Clinical trial NCT01045460
Conditions: Multiple Myeloma
Keywords: multiple myeloma
Sponsor: Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins
Start date: 2010-01-15; Completion date: 2020-06-01
Enrollment: 36 participants; Allocation: RANDOMIZED; Intervention model: PARALLEL

Brief summary:
Patient Population: Patients with active myeloma (Stage II/III) that have completed induction therapy and are eligible for an autologous stem cell transplant.

Number of Patients: Will treat a total of 32 evaluable patients in a 1:1 randomization of aMILs vs aMILs plus vaccine. An evaluable patient is defined as one which has received the activated MILs and is at least 6 months post-transplant.

Study Objectives:

Disease response as determined by the Blade' criteria will be the primary endpoint of the trial at one year.

Additional study endpoints include progression free survival, parameters of T cell reconstitution, anti-tumor immune responses as well as the effect on osteoclastogenesis and clonogenic myeloma precursor cells.

Detailed description:

Arms: arm 1: Cyclophosphamide and filgrastim will be given to mobilize peripheral blood stem cells. Leukapheresis will be performed to collect peripheral blood from which activated marrow infiltrating lymphocytes will be produced. A melphalan conditioning regimen will be used prior to autologous stem cell transplant, and the MILs product will be administered on Days 3 and 4. arm 2: Cyclophosphamide and filgrastim will be given to mobilize peripheral blood stem cells. Leukapheresis will be performed to collect peripheral blood from which activated marrow infiltrating lymphocytes will be produced. A melphalan conditioning regimen will be used prior to autologous stem cell transplant, and the MILs product will be administered on Days 3 and 4. The allogeneic myeloma vaccine will be administered on Days 21, 60, 180, and 300.
Interventions: intervention 1: Activated marrow infiltrating lymphocytes intervention 2: Allogeneic Myeloma Vaccine intervention 3: Cyclophosphamide intervention 4: Filgrastim intervention 5: Leukapheresis intervention 6: Melphalan intervention 7: Autologous stem cell transplant
Intervention descriptions: intervention 1: Administered on Days 3 and 4. intervention 2: Allogeneic granulocyte macrophage colony-stimulating factor (GM-CSF)-based myeloma cellular vaccine. Administered on Days 21, 60, 180, and 300. intervention 3: Administered at 2.5 g/m\^2. intervention 4: Administered post cyclophosphamide daily until leukapheresis. intervention 5: Performed approximately 12 days post cyclophosphamide. Exact date depends on peripheral blood CD34+ cell counts. intervention 6: 100 mg/m\^2/day given on Days -2 and -1. intervention 7: Infused on Day 0.

Protocol documents:
Prot_SAP_000.pdf: 
J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
1 
 
Randomized Trial of Activated Marrow Infiltrating Lymphocytes alone 
or in Conjunction with an Allogeneic GM-CSF-based Myeloma Cellular 
Vaccine in the Autologous Transplant Setting in Multiple Myeloma 
 
 
 
 
 
P.I.  
 
 
Ivan Borrello, M.D. 
 
Co-Investigators: 
Carol Ann Huff, M.D. 
 
 
 
 
Nilanjan Ghosh, M.D, PhD. 
 
Statistician:  
Xiaobu Ye, M.D., Ph.D. 
 
Research Nurse: 
Anna Ferguson, R.N. 
 
Data Manager:  
Katie McIntyre 
 
Regulatory:  
Patricia Rennie 
 
 
 
 
 
Protocol History: 
Amendment 1:   
March 19, 2010 
Amendment 2:  
August 06, 2010 
 
 
 
 
 
 
Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins 
Bunting-Blaustein Bldg Rm 453 
1650 Orleans St.   
Baltimore, MD 21231 
410 955 4967 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
2 
 
Trial Synopsis 
 
Protocol Title:  Randomized Trial of Activated Marrow Infiltrating Lymphocytes alone 
or in Conjunction with an Allogeneic GM-CSF-based Myeloma Cellular Vaccine in the 
Autologous Transplant Setting in Multiple Myeloma 
 
Patient Population:  Patients with active myeloma (Stage II/III) that have completed 
induction therapy and are eligible for an autologous stem cell transplant. 
 
Number of Patients:  Will treat a total of 32 evaluable patients in a 1:1 randomization of 
aMILs vs aMILs plus vaccine.  An evaluable patient is defined as one which has received 
the activated MILs and is at least 6 months post-transplant. 
 
Study Objectives:   
Primary:  
 Response rate utilizing Blade’ criteria 
 
Secondary: 
 Progression-free and overall survival 
 Feasibility 
 Safety 
 Evaluate anti-tumor immune response 
 Evaluate the effect of aMILs on osteoclastogenesis. 
 Determine the effect of aMILs on  myeloma precursors.  
 
Eligibility Criteria: 
Inclusion: 
 Durie-Salmon Stage II or III multiple myeloma 
 Newly diagnosed receiving treatment or having completed induction therapy.  
 Relapsed myeloma  not previously transplanted within the past 5 years.  
 Measurable serum and/or urine M-protein from prior to induction therapy 
documented and available.  A positive serum free lite assay is acceptable. 
 Age  18 years old  
 ECOG performance status of 0 - 2  
 Meet all institutional requirements for autologous stem cell transplantation 
 The patient must be able to comprehend and have signed the informed consent  
Exclusion: 
 Diagnosis of any of the following plasma cell disorders: 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
3 
 
o 
POEMS syndrome (plasma cell dyscrasia with polyneuropathy, 
organomegaly, endocrinopathy, monoclonal protein [M-protein] and skin 
changes) 
o 
Non-secretory myeloma (no measurable protein on Serum Free Lite 
Assay) 
o 
Plasma cell leukemia 
o 
Amyloidosis 
 Use of corticosteroids (glucocorticoids) within 21 days of pre-transplant vaccine 
or bone marrow collection 
 Use of any myeloma-specific therapy other than lenalidomide within 21 days of 
pre-transplant vaccine. 
 Infection requiring treatment with antibiotics, antifungal, or antiviral agents 
within seven  days of vaccination or bone marrow collection 
 Participation in any clinical trial, within four weeks prior to vaccination or bone 
marrow collection on this trial, which involved an investigational drug or device 
 History of malignancy other than multiple myeloma within five years of 
vaccination or bone marrow collection, except adequately treated basal or 
squamous cell skin cancer.  
 Active  autoimmune disease (e.g., rheumatoid arthritis, multiple sclerosis, 
systemic lupus erythematosis) requiring systemic treatment. Hypothyroidism 
without evidence of Grave’s Disease or Hashimoto’s thyroiditis is permitted. 
 Evidence of spinal cord compression at time of transplant 
 Positive for HTLV 1 and 2 infection  


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
4 
 
Table of Contents 
Trial Synopsis .................................................................................................................... 2 
1 
Study Overview ..................................................................................................... 8 
2 
Objectives of the Study ....................................................................................... 10 
2.1 
Primary Objective ............................................................................. 10 
2.1.1 
Evaluate Response Rates utilizing the Blade’ criteria ........................ 10 
2.2 
Secondary Objectives........................................................................ 10 
2.2.1 
Evaluate Progression-free Survival and Overall Survival .................. 10 
2.2.2 
Feasibility of clinical design ............................................................... 10 
2.2.3 
Safety .................................................................................................. 10 
2.2.4 
Determine Tumor-specific Responses ................................................ 10 
2.2.5 
Effect of aMILs on Bone Metabolism ................................................ 10 
2.2.6 
Effect of aMILs on Myeloma clonogenic precursors ......................... 11 
3 
Background ......................................................................................................... 11 
3.1 
Multiple Myeloma .............................................................................. 11 
3.2 
Current Therapies for Multiple Myeloma ........................................... 12 
3.3 
Overview of Cell-mediated Immunity ................................................ 13 
3.4 
Immune Defects in Patients with Multiple Myeloma ......................... 13 
3.5 
Rationale for Immunotherapy of Multiple Myeloma .......................... 14 
3.6 
Rationale for the Use of CD3xCD28 Bead-Activated T Cells ........... 15 
3.7 
Clinical Data Using CD3xCD28 Bead-Activated T Cells .................. 16 
3.8 
Rationale for the Use of Activated MILs in Myeloma ....................... 17 
4 
Rationale for Study Design ................................................................................ 22 
4.1 
Patient Eligibility Criteria for Marrow Collection .............................. 22 
4.2 
Eligibility Criteria for Autologous Stem Cell Transplantation ........... 24 
4.3 
Data Collected from Diagnosis, Prior to Induction Therapy .............. 24 
5 
Baseline Evaluations ........................................................................................... 25 
6 
Randomization .................................................................................................... 26 
7 
Vaccination .......................................................................................................... 26 
7.1 
  Allogeneic Myeloma Vaccine ........................................................... 27 
8 
MILs Collection ................................................................................................... 27 
8.1 
Timing of MILs Collection ................................................................. 27 
8.2 
MILs Collection Procedure ................................................................. 28 
9 
MILs Activation Process andProduct Formulation......................................... 28 
9.1 
MILs Activation Process..................................................................... 28 
9.2 
Formulation Storage of Activated MILs T Cells ................................ 28 
10 
Mobilization and Collection of Peripheral Blood Stem Cells ......................... 28 
10.1 
Mobilization Regimen ........................................................................ 28 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
5 
 
10.2 
Peripheral Blood Stem Cell Collection ............................................... 29 
10.3 
Nomenclature for Numbering of Days ............................................... 29 
10.4 
Day –2 and Day –1: Melphalan .......................................................... 29 
10.5 
Autologous Stem Cell Transplant (Day 0).......................................... 29 
10.6 
Day 1 and Day 2 ................................................................................. 29 
10.7 
Supportive Care .................................................................................. 30 
10.8 
Post-Transplant Filgrastim .................................................................. 30 
11 
Administration of Activated MILs (Day 3) ...................................................... 30 
11.1 
Timing of Infusion of Activated MILs  (aMILs) ................................ 30 
11.2 
Evaluations Prior to Activated MILs T Cell Infusion ......................... 30 
11.3 
Pre-Medication Prior to Infusion of Activated MILs ......................... 31 
11.4 
Procedure for Administering Activated MILs .................................... 31 
11.5 
Safety Monitoring During Administration of Activated MILs ........... 31 
12 
Post-transplant Vaccinations (days 21, 60, 180, 300) ....................................... 32 
13 
Observation and Evaluation After Treatment with Activated MILs ............ 34 
13.1 
Evaluations From Day 4 Through Day 28 Post Transplant ................ 34 
13.2 
Evaluations from Day 60-360 Post Transplant ................................... 35 
13.3 
Management of Progressive Disease .................................................. 36 
13.4 
Discontinuation of Evaluations after Treatment ................................. 36 
13.5 
Contraindicated Medications .............................................................. 37 
14 
Risk and Toxicity Assessment ............................................................................ 38 
14.1 
Risks of Venous Access ...................................................................... 38 
14.2 
Risks of MILs Bone Marrow Collection ............................................ 38 
14.3 
Potential Microbial Contamination of the Activated MILs ................ 38 
14.4 
Potential Toxicity of Storage Solutions .............................................. 39 
14.5 
Potential Adverse Effects Associated with the Allogeneic Myeloma 
Vaccine ........................................................................................................... 40 
14.6 
Potential Adverse Effects Associated with Activated MILs............... 40 
14.7 
Potential Adverse Events Associated with Autologous Stem Cell 
Transplant ....................................................................................................... 41 
14.7.1 Peripheral Blood Stem Cell Mobilization Chemotherapy Regimen ... 41 
14.7.2 Administration of Filgrastim (G-CSF)................................................ 41 
14.7.3 Melphalan ........................................................................................... 41 
14.7.4 Leukapheresis ..................................................................................... 41 
14.7.5 Hematopoietic Stem Cell Infusion ...................................................... 42 
 
14.8 
Risks of Interactions Between Activated MILs, Tumor Vaccination & 
Autologous Stem Cell Transplantation ....................................................................... 42 
15 
Adverse Events ............................................................................................... 43 
15.1 
Case Report Form Reporting .............................................................. 42 
15.2 
Grading of Adverse Events and Toxicities ......................................... 43 
15.3 
Attribution of Causality ...................................................................... 43 
15.4 
Adverse Events Requiring Immediate Notification of IRB, IBC, FDA and 
NIH RAC ............................................................................................ 44 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
6 
 
15.4.2  Investigator Reporting Responsibilities .............................................. 45 
  15.4.3  Report of Adverse Events to the Institutional Review Board and 
Institutional      Biosafety Committee ............................................................... 45 
15.4.4  Investigator Reporting to the FDA and RAC ....................................... 45 
 
15.5 
Definitions of Response (Blade’ Criteria) .......................................... 47 
15.6 
Survival Endpoints .............................................................................. 50 
15.7 
Definitions of Engraftment and Graft Failure ..................................... 50 
 
16 
Statistical Considerations ................................................................................... 51 
16.1 
   Sample Size ...................................................................................... 51 
16.2 
Statistical Analyses/ Secondary objectives ......................................... 52 
 
17 
Ethical, Regulatory, and Administrative Considerations ............................... 54 
17.1 
   Informed Consent............................................................................. 54 
17.2 
   Institutional Review ......................................................................... 54 
17.3 
   Tissue Use for Research Purposes ................................................... 54 
18 
Study Monitoring and Data Collection ............................................................. 55 
 
18.1 
   Study Monitoring ............................................................................. 55 
18.1.1    Completion of Case Report Forms (CRFs) ...................................... 55 
18.1.2    Cell Therapy Laboratory (CTL) Cell-Processing Facility ............... 55 
 
18.2 
   Maintenance of Study Documentation............................................. 56 
18.2.1    Retention of Records........................................................................ 56 
 
18.3 
   Final Study Report ........................................................................... 56 
 
18.4 
   Investigational Product Labeling and Accountability ...................... 56 
18.4.1    Investigational Product Labeling ..................................................... 56 
18.4.2    Investigational Product Accountability ............................................ 57 
19 
References ............................................................................................................ 58 
Appendix  ......................................................................................................................... 61 
 
 
 
 
 
 
 


J0997/ NA_00029491 
               Johns Hopkins University 
BB-IND  14117 
    
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine       August 06, 2010 
8 
 
1 
Study Overview 
This Phase II randomized clinical study is designed to examine the efficacy of 
anti-CD3/CD28 activated marrow infiltrating lymphocytes (aMILs) alone or in 
combination with an allogeneic myeloma, GM-CSF vaccine in study subjects 
undergoing an autologous stem cell transplant for the treatment of multiple 
myeloma. 16 patients will be treated with aMILs alone and 16 with aMILs plus 
the vaccine.  
 
Patients will undergo a bone marrow aspiration to obtain marrow infiltrating 
lymphocytes (MILs) that will be used to produce aMILs.  The MILs can either 
be obtained at diagnosis prior to the initiation of induction treatment or upon 
completion of induction therapy.  MILs are collected by bone marrow aspiration 
of ~200ml of marrow.  This product will be used to expand the aMILs. During 
the in vitro expansion process, T cells will be activated and ex vivo expanded by 
co-stimulation with anti-CD3 and anti-CD28 monoclonal antibodies covalently 
attached to super-paramagnetic microbeads. Patients will be treated with a 
standard high-dose chemotherapy regimen for multiple myeloma consisting of 
single agent melphalan (200mg/m2).  Patients will then receive their stem cells. 
Three days (Day 3) following stem cell infusion, patients will receive a single 
infusion aMILs.  Because of the potential negative impact of G-CSF on T cell 
trafficking to the bone marrow, patients will not receive post-transplant G-CSF.  
For patients assigned to the vaccine arm, the first vaccine will be administered 2 
weeks prior to the bone marrow collection and the post-transplant vaccines will 
be administered on days 21, 60, 180 and 300.  The vaccine will consist of two 
irradiated allogeneic myeloma cell lines, H929 and U266 admixed with 
K562/GM-CSF. 
 
