.1 for testing a null (unacceptable) rate of 
MRD of 70% against an alternative (acceptable) rate of 
85% has 20 patients acquired in the first stage, stopping for 
futility with 14 or fewer successes (MRD negative 
patients), otherwise going on to a total of 59 patients, 


 
 
 
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choosing the experimental treatment if there are at least 46 
successes (MRD negative patients) and results determined 
favorable to move to additional clinical investigation. If the 
true rate is 70%, the expected number of patients is 36, and 
the probability of stopping at stage 1 is 58%. This design 
minimizes the expected number of patients exposed to an 
ineffective treatment, given the error rates. 
 
In addition, to determine if chemotherapy regimen does 
influence MRD rates, we will stratify MRD status by 
chemotherapy regimen. 
 
Secondary Endpoints 
Secondary outcome analysis will focus on TTP (from the 
date of autoHCT). TTP among patients evaluable will be 
reported including measure of centrality and variance of the 
outcome.  
 
The immune response will also be reported descriptively at 
the completion of the trial.  
 
Correlative Biomarker Endpoints 
At the completion of the trial, we will explore new putative 
immune biomarkers of clinical response (molecular MRD 
at 1 year).  
 
Stopping rules: 
Though similar studies using PF-3512676, whole-cell 
cancer vaccines, and re-infusion of T cells post-transplant 
have shown minimal associated adverse events, we will 
assess the patient cohort in an ongoing manner and use the 
following stopping rule for the Serious Adverse Events of: 
non-engraftment or early death (within 100 days from 
transplant) from any cause.  
 
After patient number: 
 
8 
16 
24 
Non-engraftment seen in:  
2 
4 
5 
Early mortality seen in:              3      
5 
6 
 
This rule would stop the study if the statistics indicate the 
possibility with even 80% certainty that the non-
engraftment rate is > 10% or the early mortality rate is > 
15%. 
 
 


 
 
 
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Figure 1:  Treatment schema (for time interval details, see Study Calendar section 8.0) 
high dose chemo 
stem-cell rescue 
month 
 
 
  
 
induction 
chemotherapy 
       harvest primed T cells 
PBSC mobilization  
harvest stem-cells  
0 
. 
. 
.  
4 
 
 
  
8 
 
 
 
10 
 
 
 
12 
 
MCL 
biopsy 
primed T cells 
CpG 
MCL 
CpG 
MCL 
CpG 
MCL 
rituximab  


 
 
 
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TABLE OF CONTENTS 
 
 
Page 
STUDY SYNOPSIS & DESIGN .................................................................................................... i 
TREATMENT SCHEMA .............................................................................................................. iii 
 
1. OBJECTIVES ............................................................................................................................6 
 
2. BACKGROUND .......................................................................................................................6 
 
2.1 Rationale  .............................................................................................................................6 
 
2.2 Study Agents ......................................................................................................................14 
2.2.1 Autologous CpG-Activated Lymphoma (CpG-MCL)  ..........................................14 
2.2.2 Autologous Primed T Cells ....................................................................................15 
 
3. STUDY DESCRIPTION 
 
3.1   Patient Inclusion Criteria  ................................................................................................16 
3.2   Patient Exclusion Criteria  ...............................................................................................16 
 
3.3   Inclusion of Women and Minorities ................................................................................16 
 
3.4   Screening Evaluations ......................................................................................................16 
 
3.5   Enrollment ........................................................................................................................17 
 
4. TREATMENT PLAN ..............................................................................................................17 
 
4.1  
Agent Administration ..................................................................................................17 
 
4.2  
Monitoring ...................................................................................................................20 
4.3       Concomitant/excluded medications .............................................................................20 
4.4       Patient Follow-up and Adverse Event Assessment ......................................................20 
4.5       Definition of Treatment-limiting Adverse Event .........................................................20 
4.6       Supportive Care Guidelines .........................................................................................21 
4.7       Duration of Therapy .....................................................................................................21 
 
5. EXPECTED ADVERSE EVENTS/ DOSE MODIFICATIONS ............................................21 
 
5.1  
Expected Adverse Events ............................................................................................21 
 
 
5.1.1 
CpG-MCL ..............................................................................................................21 
5.1.2 Vaccine Autologous Primed T Cell Re-infusion ...................................................21 
5.1.3 Excisional Biopsy ..................................................................................................21 
 
5.2  
Dosing Delays/Dose Modifications .............................................................................22 
 
 
 
6. AGENT FORMULATION AND PROCUREMENT ..............................................................22 
 
7. CORRELATIVE/SPECIAL STUDIES ...................................................................................22  
 
8. STUDY CALENDAR..............................................................................................................24 
9. MEASUREMENT OF EFFECT ..............................................................................................27 
10.  REGULATORY AND REPORTING REQUIREMENTS ....................................................27 
 
10.1 
Adverse Event Reporting .............................................................................................27 


 
 
 
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10.1.1 Definitions ..............................................................................................................27 
 
10.2 
Adverse Event Reporting Guidelines ...........................................................................28 
 
 
10.2.1 Forms .....................................................................................................................29 
 
 
10.2.2 Secondary Malignancies ........................................................................................29 
 
10.3 
Data Reporting .............................................................................................................29 
 
 
11. STATISTICAL CONSIDERATIONS .....................................................................................29 
 
REFERENCES ..............................................................................................................................32 
 
APPENDICES 
 
 
 
APPENDIX A 
 
 
Performance Status Criteria ...............................................................................................36 
 
 
 


 
 
 
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1. OBJECTIVES 
 
The primary objective of the study is to evaluate freedom from molecular residual disease at one 
year post-autologous transplant.  
 
Secondary endpoints are Time To Clinical Progression (TTP),  and evaluation of anti-tumor 
immune responses after vaccination, and after immunotransplant.   
 
An additional consideration will be to determine feasibility; specifically what proportion of 
enrolled patients will be able to complete the entire immunotransplant protocol.  
 
 
2. BACKGROUND  
2.1 RATIONALE 
2.1.1 Mantle cell lymphoma (MCL): prognosis and treatment  
Mantle cell lymphoma (MCL) is a distinct non-Hodgkin’s lymphoma (NHL) subtype[1], 
representing 4-6% of all NHL[2]. The characteristic molecular feature of MCL, the (11;14) 
(q13;q32) translocation, places the immunoglobulin heavy chain locus upstream of the BCL1 
gene, resulting in over-expression of its cyclin D1 gene product. In a series of 1361 patients 
diagnosed from 1988 to 1990, MCL carried the worst long-term failure-free and overall survival 
rate of any of the major subtypes[3]. The poor prognosis of patients with MCL makes it 
imperative to develop more effective treatments[4]. To date, no therapy has been shown to be 
curative for a significant proportion of patients, and, thus, there is no broadly recognized 
standard of care for MCL. Unless contraindicated by co-morbid conditions or very advanced age, 
most patients are currently treated with either a R-CHOP (cyclophosphamide, doxorubicin, 
vincristine, prednisone)-like regimen followed by AHCT or by dose-intense versions of CHOP 
sometimes alternated with methotrexate and/or cytarabine. While such a hyper-CVAD+MA 
regimen has shown encouraging results in single-institution phase 2 studies, it was associated 
with significant morbidity, an 8% treatment related mortality and less satisfactory results in 
patients over age 60 years[5].  CHOP-like induction followed by AHCT resulted in a median 
progression-free survival (PFS) of 39 months, significantly longer than the 17 months for 
patients randomized to consolidation with interferon (p=0.018). Though AHCT demonstrated a 
prolongation in PFS, there was a continuous pattern of disease recurrence in this phase 3 trial[6]. 
The addition of rituximab to induction therapy has yielded significantly higher response rates in 
randomized and non-randomized trials[7-10].  In the context of AHCT for MCL, molecular 
remission as defined by quantitative PCR of clonal IgH rearrangements, was strongly predictive 
for outcome (p< 0.0001), suggesting that molecular assessment could be used to measure 
efficacy of other consolidative therapies[11].     
 