Disease response as determined by the Blade’ criteria will be the primary 
endpoint of the trial at one year. Additional study endpoints include progression 
free survival, parameters of T cell reconstitution, anti-tumor immune responses 
as well as the effect on osteoclastogenesis and clonogenic myeloma precursor 
cells. 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
 
9
 
Figure 1.1: Study Schema 
 
 
 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
10 
 
2 
Objectives of the Study 
2.1 
Primary Objective 
Evaluate the clinical efficacy of activated marrow infiltrating lymphocytes 
(aMILs) administered alone or in combination with an allogeneic myeloma cell 
vaccine combined with a GM-CSF producing bystander cell in patients 
undergoing an autologous stem cell transplantation setting for multiple 
myeloma.  
2.1.1 
Evaluate Response Rates utilizing the Blade’ criteria 
  Complete Response (CR) rate 
 Near Complete Response (nCR) rate 
 Very Good Partial Response (VGPR) rate 
 Partial Response (PR) rate 
 Minimal Response (MR) rate 
 Overall response rate (CR, VGPR, PR) 
2.2 
Secondary Objectives 
2.2.1 
Evaluate Progression-free Survival and Overall Survival 
Patients will be monitored for progression/relapse on Days 60, 180, and 360, 
and as clinically indicated. Following one year follow-up, patients will be 
followed annually for the next four years.  
2.2.2 
Feasibility of clinical design 
 
    Patient accrual 
 
   Adherence to study schema 
 
   Protocol violations 
 
   Drop-out rate 
2.2.3 
Safety 
    Treatment-related mortality 
    Grade 3 or greater hematologic toxicity 
2.2.4 
Determine Tumor-specific Responses 
 Evaluate tumor specific responses in blood and bone marrow 
 Examine T cell responses to DC-pulsed myeloma cell lines 
 Examine induction of novel antibody responses 
2.2.5 
Effect of aMILs on Bone Metabolism 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
11 
 
Parameters of bone turnover that will include: 
 RANKL/OPG ratio 
 Serum C Telopeptide levels 
 bAlkaline phosphatase and osteocalcin    
2.2.6 
Effect of aMILs on Myeloma clonogenic precursors 
 Examine side population of CD19 enriched PBLs throughout study.  
 
 
 
3 
Background 
3.1 
Multiple Myeloma 
Multiple myeloma is a plasma cell dyscrasia that is the most common cancer of 
the bone marrow.[1, 2] In 2008, the estimated incidence of myeloma in United 
States was 19,920.   
 
Multiple myeloma is most often diagnosed in middle aged and elderly 
individuals. The most common sites of disease are the bone and bone marrow. 
Malignant plasma cells arise from clonal expansion and accumulate in the bone 
marrow in masses known as plasmacytomas. These plasma cells produce large 
amounts of monoclonal immunoglobulins, most commonly IgG (50-60%) and 
IgA (20-25%) and occasionally IgD, IgM and IgE. [3]Patients often suffer from 
bone pain and skeletal fragility.[4]  Plasmacytomas are osteolytic in nature and 
often confined to the central skeleton, skull, and femur.  Bone destruction is 
usually localized but can be present throughout the skeleton.[5] 
 
The etiology remains unknown, but risk factors are thought to include chronic 
immune stimulation, autoimmune disorders, exposure to ionizing radiation, 
occupational exposure to pesticides or herbicides, occupational exposure to 
dioxin, and perhaps prolonged use of certain hair coloring products. [6, 7] 
 
The diagnosis is made using several criteria including results of radiographic 
skeletal survey, bone marrow examination, and measurement of serum and/or 
urine monoclonal protein (M-protein).[8, 9] Patients are also classified by stage 
(Stage I, II, III) according to the status of bone lesions, hemoglobin, serum 
calcium, β-2 microglobulin, C-reactive protein and M-protein.[10] The M-
protein is often used to monitor response to treatment via measurement of serum 
and/or urine analysis of Bence-Jones protein, the light chain component of the 
M-protein.  
 
Multiple myeloma is a largely incurable disorder and most patients will die of 
their disease.  A variety of chemotherapy agents have been used to treat the 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
12 
 
disease but few patients experience long-term disease-free survival with current 
therapeutic approaches.  The median overall survival for patients is 
approximately three years.[11, 12] There is an urgent need for more effective 
therapies to treat this challenging disease.   
3.2 
Current Therapies for Multiple Myeloma 
Treatment for multiple myeloma is dependent on the stage of the disease. 
Patients with Stage I disease are often monitored without treatment.  Patients 
with Stage II and III disease are usually treated with chemotherapy until a 
response is achieved.  For many years, conventional therapy has been the 
combination of melphalan and prednisone, which achieves an overall response 
in most patients.  Approximately 40% of patients demonstrate a greater than 
75% reduction in the M-protein.[13] However, nearly all patients will 
eventually fail this treatment and the median overall survival is only 3 years 
with this therapy.[13] A variety of different combination chemotherapy 
regimens have been developed in an attempt to improve on these results.  
Unfortunately, regardless of the type of initial treatment, the disease will recur 
and the 5-year survival is less than 30%.[13, 14]  
 
Dose intensification using high-dose chemotherapy followed by autologous 
stem cell transplantation has recently been used to increase the response rate 
and improve the outcome of patients with multiple myeloma.[15-26] In 1996, 
the French Myeloma Intergroup reported the results of a randomized clinical 
trial, which compared high-dose chemotherapy supported by autologous bone 
marrow support to conventional chemotherapy in patients with previously 
untreated multiple myeloma.[22] The study demonstrated the superiority of 
high-dose therapy in terms of response rate, event-free survival and overall 
patient survival.  This pivotal study has led to the widespread use of high-dose 
chemotherapy with autologous stem cell support as standard of care in multiple 
myeloma patients with good performance status. 
 
The Southwest Oncology Group study of 72 patients with chemotherapy-
refractory multiple myeloma treated with high- dose melphalan (200 mg/m2) 
followed by peripheral blood stem cell support was recently reported.[15](16) 
The regimen was well tolerated resulting in an overall response rate of 65% 
with approximately 30% of patients achieving complete remission.  The median 
progression-free survival was 11 months and the median overall survival was 19 
months.  These clinical results compare favorably to studies in this patient 
population from other clinical investigators.   
 
In the allogeneic transplant setting, long-term responses have been 
demonstrated.  However, this treatment is associated with severe graft versus 
host disease (GVHD) and substantial mortality, which has limited its use.[27]  


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
13 
 
 
A major goal of newer studies has been to increase the overall efficacy of 
autologous stem cell transplantation without added toxicity.  Clearly, the ability 
to impart a myeloma-specific immune response without the toxicity seen with 
allogeneic transplants offers significant appeal. Recent studies attempting to 
utilize vaccine approaches alone or in combination with adoptive 
immunotherapy have shed significant light into the potential efficacy of these 
approaches.  More importantly, these studies underscore the profound 
limitations of the current interventions and enabled the development of novel 
strategies with greater anti-tumor specificity.  
3.3 
Overview of Cell-mediated Immunity 
The human immune system is made up of many kinds of cells that are 
responsible for eliminating harmful invaders such as viruses or cancer from the 
body.  One type of lymphocyte, the T cell, plays a central role in orchestrating 
most immune responses.  T cells become activated when they recognize 
antigens, specific elements of microbes or tumor cells, as foreign to the body.  
This occurs when antigens are taken up, processed and presented by an antigen-
presenting cell (APC) to a molecular complex on the T cell.  This molecular 
complex contains the T cell receptor (TCR) associated with the CD3 signaling 
complex. [27] 
 
The primary signal for activating a T cell takes place when the TCR expressed 
on its surface binds to a processed antigen present on the surface of an APC.  
Each individual T cell only expresses a single TCR capable of recognizing a 
specific antigen.  However, the many billions of T cells found in the human 
body express millions of different TCRs, thereby enabling recognition of 
millions of distinct antigens.  Only a specific T cell that recognizes a particular 
antigen will become activated during a normal immune response.  
 
APCs must deliver a second signal in order to activate T cells.  This co-
stimulatory signal occurs when receptors on APCs bind to CD28 receptors on T 
cells.  Activation of T cells takes place when APCs bind to the TCR/CD3 
complex and CD28 receptor.  These activated T cells are exquisitely sensitive to 
further stimulation and also secrete a variety of chemical messengers called 
cytokines.  This process further augments the immune response both by driving 
continued activation and proliferation of T cells and recruiting and stimulating 
other cells of the immune system.  This cascade of events ultimately leads to 
destruction of pathogens such as tumor cells and viruses.   
3.4 
Immune Defects in Patients with Cancer Including Multiple Myeloma  
The inability of a patient's own immune system to respond to and control cancer 
may be due to a number of problems.  Defects in both the afferent (responding) 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
14 
 
as well as efferent (acting) arms of the immune system are well documented in 
cancer patients.  Deficits in the afferent arm of immunity include zeta chain 
defects in the TCR, which contribute to signaling problems in T cells.  In 
addition, patients with hematological malignancies including chronic 
lymphocytic leukemia and multiple myeloma demonstrate significant narrowing 
of the broad spectrum of T cell receptors present in healthy individuals.[28-30] 
This narrow T cell receptor repertoire may limit the patient’s ability to 
recognize and respond to tumor cells as well as other pathogens.  This may 
contribute not only to cancer progression, but also to the infections that are 
often observed in patients with hematological malignancies. These defects are 
both a result of the malignancy itself as well as cytotoxic therapy that can 
damage T cells.  Additionally, ineffective induction of CD40L (CD154) on T 
cells has been demonstrated in cancer patients.[31] Without CD40L signaling, 
APCs are not capable of being activated or presenting antigen to T cells.  Poor 
co-stimulation by APCs due to non-responsive elements or defects in the co-
stimulatory pathway has been observed in cancer patients.[27]   Problems with 
the effector arm of the immune system in cancer patients include the presence 
of relatively low numbers of cytotoxic T lymphocytes (CTL), which are 
required to kill the tumor cells.  Additionally, some cancers including multiple 
myeloma produce cytokines that inhibit the function of normal T cells or 
APCs.[32] The deficits in immunity may limit the ability of the patients' own T 
cells to mount an effective immune response to their own cancers.  
3.5 
Rationale for Immunotherapy of Multiple Myeloma  
Immunotherapy is one approach to improving the outcome of patients with 
multiple myeloma.  As noted above, defects in the host’s immune system are 
present in patients with cancer including multiple myeloma.  These deficits are 
thought to play a role in the patient’s inability to generate an anti-tumor 
response and control the disease.  A variety of therapeutic approaches are now 
being developed that stimulate the patient’s immune system.  Several groups are 
using idiotype vaccines to stimulate T cell-mediated responses to the patient’s 
tumor cells.[33, 34] Using this approach, patients are typically treated with an 
autologous transplant followed by vaccination with their own idiotype, which is 
derived from their unique M-protein.  Promising clinical results have been 
observed in some of these clinical trials.  However, many patients have 
weakened immune systems after the transplant and have been unable to respond 
to the vaccine[35]  
 
We have recently completed a clinical study utilizing autologous tumor 
vaccines in the autologous transplant setting.  In this study, newly diagnosed 
patients underwent a bone marrow collection to obtain autologous tumor that 
was combined with the K562/GM-CSF producing bystander cell line in the final 
vaccine formulation [36]. Patients were administered the vaccine pre-transplant 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
15 
 
in an effort to prime tumor-specific T cell responses in vivo.  The lymphocytes 
were then collected and infused at the time of transplantation in an attempt to 
impart an early anti-tumor effect.  The post-transplant vaccinations started 6 
weeks post-transplant and were administered every 3 weeks for a total of 8 post-
transplant vaccines.  The rationale for starting vaccinations this early post-
transplant is based on murine data demonstrating the existence of an early 
endogenous tumor-specific lymphocyte expansion that can then be maintained 
with vaccinations in the post-transplant setting [37]. While this study showed 
evidence of the generation of tumor-specific T cell responses, the degree of 
lymphopenia post-transplantation was greater than initially predicted with 
absolute CD4 numbers considerably below normal up to one year post-
transplant.  One attempt to increase vaccine efficacy is the ability to enhance T 
cell reconstitution and utilize vaccines at a time when maximal T cell 
responsiveness can be guaranteed. 
 
Recently, several investigators have documented the potent anti-tumor effects of 
donor lymphocyte infusions (DLI) administered to patients who relapse after 
allogeneic stem cell transplantation.[38-42] Unfortunately, a high incidence of 
GVHD has been observed with DLI, which has significantly limited its 
therapeutic application. 
 
Investigators have documented that T cells with anti-tumor activity are present 
in the blood of patients with multiple myeloma.[43] If sufficient numbers of the 
patient’s own T cells could be activated and expanded, they could be used in 
combination with an autologous stem cell transplant.  This would provide a 
potentially safer therapeutic alternative to DLI.  Patients would avoid the risks 
of GVHD as well as the substantial morbidity and mortality (up to 40%) that 
has been documented in multiple myeloma patients undergoing allogeneic stem 
cell transplantation. [44] 
 
Additionally, recent clinical data provide further rationale for the administration 
of autologous T cells in patients undergoing autologous stem cell 
transplantation.  Several clinical studies have documented improved therapeutic 
outcome in patients with multiple myeloma (as well as non-Hodgkin’s 
lymphoma, breast cancer, and ovarian cancer), who experience more rapid 
and/or complete recovery of their peripheral blood lymphocytes after 
autologous stem cell transplantation.[45-47]  
3.6 
Rationale for the Use of CD3xCD28 Bead-Activated T Cells 
Carl June and colleagues developed technology to activate T cells of the 
immune system outside of the body (ex vivo)[48].  This procedure is based on 
the roles of the CD3 signaling complex and CD28 receptor in the activation of T 
cells.  In the manufacturing process, T cells are stimulated ex vivo using 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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monoclonal antibodies that bind to the CD3 and CD28 molecules expressed on 
the surface of T cells.  The antibodies are attached to microscopic beads, 
thereby creating artificial APCs.  As a result, a universal reagent can be 
developed to activate all T cells.  Single beads, which coordinate CD3 and 
CD28 signals, optimize T cell activation and allow rapid expansion of T cells.  
Pre-clinical studies have shown that T cells can be generated ex vivo using these 
beads from patients with human immunodeficiency virus (HIV) or cancer. [49, 
50] 
 
Studies have demonstrated that T cells can be activated and expanded more than 
one hundred fold in less than 10-12 days with anti-CD3/CD28 bead activation 
with a predominant expansion of CD4 over CD8 T cells.  Further studies have 
shown that the patients’ activated T Cells express high levels of CD154 
(CD40L), CD137 (4-1BB) and other key effector molecules such as CD134 
(OX-40), CD54 (ICAM) as well as important receptors such as CD25 (IL-2 
receptor).  Finally, this process generates T cells that display a Th1 phenotype 
secreting high levels of IL-2 and interferon-gamma that are known to play 
essential roles in activating both helper T cells as well as cytolytic T cells.   
These features  demonstrate the ability to reverse tolerance in cancer patients 
and restore T cell responsiveness that may enable these T Cells to restore anti-
tumor immunity. 
3.7 
Clinical Data Using CD3xCD28 Bead-Activated T Cells 
A number of independent clinical trials have been conducted in which patients 
have been treated with T cells activated ex vivo using a CD3/CD28 bead-based 
technology. T cells activated in this manner have previously been tested in 
patients undergoing a autologous stem cell transplant for relapsed or refractory 
Non-Hodgkin’s lymphoma [51]. T cells were collected prior to high-dose 
chemotherapy. Fourteen days following the peripheral blood stem cell infusion, 
activated and expanded T cells were administered.  Three patients were treated 
at a median cell dose of 0.4 x 109, twelve patients were treated at a median cell 
dose of 1.6 x 109, and two patients were treated with a median cell dose of 9.8 x 
109. Infusion related toxicities experienced by the two patients at the highest 
dose level included transient fever, dyspnea, rigors and pulmonary edema. 
Maximal responses included five patients with complete responses, seven 
patients with partial responses, and five patients with stable disease.  
 
We recently completed a Phase I/II study in multiple myeloma in which 32 
patients were administered anti-CD3/CD28 activated T cells.  T cells were 
effectively expanded in all patients with an average fold-increase of 268 (± 
101).  T cells were then infused on day + 3 following an autologous stem cell 
transplant.  Interestingly, in addition to the in vitro expansion, patients 
experienced an additional in vivo expansion reaching maximal expansion on 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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day +21 that far exceeded the T cell numbers seen in a non-transplanted healthy 
individual.  However, despite the feasibility, safety and evidence of both in vitro 
and in vivo expansion, the overall clinical response showed CR 6%, PR 34% 
with an overall response rate of 40% which is no better than standard 
autologous stem cell transplants.  There were no significant toxicities related to 
the infusion of activated T cells and the majority of adverse events related to T-
cells were mild in severity and included fever (19%), chills (17%), asthenia 
(10%), headache (10%), and nausea (10%). 
 