There is evidence that MCL may be responsive to active immunotherapy.  Immunologically 
mediated graft-versus-lymphoma effect is evidenced after allogeneic stem cell transplantation by 
low relapse rates in patients already having failed AHCT[12, 13], clinical responses occurring 
after development of graft-versus-host disease (GVHD)[14], or after donor lymphocyte 
infusion[13].  In accord with the potential immunogencity of MCL, there have been several 
clinical trials of active immunization of MCL patients[15-17] with good evidence of immune 


 
 
 
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responses and some patients with remarkably good clinical courses. MCL tumor cells have 
demonstrated sensitivity in vitro to the TLR9 agonist CpG[18, 19] 
 
2.1.2 CpG Basic Biology and Vaccines  
Consistent with their B-cell lineage, MCL and other B-cell NHLs express Toll-Like Receptor 9 
(TLR9).  Ligation of TLR9 by its ligand oligodeoxynucleotides (usually 20-30 bases long) 
enriched for hypomethylated Cytosine-Guanosine repeats (CpG) activates a broad signal 
transduction network culminating in the upregulation of costimulatory molecules such as CD80, 
CD86, and CD54 as well as improved APC function with upregulation of MHC molecules[20, 
21].  Additionally, TLR9 ligation can lead to upregulation of fas, which transmits a pro-apoptotic 
signal to the NHL cell upon exposure to fasL expressing cells such as NK- or T-cells.  TLR9 
ligation by CpG molecules can also induce activation of antigen presenting cells such as 
dendritic cells that can then more effectively present tumor antigens from nearby apoptotic tumor 
cells.   
 
Hence, there are two mechanisms by which subcutaneous injection of CpG along with CpG-
activated NHL can induce systemic, tumor-specific, T cell mediated immunity: 
 
- CpG-NHL can directly present tumor antigens to T cells; 
- CpG-NHL undergoing apoptosis can transfer tumor antigens to nearby dendritic cells that 
can, in turn, be activated by CpG and more effectively present antigens to T cells. 
   
2.1.3 CpG in Murine Models of B-cell Malignancies 
We have recently published our initial findings of an in situ vaccination strategy using intra-
tumoral injection of CpG combined with cytotoxic therapies[22] and these data have shown the 
importance of co-localization of tumor antigens with the immuno-stimulant CpG.  In more recent 
studies, we have tested lymphoma cells cultured ex vivo with CpG as a therapetic vaccine.  Anti-
tumor T cells can be generated by such a vaccine. These T cells could be transferred to syngeneic 
recipients in conjunction with a hematopoietic stem cell transplant and mediate the cure of large 
established tumors [23]. This result provides the rationale for the design of the current clinical 
trial.  
 
2.1.4 CpG in Clinical Trials of B-cell Malignancies 
There have been numerous studies of CpG in patients with cancer[24-29] these studies have 
demonstrated the safety profile of CpG in over 1500 patients.  Based on our initial studies of in 
situ CpG vaccination[22], demonstrating the importance of co-localization of tumor and 
immuno-stimulant, we initiated a phase 1/2 trial of intra-tumoral CpG with low dose (2x200cGy) 
external beam irradiation for patients with recurrent, low-grade lymphoma.  Therapy was 
extremely well tolerated in all 15 patients with none experiencing adverse reactions greater than 
grade 2.  The only significant reactions were fever and flu-like symptoms lasting 1-4 days after 
injections.  Notably, there was a proof of the anti-tumor efficacy with one patient achieving a 
complete response, two patients with partial response, and seven patients with stable disease.  
We have also demonstrated the induction of tumor-specific, memory CD8 T cell responses 
resulting from vaccination, correlating temporally with the development of clinical tumor 
regressions (Figure 2)[30]. 
 


 
 
 
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Figure 2: A phase 1/2 clinical trial of in situ CpG vaccination for low-grade lymphoma. A) Schema of trial, B) 
Memory (CD45ROhi) CD8 tumor-specific immune response as seen by up-regulation of the activation marker 
CD137 upon co-culture with autologous tumor, induced by vaccination in a patient demonstrating a C) objective 
clinical response with resolution of retroperitoneal (and additional sites of) adenopathy. 
 
 
2.15CpG in MCL 
There have been several studies of in vitro CpG-activation of primary MCL tissues, which 
suggest an increase in antigen-presenting features[18, 19].   Together, these studies demonstrate 
up-regulation of CD40, CD54, CD80, CD86, MHC-I, and MHC-II as well as the well-described 
target of passive immunotherapy CD20. 
  
We have confirmed these findings using primary tumor samples from patients with newly 
diagnosed or recurrent classic MCL.  Culture with varying doses of CpG- oligodeoxynucleotides 
induced upregulation of MHC class I and II molecules as well as the co-stimulatory molecules 
CD80 and CD86 (Figure 3). 
 


 
 
 
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Figure 3:  CpG oligodeoxynucleotides induce an immunogenic pheynotype in primary MCL cells.  Single-cell 
suspensions of primary MCL cells from four specified patients were cultured with media alone (red line) or 2,10,or 
50g/ml of PF-3512676 (black, blue, and green lines respectively) at 37C, 5%CO2 for 72 hours, then flow 
cytometrically assessed for surface expression of the indicated antigen presentation or co-stimulatory molecules. 
 
2.1.6 CpG-MCL Vaccination Dose, Route, and Frequency 
CpG-MCL vaccination will be administered within a period of 21 days  (4-7 days between 
treatments is allowed), prior to leukapheresis for collection of primed T cells. The three 
‘priming’ vaccinations of CpG-MCL 108 cells are administered subcutaneously (s.c.) together 
with PF-3512676 18mg s.c.  The 18mg dose is well tolerated and was used in the majority of 
prior clinical studies (Investigator Brochure). The timing of the priming vaccinations is intended 
to maximize the amount of time since the immunosuppressive effects of induction 
chemotherapy, but minimize the delay time prior to myeloablative therapy and AHCT.  There is 
some evidence that a cancer vaccine can induce a tumor-specific immune response in MCL 
patients at this time point after induction therapy[17].    
       
Within 3 days post-transplant time-point, the vaccine primed T-cells will be administered i.v. 
(immunotransplant), followed by another s.c. dose of CpG-MCL vaccine with 18 mg of PF-
3512676. The final, post-transplant vaccination is given s.c. along with 18 mg of PF-3512676 at 
≥3 months post-AHCT, as medically feasible with resolution of interfering morbidities and 
medications. 


 
 
 
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The timing of the  post-transplant ‘boost’ vaccination is to coincide with the infusion of vaccine-
primed PBMCs.  The timing of the final, post-transplant vaccination is intended to allow an 
opportunity for priming of the reconstituted immune system, as CD8 T cell levels are generally 
regained by this time[31]. 
 
2.1.7 Vaccination During a State of Minimal Residual Disease 
Vaccination in the presence of tumor or tumor antigens specifically decreases immune response 
to those antigens[32-34]. Tumors secrete factors and recruit cells that impede tumor 
immunity[35-37]. In our recent trial of DC-Id vaccination for lymphoma, we found that 53% of 
patients with minimal residual disease mounted cell-mediated immune responses as opposed to 
0% in those with residual disease[38]. In the European MCL phase 3 study, 81% of MCL 
patients were in complete remission (CR) after the completion of high dose chemotherapy and 
AHCT[6]. These data suggest that vaccination in a state of minimal residual disease such as the 
post-AHCT setting will be possible and that the absence of tumor will facilitate the development 
of an anti-tumor immune response.  
 