3.8 
Rationale for the Use of Activated MILs in Myeloma 
aMILS have significant anti-myeloma activity  
 
From the above mentioned clinical studies, we have shown the ability of this 
technology to effectively 
overcome the inherent 
unresponsiveness seen in 
T 
cell 
from 
tumor-
bearing hosts and to 
expand 
upon 
anti-
CD3/CD28 stimulation.  
This 
data 
also 
underscores a significant 
limitation 
of 
the 
polyclonal 
expansion 
that 
lacks 
antigen 
specificity. 
 
To 
this 
effect, 
we 
have 
attempted 
to 
develop 
strategies 
aimed 
at 
increasing 
the 
tumor 
specificity 
of 
this 
approach.  Specifically, 
we have discovered that marrow infiltrating lymphocytes can be effectively 
activated and expanded with properties suggestive of an effector/memory 
population.  More importantly, they possess several critical features required for 
effective anti-tumor adoptive immunotherapy:  1) they can be activated and 
expanded to reasonable numbers; 2) they demonstrate significant specificity 
against  mature plasma cells.  PBLs or MILs were activated for 5 days in vitro 
and tumor specificity was assessed by determining their proliferative response 
to autologous tumor. As shown in Figure 1, whereas activated PBLs (aPBLs) 
failed to show measurable tumor specificity, activated MILs (aMILs) exhibited 
marked tumor reactivity.  Interestingly, no reactivity was appreciated against 
Figure 1: aMILs exhibit marked anti-myeloma activity.  
T cells from blood or marrow were either activated with anti-
CD3/CD28 beads or left unstimulated.  Tumor specificity was 
determined by incubating the cells either with autologous plasma 
cells (CD138), autologous non-malignant myeloid cells (CD33) or 
nothing and determining H-thymidine incorporation 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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normal hematopoietic elements.  Interestingly, in addition to their significant 
activity against mature plasma cells, aMILs were also capable of significantly 
limiting the outgrowth of clonogenic myeloma precursors suggesting a broad 
range of tumor antigen recognition. 
 
Another critical aspect of effective adoptive immunotherapy is the ability of T 
cells to traffic to the tumor microenvironment.  SDF-1 (stromal derived factor -
1) and its cognate chemokine receptor, CXCR4 are critical factors in cell 
trafficking in the marrow.  We have shown that a significantly higher 
percentage of MILs express CXCR4 as compared to PBLs thus increasing the 
likelihood of trafficking of these cells to the appropriate compartment.  [52].  To 
confirm this, we performed an experiment utilizing NOD/SCID mice.  Mice 
were irradiated and challenged with the H929 myeloma cell line and then 18 
days later either given activated MILs, activated PBLs or no T cells.  The T cell 
doses used corresponded to 
doses ranging from 3 – 20 x 
107 CD3/kg (doses easily 
achievable in humans).  As 
shown 
in 
Figure 
2, 
activated PBLs imparted no 
measurable 
anti-tumor 
effect compared to no T 
cells 
whereas 
the 
mice 
receiving activated MILs 
demonstrated 100% survival 
with 
no 
evidence 
of 
detectable 
tumor.  
Furthermore, T cells were 
detectable in the marrows of 
mice having received aMILs 
wereas no T cells but 
CD138+ plasma cells were 
seen in the bone marrows of 
mice treated with HBSS or 
aPBLs.  These findings confirm our in vitro data and suggest the overall 
efficacy of this approach. 
 
aMILs inhibit plasma cell colony outgrowth of myeloma progenitors  
The CD138+ plasma cell represents a terminally differentiated cell with minimal 
self-renewal potential.  
As such, it unlikely 
represents 
the 
clonogenic precursor of 
the malignant myeloma 
Fig 2: aMILs confer significant tumor-free survival 
advantage.  NOD/SCID mice were challenged with the human 
myeloma cell line, H929.  18 days later they either received 
HBSS, aPBLs or aMILs at doses ranging from 1-5 x 106 /mouse 
and were followed for tumor-free survival. 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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cell.  Complete eradication of mature plasma cells will seldomly result in 
sustained remissions.  When marrow from myeloma patients is depleted of 
CD34+ hematopoietic stem cells as well as CD138+ plasma cells and CD3+ T 
cells and plated on methylcellulose, there is an outgrowth of CD138+ plasma 
cells with the same light chain restriction of the initial myeloma clone. Early 
evidence that infiltrating T cells may mediate an anti-MM effect was provided by 
the observation that the number of MM colonies arising in the clonogenic assay 
were significantly increased if T cells were first removed from the marrow 
cultures prior to plating. With this knowledge and our data demonstrating a 
greater memory/effector phenotype of MILs over PBLs, we compared the 
number of colonies arising from CD34/CD138 marrow cells depleted of T cells 
to those seen when the same marrow precursors were co-incubated with either 
resting or previously activated PBLs or MILs. These studies showed a clear 
reduction of MM colony growth upon co-incubation with T cells.  Interestingly, 
while aMILs exerted the most potent effect on inhibiting plasma cell colony 
outgrowth, even the add-back of purified, unstimulated MILs demonstrated 
measurable anti-tumor efficacy (Fig 3). It should be noted that CFU-GM colony 
outgrowth was unaltered with the addition of aMILs suggesting the myeloma 
specificity of aMILs with no reactivity towards normal hematopoietic elements. 
These results demonstrate the ability of in vitro anti-CD3/CD28 stimulation to 
enhance antigen recognition and are consistent with the hypothesis that T cell 
recognition by aMILs occurs not only against antigens expressed on CD138+ 
plasma cells, but also on their progenitors – an observation that if true will 
significantly enhance the therapeutic efficacy of these cells. 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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In summary, several conclusions can be drawn from this data that make adoptive 
therapy utilizing aMILs an approach worthy of pursuing clinically:  1) the tumor 
micro-environment 
of myeloma patients 
contains 
lymphocytes (MILs) 
with greater anti-
tumor specificity 
than is observed in 
peripheral 
lymphocytes; 2) the 
kinetics of T cell 
activation of MILs 
compared to PBLs is 
consistent with 
expansion of a 
memory T cell 
population; 3) anti-
CD3/CD28 beads are 
capable of reversing 
tolerance and 
restoring T cell 
responsiveness to 
tumor-specific antigens; 4) the tumor specificity of aMILs greatly exceeds that of 
aPBLs; 5) aMILs offer the prospect of significantly impairing 
osteoclastogenesis; and 6) MILs exhibit specificity towards a broad range of 
antigens present on both mature plasma cells as well as their clonogenic 
precursors thus offering the theoretical possibility of eradicating the disease. 
aMILs inhibit osteoclastogenesis 
CD4+CD25+ regulatory T cells (Tregs) are responsible for many aspects of 
immune regulation which limit responses to auto-antigens in preventing 
autoimmunity, to allo-antigens in transplantation tolerance, and even pregnancy.  
In the tumor-bearing host, this regulatory T cell population plays a dominant role 
in suppressing tumor specific immunity and is likely key to the induction of 
tumor antigen associated tolerance. We have shown that the marrow of myeloma 
patients fails to contain Tregs - a finding with significant implications for 
adoptive T cell transfer.  
Fig 3: Inhibition of Clonogenic Outgrowth.  Bone 
marrow from myeloma patients was depleted of CD34+, 
CD138+ plasma cells, and CD3+ cells, resuspended in 
methylcellulose overnight and then plated out and scored for 
the development of CD138+ colonies cultured in the presence 
of various T cell populations. 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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In myeloma, both the tumor and the stroma are a major source of IL-6 which in 
combination with TGF induces an effector Th17 phenotype [53].  Recent 
evidence has suggested that a reciprocal relationship exists between Tregs and 
Th17 cells[54]. A significant amount of literature has recently emerged 
highlighting the role of Th17 cells in autoimmune processes including 
experimental 
autoimmune 
encephalitis, 
inflammatory 
bowel 
disease and rheumatoid 
arthritis. In attempting 
to 
understand 
the 
function 
of 
Th17 
effector cells within 
the myelomatous bone 
marrow, we examined 
the role of MILs on 
osteoclastogenesis 
whose 
activation 
is 
responsible 
for 
the 
lytic bone lesions seen 
in 
this 
disease.  
Osteoclasts 
were 
generated from bone 
marrow 
of 
normal 
donors or myeloma patients in the presence of M-CSF and Rank-L.  As shown in 
Fig 4, the addition of either IL-17 or Th17-skewed MILs significantly increased 
the number of osteoclasts generated whereas, the addition of IFN dramatically 
reduced these numbers.  Interestingly, co-culture of osteoclast precursors with 
aMILs (IFN producing Th1 cells) reduced osteoclast formation to a similar 
degree as IFN alone.  Taken together, these findings provide a biologic rationale 
as to the genesis and function of Th17 MILs in the myelomatous bone marrow 
and also explain the absence of Tregs in this disease as compared to normal 
marrow.  Furthermore, in addition to anti-myeloma activity of aMILs, they may 
also play a significant role in limiting osteoclast-induced bone destruction - a 
prospect that could have considerable therapeutic impact and justify the rationale 
for utilizing autologous marrow infiltrating lymphocytes as a means to augment 
myeloma-specific anti-tumor immunity.  
 
 
 
Fig 4:  The Th17 phenotype of myeloma MILs induces 
osteoclastogenesis.  BM from normal or myeloma patients was 
incubated with M-CSF/Rank-L alone or with the addition of IL-17, 
Th17 MILs, IFN, or activate MILs.  Osteoclasts were scored on 
day 21. 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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4 
Rationale for Study Design 
Current treatments for relapsed or refractory multiple myeloma are 
unsatisfactory. Conventional therapy using alkylating agents as well as the 
newer agents such as bortezomib, thalidomide or lenalidomide can generate 
responses, but disease recurrence and progression are inevitable.  More 
aggressive treatment regimens incorporating high-dose chemotherapy with stem 
cell support have led to increased response rates and improved survival.  
Nevertheless, only about 30% of patients achieve complete remissions [20] and 
few patients achieve durable remissions with this form of therapy.  There is 
increasing interest in using immunotherapeutic approaches in combination with 
high-dose chemotherapy and autologous stem cell transplantation to improve 
upon these results.  
 
Utilizing peripheral blood lymphocytes in a previously conducted clinical trial, 
T cells were reproducibly generated from all patients. Delivery of activated T 
cells in the transplant setting is attractive in light of data from both animal and 
human studies demonstrating the enhanced activity of T cells when 
administered in a lymphopenic setting [37, 55].  Furthermore, clinical data 
suggests that patients experiencing more rapid recovery of their peripheral 
blood lymphocytes after autologous stem cell transplantation have improved 
survival. [45-47] Based on these data, we propose to evaluate the safety, 
feasibility and therapeutic activity of aMILs after high-dose chemotherapy and 
autologous stem cell transplantation in patients with multiple myeloma. 
4.1 
Patient Eligibility Criteria for Marrow Collection 
Patients interested in participating on this study will sign consents.  Eligibility 
will subsequently be determined based on the inclusion and exclusion criteria 
listed below. Please refer to Patient Study Calendar, Appendix E.  It should be 
noted, that bone marrow may be collected prior to the initiation of therapy. 
MILs will not be expanded until the eligibility for the trial has been determined. 
 
To be eligible, the patient must meet all of the following inclusion criteria:   
 Previous diagnosis of multiple myeloma based on standard criteria as 
defined in Appendix A (Diagnostic Criteria for Multiple Myeloma). 
Tests need not be peformed within 30 days of vaccination or bone 
marrow collection. 
 Durie-Salmon Stage II or III disease as defined in Appendix B 
(Durie Salmon Staging for Multiple Mylemoma) at any time since 
diagnosis 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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 Measurable serum and/or urine M-protein from prior to induction 
therapy documented and available.  A positive serum free lite assay 
is acceptable. 
 Age  18 years old  
 ECOG performance status of 0 -2 (refer to Appendix C) 
 Life expectancy  6 months 
 Corrected serum calcium < 11 mg/dL, and no evidence of 
symptomatic hypercalcemia. (Corrected serum calcium is calculated 
by adding 0.8 mg/dL to the measured serum calcium for every 1 
g/dL that the serum albumin falls below 4.0 g/dL.) 
 Serum total bilirubin and SGPT (ALT) < 2.0 times the upper limit of 
normal 
 Serum creatinine < 2.0 mg/dL 
 The patient must be able to comprehend and have signed the 
informed consent  
At the time of consent signing, patients will be ineligible for the study if any of 
the following exclusion criteria apply: 
 
 Diagnosis of any of the following cancers: 
o 
POEMS syndrome (plasma cell dyscrasia with polyneuropathy, 
organomegaly, endocrinopathy, monoclonal protein [M-protein] 
and skin changes) 
o 
Non-secretory myeloma (no measurable protein on Serum Free 
Lite Assay) 
o 
Plasma cell leukemia 
 Diagnosis of amyloidosis 
 Previous hematopoietic stem cell transplantation within the past 5 
years. 
 Known history of HIV infection. 
 Use of corticosteroids (glucocorticoids) within 21 days of 
vaccination. 
 Use of any myeloma-specific therapy within 21 days of vaccination. 
 Infection requiring treatment with antibiotics, antifungal, or antiviral 
agents within seven  days of consent signing 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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 Participation in any clinical trial, within four weeks prior to 
vaccination or bone marrow collection on this trial, which involved 
an investigational drug or device 
 History of malignancy other than multiple myeloma within five 
years of consent signing, except adequately treated basal or 
squamous cell skin cancer.  
 Active autoimmune disease (e.g., rheumatoid arthritis, multiple 
sclerosis, 
systemic 
lupus 
erythematosis) 
requiring 
systemic 
treatment. Hypothyroidism without evidence of Grave’s Disease or 
Hashimoto’s thyroiditis is permitted. 
 Positive for HTLV1 and/or HTLV2 
4.2 
Patient Eligibility Criteria for Autologous Stem Cell Transplantation 
Prior to transplant, the patients must meet all of the following inclusion criteria: 
 Myeloma specific therapy with a minimum of 3 cycles.  
 Institutional critieria for and has institutional approval to undergo 
autologous stem cell transplantation as per CP1 and CP2 
(http://www.rig.onc.jhmi.edu/oncres/Protocols/CP1.pdf; 
http://www.rig.onc.jhmi.edu/oncres/Protocols/CP2.pdf) 
 Females of child-bearing potential must have a negative serum 
HCG test and be willing to use effective contraception (i.e. a 
hormonal contraceptive, intra-uterine device, diaphragm with 
spermicide, or condom with spermicide, or abstinence) up to Day 
180. 
Prior to transplant, patients will be ineligible for the study if any of the 
following exclusion criteria apply: 
 
 Evidence of spinal cord compression 
 Major organ system dysfunction including (but not limited to): New 
York Heart Association Class III or IV (Appendix D), pulmonary 
disease requiring the use of inhaled steroids or bronchodilators, 
renal, hepatic, gastrointestinal, neurologic, or psychiatric dysfunction 
which would impair patient’s ability to participate in the trial 
 
HIV infection 
4.3 
Data Collected from Time of Diagnosis, Prior to Induction Therapy 
M-protein results from the time of diagnosis, prior to induction therapy, will be 
recorded, These M-protein results are required for study eligibility, as stated in 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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25 
 
Section 4.1, Patient Inclusion Criteria. Additional data will be collected when 
available, but will not be required for entry into the study. This will include, 
e.g., date of diagnosis, type and duration of induction or other myeloma therapy, 
as well as data from the time of diagnosis, such as Durie-Salmon stage, beta-2 
microglobulin level, albumin, cytogenetics or FISH, bone marrow results, 
skeletal survey results, and measurement of any known soft tissue 
plasmacytomas.   
 