2.1.8 Vaccination and Primed T Cell Re-Infusion post-AHCT (Immunotransplant) 
The period immediately following AHCT is a setting of severe lymphodepletion. Though 
myeloid reconstitution occurs within weeks after AHCT, lymphoid reconstitution, particularly 
that of B-cells and CD4 T-cells, takes several months[31, 39]. This time period has been 
considered a window of opportunity for adoptive transfer of primed T-cells to be more effective 
in eliminating cancer[40]. Levitsky et al, have clearly shown that adoptive transfer of otherwise 
unstimulated T cells can cure a majority of mice in a model system of non-Hodgkin’s 
lymphoma[41].  Dudley et al. have demonstrated the impressive clinical benefit of 
immunotherapy with adoptive T-cell transfer in the context of a lymphodepleted host with 
achievement of objective response rates as high as 50% in metastatic melanoma patients[42], 
including several instances of massive tumor reduction. The mechanisms suggested to explain 
this effect are two-fold. First, the relative reduction of T regulatory (Treg) cells with 
lymphodepleting chemotherapies[43, 44]) prevents inhibition of the administered vaccine. 
Secondly, there is decreased competition for the homeostatic proliferation cytokines (e.g. IL-7 
and IL-15), which allow for expansion of the administered effector cells[41, 45]. A recent 
randomized study of vaccination in the post-AHCT setting demonstrated that specific immunity 
was induced after AHCT only with adoptive transfer of primed T-cells (collected after priming 
vaccinations)[46]. Our earlier pilot trial[47] was based on these advantages provided by the post-
AHCT setting, and others have initiated vaccine strategies post-AHCT in AML and myeloma as 
well as indolent lymphoma.  
 
Our recent pre-clinical studies have shown that the homeostatic proliferation induced by the 
“empty” post-transplant recipient induces qualitative as well as quantitative changes in the 
population of transferred T cells.  Specifically, we have shown that there is a proportional 
increase in the proliferation of NK cells and CD4 or CD8 effector T cells relative to that of Tregs - 
as defined by foxP3 expression (Figure 4).  


 
 
 
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Figure 4:  Homeostatic proliferation of transferred splenocytes preferentially expands Teffector cells over 
Tregulatory cells  50 x 106 balb/c splenocytes were labeled with CFSE and injected by tail vein along with 5 x 106 bone 
marrow cells into A) recipient mice or B) recipient mice that had received lethal (900cGy) irradiation.  14 days later 
spleens were harvested and stained (above results gated for CD3+, CD4+ cells). 
 
Consistent with our finding that immunotransplant induces preferential proliferation of Teffectors 
over Tregs, we have shown that CpG-based vaccination, such as described in Section 2.1.3. is 
significantly enhanced by immunotransplant.  Specifically, the proportion of tumor-specific T 
cells induced in immunized donors is increased nearly ten-fold upon immunotransplant into 
lymphodepleted (‘empty’) recipients (Figure 5).   
 
Figure 5: Immunotransplant enhances the vaccine-induced tumor-specific T-cell response.  Mice received 
either: no vaccine, CpG-based lymphoma vaccine, or vaccinated-donor bone marrow and splenocytes after no 
irradiation (‘full’ recipients) or 900cGy TBI (‘empty’ recipients).   On day 15 post-transplant, peripheral blood 
lymphocytes were tested for lymphoma-specific IFN production.  Graphs gated for CD3(+) lymphocytes and 
statistics are IFN(+)cells as a percentage of all CD44hi cells.  
 


 
 
 
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Additionally, protection from subsequent lymphoma challenge increased from 70% with 
vaccination to 100% with immunotransplant.  Further, we have shown that the addition of a post-
transplant CpG-A20 vaccine boost further enhances anti-tumor immunity insofar as the transient 
tumor growth otherwise seen was eliminated with the addition of the vaccine boost (Figure 6).  
 
 
Figure 6: Immunotransplant of CpG-A20 vaccinated donors increases anti-tumor immunity.  Donor mice 
were vaccinated with irradiated CpG-A20 cells on days 1-6.  On day 13, one cohort of donors was left intact, one 
cohort had bone marrow and splenocytes transferred i.v. to recipients irradiated to 900cGy (immunotransplant), one 
cohort received the same immunotransplant plus an additional i.v. CpG-A20 ‘boost’ vaccination at the time of 
transplant.  On day 16 all cohorts were tumor-challenged s.c. and followed for tumor growth.  Statistics shown are 
proportion of tumor-free mice at day 30. 
 
Together, Figures 5 and 6 demonstrate that CpG-based vaccination induces a tumor-specific, 
memory T-cell immune response that is enhanced by immunotransplant.  Our recent 
demonstration of a tumor-specific T-cell immune response induced by CpG-based vaccination of 
low-grade lymphoma patients (Figure 2B) suggests that immunotransplant could have the same 
benefit in patients. 
 
 
2.19 Clinical Experience with CpG-MCL Vaccine + Immunotransplant 
This trial has now been open for over 6 years with the original primary end point of anti- tumor 
immune response. The study presently has a total of 59 patients consented, 58 vaccines prepared, 
44 vaccinated and 43 transplanted.  The current projection is that it will require another year to 
fully accrue the trial to eventually give sufficient power to determine the end point of MRD at 1 
year post transplant. So far patients are tolerating all the procedures well, with the possible 
exception of pneumonitis post transplant at a rate of approximately 50% that in some cases is 


 
 
 
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associated with a recovered viral organism.  In the majority of cases the pneumonitis has 
responded to steroid therapy and is reversible. This rate may be higher than historically seen with 
conventional stem cell transplant.  Notably, the BCNU dose in the preparative regimen was 
found as the major contributor and have now reduced this dose. A swimmer plot, time and event 
tracking, of all the patients accrued as December 2015 is shown below.  
 
 
Figure 7: Clinical Experience: Summary of Results to December 2015 
 
23 patients have been evaluated for MRD status at the landmark of 12 months post transplant by 
the technique of high throughput DNA sequencing on peripheral blood lymphocytes. Serial 
measurements on each of these patients are shown in the graph with each patient designated by 
the symbol/color. 19/23 patients are free of tumor DNA signal at a sensitivity of 1/100,000 
(dotted line on the graph).  Values below this level are of questionable significance. At least one 
of these patients with signals below the dotted line (blue symbol) as disappeared on further 
follow up. This value of 83% MRD at one year post transplant is trending to be superior to our 
benchmark historical rate of non vaccinated patients of 60% that we are trying to beat.  But this 
claim will require additional cases to achieve statistical significance.   
A plot the MRD measurements on each patient, up until December 2015 is shown below. 
 


 
 
 
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Figure 8: Molecular Residual Disease of all patients completing AHCT to Dec 2015 
 
The full immune response testing against the autologous tumor  has been completed on 21 
patients.  The data indicate that most, but not all, patients have made an anti-tumor immune 
response detected by at least one of the readouts of CD4 or CD8 T cells as detected in the PBL 
after recovery for stem cell transplant as shown below.  A new objective of this study is to relate 
these immune responses to other outcomes, such as MRD and PFS.  In parallel with these 
cellular measurements of immune response, high throughput sequencing of the T cell receptor 
repertoire at matching time points will be conducted.  T cell clones that rise with initial 
immunization and rise again on subsequent boosting after transplant are observed.  In some cases 
it has been possible to match the T cell clones that responded in vitro to those that responded in 
vivo.  An additional objective is to validate the in vivo high throughput TCR sequencing as a 
substitute for the in vitro cell stimulation test.     
 