5 
Baseline Evaluations 
 
After the patient has been registered in the study, the following evaluations will 
be performed to determine the patient’s baseline status. All tests must be 
completed within 30 days of the MILs bone marrow collection.  
 Complete clinical exam, including complete medical history, review 
of systems, ECOG performance status, vital signs, weight, physical 
examination 
 Current medications 
 Serum chemistries: sodium, potassium, chloride, bicarbonate, BUN, 
creatinine, calcium, ALT, AST, total bilirubin, alkaline phosphatase, 
albumin  
 CBC with differential and platelet count 
 Beta-2 microglobulin 
 M-protein,  serum: 
- Protein electrophoresis 
- Immunofixation 
- Freelite (sensitive assay for kappa and lambda light chains) only if 
most recent immunofixation result showed no evidence of M-
protein 
 M-protein, urine (24 hour collection) for: 
Total protein 
Protein electrophoresis 
Immunofixation 
 Quantitative immunoglobulins (IgG, IgM, IgA) 
 Study blood (100ml of peripheral blood for research purposes) and 2 
green top tubes (for the evaluation of myeloma precursors) Bone 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
26 
 
marrow aspirate and biopsy.* Additional bone marrow sample (10-
15 cc) will be obtained for research purposes 
 Skeletal survey* 
 Bone turnover markers: Serum C telopeptide levels,  bAlkaline 
phosphatase and osteocalcin   
 *Acceptable if performed prior to the autologous stem cell 
transplant 
 
 
6 
Randomization 
There will be a 1:1 randomization to either aMILs alone or aMILs with the 
myeloma vaccine.  Randomization will be determined based on the following 
parameters: 
1. De novo vs relapsed myeloma 
2. International Staging System (ISS) stage 1 or 2 vs. 3 
3. Patients in complete remission vs not. 
 
7 
Vaccination 
Patients will be administered the allogeneic myeloma cellular vaccine pre- and 
post-transplantation.   
 
Vaccines will be numbered as follows: 
1 – pre-transplant vaccine (administered roughly 2 weeks prior to bone marrow 
collection) 
2 – d+21 post-transplant (+/- 3 days) 
3 – d +60 (+/- 10 days) 
4 – d+180 (+/- 2 weeks) 
5 – d+300 (+/- 4 weeks) 
 
 
CBC with differential will be performed on vaccine days  
 
Research Blood Collection: 
Vaccine arm: 
o  Patients will have 100ml of blood collected in heparinized 
syringes at the time of first vaccine administration and another 
100 mls on the day of the bone marrow collection.  This will 
enable us to determine the effects of vaccine priming on T 
cells. 
o Patients will have 10 mls of blood collected in tiger top tubes 
to be used for analysis of GM-CSF serum levels.  Blood will be 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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27 
 
collected prior to administering the vaccine and 2 days post 
vaccine at the following vaccine timepoints: 
 Vaccine 1 (pre-transplant vaccine) 
 Vaccine 2 
 Vaccine 4 
7.1 
  Allogeneic Myeloma Vaccine 
The allogeneic myeloma vaccine will consist of 3 cells: 
H929 – is an unaltered myeloma cell line grown in serum-free medium.  This is 
an IgA cell line isolated from the pleural effusion of a patient with end-stage 
myeloma.  The cell line possesses the t(4;14) as well as the N-13 ras mutation.  
It also expresses MAGE-A2 and NY-ESO-1. 
U266 – is an unaltered myeloma cell line isolated from a patient with plasma 
cell leukemia.  The line contains the t(11;14), overexpression of cyclin D1 and 
amplication of bcl-2.  It also expresses NY-ESO-1 and GAGE-3. 
K562/GM-CSF – is an erythroleukemia cell line modified to express stable, 
high levels of GM-CSF to be used as the GM-CSF-producing bystander.  This 
cell line was chosen in that it lacks HLA surface expression.  The cell line 
produces approximately 1500ng/106 cells/24hrs of GM-CSF. 
The actual vaccine formulation will consist of 5x107 H929 cells, 5x107 U266 
cells and 5x106 K562/GM-CSF cells.  All cell lines will be irradiated prior to 
cryopreservation.  Upon thawing, the appropriate volume of each cell line will 
be mixed and then multiple syringes will be prepared for injection.  Five 
intradermal injections will be administered over 3 limbs to exclude the 
dominant arm. 
The pre-transplant vaccine will be administered 2 weeks prior to the bone 
marrow collection.  
 
8 
MILs Collection 
The bone marrow collection will be performed to obtain the marrow infiltrating 
lymphocytes (MILs) required for the in vitro activation. The patient’s treatment 
schedule is summarized in Appendix E, Patient Study Calendar. 
8.1  
Timing of MILs Collection 
The MILs collection can occur at two separate times.  Either at diagnosis prior 
to the initiation of induction therapy or upon completion of induction therapy.  


J0997/ NA_00029491                                                            
Johns Hopkins University 
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8.2 
MILs Collection Procedure 
Each eligible patient will have approximately 200cc of bone marrow collected 
in syringes containing 2ml of 10,000 units/ml heparin under steady state 
conditions. The marrow will be collected in 60cc syringes.  Sedation and 
analgesia will be administered per standard institutional practices.   
 
 
9  
MILs Activation Process and Product Formulation 
9.1  
MILs Activation Process 
The MILs bone marrow product will undergo ex vivo activation and expansion 
of T cells in the Johns Hopkins Cell Therapy Lab (CTL) for up to 10 days. In 
the activation process, the T cells are activated and expanded by co-stimulation 
of T cells with anti-CD3 and anti-CD28 antibodies conjugated to super-
paramagnetic microbeads (ClinExVivo™ CD3/CD28 Paramagnetic Particles).  
The super-paramagnetic microbeads are removed at the completion of the 
process.  The product will not be released, unless acceptance criteria have been 
met. 
9.2  
Formulation Storage of Activated MILs T Cells 
The aMILs products will be formulated in multiple cryopreservation bags.  Each 
bag will contain aMILs formulated in a cryoprotectant consisting of 6% 
Hetastarch in 0.9% sodium chloride injection supplemented with 2% Human 
Serum Albumin (HSA) and 5% dimethylsulfoxide (DMSO),  The aMILs 
product will be frozen in a controlled-rate freezer until it reaches –80ºC.  The 
product will then be stored in the vapor phase of a liquid nitrogen freezer at less 
than -135ºC. 
 
 
10 
Mobilization and Collection of Peripheral Blood Stem Cells 
Following MILs collection, the patient may begin the mobilization regimen, 
which should be performed according to institutional standards.  The autologous 
stem cell transplant procedures will be carried out as per the institutional 
standard of care. 
10.1 
Mobilization Regimen 
The mobilization regimen will be cyclophosphamide 2.5-3 g/m2.  Filgrastim 
(Neupogen; G-CSF) will be administered following cyclophosphamide as per 
institutional standards. Hydration, mesna, and diuretics should be administered 
with the cyclophosphamide per institutional standards. 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
29 
 
10.2 
Peripheral Blood Stem Cell Collection 
Peripheral blood stem cell leukapheresis will be performed approximately 12 
days following cyclophosphamide depending on the peripheral CD34+ cell 
counts as per standard institutional practice. Leukapheresis will be performed to 
obtain a minimum of 2.0 x 106 CD34+ cells/kg.  Standard leukapheresis will be 
performed according to institutional guidelines for the collection of peripheral 
blood stem cells.  Up to 2% of the leukapheresis product will be stored for 
immune assays as long as the removal of these cells does not decrease the 
number of CD34+ cells frozen to below 2.0 x 10e6 CD34+ cells/kg. 
For patients failing to mobilize the peripheral stem cells, they will be offered the 
option of undergoing a standard bone marrow collection.   
10.3 
Nomenclature for Numbering of Days 
The day of infusion of the stem cells is called Day 0.  Days following 
autologous stem cell transplant are numbered accordingly, e.g., the first day 
after transplant is designated as Day 1. Days before the transplant are 
designated in the negative, e.g., the day prior to transplant is referred to as Day 
–1.     
10.4 
Day –2 and Day –1: Melphalan 
Melphalan will be administered on two consecutive days (Day –2 and Day –1) 
at a dose of 100 mg/m2/day intravenously in sterile water (5mg/ml) over 20-30 
minutes.  
 
CBC with differential will be obtained on Day –2 and Day –1 prior to 
Melphalan infusion. 
10.5 
Autologous Stem Cell Transplant (Day 0) 
A CBC with differential and platelets will be obtained prior to stem cell 
infusion. 
 
A minimum of 2.0 x 106 CD34+ cells/kg of stem cells will be infused. The 
infusion will occur the day after the last dose of melphalan. Corticosteroids may 
have an adverse effect on the activity of T cells contained in the stem cell 
product, and should therefore be avoided.  See Section 0, Contraindicated 
Medications. 
10.6 
Day 1 and Day 2 
A CBC with differential and platelet count will be obtained on Day 1 and Day 
2.  


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
30 
 
10.7 
Supportive Care 
All patients will receive supportive care according to institutional clinical 
guidelines for autologous stem cell transplantation. This may include but not be 
limited to allopurinol, menstrual suppression, prophylactic antibiotics, empiric 
antibiotics, intravenous immunoglobulin (IVIG), transfusion of blood products, 
hyperalimentation, and erythropoietin.   
10.8 
Post-Transplant Filgrastim 
Post-transplant filgrastim (Neupogen; G-CSF) will NOT be permitted on this 
trial because of the known ability to downregulate CXCR4 which may 
negatively impact on the ability of aMILs to traffic to the bone marrow 
following infusion. 
 
 
11 
Administration of Activated MILs (Day 3) 
11.1 
Timing of Infusion of Activated MILs  (aMILs)  
Activated MILs infusions will be scheduled for three days (Day 3) after the 
administration of the stem cell product.  Infusion may be delayed if required by 
cell processing issues or if warranted by the patient’s clinical condition.  
11.2 
Evaluations Prior to Activated MILs T Cell Infusion 
The following evaluations will be made prior to infusion of activated MILs: 
 
 Clinical assessment: review of systems, vital signs 
 CBC with differential and platelet count 
 Blood in a heparinized green top tube.  
 Serum chemistries: sodium, potassium, chloride, bicarbonate, BUN, 
creatinine, calcium,  ALT, AST, total bilirubin, alkaline phosphatase, 
albumin 
 Concurrent medication documentation 
 Adverse event documentation 
 
aMILs will not be infused if: 
 Temperature >38.5oC 
 Oxygen saturation < 90% on room air 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
31 
 
 Patient is felt to be clinically unstable by the medical team 
If aMILs infusion is delayed because of these issue, the cells will be 
infused after at least 24 hours from when the issues have resolved. 
11.3 
Pre-Medication Prior to Infusion of Activated MILs 
The patient will be pre-medicated with acetaminophen 650 mg by mouth and 
diphenhydramine hydrochloride 25-50 mg by mouth or IV, no greater than 30 
minutes prior to the infusion of activated MILs. These medications may be 
repeated every six hours as needed. A course of non-steroidal anti-inflammatory 
medication may be prescribed if the patient continues to have a high fever not 
relieved by acetaminophen.  
 
Intravenous diphenhydramine hydrochloride, epinephrine, hydrocortisone and 
supplemental oxygen will be available at the patient’s bedside in the event of 
anaphylaxis or an infusion reaction. 
 
Other medications may also be prescribed to treat additional side effects during 
the course of treatment.  However, there are some medications and therapies 
that may have adverse effects on the activity of activated MILs T Cell therapy 
and therefore are contraindicated (see Section 0, Contraindicated Medications). 
In particular, patients should not receive systemic corticosteroids such as 
hydrocortisone, prednisone, prednisolone (Solu-Medrol) or dexamethasone 
(Decadron) at any time, except in the case of a life-threatening emergency, since 
this may have an adverse effect on activated MILs. If steroids are required for 
an acute infusional reaction, an initial dose of hydrocortisone 100 mg is 
recommended.  
11.4 
Procedure for Administering Activated MILs  
For the administrating of the activated MILs, the patient will be hydrated with 
D5½NS at approximately 200 ml per hour for at least one hour. If the activated 
MILs product appears to have a damaged or leaking bag, or otherwise to be 
compromised, it will not be infused. Otherwise the activated MILs will be 
thawed and infused through standard blood tubing without an additional filter 
into a peripheral or central IV site. Each of the bags will be infused at a rate of 
approximately 10 ml per minute. Following aMILs infusion, the patient will be 
hydrated with D5W½NS at approximately 200 ml per hour for two hours.   
11.5 
Safety Monitoring During Administration of Activated MILs  
During the infusion of activated MILs, the patient’s blood pressure, heart rate, 
respiratory rate, temperature, and peripheral oxygen saturation will be 
monitored approximately as frequently as the following time intervals: 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
32 
 
 
 Immediately prior to infusion of first bag of activated MILs 
 15 minutes after initiation of first  activated MILs 
 30 minutes after initiation of first  activated MILs 
 1 hour after initiation of first  activated MILs 
 2 hours after initiation of first  activated MILs 
In the event of a severe allergic reaction (e.g., anaphylaxis, bronchospasm, 
hypotension) or other serious adverse reaction (i.e., cardiac failure, respiratory 
failure, severe nausea and vomiting) to the, activated MILs the infusion of the 
activated MILs will be stopped immediately and the patient will be provided 
with the supportive care deemed necessary by the medical staff at the clinical 
site.  The reaction should be noted as a serious adverse event, and the IRB 
should be notified within 24 hours.   
 
12 
Post-transplant Vaccinations (days 21, 60, 180, 300) 
 
The allogeneic myeloma vaccine will be administered on day +21, 60 (± 7 
days), 180 (± 14 days) and 300 (± 14 days) post-transplant assuming that 
patients show no evidence of ongoing transplant related toxicity. 
 
The day 21 post-transplant vaccine will be administered assuming the following 
criteria are met: 
 No active uncontrolled infection 
 Platelet count > 20,000/mm3* 
 Hemoglobin > 8 g/dL* 
 AST/ALT, total bilirubin < 3-fold normal 
*The patient need not be RBC or platelet-transfusion independent but 
must be able to sustain a Hgb >8g/dL or platelet >20,000 with 
transfusion 
 
At the post-transplant vaccination screen, patients will be evaluated for 
eligibility to proceed with vaccination.  The post-transplant vaccination will not 
be administered to patients with Grade 2 or higher hematologic toxicities based 
on the National Cancer Institute Common Toxicity Criteria (NCI-CTC) for bone 
marrow transplant studies (platelet count < 20,000/mm3, and hemoglobin 
<8g/dL).   
 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
33 
 
Patients who do not meet the eligibility for vaccination will be evaluated weekly 
until the blood counts improve to Grade 1 or better (platelet count > 
20,000/mm3 and hemoglobin >8g/dL) at which time they will receive the 
vaccine. 
 
The cell lines used for the vaccine preparation have undergone extensive 
regulatory testing and have been determined to be sterile and free of mycoplasma 
and viral contamination. The vaccine clinical lots were manufactured and released 
by the Cell Processing and Gene Therapy Facility, a cGMP compliant facility, at 
the Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins.  Dr. 
Borrello will submit the manufacturing and product release data to the FDA prior 
to initiating the study. The vaccine cells are irradiated prior to cryopreservation 
and stored frozen in the vapor phase of liquid nitrogen until the day of use. The 
details of production, irradiation, freezing, and preparation for administration are 
discussed in detail in the Investigational New Drug (IND) submission. Equal 
numbers (5 x 107 each) of the myeloma cell lines H929 and U266 cells will be 
combined with the bystander cell line K562/GM-CSF (5x106) to prepare the 
vaccine. Vaccine cells frozen in an injectable formulation Pentaspan® (10% 
Pentastarch in 0.9% sodium chloride with 2% human serum albumin and 5% 
DMSO) will be thawed on the day of vaccination in a 37ºC water bath, mixed,  
and then taken up into five labeled syringes prior to administration.  The syringes 
are kept on ice until administration.  Injections should be started within 60 
minutes of the thaw. 
 
Toxicities associated with the intravenous administration of Pentaspan® include 
circulatory overload, abnormalities of coagulation (prolonged prothrombin, partial 
thromboplastin and clotting times), and hypersensitivity reactions characterized 
by wheezing, urticaria, hypotension, and anaphylactic/anaphylactoid reactions.  
Additionally, patients who are allergic to corn can also develop hypersensitivity to 
Pentaspan®.    
 
Each vaccination will consist of five total intra-dermal injections, two each in the 
right and left anterior upper thighs, and one in the non-dominant upper arm 
(unless contraindicated). In the event that the specified limb is contraindicated, 
the dominant arm may be used. Vaccine injections sites shall be at least 5 cm at 
needle entry from the nearest neighbor. The approximate volume of each 
vaccination injection is approximately 0.3-0.4ml.  
 
A lidocaine based topical anesthetic (may include but is not limited to EMLA or 
ELA-MAX) cream will be recommended being applied to each injection site 
approximately 1-2 hours prior to vaccination to diminish the discomfort 
associated with the intra-dermal injections. “Approximately” allows for the 
topical anesthetic to be placed up to two and a half hours prior to the vaccine 
administration. The topical anesthetic is optional and participants are allowed to 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
34 
 
decline its use. Self-application of the topical anesthetic is allowed after the 
research participant receives instruction. If the research participant is non-
compliant with topical anesthetic administration instructions, the topical 
anesthetic will be applied by the research nurse. If the subject is allergic to 
lidocaine, the topical anesthetic will not be used. All research participants will be 
seen in the oncology outpatient center for vaccine administration and monitoring. 
 
Each dose will be administered on an outpatient basis. The subject must be 
observed in the clinic for at least 30 minutes after vaccination is completed. 
 
 
 
13 
Observation and Evaluation After Treatment with Activated 
MILs 
Follow-up evaluation of the patient will be carried out according to standard 
institutional guidelines for autologous stem cell transplantation.  In addition, the 
patient will have a limited number of tests performed specifically for this 
protocol.  Any toxicities felt to be related to the infusion of aMILs will be 
reported as adverse events. 
 
See Appendix E, Patient Study Calendar 
13.1 
Evaluations from Day 4 through Day 28 after Autologous Stem Cell Transplant 
The following tests will be performed daily from Day 4 until the day of 
confirmed engraftment for both neutrophils and platelets (See section 
13.7.1 for definition of engraftment) 
 
 Daily CBC (± 24 hours) with differential and platelet count 
(differential will only be performed if the total WBC > 100/µl as per 
standard institutional practice).   
 
The following tests will be performed on Days 7, 14, and 21  
 Serum chemistries: sodium, potassium, chloride, bicarbonate, BUN, 
creatinine, calcium,  ALT, AST, total bilirubin, alkaline phosphatase, 
albumin  
 Study blood draw (100ml) in heparinized syringes 
 Tiger top tube 10 cc  
 
 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
35 
 
The following will be performed on Day 28 (± 2 days) 
 Bone marrow biopsy and aspirate (collect 20ml for research 
purposes) 
 M-protein,  serum and urine: 
- Protein electropheresis 
- Immunofixation (if no measurable M-spike in serum or urine) 
- Freelite (sensitive assay for kappa and lambda light chains) only if 
most recent immunofixation result showed no evidence of M-
protein 
 Quantitative immunoglobulins (IgG, IgM, IgA) 
 Clinical assessment to include disease response/progression/relapse 
(the clinical assessment can be performed up to 10 days after the 
laboratory evaluation)  
 Concurrent medication documentation 
 Bone turnover markers: Serum C telopeptide levels and bAlkaline 
phosphatase and osteocalcin    
 Green top tubes  (total of 20 cc) 
 Study blood draw (100ml) in heparinized syringes 
 Tiger top tube 10 cc  
 
13.2 
Evaluations from Day 60-360 after Autologous Stem Cell Transplant 
The patients will be evaluated post transplant on day 60 ± 7 days, day 180 ± 14 
days, day 300 ± 14 days, and day 360 ± 30 days.  The day 300 visit will only be 
required of patients randomized to the vaccine arm of the protocol. 
 
 Clinical assessment: review of systems, ECOG performance status, 
vital signs 
 CBC with differential and platelet count 
 Serum chemistries: sodium, potassium, chloride, bicarbonate, BUN, 
creatinine, calcium,  ALT,  AST, total bilirubin, alkaline 
phosphatase, albumin 
 M-protein,  serum and urine: 
- Protein electrophoresis 
- Immunofixation (if no measurable M-spike in serum or urine) 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
36 
 
- Freelite (sensitive assay for kappa and lambda light chains) only if 
most recent immunofixation result showed no evidence of M-
protein 
 Quantitative immunoglobulins (IgG, IgM, IgA) 
 Disease response/progression/relapse assessment (the clinical 
assessment can be performed up to 10 days after the laboratory 
evaluation) 
 Concurrent medication documentation 
 Adverse event documentation until progression/relapse or Day 360, 
whichever occurs first 
 Bone marrow aspirate and biopsy.  The bone marrow examination 
will be performed on days 28, 60 ± 7 days, 180 ±  14 days and 360 + 
30 days.  Additional bone marrow (at least 20ml) will be obtained to 
assess for T cell subsets, as well as evaluation of tumor specificity.  
The samples need to be collected prior to vaccination. 
 Study Bloods:  
o Green top tubes  (total of 20 cc) 
o Study blood draw (100ml) in heparinized syringes 
o Tiger top tube 10 cc Bone marrow turnover markers: serum C 
telopeptide and bAlkaline phosphatase and osteocalcin These 
tests will not be performed on day 300. 
13.3 
Management of Progressive Disease 
Patients with disease progression/relapse shall have this documented.  Once the 
patient has progressed/relapsed, treatment will not be dictated or limited by the 
protocol, but the patient will continue to be followed for survival for five years 
from Day 0. 
 
It should be noted that since the purpose of this study is to examine the anti-
tumor efficacy of aMILs combined with a myeloma-specific vaccine, no post-
transplant maintenance therapy is permitted while on the study. 
13.4 
Discontinuation of Evaluations after Treatment  
There are several reasons that a patient may be discontinued from the post-
treatment protocol evaluations. These include, but are not limited to: 
 
 Patient decision to withdraw from the study 
 Protocol non-compliance 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
37 
 
 Patient lost to follow-up 
 Activated MILs product failed to meet specifications 
 Activated MILs product was not infused 
The reason for discontinuation of the protocol evaluations will be documented.  
The patient should continue to be followed annually for survival when possible. 
 
As described in Section 2.2.2, patients will be monitored for progression/relapse 
on Days 60, 180, and 360, and as clinically indicated. Following one year 
follow-up, patients will be followed annually for the next four years. 
13.5 
Contraindicated Medications 
There are some medications that may have adverse effects on the activity of 
activated MILs.  These medications and/or therapies should NOT be given to 
patients at any time during the entire study unless indicated in the protocol or to 
treat life-threatening conditions.  Patients should not be placed on any 
maintenance therapy following transplant. Contraindicated medications and 
therapies include: 
 
 Corticosteroids (e.g., hydrocortisone, prednisone, prednisolone [56], 
dexamthasone (Decadron), etc.). Inhaled steroids for the use of 
allergic rhinitis or pulmonary disease are allowed and not 
contraindicated.  
 Thalidomide 
 Interferon 
 Growth factors, interleukins, or other cytokines (except filgrastim as 
outlined in the protocol, or erythropoietin) 
 Cytotoxic agents 
(except 
cyclophosphamide for stem 
cell 
mobilization and high-dose melphalan as outlined in the protocol)   
 Other immunosuppressive drugs 
 Other experimental therapies 
 Radiation therapy  
 
NOTE: Corticosteroids treatment may have an adverse effect on the 
activity of activated MILS. Each patient’s hospital and clinic chart will be 
labeled 
“NO 
STEROID 
MEDICATIONS 
EXCEPT 
FOR 
EMERGENCIES” on the cover. 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
38 
 
After disease progression, use of corticosteroids and other medications are not 
dictated by the protocol.  
 
 
14 
Risk and Toxicity Assessment 
If the Principal Investigator determines that an adverse event is sufficiently 
severe to remove the patient from the study, an assessment should be performed 
and the patient should be given appropriate treatment under medical 
supervision. 
 
Patients will be subject to the risks associated with high-dose chemotherapy and 
autologous stem cell transplant.  Because autologous stem cell transplantation is 
considered a standard treatment option in multiple myeloma, this section will 
focus on the risks associated with the infusion of activated MILs, as well as 
unforeseen adverse events that could result from the interaction of activated 
MILs with the stem cell transplantation therapy.  
14.1 
Risks of Venous Access  
Complications associated with venous access for leukapheresis of stem cells and 
the infusion of stem cells and activated MILs include discomfort, bruising, 
and/or bleeding at the catheter insertion site, thrombosis of the accessed vessel, 
and infection. Complications from placement of a central line required for the 
high-dose chemotherapy and stem cell transplantation procedure and activated 
MILs include increased risk and severity of the above complications, as well as 
pneumothorax. 
14.2 
Risks of MILs Bone Marrow Collection 
The patients will undergo a standard bedside bone marrow aspiration with 
sedation and sterile technique to collect the marrow infiltrating lymphocytes for 
subsequent T cell activation. There is a potential risk of excessive pain that will 
be handled with analgesia as needed.  There is also the possibility of local 
and/or systemic infection.    
14.3 
Potential Microbial Contamination of the Activated MILs 
There is the potential that microorganisms inadvertently introduced during cell 
collection and processing could cause an infection in the patient following 
infusion of the activated MILs.  All precautions to maintain sterility will be 
taken.  Cultures will be obtained prior to and after completing the T cell 
activation process.  
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
39 
 
The activated MILs product will not be released until 14-day sterility testing is 
complete.  In the event that there is a sterility failure during final product release 
testing, the activated MILs will not be administered.   
14.4 
Potential Toxicity of Storage Solutions 
14.4.1 Dimethyl Sulfoxide (DMSO)  
aMILs products and the Myeloma Vaccine are cryopreserved in a solution 
containing approximately 5% DMSO.  DMSO is a cryoprotectant used to store 
a variety of infused cell products including hematopoietic stem cells and bone 
marrow.  After cells are thawed, DMSO contained in the freezing solution is 
infused along with the cells.  Toxicity associated with DMSO infusion is 
usually limited to minor changes in heart rate, blood pressure, fever, chills, 
sweats, nausea, vomiting and headaches.  These are usually self-limited, lasting 
no more than a few hours.  The most notable effect is a garlicky odor due to 
exhalation of a DMSO metabolite that may last up to one day.  More severe 
adverse effects include severe acute allergic reactions including anaphylactoid 
reactions, pulmonary embolism, respiratory failure, renal failure, cardiac 
arrhythmias, seizures, and death.[57, 58] These side effects are rarely observed 
and occur primarily in patients receiving much higher volumes of cryopreserved 
cell products than being infused in the present study.  Nevertheless, patients will 
be carefully monitored for any adverse effects as outlined in Section 0, Adverse 
Events.  Any adverse events should be documented and recorded on the 
appropriate Case Report Form.  
14.4.2   Human Serum Albumin (HSA) 
aMILs products and the Myeloma Vaccine are formulated in a cryoprotectant 
containing 2% human serum albumin.  Albumin solutions are FDA approved 
for volume resuscitation (treatment of hypovolemia, shock, and burns).  
Although hypersensitivity reactions are possible, reactions are not anticipated to 
the small amount of albumin administered.  
 
14.4.3   6% Hetastarch 
 
aMILs are formulated in a cryoprotectant containing 6% Hetastarch in 0.9% 
sodium chloride.  Toxicities associated with the intravenous administration of 
6% Hetastarch include circulatory overload , abnormalities of coagulation 
(prolonged prothrombin, partial thromboplastin and clotting times), and 
hypersensitivity reactions characterized by wheezing, urticaria, hypotension, 
and anaphylactic/anaphylactoid reactions.   
 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
40 
 
14.4.4  Pentaspan® 
 
The cell lines used to prepare the Myeloma Vaccine are formulated in a 
cryoprotectant containing Pentaspan® (10% pentastarch in 0.9% sodium 
chloride).  Toxicities associated with the intravenous administration of 
Pentaspan® include circulatory overload, abnormalities of coagulation 
(prolonged prothrombin, partial thromboplastin and clotting times), and 
hypersensitivity reactions characterized by wheezing, urticaria, hypotension, 
and anaphylactic/anaphylactoid reactions.  Additionally, patients who are 
allergic to corn can also develop hypersensitivity to Pentaspan®.    
 
14.5 
Potential Adverse Effects Associated with the Allogeneic Myeloma Vaccine 
 
The vaccine will consist of two allogeneic myeloma cell lines, H929 and U266 
admixed with a GM-CSF - secreting bystander cell line, K562/GM-CSF.  GM-
CSF secreting vaccines have been administered to over 300 patients and 
toxicities have been limited to localized vaccine site reactions involving 
erythema and tenderness.  Rarely, systemic reactions have been reported such as 
low-grade fevers and Grover’s syndrome (personal communication, Dr. 
Elizabeth Jaffee).   
 
These two allogeneic cell lines have never been used in a vaccine formulation 
prior to this study.  However, considering the vast experience at Johns Hopkins 
alone with other allogeneic vaccine formulations: breast, prostate and pancreas 
cancer, and the absence of any rate limiting toxicities, we cannot anticipate any 
possible toxicities from this vaccine formulation. 
 
14.6 
Potential Adverse Effects Associated with Activated MILs   
 
Previous clinical trials using CD3xCD28 bead-activated and expanded 
peripheral blood T cells were described above. Eighty-nine HIV patients have 
been treated, with the most common toxicities being fevers, chills, fatigue, rash, 
sinusitis, asthenia, headaches, and nausea. [59-62]Grade 3 and 4 toxicities were 
predominantly associated with IL-2, which was infused in some of the patients 
in these trials.  T cells activated with CD3xCD28 beads have also been 
administered to patients undergoing autologous stem cell transplantation for 
relapsed or refractory Non-Hodgkin’s lymphoma.[51] T cells were collected 
prior to high-dose chemotherapy. Fourteen days following peripheral blood 
stem cell infusion, activated and expanded T cells were administered.  Three 
patients were treated at a median cell dose of 0.4 x 109, twelve patients were 
treated at a median cell dose of 1.6 x 109, and two patients were treated with a 
median cell dose of 9.8 x 109.  The latter two patients experienced infusion 
related toxicities including transient fever, dyspnea, rigors and pulmonary 
edema (one patient).    
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
41 
 
It is possible that activated T cells could target the patient’s own tissues, 
resulting in autoimmune disorders.  However, this was thought to be due to 
high-dose IL-2, a known cause of this disorder, which was given with the cell 
therapy.  Autoimmune disease has not been observed in previous studies using 
CD3xCD28 bead-activated T cells. Considering that in this case, the 
lymphocytes are obtained from the marrow microenvironment, it is possible that 
an immune response could develop towards normal hematopoietic elements.  
For this reason, hematologic engraftment will be closely monitored throughout 
the trial.  
 
14.7 
Potential Adverse Events Associated with Autologous Stem Cell Transplant 
 
Patients will be at risk of all of the complications associated with the autologous 
stem cell transplant procedure. The patients will be signing a separate consent 
form for the autologous stem cell transplant utilizing the standard of care 
protocol currently in use in our institution.  This will be done per institutional 
practice by the attending physician on the Bone Marrow Transplant Service. 
The Investigator is responsible for discussing these risks in detail with the 
patient. The following is a brief overview of the major risks associated with 
autologous stem cell transplantation. 
 
14.7.1 Peripheral Blood Stem Cell Mobilization Chemotherapy Regimen 
 
Patients receive cyclophosphamide chemotherapy and filgrastim to mobilize 
peripheral blood stem cells for collection.  Cyclophosphamide can cause alopecia, 
nausea, vomiting, bladder irritation and hemorrhage, and myelosuppression 
including anemia, leukopenia, and thrombocytopenia.  Patients are at risk of 
bleeding and infection.   
 
14.7.2 Administration of Filgrastim (G-CSF) 
   
Filgrastim is given in the peripheral blood stem cell mobilization regimen and can 
cause significant bone pain in many patients. 
 
 
14.7.3 Melphalan 
 
High doses of melphalan can cause nausea, vomiting, fatigue, anemia, and 
alopecia. Patients are at significant risk of serious and life-threatening infections 
during the neutropenic period, which usually lasts for approximately two weeks.  
Patients will likely require red blood cell and platelet transfusions, both of which 
are associated with infectious risks such as hepatitis and HIV. 
 