2.2   STUDY AGENTS-Vaccine Preparation 
  
2.2.1 Autologous CpG-Activated Lymphoma (CpG-MCL) 
Patients with MCL will undergo excisional biopsy at Stanford University Medical Center prior to 


 
 
 
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initiation of induction therapy to yield at least 1.5x109 cells.  Alternatively, patients with 
significant peripheral blood involvement may undergo leukapheresis to yield the same number of 
MCL cells.  Biopsy samples will be dissociated into single cell suspension under sterile 
conditions, placed directly into Culture Media as per CpG-MCL SOP and split into two 
fractions: 1x109 cells for CpG-MCL production and the remainder will be stored in the vapor 
phase of liquid N2 for immune assays.  An aliquot from the immune assay fraction will be used 
for the development of patient-specific assays of molecular residual disease (MRD).  
 
Production of CpG-MCL 
CpG-MCL production is carried out per SOP included with this product’s approved IND, which 
is maintained on file with the Principal Investigator.  Briefly, a sterile aliquot of the MCL single-
cell suspension totaling 1x109 cells will be suspended in medium containing 3g/ml PF-3512676 
and cultured at 37C, 5%CO2 for 72 hours[18] to allow for up-regulation of antigen-presenting 
and co-stimulatory molecules, then irradiated to 200Gy[48], split into aliquots of 1x108 cells in 
Cryopreservation Media, composed of saline, human serum albumin, hydroxyethyl starch, 10% 
DMSO, and cryopreserved in the vapor-phase of liquid N2.   
 
Quality testing criteria of CpG-MCL is conducted as per SOP.  Briefly, CpG-MCL cells and an 
aliquot of 5x106 MCL cells cultured in parallel without PF-3512676 are assessed by flow 
cytometry for their ‘identity’ per non-expression of CD3, 4, 8, and co-expression of CD5 and 20 
and for their ‘activation’ per expression of the activation markers, MHC class I, II, CD25, 40, 54, 
69, 70, 80, and 86.  Identity is confirmed by the phenotype of CD5(+)CD20(+)CD3(-)and 
effective potency is defined by a > 2-fold up-regulation of at least 3 of the activation markers. 
 
Lot release criteria are as follows:  upon cryopreservation of at least 7 CpG-MCL cell aliquots in 
1mL, a sample (of less than 10% of volume of a vial) from one vial of the cells will be set aside 
and tested for bacterial, fungal, and mycoplasma contamination, endotoxin content (all pre-
freeze). If any of the above tests is positive, and at least 2 extra aliquots are available, each of 
those will be thawed and tested for all tests described.  CpG-MCL product will be deemed 
acceptable only if initial tests or if both duplicate repeat tests are negative.   
 
Immunization with CpG-MCL plus PF-3512676 
At each vaccination time point, one aliquot of CpG-MCL 1x108 cells will be thawed, aspirated 
into a 1ml sterile syringe, and given as a subcutaneous injection in the patient’s lateral thigh.  
Patients receive simultaneous, adjacent injections of the adjuvant PF-3512676 18mg s.c.  
Patients experiencing grade < 2 injection site reactions to prior vaccinations can receive 
subsequent vaccinations to alternate sites including the opposite lateral thigh or either deltoid. 
 
   
2.2.2  Autologous Vaccine-Primed T Cells 
Within approximately four weeks after the third priming CpG-MCL vaccination, patients receive 
standard rituximab 375mg/m2 (used here as an in vivo purge) followed within 72 hours by 
leukapheresis to obtain in vivo vaccine-primed T cells.  Leukapheresis is continued until 
sufficient PBMCs to obtain approximately 1 x 1010 CD3+ T cells are collected and these are 
immediately cryopreserved as per SOP.  This number of PBMCs is comparable to that obtained 


 
 
 
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after G-CSF mobilization of PBPC as is standard of care prior to autologous transplant, though 
this leukapheresis will be performed separately from and in addition to the standard one.  
 
 
3. STUDY DESCRIPTION 
 
Patient Selection: 
This is a single institution, single-arm, phase 1/2 study to evaluate the safety and efficacy of 
incorporating CpG-MCL vaccination, vaccine-primed T-cell infusion into a standard AHCT for 
newly diagnosed adult MCL patients. The enrollment goal is the number sufficient to obtain 59 
treated patients. 
 
 
3.1      Patient Inclusion criteria: 
At time of enrollment: 
 
Patients must be newly diagnosed with mantle cell lymphoma, have an accessible disease site 
for excisional biopsy or have sufficient peripheral blood tumor to leukapherese at least 1.5 x 
109 lymphoma cells in a single session. 
 
By standard clinical criteria, be medically appropriate to receive rituximab and standard 
induction chemotherapy and high-dose chemotherapy with AHCT. 
 
Must be between 21 to 70 years of age. 
 
Patients must be HIV negative. 
 
ECOG performance status 0, 1, or 2 or Karnofsky performance scale 50-100%. 
 
Patients must be capable of signing an informed consent. 
 
3.2      Patient Exclusion criteria: 
 
Patients who are currently taking immunosuppressive medications. 
 
Patients with severe psychological or medical illness. 
 
Pregnant or lactating women. 
 
At the discretion of the principal investigator if he/she feels that the patient is unable to safely 
complete the study.  Specifically, patients must be considered medically eligible to undergo 
standard high dose chemotherapy and autologous stem cell transplantation. 
 
 
3.3     Inclusion of Women and Minorities 
 
Both men and women of all ethnic groups are eligible for this trial.  
 
3.4 Screening Evaluations 
 
At the time of screening the following will be evaluated to verify eligibility: 
 History and Physical Exam 
 Performance Status (ECOG or Karnofsky) 
 CBC with differential, comprehensive metabolic panel (electrolytes, BUN, creatinine, 
calcium, total protein, AST, ALT) 
 Medically indicated imaging studies (e.g. PET-CT) 
 Bone marrow biopsy per standard of care, but required on follow-up only if results would 
distinguish between partial response (PR) and complete response (CR)  
 Informed Consent 


 
 
 
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3.5 Enrollment 
 
Eligible MCL patients will undergo excisional biopsy or leukapheresis to collect circulating 
tumor cells to cryopreserve at least 1.5 x 109 malignant cells.  This specimen will be used to 
make the patient specific CpG-MCL vaccine.  The following tests will be done prior to tumor 
cell collection: 
 
 Infectious Disease Testing:  HBV S Ag, HBV Core Ab, HCV Ab, HCV RNA, HTLV I/II 
Ab, HIV 1/2 Ab, HIV-1 RNA, syphilis (RPR) 
 
4. TREATMENT PLAN 
 
4.1  Agent Administration 
Patients will receive induction therapy by their primary oncologist with any of the accepted 
standard-of-care regimens for Mantle Cell Lymphoma and they must attain a PR or a CR and be 
eligible to proceed to AHCT.   
 
All investigational treatments as well as the AHCT will be administered at Stanford University 
Medical Center.  
 
Investigational treatments will be administered on an outpatient basis except for those doses 
administered while patients are in the hospital during AHCT. Expected adverse events and 
appropriate dose modifications for CpG-MCL vaccine are described in Section 5.  No 
investigational agents or therapies with the intent to treat the malignancy other than those 
described below may be administered while the patient is on this study.  
 
Post Chemotherapy Assessment 
Once in remission and after recovery of sufficient number of T cells, patients will be evaluated 
prior to receiving priming vaccinations.  Study related tests, procedures and imaging must be 
done within 30 days before starting study agent administration including:   
 
 History and Physical Exam  
 Performance Status (ECOG or Karnofsky) 
 CBC with differential; comprehensive metabolic panel (CMP) (electrolytes, BUN, 
creatinine, calcium, total protein, AST, ALT) 
 Medically Indicated Imaging Studies (e.g. PET-CT)  
 Obtain baseline research blood samples for immune assays and molecular residual 
disease testing. Leukapheresis [60 minute buffy coat] OR - 10 green top tubes  [100 mls] 
of blood will be drawn for immune assays; the ClonoSEQ® MRD test (Adaptive 
Biotechnologies) will be used to measure molecular residual disease 
 
Once it is considered medically feasible and any interfering morbidities and/or medications are 
resolved, patients will go on to receive priming vaccinations. 


 
 
 
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Priming Vaccinations 
Patients will receive three priming vaccinations administered within a 21 day period 
(approximately 4-7 days apart). Assessments and procedures are as follows: 
 
Within a period of 21 Days:  
 Performance Status (ECOG or Karnofsky) 
 Adverse event assessment  
 Vital signs – prior to and 60 minutes post vaccination  
 CpG-MCL vaccine s.c. injection (1e108 cells) 
 PF-3512676 – s.c. injection at the same site as vaccine (18 mg in a volume of 1.2 ml)  
 
Post-Priming Vaccinations 
Within approximately  four weeks after the third CpG-MCL priming vaccination, patients will 
receive an in vivo purge with rituximab 375 mg/m2 intravenously as per institutional standard of 
care. Within 72 hours of rituximab purge, patients will undergo leukapheresis to obtain vaccine-
primed T cells.  Assessments and procedures at the time of vaccine-primed T-cell collection are 
as follows: 
 
 Performance Status (ECOG or Karnofsky) 
 Adverse event assessment  
 CBC with differential, CMP Includes: electrolytes, BUN, Creatinine, Ca++, total protein, 
LFTs, within 72 hours of leukapheresis 
 Within 72 hours of rituximab dose, begin leukapheresis to obtain sufficient PBMC to 
contain approximately 1x1010 T cells  or goal identified by attending physician to ensure 
patient safety 
 Ten percent of the total T cell product will be retained for immune assays. 
 ClonoSEQ® MRD test will be drawn to assess molecular residual disease (MRD.) 
 
Patients then proceed to PBPC mobilization/harvesting followed by high dose chemotherapy and 
AHCT per standard institutional protocol.  
 
Post-AHCT Immunotransplant and CpG-MCL Vaccination 
Within 72 hours following AHCT, (day +1, +2 or +3 post-AHCT) patients will receive their 
vaccine-primed T cells and CpG-MCL vaccine.  Vaccine-primed T cells (PBMC containing 
approximately  0.5-1.5x1010 T cells) are thawed and re-infused over 15 minutes (as per standard 
practice of PBPC re-infusion). Approximately one hour following T-cell re-infusion, a dose of 
CpG-MCL is administered s.c. along with PF-3512676 18mg s.c. at the same site.  During 
inpatient therapies, patients receive continuous monitoring as per standard of care of patients 
undergoing AHCT.  Vaccinations and cell infusions will be initiated within one hour after cells 
are thawed. If no adverse reactions are observed, patients in the outpatient setting will be 
discharged.  Assessments and procedures are as follows: 
 
 Record Performance status (ECOG or Karnofsky) 


 
 
 
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 Draw blood for CBC with differential and comprehensive metabolic panel 
 Adverse event assessment 
 Infusion of vaccine-primed T cells 
 Record Vital signs – prior to infusion 
 CpG-MCL vaccine s.c. injection (approximately 1x108 cells) - approximately one hour  
post infusion of T cells  
 PF-3512676 – s.c. injection at the same site as vaccine (18 mg in a volume of 1.2 ml)  -  
 Record vital signs – approximately one hour after injections 
 
Post-AHCT, Immunotransplant and CpG-MCL Vaccine Assessment 
 
When considered medically feasible and any interfering morbidities and/or medications are 
resolved: 
 
 Record Performance status (ECOG or Karnofsky) 
  Adverse event assessment 
 Leukapheresis OR 10 green top tubes (100 mls), to obtain post-immunotransplant blood 
samples will be collected for immune assays 
 ClonoSEQ® test will be drawn to assess MRD. 
 
If patients are medically unable to undergo these procedures within 30 days post-
immunotransplant and the procedures will be delayed, the delay will be noted in the records and 
in the analysis of the data. 
 
Post-AHCT Final Vaccination 
Patients will receive a final vaccination with CpG-MCL s.c. along with PF-3512676 18mg s.c. at 
≥3 months post-AHCT as medically feasible upon resolution of any interfering morbidities and 
medications.  Evaluations as follows: 
 
 Record Performance Status (ECOG or Karnofsky) 
 Adverse event assessment  
 Obtain blood samples for immune assays (8 green tops [80 mls]) 
 Vital signs – prior to injections 
 CpG-MCL vaccine - s.c. injection (1e108 cells) 
 PF-3512676 – s.c. injection at the same site as vaccine (18 mg in a volume of 1.2 ml)  
 Vital Signs approximately one hour post injections 
 ClonoSEQ® test will be drawn to assess MRD 
 
Approximately 2 weeks following the final vaccination: 
 Record Performance Status (ECOG or Karnofsky) 
 Adverse event assessment  
 
Obtain Blood samples for immune assays (8 green tops [80] mls ) 
 
ClonoSEQ® test will be drawn to assess MRD 
  


 
 
 
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Follow-up  
Patients will be followed clinically for relapse with CT imaging per standard of care.  Molecular 
residual disease will be assessed at 1 year post-AHCT using the ClonoSEQ® test, or until disease 
progression.  Patients will be followed at the time of their standard care visits or with phone calls 
every 3 months the first year, and every 6 months for years 2-3 years post-AHCT or until 
progression.   
 
4.2  Monitoring 
A nurse and/or a physician will supervise all treatments.  During inpatient therapies (primed T 
cell infusion and CpG-MCL vaccinations) patients receive continuous monitoring as per standard 
of care of patients undergoing AHCT.  After all outpatient vaccine administrations (“priming” 
and 3 months post-transplant), patients will be monitored for approximately one hour for any 
acute adverse reactions.  Vital signs will be monitored prior to and approximately one hour 
following vaccination.   
 
4.3  Concomitant/Excluded medications 
All concomitant medications administered during the study will be recorded.   
 
 
4.4   Patient Follow-up and Adverse Event Assessment during Study 
 
Before each vaccine treatment, adverse event  and performance status will be assessed.   A 
physical exam will be done within a reasonable time period prior to the first priming 
vaccinations, prior to the immunotransplant, and at the time of the final CpG-MCL vaccine 
injection. Laboratory evaluations (including CBC with differential, LFT’s, creatinine, BUN) will 
be done within 7 days prior to the first priming vaccine, and a CBC with differential will be done 
within one day of the primed T-cell collection.  Concomitant medications will be monitored.   
 
Vital signs (BP, HR, RR, Temp) will be checked prior to and approximately one hour after 
administration of vaccine.  Study nurses under the supervision of a physician will monitor 
patients.  Adverse events will be graded using the NCI Common Toxicity Criteria for Adverse 
Events version 3.0. 
 
 
 
4.5   Definition of Treatment-Limiting Adverse Event  
The Investigators and the Stanford Comprehensive Cancer Center Data and Safety Monitoring 
Board (DSMB) will evaluate outcomes with respect to patient safety.  The primary adverse 
events of PF-3512676 as described in the Investigator’s Brochure include systemic reactions 
such as Grade < 2 flu-like symptoms, fevers, myalgias, and arthralgias lasting 12-72 hours and 
Grade < 2 injection site reactions including pain, erythema, and induration.  The primary adverse 
events of whole-cell cancer vaccines include primarily injection site reactions[48-57].  The 
primary adverse events seen in other studies of T cell re-infusion post-transplant[46] were a less 
than 20% incidence of transient flu-like symptoms, but no adverse events were treatment-
limiting.  In this study, unacceptable adverse events, including local or systemic grade > 3 
adverse event or toxicity thought to be related to the vaccine or immunotransplant, will result in 
discontinuation of vaccine therapy.  Grade < 3 adverse events or toxicities may prompt treatment 


 
 
 
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modifications, delay or discontinuation at the discretion of the investigators if there is evidence 
that unmodified continuation would increase the risk of adverse events. 
 