14.7.4 Leukapheresis 
 
Patients will require leukapheresis to collect the peripheral blood stem cells 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
42 
 
 
14.7.5 Hematopoietic Stem Cell Infusion 
 
The hematopoietic stem cells may become contaminated with microbes during 
their collection or processing.  Infusion of hematopoietic stem cells contaminated 
with microbes such as bacteria could cause an infection.  The hematopoietic stem 
cells are stored in DMSO.  Similar to the administration of activated MILs, the 
DMSO is infused along with the cells.   
 
 
14.8 
Risks of Interactions Between Activated MILs, Tumor Vaccination & 
Autologous Stem Cell Transplantation 
 
It is possible there may be unforeseen interactions between the activated MILs 
and high-dose chemotherapy followed by autologous stem cell transplantation. 
Such interactions could cause, for example, delays in neutrophil or platelet 
recovery, the development of autoimmune disease, or exacerbations of known 
transplant toxicities, such as mucositis.  Patients will be monitored closely for 
hematopoietic engraftment, and adverse events.  
 
 
15 
Adverse Events 
 
15.1 
Case Report Form Reporting 
Only the adverse events (AEs) occurring during the study from the time of bone 
marrow collections of MILs and myeloma vaccine through Day 360 that are felt 
to be related to the experimental products (collection or infusion of aMILs as 
well as vaccine) and not a result of the standard and accepted toxicity associated 
with autologous stem cell transplantation will be documented and recorded on 
the AE Case Report Form. AEs will not be collected past Day 360 or after the 
patient progresses.  AEs that occur after Day 360 or disease progression/relapse 
would likely reflect the disease process, and not the activated MILs or 
vaccination. 
 
Clinically significant laboratory results considered to be possibly related to 
aMILs and not related to autologous stem cell transplant will also be recorded 
as AEs.  For purposes of this study, clinical significance is defined as any 
laboratory result that is outside the range of normal, as defined by each 
laboratory’s recorded reference range and is deemed clinically significant by the 
Principal Investigator. 
 
Adverse events and con-meds will be assessed on an on-going basis during 
patient treatment. Serious adverse events and unexpected adverse events will be 
reported per section 15.4. All adverse events documentation related to the 
infusion of aMILs will continue until resolution of those events.  


J0997/ NA_00029491                                                            
Johns Hopkins University 
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Reporting will include a description, onset and resolution date, severity, and 
assessment of relatedness to activated MILs, vaccination or other suspect cause. 
 
15.2 
Grading of Adverse Events and Toxicities 
AEs and toxicities will be graded using the NCI Common Terminology Criteria 
for Adverse Events v3.0 (CTCAE) (http://ctep.cancer.gov/reporting/ctc.html).  
AEs that are not included in the NCI Common Toxicity Criteria will be graded 
as follows: 
 
 
 Mild (Grade 1) – near the lowest intensity (or within the lower one-
third) typically seen with the observed sign or symptom 
 Moderate (Grade 2) – average intensity typically seen with the 
observed sign or symptom 
 Severe (Grade 3) – near the highest intensity (or within the top third) 
typically seen with the observed sign or symptom 
 Life-threatening or disabling (Grade 4) – any AE that places the 
patient, in the view of the investigator, at immediate risk of death 
from the reaction as it occurred.  It does not refer to an event which 
hypothetically might have caused death if it were more severe. 
 Fatal (Grade 5)  
Note:  These terms are used to describe intensity of a specific event; the 
event itself may be of relatively minor medical significance (such as severe 
headache). 
 
15.3 
Attribution of Causality  
The association or relationship of the AEs to activated MILs or myeloma 
vaccine will be defined according to the NCI Common Toxicity Criteria 
guidelines as follows: 
 
 Definite – The AE is clearly related to the study product, e.g., an 
event that follows a reasonable temporal sequence from 
administration of the drug or in which the drug level has been 
established in body fluids or tissues, that follows a known or 
expected response pattern to the suspected drug, and that is 
confirmed by improvement on stopping or reducing the dosage of 
drug 
 Probable – The AE is likely related to the study product, e.g., an 
event that follows a reasonable temporal sequence from 
administration of the drug, that follows a known or expected 
response pattern to the suspected drug, that is confirmed by stopping 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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or reducing the dosage of the drug, and that could be reasonably 
explained by the known characteristics of the subject’s clinical state 
 Possible – The AE may be related to the study product, e.g., an event 
that follows a reasonable temporal sequence from administration of 
the drug, that follows a known or expected response pattern to the 
suspected drug, but that could readily have been produced by a 
number of other factors 
 Unlikely – The AE is doubtfully related to the study product  
 Unrelated – The AE is clearly not related to the study product  
 Not applicable – The AE occurred prior to administration of the 
study product 
 Unknown – No evaluation for causality can be made 
 
15.4 
Adverse Events Requiring Immediate Notification of IRB, IBC, FDA and 
NIH RAC 
The Principal Investigator is obligated to report certain AEs to the FDA (see 21 
CRF, part 312.32)   associated with the activated MILs or vaccination that are 
“serious” or “unexpected” (see definitions below):   
 Serious Adverse Event - any adverse experience which is fatal or life-
threatening, permanently disabling, requires inpatient hospitalization, or is a 
congenital anomaly, cancer, or overdose 
 Unexpected Adverse Event - any adverse experience that is not identified in 
nature, severity or frequency in the current Investigator Brochure 
The Principal Investigator will notify the Institutional Review Board (IRB) and 
Institutional Biosafety Committee (IBC) as per institutional guidelines.    
 
15.4.1 Adverse Drug Reaction Reporting 
 
TOXICITY WILL BE SCORED USING CTCAE VERSION 3.0 FOR 
TOXICITY AND ADVERSE EVENT REPORTING.  A COPY OF 
THE CTC VERSION 3.0 CAN BE DOWNLOADED FROM THE 
CTEP HOMEPAGE (http://ctep.info.nih.gov).  ALL APPROPRIATE 
TREATMENT AREAS HAVE ACCESS TO A COPY OF THE 
CTCAE VERSION 3.0.ALL ADVERSE CLINICAL 
EXPERIENCES, WHETHER OBSERVED BY THE STUDY STAFF 
OR REPORTED BY THE PATIENT, MUST BE RECORDED, 
WITH DETAILS ABOUT THE DURATION AND INTENSITY OF 
EACH EPISODE, THE ACTION TAKEN WITH RESPECT TO 
THE TEST DRUG, AND THE PATIENT’S OUTCOME.  THE 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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INVESTIGATOR MUST EVALUATE EACH ADVERSE 
EXPERIENCE FOR ITS RELATIONSHIP TO THE TEST DRUG 
AND FOR ITS SERIOUSNESS. 
 
 
 
 
 15.4.2 Investigator Reporting Responsibilities 
The conduct of the study will comply with all FDA safety reporting 
requirements.   
All adverse experience reports must include the patient number, age, 
sex, weight, severity of reaction (mild, moderate, severe), relationship 
to study drug (probably related, unknown relationship, definitely not 
related), date and time of administration of test medications and all 
concomitant medications, and medical treatment provided.   The 
investigator is responsible for evaluating all adverse events to 
determine whether criteria for “serious” and “unexpected” as defined 
above are present. 
15.4.3 Report of Adverse Events to the Institutional Review Board and 
Institutional Biosafety Committee 
The Principal Investigator is required to notify his/her Institutional Review 
Board (IRB) of events that meet the definition of “unanticipated involving 
the risks to participants and others” within 10 working days of observing 
or learning of the serious and unexpected adverse event. The report should 
identify all safety reports previously filed with the IND concerning a 
similar adverse reaction, and should analyze the significance of the 
adverse reaction in light of the previous, similar reports.   
 
15.4.4 Investigator Reporting to the FDA and RAC 
Adverse drug reactions that are serious, unlisted/unexpected, and at 
least possibly associated to the drug, and that have not previously been 
reported should be reported promptly to the Food and Drug 
Administration (FDA) in writing on a Med Watch (FDA Form #3500A) 
by each investigator/physician engaged in clinical research.  A clear 
description of the suspected reaction should be provided along with an 
assessment as to whether the event is drug or disease related.   
 
7 Calendar-Day Telephone or Fax Report   
The Sponsor-Investigator is required to notify the FDA of any fatal or life-
threatening adverse event that is unexpected and assessed by the 
investigator to be possibly related to the use of the investigational 
product(s): activated myeloma infiltrating lymphocytes and/or myeloma 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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vaccine.  An unexpected adverse event is one that is not already described 
in the Investigator Brochure or the approved product insert.  Such reports 
are to be telephoned or faxed to the FDA and   within 7 calendar days of 
first learning of the event.  Each telephone call or fax transmission should 
be directed to the FDA   product review division for the Center for 
Biologics Evaluation and Research (This will be identified on the FDA 
letter assigning the IND # 14117), which is responsible for the review of 
the IND. 
 
15 Calendar-Day Written Report   
The Sponsor-Investigator is also required to notify the FDA and all 
participating investigators, in a written IND Safety Report, of any serious, 
unexpected AE that is considered reasonably or possibly related to the use 
of activated marrow infiltrating lymphocytes and myeloma cellular 
vaccine.  An unexpected adverse event is one that is not already described 
in the reference safety information. 
 
Written IND Safety Reports should include an Analysis of Similar Events 
in accordance with regulation 21 CFR § 312.32.  All safety reports 
previously filed with the IND concerning similar events should be 
analyzed.  The new report should contain comments on the significance of 
the new event in light of the previous, similar reports.   
 
Written IND safety reports with Analysis of Similar Events are to be 
submitted to the FDA and all participating investigators within 15 calendar 
days of first learning of the event.  The FDA prefers these reports on a 
MedWatch 3500A Form but alternative formats are acceptable (e.g. 
summary letter). 
  
The address of the Food and Drug Administration is: 
 
Center for Biologics Evaluation and Research 
 
Food and Drug Administration  
 
HFM-99, Room 200 
 
1401 Rockville Pike 
 
Rockville, MD  20852-1448 
 
 
The address of the NIH RAC is:  
Recombinant DNA Advisory Committee 
NIH Office of Biotechnology Activities 
National Institutes of Health 
6705 Rockledge Drive, Suite 750 
Bethesda, MD  20892-7985  
 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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15.5 
Definitions of Response (Blade’ Criteria) 
 
Definitions of response are adapted from the combined criteria developed for 
patients with multiple myeloma treated by high-dose chemotherapy and 
hematopoietic stem cell transplantation.[63] If confirmation of a measurement is 
required, the date of response/progression/relapse will be the date at which the M-
protein measurement first meets the required criterion. M-protein measurements 
by serum protein electrophoresis (SPEP) will take precedence over quantitative 
immunoglobulins 
for 
determining 
response. 
The 
date 
of 
response/progression/relapse will be determined by the date when the relevant 
tests were obtained, and not by the date when the investigator reviews the test 
results. 
 
Criteria for Evaluation and Endpoint Definitions  
The serum and urine M-protein levels obtained at baseline will be used as the 
reference baseline for calculation of increases or decreases in the M-protein level. 
The M-protein levels from the time of diagnosis will also be obtained, and 
response calculations using that value as baseline will also be performed in 
secondary analyses. 
 
15.5.1 Complete Response (CR) 
 
Requires all of the following: 
 Absence of the original M-protein in serum by immunofixation, maintained for a 
minimum of 6 weeks.  Presence of M-protein by Freelite assay does not exclude 
CR. The presence of oligoclonal bands consistent with oligoclonal immune 
reconstitution does not exclude CR. Normalization of serum concentrations of 
normal immunoglobulins is not required 
 If a urine M-protein is present at baseline or other prior evaluation, absence of this 
M-protein in urine by immunofixation, maintained for a minimum of 6 weeks 
 < 5% plasma cells in a bone marrow aspirate and on bone biopsy, if biopsy 
performed  
  No increase in the size or number of lytic bone lesions on skeletal x-rays, if 
performed. Development of a compression fracture does not exclude response 
 Disappearance of any soft tissue plasmacytomas 
 
15.5.2 Near Complete Response (nCR) 
 
Requires all of the following: 
 Absence of the original M-protein in serum but still identifiable by 
immunofixation, maintained for a minimum of 6 weeks.  Presence of M-protein 
by Freelite assay does not exclude CR. The presence of oligoclonal bands 
consistent with oligoclonal immune reconstitution does not exclude CR. 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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Normalization of serum concentrations of normal immunoglobulins is not 
required 
 If a urine M-protein is present at baseline or other prior evaluation, absence of this 
M-protein in urine but still present by immunofixation, maintained for a minimum 
of 6 weeks 
 < 5% plasma cells in a bone marrow aspirate and on bone biopsy, if biopsy 
performed  
  No increase in the size or number of lytic bone lesions on skeletal x-rays, if 
performed. Development of a compression fracture does not exclude response 
 Disappearance of any soft tissue plasmacytomas 
 
15.5.3     Very Good Partial Response (VGPR) 
 
Requires all of the following: 
 A decrease in the serum M-protein by  90%, maintained for a minimum of 6 
weeks 
 If a urine M-protein is present at baseline or other prior evaluation, absence of 
this protein by immunofixation, maintained for a minimum of 6 weeks 
 < 5% plasma cells in a bone marrow aspirate and on bone biopsy, if biopsy 
performed 
  No increase in the size or number of lytic bone lesions on skeletal x-rays, if 
performed. Development of a compression fracture does not exclude response 
 Disappearance of any soft tissue plasmacytomas 
 
15.5.4    Partial Response (PR) 
 
Requires all of the following: 
 A decrease in the serum M-protein by  50%, maintained for a minimum of 6 
weeks 
 If a urine M-protein is present at baseline or other prior evaluation, a  > 90% 
reduction in this protein, or a decrease to less than 200 mg in a 24 hour urine 
collection. This response must be maintained for a minimum of 6 weeks 
  No increase in the size or number of lytic bone lesions on skeletal x-rays, if 
performed. Development of a compression fracture does not exclude response 
 Decrease of  50% in the size of any soft tissue plasmacytomas (by 
radiography or clinical examination) 
 
15.5.5    Minimal Response (MR) 
 
Requires all of the following: 
 A decrease in the serum M-protein by  25%, maintained for a minimum of 6 
weeks 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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 If a urine M-protein is present at baseline or other prior evaluation, a  > 50% 
reduction in this protein in a 24 hour urine collection. This response must be 
maintained for a minimum of 6 weeks 
  No increase in the size or number of lytic bone lesions on skeletal x-rays, if 
performed. Development of a compression fracture does not exclude response 
 Decrease of  25% in the size of any soft tissue plasmacytomas (by 
radiography or clinical examination) 
 
15.5.6    Stable Disease 
 Not meeting the criteria of CR, VGPR, PR, MR, progression, or relapse from 
CR 
 
15.5.7    Relapse from CR 
 
Requires one or more of the following:  
 Reappearance of serum or urinary M-protein on immunofixation or routine 
electrophoresis, confirmed by at least one further investigation, and excluding 
oligoclonal immune reconstitution 
 > 5% plasma cells in a bone marrow aspirate or on bone biopsy 
 Development of new lytic bone lesions or soft tissue plasmacytomas or 
definite increase in the size of residual bone lesions. Development of a 
compression fracture does not exclude continued response and may not 
indicate progression 
 Development of hypercalcemia (corrected serum calcium > 11.5 mg/dL or 2.8 
mmol/L) not attributable to any other cause 
 
15.5.8    Progression 
 
For patients not in CR. Requires one or more of the following: 
 A > 25% increase in the level of serum M-protein, which must also be an 
absolute increase of at least 5 g/L and confirmed by at least one repeat 
investigation.  
 A > 25% increase in the 24 hour urinary light chain excretion, which must 
also be an absolute increase of at least 200 mg/24 hours and confirmed by at 
least one repeated investigation 
 A > 25% increase in plasma cells in a bone marrow aspirate or on biopsy, 
which must also be an absolute increase of at least 10% 
 Definite increase in the size of existing bone lesions or soft tissue 
plasmacytomas 
 Development of new bone lesions or soft tissue plasmacytomas at any time 
since baseline. Development of a compression fracture does not exclude 
continued response and may not indicate progression 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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 Development of hypercalcemia (corrected serum calcium > 11.5 mg/dL or 2.8 
mmol/L) at any time since baseline, which is not attributable to any other 
cause. (Corrected serum calcium is calculated by adding 0.8 mg/dL to the 
measured serum calcium for every 1 g/dL that the serum albumin falls below 
4.0 g/dL.) 
 
Reference level for increase in M-protein, plasma cells, or lesion sizes is the 
lowest level documented since (and including) baseline evaluation. 
 
15.6 
Survival Endpoints 
 
15.6.1    Overall Survival 
Measured as the time from Day 0 (day of stem cell infusion) to death from any 
cause.  This will be monitored annually for up to 4 years. 
During this annual visit, the following will occur for the patients that opt to 
come here for their follow-up.  Alternatively, survival will be determined by the 
medical records obtained from the locally treating physician: 
 Disease response/progression/relapse assessment (the clinical 
assessment can be performed up to 10 days after the laboratory 
evaluation) 
 100ml heparinized syringes and 20 mls total in green top tubes of 
blood will be collected from the patient for immune monitoring 
assays. 
 Bone marrow biopsy and aspirate will be performed.  20ml of 
marrow will be collected for immune monitoring. 
 
15.6.2    Progression-Free Survival 
Measured as the time from initial enrollment to progression/relapse of disease 
or death from any cause, whichever occurs first. 
 
15.7 
Definitions of Engraftment and Graft Failure 
 
15.7.1    Engraftment 
Neutrophil engraftment is defined as the day post-transplant when the absolute 
neutrophil count is > 500/mm3 for two consecutive days post-transplant (note: 
bands as well as neutrophils are included in calculations).  The first day on 
which this criterion is met is considered the day of neutrophil engraftment. 
 
Platelet engraftment is defined as the first of three consecutive counts post-
transplant when the platelet count is > 20,000/mm3 without platelet transfusion 
support.  The first day on which this criterion is met is considered the day of 
platelet engraftment. 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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15.7.2    Failure of Engraftment 
Primary graft failure is defined as failure to recover an absolute neutrophil count 
> 500/mm3 or platelet count > 20,000/mm3 by Day 28 post-transplant.  
Secondary graft failure is defined as a fall to less than these levels for three or 
more consecutive days after Day 28 without another cause.  
 
 
 
16 
Statistical Considerations 
 
Study Design  
 
This is a continuation study stemming from two previous early stage clinical 
evaluations: J0115 – autologous myeloma vaccine;  and J0770 – activated MILs 
with regard to treatment effect, and tolerability of allogeneic myeloma cell 
vaccine and aMILs in this patient population . In the aMILs trial (J0770) we have 
observed the early results shown 5 out of 19 (26%) patients had complete 
response (CR) based on Blade’ Criteria. The current trial intends to build a 
treatment regime using a modified higher dose of aMIL, adoptive 
immunotherapy, and concurrent with vaccination which had been tested in J0115 
trial to enhance overall immune anti-tumor responses. Due to alterations in ex-
vivo MILs expansion and the possibility that tumor specific vaccination could 
further augment anti-tumor immune responses, the study is designed to 
randomize patient onto either a high dose aMILs alone or a high dose aMILs + 
vaccine with 1:1 randomization ratio. The purpose of the randomization is not 
for hypothesis testing, but to have initial estimation of the CR rate for the two 
treatment regimens in a parallel fashion, second to the primary objective of an 
overall response assessment. 
 
16.1 
   Sample Size 
A total 32 evaluable patients will be needed in this clinical trial. If patients are 
registered but do not receive the activated T cells for any reason, then additional 
patients will be registered to ensure 32 patients are treated.  
 
The primary clinical activity is defined as completed response based on Blade’ 
Criteria. The study is designed to have initial estimation of the CR rate among 
those treated either with aMILs alone or aMILs+ the vaccine. Our early clinical 
experience using aMILs alone shows the CR rate of 26% compared to an 
estimated 20% CR from standard transplant. The total of 32 patients will yield a 
minimum of 40% CR rate with a lower bound of 95% confidence interval 
excess 20%. The 32 patients will be randomly assigned to either aMILs alone or 
aMILs +vaccine group with 1:1 randomization ratio stratified by the clinical 
status (newly diagnosed vs relapsed, ISS staging 1-2 vs. 3). The sample size 
justification based on precision of estimation is tabulated below: 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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No. of Patients 
No. of patients 
with CR 
% CR rate 
95% Confidence 
Intervals 
32 
13 
40% 
24%  -- 59% 
32 
16 
50% 
32%  -- 68% 
16 
7 
44% 
20%  -- 70% 
16 
8 
50% 
25%  -- 75% 
 
The clinical efficacy is defined as complete response based on Blade’ Criteria.  
Overall complete response rate and individual treatment response rate will be 
estimated using the binomial distribution (exact method) with 95% confidence 
intervals. However, other response rates such as nCR , VGPR, PR, MR,  and a 
overall response rate (CR, VGPR, PR, MR definition see section 13.5) at 360 
days post transplant will be calculated as proportions and reported as a 
combined response for the 32 patients and separately by group as well.    
 
16.2  Statistical Analyses/ Secondary objectives 
 
Progression-free and overall survival:  
The safety endpoints will be treatment-related mortality and the incidence of 
Grade 3 hematologic toxicity. Toxicity will be graded using the NCI Common 
Toxicity Criteria (Version 3.0; http://ctep.cancer.gov/reporting/ctc.html; see 
Appendix G). Treatment related mortality is defined as death not attributable to 
disease progression/relapse (and excluding the ‘unrelated’ and ‘not applicable’ 
categories in attribution of causality per Section 15.3) that occurs within the 
first 100 days following transplantation.  Previous randomized studies involving 
high-dose chemotherapy and hematopoietic stem cell transplantation in patients 
with multiple myeloma have generally reported treatment related mortality in 
the 4-8% range, [64-66]although rates as high as 14-20% have been reported. 
[67, 68]Treatment related mortality in the current era is likely lower than these 
reported rates. 
 
The toxicity will be monitored closely.  If any patient develops Grade 4 
hematologic toxicity of 3 weeks or longer that represents greater than a 50% 
decrease from their post-transplant screen, the trial will be suspended until 
appropriate analysis and discussion with the FDA and IRB has been conducted. 
The study will be terminated if there are one or more deaths, which are 
definitely or probably attributable specifically to the activated T cell therapy, as 
defined in Section 13.3, Attribution of Causality. However, given the toxicities 
associated with high-dose therapy and hematopoietic stem cell transplantation, 
attribution of death specifically to the experimental therapies may be difficult. 
Therefore the study will also be terminated and the treatment approach deemed 
not worthy of further investigation in this setting if at any time 4 patients die of 
treatment related causes or experience Grade 3 hematologic toxicity.   
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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Disease progression is defined in the protocol section 15.5 Progression-free 
survival is defined as time from date of randomization to the date of first 
observation of disease progression or death due to disease progression. Overall 
survival is defined as the time from the date of randomization to date of death 
due to all causes.  The Probability of progression-free survival (PFS) and overall 
survival (OS) will be estimated using the Kaplan-Meier method and displayed 
graphically. Median PFS and OS will be reported along with 95% confidence 
intervals. Proportion of patients who were PFS and OS at specific time of 
follow-up will be reported as well.  
 
Statistical analyses of other secondary objectives such as T cell reconstitution, 
anti-tumor immune response, tumor-specific vaccine response, effect of aMIL 
on osteclastogenesis, and Myeloma clonogenic precursors will be considered 
exploratory. Descriptive statistics will be provided. All data will be summarized 
based on nature distribution of data and presented separately by treatment group 
at clinical defined time points, or by primary clinical response. 
The association between clinical responses and in vitro measurements of T cell 
laboratory correlates, including T cell proliferation to tumor and ELISPOT 
analysis of both peripheral as well as marrow infiltrating lymphocytes will be 
explored by using multivariate regression models as appropriate.   
 
Evaluation of hematopoietic engraftment: 
The mean and median time to neutrophil and platelet engraftment (absolute 
neutrophil count >500/mm3 and platelet >20,000/mm3) will be summarized 
among those with successful engraftment, along with the rate of primary graft 
failure.   It should be remember that in the absence of post-transplant G-CSF, 
we would expect neutrophil engraftment to be slightly delayed.  However, 
should red cell, platelet and neutrophil engraftment all show evidence of 
significant delay in more than 2 patients, the study will be placed on hold until a 
more formal evaluation can be performed.  
 
All patients demographic information, disease, and baseline characteristics will 
be summarized for each of the study groups independently.   
 
 
16.4.1 Subject Accountability 
The following data will be summarized: 
 Number (%) of subjects registered 
 Number (%) of subjects undergoing autologous stem cell transplant 
 Number (%) of subjects who receive activated MILs 
16.4.3 Protocol Compliance 
The registered study subject population will be described in reference to the 
study criteria.  At a minimum, the following tabulations will be provided: 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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 Number of subjects with violations of inclusion/exclusion criteria 
 Number of subjects with major on-study deviations or violations  
 
 
17 
Ethical, Regulatory, and Administrative Considerations 
 
17.1 
   Informed Consent 
The principles of informed consent are described in the Code of Federal 
Regulations 21 CFR, part 50. 
 
Patients or their legal guardians must be made aware of the investigational 
nature of this treatment protocol, and have the possible risks, hazards and 
benefits of the protocol explained to them.  The information that is given shall 
be in a language understandable to the patient.  No informed consent, whether 
oral or written, may include any exculpatory language through which the patient 
is made to waive or appear to waive any of their legal rights, or releases or 
appears to release the Principal Investigator, the institution, or its agents from 
liability for negligence. 
 
The patient, or legal guardian, must be able to comprehend the informed 
consent and must sign the document prior to consent signing on study.  The 
patient will receive a copy of the respective signed consent form. 
 
17.2 
   Institutional Review 
The principles of Institutional Review Board (IRB) are described in the Code of 
Federal Regulations 21 CFR, part 56. 
 
The Principal Investigator will obtain approval for the study from the IRB. The 
Principal Investigator must notify the IRB within 5 days of protocol deviations 
in emergency situations regarding patient safety. The Principal Investigator will 
be responsible for obtaining annual IRB renewal through the duration of the 
study, or more frequently if required by the IRB.  Copies of the Principal 
Investigator's report and copies of the IRB’s continuance of approval must be 
maintained in the regulatory binder located at the clinical site.   
 
17.3 
   Tissue Use for Research Purposes 
A portion of the MILs bone marrow product and/or final activated MILs 
product may remain unused at the conclusion of the study. Patients will have the 
option on the study consent form to allow this tissue to be used for further 
research purposes by the Principal Investigator, or to be destroyed one year after 
collection. 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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55 
 
In addition, various patient tissues, including peripheral blood and bone marrow 
will be collected from patients during the course of the study expressly for 
research assays. Baseline bone marrow specimens with adequate tumor 
involvement will be used to develop primers required for a PCR based assay for 
minimal residual tumor in the final activated MILs product. Quantification of 
minimal residual disease will also be performed in bone marrow from patients 
in complete remission for whom primers can be developed. Peripheral blood 
and tumor containing marrow specimens may also be used in immune assays.  
Results from assays performed for research purposes will not be provided to 
patients. At the conclusion of the study and follow-up period (five years from 
Day 0), some of this tissue may remain unused. Patients will have the option in 
the study consent form to allow this tissue to be used for further research 
purposes or to be destroyed. 
 
18 
Study Monitoring and Data Collection 
 
18.1 
   Study Monitoring 
This is a Level III study under the SKCCC CRO Data and Safety Monitoring 
Program.  Data Monitoring of this protocol will occur on a regular basis with 
the frequency dependent on the rate of subject accrual and the progress of the 
study.  The protocol will be monitored internally by the principal investigator. 
 
External monitoring will be performed by the SKCCC CRO in accordance with 
SKCCC and federal guidelines.  Trial monitoring and reporting will be done 
through the Medical Expert Committee when approximately half the patients 
have been enrolled and upon completion of enrollment in the study.   
 
18.1.1    Completion of Case Report Forms (CRFs) 
The Principal Investigator or his designee will be responsible for completing, in 
a timely manner, a CRF for each patient who is registered to participate in this 
study.  CRFs will be completed, as information becomes available.   
 
The Principal Investigator or a sub-investigator will sign and date the indicated 
places on the CRF.  This signature will indicate that a thorough inspection of 
the audited data therein has been made and will thereby certify the contents of 
the form. 
 
18.1.2    Cell Therapy Laboratory (CTL) Cell-Processing Facility 
The Cell Therapy Laboratory (CTL) is the processing lab for all cellular therapy 
products (bone marrow, vaccines, and adoptive T cell products) administered in 
the Cancer Center at Johns Hopkins.  It operates according to Good Tissue and 
Manufacturing Practices and is accredited by FACT. 
 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
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The results of cell processing using the activated T cell process will be recorded 
on CRFs or in the Batch Production Records at the Johns Hopkins CTL facility.  
CRFs and other study records will be reviewed in detail by the CRA, or 
designee, who will have access to all patient medical records, laboratory data, 
and other source documentation to verify the accuracy of the data recorded on 
these documents. 
 
18.2 
   Maintenance of Study Documentation 
It is the responsibility of the Principal Investigator and study staff to maintain a 
comprehensive and centralized filing system of all study-related documentation. 
This filing system must be suitable for inspection at any time by the CRO or 
Quality Assurance (QA) designee of Johns Hopkins and the FDA.  Elements 
should include: 
 
 
Subject Files - containing the completed patient CRFs, supporting source 
documentation, and a signed and dated Informed Consent 
 
Regulatory Binder - containing the protocol with all amendments, current 
Consent Form, the Investigator Brochure, clinical site logs, accountability 
records and laboratory documents (e.g., certification, norms/ranges, etc.)  
 
18.2.1    Retention of Records 
 
The FDA requires that each Principal Investigator retain records for a period of 
two (2) years from the date of FDA approval to market the product for this 
indication, or two (2) years from the date that the Sponsor withdraws the 
application for approval. 
 
18.3 
   Final Study Report 
Upon completion of the study, the Principal Investigator is required to submit a 
final study summary report to the Institution.  This report must also be 
submitted to the IRB. 
 
18.4 
   Investigational Product Labeling and Accountability 
18.4.1    Investigational Product Labeling 
 
aMILs products will be labeled at a minimum with the patient’s name and 
history number, the unique product number, and the volume within the bag.   
 
The Myeloma Vaccine will be labeled with the lot numbers of the cell lines 
used to prepare the vaccine in addition to the preparation date, product 
expiration time, and the patient’s name and history number. 
. 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
57 
 
18.4.2    Investigational Product Accountability 
The Investigator should take adequate precautions, including storage of the 
investigational products, to prevent theft or diversion of the products into 
unauthorized channels of distribution.  The processing of each product will be 
documented on a Batch Production Record.  Release test results will be 
summarized on the Product Specification Report.  