 
4.6   Supportive Care Guidelines 
Best supportive care will be administered. 
 
4.7   Duration of Therapy 
Treatment will consist of three priming CpG-MCL vaccinations, primed T cell infusion and two 
post-transplant CpG-MCL vaccinations unless one of the following occurs: 
 
 Lymphoma progression 
 
 Intercurrent illness that prevents administration of treatment 
 
 Unacceptable adverse events including local or systemic > grade 3 toxicity that prevents 
patient from continuing to participate 
 
 Patient is no longer eligible for AHCT 
 
 Patient decides to withdraw from the study 
 
 Non-compliance with the protocol, defined as inability to have all treatments, follow-up 
appointments and tests 
 
 General or specific changes in the patient's condition that render the patient medically 
unacceptable for further treatment in the judgment of the investigator. 
 
 Patient does not engraft (ANC <500 at Day 35 post-AHCT) 
 
Reasons for early trial discontinuation may include, but are not limited to, unacceptable adverse 
event or toxicity of study drug, a request to discontinue the trial from a regulatory authority, 
protocol violations, or poor enrollment.  
 
 
5. 
EXPECTED ADVERSE EVENTS/DOSE MODIFICATIONS 
 
5.1  Expected Adverse Events 
 
5.1.1  CpG-MCL (given with adjuvant PF-3512676).   
Local skin reaction at injection site may occur including pain, erythema, and swelling. Low-
grade transient flu-like reactions such as fevers, myalgias and arthralgias may occur. Serious side 
effects such as anaphylaxis and respiratory distress may occur but are unlikely.    
 
5.1.2  Vaccine-primed T-cell Re-infusion.  
Anaphylaxis, hypotension, or respiratory distress, all primarily related to the included 
cryopreservant (DMSO) may occur but are unlikely. 


 
 
 
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5.1.3  Excisional biopsy:  
Reactions common to all surgical procedures, e.g. risk of bleeding, scarring, infection, pain, and 
subsequent development of lymphedema. 
 
5.2 Dosing Delays/Dose Modifications 
Once in remission and after recovery of sufficient number of T cells, patients will receive 
priming vaccinations; the immunotransplant and CpG-MCL vaccine within 72 hours post-
AHCT, and the CpG-MCL vaccine >/= 3 months post AHCT.  If a patient develops intercurrent 
illness or adverse event during the study, the CpG-MCL vaccinations and immunotransplant 
infusion may be given once it is medically feasible and any interfering morbidities and/or 
medications are resolved.   If there is failure of lot release criteria for the first tested dose of a 
vaccine batch, and two additional lots are available, those will both be re-tested for all lot release 
criteria and if both of those additional lots pass all criteria, they may be used. 
 
All adverse events thought to be related to the study treatment will be followed until reasonable 
resolution.  All intercurrent illnesses and adverse events temporally associated with the 
vaccinations will be collected and documented. 
 
Dose modifications of the investigational agents are not planned. 
 
Patients experiencing local injection site reactions of grade 3 or higher will be taken off study.  
Patients experiencing local injection site reactions of grade 1 or 2 may receive subsequent 
injections in an alternate site (alternate thigh or either deltoid), or, per the discretion of the 
investigator, may be taken off study if additional vaccine doses are felt to be medically contra-
indicated.  
 
With evidence of grade < 2 allergic reaction during any vaccination or cell infusion, patients will 
receive additional acetaminophen and diphenhydramine unless contraindicated.  Should the 
patient develop grade 3 allergic reaction or greater, hypotension refractory to fluids, or 
bronchospasm, any incomplete vaccine or cell infusion will be terminated and the patient will be 
treated according to the standard of supportive care. 
 
6. AGENT FORMULATIONS AND PROCUREMENT 
 
Described under study drug section 2.2 above. 
 
 
7. CORRELATIVE/SPECIAL STUDIES 
 
CpG-MCL Phenotype and Functional Assessment: An aliquot of CpG-MCL will be set aside for 
flow cytometric analysis of surface markers including CD5,20,25,40,70,80,86, HLA-A,B,C, and 
HLA-DR as described in section 2.2.  
 
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and surface activation marker up-regulation are measurements of vaccine efficacy.   
 
An aliquot of each patient’s tumor cells are thawed and activated for 3 days with PF-3512676.  
Patient PBMCs (5 x 105) will be cultured with either medium alone or autologous tumor cells (5 x 
105) at 37C.  Afterwards, cells are stained with a panel of surface antibodies including CD4, 8, 
45RO, CD137 and CD278.  In other assays, brefeldin-A is added for the last 8 hours. Cells are 
fixed, permeabilized and stained for intra-cellular cytokines and enzymes such as IFN, TNF and 
, IL-2, perforin and granzyme.  
 
Handling of Serum samples:   Serum will be centrifuged at 2000 RPM for 10 minutes and stored 
at – 80 C for evaluation. 
 
Handling of PBMC:    PBMC are isolated by Ficoll-Hypaque centrifugation, washed and 
cryopreserved in: RPMI, 20% FCS, 10% DMSO and stored in liquid N2. 
 
MRD Testing: Tumor cells, PBMC and bone marrow cells and serum specimens will be 
subjected to high throughput sequencing of Ig variable region and T cell receptor genes, or 
alternatively real time PCR testing of Ig and T cell receptor genes with ClonoSEQ® (Adaptive 
Biotechnology, USA)


 
 
 
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8. 
STUDY CALENDAR 
         
           Procedures 
Screen Induction  
Chemo 
 
Pre-tx 
Visit / 
post- 
chemo 
Priming 
Vaccine 
within 
21 days 
Post-
Primin
g 
Vaccin
e 
within 
30 days 
 Pre-
AHCT 
within 
30-60 
days 
AHCT 
Day 0 
Post 
AHCT 
 withn 
3 days 
Post –
AHCT 
within 
30 days  
Final 
Vaccine 
≥ 3 mos  
Post -
AHCT 
Post-Final 
Vaccine  
Follow-up 
2 weeks  
 
Post-
AHCT 
Follow-up  
Months to 
3 years 
post AHCT  
History & Physical 
X 
  
X 
  
 
 
  
X  
 
X 
 
X  
(standard of 
care) 
Adverse Event Assessment 
 
 
X1 
X1 
X1 
X1 
X1 
X1 
X1 
X1 
X1 
 
Performance Status 
X 
  
X 
X 
X 
 
X 
X 
X 
X 
X 
 
Virology Labs (Standard of Care) 
X2 
 
 
 
 
X2 
 
 
 
 
 
 
CBC, CMP3 
X 
  
X 
  
X 
 
X 
 
 
 
 
 
Imaging studies review 
(Standard of Care)  
X14 
 
X14 
 
 
 
 
 
  
X14 
 
X14 
Lymph node biopsy or apheresis 
for peripheral blood tumor cell 
collection  
X 
 
 
 
 
 
 
 
 
 
 
 
BM biopsy (as needed) 
X4 
 
 
 
 
 
 
 
 
 
 
 
Induction Chemotherapy 
(Standard of Care) 
 
X5, 12 
 
 
 
 
 
 
 
 
 
 
CpG-MCL vaccine 108 cells 
 
 
 
s.c. 12, 15 
 
 
 
s.c. 11 
 
s.c. 12 
 
 
PF-3512676 18mg  
 
 
 
s.c. 12, 15 
 
 
 
s.c. 11 
 
s.c. 12  
 
 
Vaccine-primed T cells infusion 
(0.5-1.5x1010 PBMC)  
 
 
 
  
 
 
 
X 11 
 
 
 
 
Rituximab 375mg/m 
(Standard of Care)  
 
 
 
 
X8 
 
 
 
 
 
 
 
Leukapheresis  
 
 
X7 
 
X8 
X9 
 
 
X7, 
 
 
 
AHCT Preparation: 
mobilization, chemo  
(Standard of Care) 
 
 
 
 
 
 
X9 
 
 
 
 
 
 
AHCT (Standard of Care) 
 
 
 
 
 
 
X 
 
 
 
 
 
Immune assays  
 
 
X7 
 
X8 
 
 
 
X7, 
X10 
X10 
 
Molecular Residual Disease (MRD) 
with ClonoSEQ® 
 
 
X13 
 
X13 
 
 
 
X13 
X13 
X13 
X13  
MRD 
required  
only at 1 
year post-


 
 
 
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AHCT 
 


 
 
 
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STUDY CALENDAR FOOTNOTES 
 
1 Adverse event assessment.. 
2 Infectious disease screening panel to include HBV S Ag, HBV Core Ab, HCV Ab, HCV RNA, 
HTLV I/II Ab, HIV 1/2 Ab, HIV-1 RNA, syphilis (RPR) to be done within 7 days of tumor cell 
collection. 
3 CBC with differential, CMP Includes: electrolytes, BUN, Creat, Ca++, total protein, LFTs, 
Urine pregnancy test (if applicable).  
4 BM biopsy on enrollment per standard of care, but required on follow-up only if results would 
distinguish between PR and CR. 
5 May be at Stanford or with other primary oncologist (per standard of care).  
6   Footnote #6 deleted 
7 Leukapheresis (1 hour) OR 10 green top tubes (100 mls) to obtain cells for immune assays and 
MRD testing, plus 10 mls peripheral blood typically in one red top tube.   Post AHCT 
Leukapheresis will have a target date of within 30 days post-immunotransplant; if patient is 
medically unable to undergo the leukapheresis within 30 days post-immunotransplant and it will 
be delayed, the delay will be noted in the records and in the analysis of the data. 
8 A one time dose of Rituximab as per institutional standard of care, given within approximately 
four weeks after the third CpG-MCL priming vaccination.  
Within 72 hours of rituximab purge, begin Leukapheresis to obtain approximately 1 x 1010 CD3+ 
T-cell product or a collection goal identified by attending physician to ensure patient safety.  Ten 
percent of primed T-cell product will be retained for immune assays; CBC with differential, 
CMP (Includes: electroytes, BUN, Creat, Ca++, total protein, LFTs) within 72 hours of 
leukapheresis.   
 
9 AHCT preparation (mobilization, stem cell harvest and myeloablative chemotherapy) as per 
SUMC current protocol; (per standard of care – exact time period is not proscribed).  
10 Blood for immune assays is 8 green tops (approximately 80 mls) 
11 Post-AHCT immunotransplant and vaccination to be administered within 72 hours of AHCT. 
 
12 Once it is medically feasible and any interfering morbidities and/or medications are resolved, 
patients will receive the CpG-MCL vaccine and PF-03512676 injections. 
 
13Molecular Residual Disease (MRD) – approximately 15 mls of blood be collected for the 
ClonoSEQ® (Adaptive Biotechnology, USA) test at the following time points:  
 At the time of Leukapheresis or collection of 8GTTs for immune assays prior to vaccine 
administration 
 At the time of Leukapheresis for T cell collection 
 post-AHCT at 1 month 
 post-AHCT twice at approximately 3 months: once prior to vaccine #5, and once two 
weeks after vaccine #5 


 
 
 
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 post-AHCT at 1 year 
 at additional time points per investigator discretion. 
 
14Imaging studies will be reviewed as clinically indicated and available. 
15Priming vaccinations may be given 4-7 days apart (within a period of 21 days) 
 


 
 
 
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9. MEASUREMENT OF EFFECT 
 
The primary endpoint of this study is to determine freedom from molecular residual disease at 12 
months post autologous transplant, as has been previously validated to be predictive for 
subsequent clinical outcome in mantle cell lymphoma[6].   
 
A Secondary endpoint will be anti tumor immune response, as measured by intracellular 
cytokines and/or intracellular perforin/granzyme in CD8+ T cells, and/or CD137 induction on 
CD4+ T cells. 
 
Our hypothesis is that immunotransplant will significantly increase the proportion of tumor-
specific T cells relative to that induced by vaccination alone. 
 
Another Secondary endpoint will be the clinical outcomes of progression-free and overall 
survival.  Previous studies of ‘standard’ auto HCT for MCL patients have demonstrated median 
PFS and OS of 3.7 and 7.5 years, respectively. 
 
Progression is defined as per 2008 modification of standard “Cheson” criteria[58]. Additional 
clinical endpoints will be overall survival, The initial time point for all endpoints will be the date 
of transplant. 
 
Measurable disease will be detected at least by CT imaging, usually of the neck, chest, abdomen 
and pelvis and compared to baseline imaging.  In the event that a patient has additional sites of 
adenopathy that are best measured by other imaging modality, those sites will also be included in 
baseline and follow-up imaging.  All patients will require bone marrow (BM) biopsy at initial 
diagnosis as per standard of care.  Those patients with evidence of BM involvement at any time 
point may require follow-up biopsy to determine extent of clinical response per standard of care.  
 
10.  
REGULATORY AND REPORTING REQUIREMENTS  
 
10.1   Adverse Event Reporting 
 
10.1.1 Definitions: 
 
Adverse Events (AEs): An adverse event is any untoward medical occurrence in a patient 
treated with the vaccine during treatment and post-treatment follow-up period regardless of 
causality assessment.  This includes adverse clinical or laboratory findings, intercurrent illness or 
an exacerbation or progression of a disease/condition present at the time the study was initiated, 
other than signs or symptoms resulting from the disease being treated (MCL), which are 
considered lack-of-efficacy as opposed to an adverse event.  The study will collect adverse event 
information during the CpG-MCLvaccine treatments and for 2 weeks following the final study 
related procedure.  Adverse events will be followed until reasonable resolution.  Adverse Events 
will be graded according to the NCI CTCAE v3. Adverse events occurring during the patient’s 
induction chemotherapy will not be collected.  Adverse events occurring during standard of care 
treatments (i.e. stem cell transplant) and that are temporally associated with study related 


 
 
 
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treatment will be collected during the study.  
 
Serious Adverse Event (SAE): An adverse event which meets one or more the following 
criteria is considered serious. 
 Results in death 
 Is life-threatening 
 Requires or prolongs inpatient hospitalization 
 Is disabling 
 Is a congenital anomaly/birth defect 
 Is medically significant or requires medical or surgical intervention to prevent one of the 
outcomes listed above. 
 
Unanticipated Problem Involving Risks to Participants or Others (UPs) 
Serious Adverse Events that are: 
 Unexpected: Not in the consent form, investigator brochure, protocol, package insert, 
or label, or unexpected in its specificity, severity or frequency AND 
 Related to the research: Caused by, or probably caused by research activity. Events 
caused by progression of underlying disease are NOT related. If a device is involved, 
caused by, or associated with the device.  
 Caused harm or increased risk of harm: Involves harm to participants or others, or 
places participants or others at increased risk of harm. 
 
Unanticipated problems involving risks to participants or others (UP), that are unexpected and 
related and harmful must be reported to the IRB and to the FDA. 
 
 
10.2  Adverse Event Reporting Requirements 
 
Adverse Events 
Adverse events should be captured in the CRF. The research team will meet regularly to discuss 
AEs being experienced by the participants.  
 