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
58 
 
19 
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Harousseau, J.L., et al., Autologous stem cell transplantation after first remission induction 
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J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
59 
 
27. 
Bensinger, W.I., D. Maloney, and R. Storb, Allogeneic hematopoietic cell transplantation for 
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30. 
Frohlich, M., Grosmaire, L., Xu, J., Rasmussen, A., Roehrs, H., Lindren, R., Ferrand, C., 
Tiberghien, P., Leis, J., and Bonyhadi, M. L. Xcellerate:  a novel autologous T cell 
immunotherapeutic approach for the treatment of B-cell chronic lymphocytic leukemia (B-CLL). 
in The IX International Workshop on CLL. 2002. 
31. 
Kato, K., et al., Gene transfer of CD40-ligand induces autologous immune recognition of chronic 
lymphocytic leukemia B cells. J Clin Invest, 1998. 101(5): p. 1133-41. 
32. 
Sotomayor, E.M., I. Borrello, and H.I. Levitsky, Tolerance and cancer: a critical issue in tumor 
immunology. Crit Rev Oncog, 1996. 7(5-6): p. 433-56. 
33. 
Gorschluter, M., et al., Current clinical and laboratory strategies to augment the efficacy of 
immunotherapy in multiple myeloma. Clin Cancer Res, 2001. 7(8): p. 2195-204. 
34. 
Liso, A., et al., Idiotype vaccination using dendritic cells after autologous peripheral blood 
progenitor cell transplantation for multiple myeloma. Biol Blood Marrow Transplant, 2000. 6(6): 
p. 621-7. 
35. 
Titzer, S., et al., Vaccination of multiple myeloma patients with idiotype-pulsed dendritic cells: 
immunological and clinical aspects. Br J Haematol, 2000. 108(4): p. 805-16. 
36. 
Borrello, I., et al., A universal granulocyte-macrophage colony-stimulating factor-producing 
bystander cell line for use in the formulation of autologous tumor cell-based vaccines. Hum Gene 
Ther, 1999. 10(12): p. 1983-91. 
37. 
Borrello, I., et al., Sustaining the graft-versus-tumor effect through posttransplant immunization 
with granulocyte-macrophage colony-stimulating factor (GM-CSF)-producing tumor vaccines. 
Blood, 2000. 95(10): p. 3011-9. 
38. 
Tricot, G., et al., Graft-versus-myeloma effect: proof of principle. Blood, 1996. 87(3): p. 1196-8. 
39. 
Verdonck, L.F., et al., Graft-versus-myeloma effect in two cases. Lancet, 1996. 347(9004): p. 800-
1. 
40. 
Alyea, E., et al., T-cell--depleted allogeneic bone marrow transplantation followed by donor 
lymphocyte infusion in patients with multiple myeloma: induction of graft-versus-myeloma effect. 
Blood, 2001. 98(4): p. 934-9. 
41. 
Cabrera, R., et al., Infusion of lymphocytes obtained from a donor immunised with the paraprotein 
idiotype as a treatment in a relapsed myeloma. Bone Marrow Transplant, 2000. 25(10): p. 1105-8. 
42. 
Salama, M., et al., Donor leukocyte infusions for multiple myeloma. Bone Marrow Transplant, 
2000. 26(11): p. 1179-84. 
43. 
Li, Y., et al., Tumor-specific recognition of human myeloma cells by idiotype-induced CD8(+) T 
cells. Blood, 2000. 96(8): p. 2828-33. 
44. 
Gahrton, G., Allogeneic bone marrow transplantation in multiple myeloma. Pathol Biol (Paris), 
1999. 47(2): p. 188-91. 
45. 
Porrata, L.F., et al., Early lymphocyte recovery predicts superior survival after autologous 
hematopoietic stem cell transplantation in multiple myeloma or non- Hodgkin lymphoma. Blood, 
2001. 98(3): p. 579-85. 
46. 
Pierelli, L., et al., The role of growth factor administration and T-cell recovery after peripheral 
blood progenitor cell transplantation in the treatment of solid tumors: results from a randomized 
comparison of G-CSF and GM-CSF. Transfusion, 2001. 41(12): p. 1577-85. 
47. 
Porrata, L.F., et al., Prolonged survival associated with early lymphocyte recovery after 
autologous hematopoietic stem cell transplantation for patients with metastatic breast cancer. Bone 
Marrow Transplant, 2001. 28(9): p. 865-71. 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
60 
 
48. 
Levine, B.L., et al., Effects of CD28 costimulation on long-term proliferation of CD4+ T cells in 
the absence of exogenous feeder cells. J Immunol, 1997. 159(12): p. 5921-30. 
49. 
Levine, B.L., et al., Large-scale production of CD4+ T cells from HIV-1-infected donors after 
CD3/CD28 costimulation. J Hematother, 1998. 7(5): p. 437-48. 
50. 
Liebowitz, D.N., K.P. Lee, and C.H. June, Costimulatory approaches to adoptive immunotherapy. 
Curr Opin Oncol, 1998. 10(6): p. 533-41. 
51. 
Laport, G.G., et al., Adoptive transfer of CD3/CD28 ex vivo costimulated T cells in patients with 
relapsed/refractory Non-Hodgkin's lymphoma (NHL) following high dose chemotherapy (HDC) 
with CD34-selected peripheral blood stem cell (PBSC) support. Blood, 2000. 96((Suppl 1)). 
52. 
Noonan, K., et al., Activated marrow-infiltrating lymphocytes effectively target plasma cells and 
their clonogenic precursors. Cancer Res, 2005. 65(5): p. 2026-34. 
53. 
Steinman, L., A brief history of T(H)17, the first major revision in the T(H)1/T(H)2 hypothesis of 
T cell-mediated tissue damage. Nat Med, 2007. 13(2): p. 139-45. 
54. 
Bettelli, E., et al., Reciprocal developmental pathways for the generation of pathogenic effector 
TH17 and regulatory T cells. Nature, 2006. 441(7090): p. 235-8. 
55. 
Dudley, M.E., et al., Cancer regression and autoimmunity in patients after clonal repopulation 
with antitumor lymphocytes. Science, 2002. 298(5594): p. 850-4. 
56. 
Boon, T. and L.J. Old, Cancer Tumor antigens. Curr Opin Immunol, 1997. 9(5): p. 681-3. 
57. 
Shpall, E.J., et al., A prospective randomized trial of buffy coat versus CD34-selected autologous 
bone marrow support in high-risk breast cancer patients receiving high-dose chemotherapy. Blood, 
1997. 90(11): p. 4313-20. 
58. 
Davis, J., S.D. Rowley, and G.W. Santos, Toxicity of autologous bone marrow graft infusion. 
Prog Clin Biol Res, 1990. 333: p. 531-40. 
59. 
Mitsuyasu, R.T., et al., Prolonged survival and tissue trafficking following adoptive transfer of 
CD4zeta gene-modified autologous CD4(+) and CD8(+) T cells in human immunodeficiency 
virus-infected subjects. Blood, 2000. 96(3): p. 785-93. 
60. 
Walker, R.E., et al., Long-term in vivo survival of receptor-modified syngeneic T cells in patients 
with human immunodeficiency virus infection. Blood, 2000. 96(2): p. 467-74. 
61. 
Levine, B.L., et al., Adoptive transfer of costimulated CD4+ T cells induces expansion of 
peripheral T cells and decreased CCR5 expression in HIV infection. Nat Med, 2002. 8(1): p. 47-
53. 
62. 
Deeks, S.G., et al., A phase II randomized study of HIV-specific T-cell gene therapy in subjects 
with undetectable plasma viremia on combination antiretroviral therapy. Mol Ther, 2002. 5(6): p. 
788-97. 
63. 
Blade, J., et al., Criteria for evaluating disease response and progression in patients with multiple 
myeloma treated by high-dose therapy and haemopoietic stem cell transplantation. Myeloma 
Subcommittee of the EBMT. European Group for Blood and Marrow Transplant. Br J Haematol, 
1998. 102(5): p. 1115-23. 
64. 
Attal, M., et al., A prospective, randomized trial of autologous bone marrow transplantation and 
chemotherapy in multiple myeloma. Intergroupe Francais du Myelome. N Engl J Med, 1996. 
335(2): p. 91-7. 
65. 
Barlogie, B., et al., Superiority of tandem autologous transplantation over standard therapy for 
previously untreated multiple myeloma. Blood, 1997. 89(3): p. 789-93. 
66. 
Lenhoff, S., et al., Impact on survival of high-dose therapy with autologous stem cell support in 
patients younger than 60 years with newly diagnosed multiple myeloma: a population-based study. 
Nordic Myeloma Study Group. Blood, 2000. 95(1): p. 7-11. 
67. 
Alexanian, R., et al., Limited value of myeloablative therapy for late multiple myeloma. Blood, 
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1995. 6(1): p. 11-8.


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
aMILs with or without an Allogeneic GM-CSF-based Myeloma Cellular Vaccine  August 06, 2010 
 
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20 
Appendices 
 
Appendix A:  Diagnostic Criteria for Multiple Myeloma 
 
o 
Major criteria: 
1. Plasmacytomas on tissue biopsy 
2. Bone marrow plasmacytosis (>30% plasma cells) 
3. Monoclonal immunoglobulin spike on serum electrophoresis IgG >3.5 g/dL or IgA >2.0 g/dL; kappa 
or lambda light chain excretion > 1 g/day on 24 hour urine protein electrophoresis 
 
o 
Minor criteria: 
a. 
Bone marrow plasmacytosis (10 to 30% plasma cells) 
b. Monoclonal immunoglobulin present but of lesser magnitude than given under major criteria 
c. 
Lytic bone lesions 
d. Normal IgM <50 mg/dL, IgA <100 mg/dL or IgG <600 mg/dL 
 
Any of the following sets of criteria will confirm the diagnosis of multiple myeloma 
o 
Any two of the major criteria 
o 
Major criterion 1 plus minor criterion b, c, or d 
o 
Major criterion 3 plus minor criterion a or c 
o 
Minor criterion a, b and c or a, b and d 
 
Reference: Durie, B. G. 1986. Staging and kinetics of multiple myeloma. Semin.Oncol. 13[3], 300-309. 


J0997/ NA_00029491                                                            
Johns Hopkins University 
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Appendix B:  Durie-Salmon Staging of Multiple Myeloma 
 
Stage I 
 
All of the following must be present 
 
 Hemoglobin > 10.5 g/dL or hematocrit > 32% 
 Serum calcium level normal 
 Low serum myeloma protein production rates as evidenced by all of the following: 
- IgG peak < 5g/dL 
- IgA peak < 3g/dL 
- Bence Jones protein < 4g/24 h 
 No bone lesions 
 
Stage II 
 
All patients who do not meet criteria for Stage I or III are considered to be Stage II. 
 
Stage III 
 
One of the following abnormalities must be present: 
 
 Hemoglobin < 8.5 g/dL, hematocrit < 25% 
 Serum calcium > 12 mg/dL 
 Very high serum or urine myeloma protein production rates as evidenced by one or more of the 
following: 
- IgG peak > 7g/dL 
- IgA peak > 5g/dL 
- Bence Jones protein > 12 g/24 h 
 > 3 lytic bone lesion on bone survey (bone scan not acceptable) 
 
Subclassification 
 
A: Serum creatinine < 2.0 mg/dL 
B: Serum creatinine > 2.0 mg/dL 
 
 
Adapted from: Durie BGM, Salmon SE. A clinical staging system for multiple myeloma. Correlation of 
measured myeloma cell mass with presenting clinical features, response to treatment and survival. Cancer 
1975;36:842-54. 


J0997/ NA_00029491                                                            
Johns Hopkins University 
BB-IND  14117 
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Appendix C:  ECOG Performance Status 
 
Grade 
ECOG 
0 
Fully active, able to carry on all pre-disease performance 
without restriction 
1 
Restricted in physically strenuous activity but ambulatory and able to 
carry out work of a light or sedentary nature, e.g., light house work, 
office work 
2 
Ambulatory and capable of all self care but unable to carry out any work 
activities. Up and about more than 50% of waking hours 
3 
Capable of only limited self care, confined to bed or chair more than 50% 
of waking hours 
4 
Completely disabled. Cannot carry on any self care. Totally confined to 
bed or chair  
5 
Dead 
 
AS PUBLISHED IN AM. J. CLIN. ONCOL. (CCT) 5:649-655, 1982 
 


J0997/NA_00029491 
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Appendix D:  New York Heart Association Classification of Patients with    
Diseases of the Heart 
 
Functional Classification 
Class l 
 
 
 
Class ll 
 
 
 
Class lll 
 
 
 
Class lV 
Patient with cardiac disease but without resulting limitations of physical activity. 
Ordinary physical activity does not cause undue fatigue, palpitation, dyspnea, or 
anginal pain. 
 
Patients with cardiac disease resulting in slight limitation of physical activity. They are 
comfortable at rest. Ordinary physical activity results in fatigue, palpitation, dyspnea, 
or anginal pain. 
 
Patients with cardiac disease resulting in marked limitation of physical activity. They 
are comfortable at rest. Less than ordinary physical activity causes fatigue, 
palpitation, dyspnea, or anginal pain. 
 
Patients with cardiac disease resulting in inability to carry on any physical activity 
without discomfort. Symptoms of cardiac insufficiency or of the anginal syndrome 
may be present even at rest. If any physical activity is undertaken, discomfort is 
increased. 
(New York Heart Association Criteria Committee: Diseases of the Heart and Blood Vessels: 
Nomenclature and Criteria for Diagnosis, 6th ed. Boston, Little, Brown, & Co. 1964) 


Appendix E: Patient Study Calendar 
 
 
 
66 
 
 
 
 
 
  
Consent Signing Eligibility 
MM Vaccination 
Marrow Collection 
Stem Cell Mobilization  
Stem  Cell Collection 
Day -2 (Melphalan) 
Day -1 (Melphalan 
Day 0 (Stem Cell Infusion) 
Therapy Week 1 
  
  
  
Follow-up 
  
  
  
Day 1 
Day 2 
Day 3  aMILs Infusion* 
Day 4 
Day 5 
Day 6a 
Day 7a 
Day 14 
Day 21 Vaccine 
Day 28 (±2 days) 
Day 
60 
(±7 
days) 
Vaccine 
Day 180 (± 30 days) 
Vaccine 
Day 300 (± 30 days) 
Vaccine12 
Day 360 (± 30 days)  
Annually  for 4 
years  
Off Study 
Informed Consent 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Inclusion/Exclusion criteria 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Clinical Assessment1 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
 
 
Bone marrow examination2 
 
 
Xa 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
 
X 
 
 
Beta2-microglobulin 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
 
 
HTLV1 and 2  
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
CBC w/diff, platelets3 
X 
X 
X 
 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
Serum Chemistries3 
X 
 
 
 
 
 
 
 
 
 
X 
 
 
 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
Serum myeloma studies4 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
 
 
M-protein, urine5 
 
X 
(X) 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
 
 
Bone Turnover Labs6 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
 
 
Myeloma Vaccine7 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
X 
X 
 
 
 
Study Blood (100ml) & Tiger top tube (10ml)8 
 
  
Xb,c 
X 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
Xc 
X 
X 
Xc 
X 
X 
 
 
Study Blood  in Green top tubes (20ml) )9 
 
Xd 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
 
 
Disease Response/Progression 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
 
Survival Status10 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
X 
X 
X 
Current Medications 
 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
 
 
X 
X 
X 
X 
 
 
 
Adverse Events11 
 
 
X 
 
 
 
 
 
 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
1 Clinical assessment: history, review of systems,, weight, vital signs, physical exam, ECOG. For day 28 and 60 this can be done up to 10 days later 
 
 
2 Collect 20ml of marrow in a heparinized 60ml syringe. ( aOn day of marrow collection, patients will have 200ml of BM collected in heparinized syringes) 
3 Chemistries: sodium, potassium, chloride, bicarbonate, BUN, creatinine, calcium, AST, ALT, alk. phosph., total protein, albumin.  CBC differential will only be performed if total WBC >100/ul.  Labs may be drawn ± 24hrs. 
 
 
4 M-protein, serum: serum protein electropheresis & immunofixation, Serum free light assay, quantitative immunoglobulins 
 
 
5 M-protein, urine: 24 hour urine collection for total protein, protein electropheresis & immunofixation (should be obtained either with pre-transplant vaccine or at time of bone marrow collection)     
6 Bone Turnover to include: Blood for C telopeptide  and bAlkaline phosphatase in yellow top tube  
7 Myeloma Vaccine consists of 3 cell lines admixed, irradiated and administered over 3 limbs.  Pre-transplant vaccine to be administered 14 days (± 2 days)  
prior  to the bone marrow collection.  
 
 
8 100ml of blood in heparinized syringes and 10 ml in tiger top tube will be collected at the time of bone marrow collection.  b  Vaccine Arm:  Blood will be collected  immediately before pre-transplant vaccination as well as at the time of the bone marrow collection.  
 c  Vaccine Arm: 10 ml tiger top tube collected  2 days later. 
9  20ml of blood in green top tubes will be obtained for evaluation of myeloma precursors. d Vaccine Arm: This blood can be collected either immediately before pre-transplant vaccine or at time of bone marrow collection)       
10  Survival and disease status will be followed for4 years from Day 0        
11  Capture adverse events are those felt to be related to the infusion of aMILs. Standard stem cell transplant toxicities will not be captured. 
12  The day 300 visit will only apply to the patients randomized to the vaccine arm. This will include a CBC with differential.                                                                                     
 
 
* For aMILs infusion, patients must have T<38.5oC and room air O2 saturation of >90%.