Serious Adverse Events 
The Sponsor-Investigator shall promptly notify the IRB and the FDA in writing of the 
occurrence of any unexpected and “serious” adverse experience in accordance with GCP, 
regulatory agencies and institutional procedures and guidelines.  All serious adverse events 
occurring in patients at Stanford will be reported to the Sponsor- Investigator (also the PI) within 
10 days (5 days if the event is life-threatening or resulted in death). The PI will then report the 
events to Stanford Cancer Clinical Trials Office (CCTO) in accordance with the CCTO Standard 
Operating Procedures.  Deaths within 30 days of a patient being treated with the study drug will 
be reported within 5 days, all other SAEs will be reported within 10 days of the PI becoming 
aware of the event. If the event qualifies as an Unanticipated Problem Involving Risks to 
Participants or Others (UP), the PI will report this event to the Stanford IRB as per policy and to 
the FDA.  
 


 
 
 
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SAEs which are considered suspected adverse reactions will be reported by the Investigator to 
Pfizer (the provider of PF-3512676) using the MedWatch 3500a form. The Medwatch Report 
will be faxed to Pfizer U.S. Clinical Safety (using the Pfizer provided SAE Fax Cover sheet) at 
fax number 866-997-8322 within 24 hours of the PI being informed of the event. 
 
 
10.2.1 Forms 
 
FDA Form 3500a (MedWatch) will be used to report serious adverse events and UPs.  The form 
is available for download from the FDA website: 
http://www.fda.gov/Safety/MedWatch/HowToReport/DownloadForms/default.htm. 
 
 
10.2.2 Secondary Malignancies 
 
Investigators will report secondary malignancies occurring on or following treatment using the 
appropriate form noted above.  Exception:  Cases of secondary AML/MDS will be reported 
using the NCI/CTEP Secondary AML/MDS Report Form.  
 
10.3 
Data Reporting 
 
 
The data will be monitored continuously over the accrual and follow-up periods by the principal 
investigator (PI) and the study coordinator.  The PI is responsible for maintaining the clinical 
protocol, reporting adverse events, assuring that consent is obtained and documented, reporting 
unexpected outcomes, and reporting the status of the trial to the IRB and the data monitoring 
committee provided by the Stanford University Cancer Center Data Safety Monitoring Board 
(DSMB).  The DSMB will review the study data at scheduled intervals and when the data 
suggests safety threats to the patients.  DSMB responsibilities will be to review the general 
process and conduct of the study, including accrual, eligibility, data completeness, data 
timeliness, adverse events, consent forms, and trial renewal and amendments. 
There are no well-described toxicities of the CpG-MCL vaccinations in the above described 
studies[48, 56, 57]. 
 
11. STATISTICAL CONSIDERATIONS 
 
 
Study Design/Endpoints 
Primary Endpoint 
Statistical evaluation of our study is based on a Simon Two Stage Optimum Design powered to 
the endpoint of Minimal Residual Disease at the time point of 1 year post transplant. The rate of 
molecular remission at the landmark of 1 year post transplant is based on the literature that has 
validated this endpoint for prediction of continued clinical remission after autologous stem cell 
transplantation for Mantle Cell lymphoma (24-26).  To allow for early stopping if no clinical 
effect is observed, a 2-stage optimum design, following pre-specified probability of the null 
hypothesis (no effect on MRD rate) of 70% (p0=0.7) and alternative hypothesis of 85% (an 
increase in molecular MRD of 15% with vaccination; p1=0.85) with desired significance level 
(α) and desired power (1-β) of 0.1 and 0.9, respectively was used for sample size assessment.  
 


 
 
 
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Based on the 2-Stage Simon Optimal Design with both error rates below 0.1 for testing a null 
(unacceptable) rate of MRD of 70% against an alternative (acceptable) rate of 85% has 20 
patients acquired in the first stage, stopping for futility with 14 or fewer successes (MRD 
negative patients), otherwise going on to a total of 59 patients, choosing the experimental 
treatment if there are at least 46 successes (MRD negative patients) and results determined 
favorable to move to additional clinical investigation. If the true rate is 70%, the expected 
number of patients is 36, and the probability of stopping at stage 1 is 58%. This design 
minimizes the expected number of patients exposed to an ineffective treatment, given the error 
rates. 
 
The goal is to enroll patients sufficient to obtain a total of 59 treated patients over 60 months.   
 
Our current rate of accrual predicts that we will equal or exceed that goal. As recent data 
suggests RCHOP/DHAP alternating chemotherapy is superior to RCHOP or modified 
RHyperCVAD. As our null, or baseline response rate is based on MRD rates using RCHOP or 
modified RHyperCVAD, our endpoint analysis will exclude patients who receive 
RCHOP/DHAP chemotherapy (currently 3 of all enrolled patients will be excluded from 
analysis). Finally, to determined if chemotherapy regimen does influence MRD rates, we will 
stratify MRD status by chemotherapy regimen. 
 
Secondary Endpoints 
Secondary outcome analysis will focus on TTP (from the date of autoHCT).  In the 
aforementioned European MCL Network randomized trial, the median TTP was 39 months in 
patients consolidated with autoHCT. This is consistent with our own outcomes with MCL 
patients treated at Stanford with standard autoHCT. The trial is not powered to secondary 
endpoints. TTP among patients evaluable will be reported including measure of centrality and 
variance of the outcome.  
 
The immune response will also be reported descriptively at the completion of the trial. Note that 
this was the original endpoint that has already been reached and therefore a clinical endpoint has 
now replaced immune response as the primary endpoint to which the study is powered. Our prior 
primary endpoint of immune response was assessed using a 2-stage optimum design allowing 
early stopping for immunologic futility, following pre-specified probability of the null 
hypothesis (no immune response) of 5% (p0=0.05) and alternative hypothesis of 35% (presence 
of an immune response with vaccination; p1=0.35) with desired significance level (α) and 
desired power (1-β) of 0.05 and 0.9, respectively. Three of the first 6 patients showed an immune 
response as defined by a >10% change in 1 measure of the immune response. Currently 5 of the 
first 10 patients showed an in immune response. Therefore we previously determined early 
success of stage 1 and stage 2 (stage 1 required at least 1 immune response among 6 patients, and 
stage 2 required at least 3 immune responses among 17 patients). 
 
Correlative Biomarker Endpoints 
At the completion of the trial, we will explore new putative immune biomarkers of clinical 
response (molecular MRD at 1 year). As there will be approximately 300 potential comparisons 
to consider, we will evaluate each based on a desired false discovery rate (FDR) of less than 
20%. Putative biomarkers with a FDR of <20% will be selected for further study and validation 


 
 
 
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in subsequent clinical trials. For secondary analysis, we will model the relationship between the 
strength of clinical response to the vaccination and the change in the novel biomarkers with an 
FDR<20%. This is secondary, because it is not possible, at this point, to specify the distribution 
of response and whether the relationship between the response measure and the putative 
biomarkers will be strong or weak. 
 
Stopping rules: 
Though similar studies using PF-3512676, whole-cell cancer vaccines, and re-infusion of T cells 
post-transplant have shown minimal associated adverse event or toxicity, we will assess the 
patient cohort in an ongoing manner and use the following stopping rule for the Serious Adverse 
Events of: non-engraftment or early death (within 100 days from transplant) from any cause.  
 
After patient number:  
8 
16 
24 
Non-engraftment seen in:  
2 
4 
5 
Early mortality seen in: 
3 
5 
6 
 
This rule would stop the study if the statistics indicate the possibility with even 80% certainty 
that the non-engraftment rate is > 10% or the early mortality rate is > 15%. 
 
 
Sample Size/Accrual Rate 
The goal is to enroll patients sufficient to obtain a total of 59 evaluable patients over 60 months.   
 
 


 
 
 
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APPENDIX A: 
 
Performance Status Criteria 
 
 
 
ECOG Performance Status Scale 
 
 
Karnofsky Performance Scale 
Grade 
Descriptions 
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 
Deceased. 
0 
Deceased.