Immunization With 8 Peptides Mixed With CpG 7909 or Montanide ISA51 in Patients With Metastatic Cutaneous Melanoma

Study Overview
=================
Brief Summary
-----------------
The purposes of this study are to describe the immune response to individual peptides after immunization with a combination of 8 peptides and CpG 7909 or Montanide ISA51; to determine the safety of the vaccines and; to document the tumor response in patients receiving the vaccines.

Detailed Description
-----------------
Patients received six sequential immunizations with 8 peptides presented by HLA-A2 and mixed with either CpG 7909 or Montanide ISA51, at 2-week intervals. The 8 peptides were injected at 8 distinct injection sites. These peptides are the following: MAGE-1.A2, MAGE-3.A2, MAGE-4.A2, MAGE-10.A2, MAGE-C2.A2 (ALKD), and NA17.A2 (20% intradermally and 80% subcutaneously); NY-ESO-1.A2 and Tyrosinase.A2 (100% subcutaneously). 300 µg of each peptide (except MAGE-10.A2 150 µg) was mixed with 4 mg CpG 7909 (Cohort 1) or 0.5ml of Montanide ISA51 (Cohort 2). In Cohort 2, the Tyrosinase.A2 was administered without Montanide ISA51. Tumor staging was performed before inclusion and at week 13. Peripheral Blood Lymphocytes (PBL) collections were performed before starting the treatment, and at weeks 3, 7 and 13. They provided the T lymphocytes for the immunological analysis. At week 13, the PCR results of the pre-immune tumor biopsy must be available. Additional cycles of immunization, ONLY with the peptides expressed by the tumor, mixed with Montanide ISA51, will be proposed to patients without tumor progression requiring another treatment. A second cycle of 3 injections at 6-week intervals will be started at week 17, followed by a third cycle of 12 injections at 3-month intervals, starting at month 11. At any time, progression of the disease necessitating any treatment not allowed during the study will result in withdrawal. The immune response may well be a limiting factor to the therapeutic efficacy of the vaccine. If this is the case, it then becomes crucial to understand why some patients develop a cytotoxic t-lymphocyte (CTL) response against the vaccine, while the majority of them do not. One possible explanation for the low frequency of clinical responses is that each injection of a single peptide has a low probability to provide the adequate stimulus to activate very rare CTL precursors. This probability should be increased if several peptides known to be undoubtedly associated with tumor regressions were used together to immunize patients.

Official Title
-----------------
Phase I/II Study of Immunization With Multiple Peptides Mixed With the Immunological Adjuvant CpG 7909 or Montanide ISA51 in HLA-A2 Patients With Metastatic Cutaneous Melanoma

Conditions
-----------------
Malignant Melanoma

Intervention / Treatment
-----------------
* Biological: 8 HLA-A2-restricted peptides and Montanide ISA51
* Biological: 8 HLA-A2-restricted peptides and CpG 7909


Participation Criteria
=================
Eligibility Criteria
-----------------
Inclusion Criteria Histologically proven cutaneous melanoma, or clear cell sarcoma, which is considered as a subtype of melanoma. Melanoma must be at one of the following AJCC 2002 stages: Regional metastatic disease (any T; N2b, N2c or N3; M0). Distant metastatic disease (any T; any N; M1a, M1b or M1c), except brain or leptomeningeal localizations, and except elevated LDH. Patients must be HLA-A2. A pre-immune tumor biopsy must be kept frozen for post-study PCR analysis. Presence of at least one measurable or non-measurable tumor lesion. Expected survival of at least 3 months. Karnofsky performance scale ≥70 or WHO performance status of 0 or 1. Within the last 4 weeks prior to study day 1, vital laboratory parameters should be within normal range, except for the following laboratory parameters, which must be within the ranges specified: Lab Parameter Range Hemoglobin ≥ 10 g/dl or ≥ 6,25 mmol/l Granulocytes ≥ 1,500/µl Lymphocytes ≥ 700/µl Platelets ≥ 100,000/µl Serum creatinine ≤ 2.0 mg/dl or ≤ 177 μmol/l Serum bilirubin ≤ 2.0 mg/dl or ≤ 34.2 μmol/l ASAT and ALAT ≤ 2 x the normal upper limits LDH ≤ the normal upper limit. Viral tests: HIV (human immunodeficiency virus): negative antibodies. HBV (hepatitis B virus): negative antigens; antibodies may be positive. HCV (hepatitis C virus): negative antibodies. Age ≥ 18 years. Able and willing to give valid written informed consent. Exclusion Criteria Previous treatment with more than one regimen of systemic chemotherapy, combined or not with non-specific immunotherapy such as interferon alpha or interleukins. Chemoimmunotherapy or radiotherapy must be stopped within the preceding 4 weeks (6 weeks for nitrosoureas and mitomycin C). Clinically significant heart disease (NYHA Class III or IV) i.e. NYHA class 3 congestive heart failure; myocardial infarction within the past six months; unstable angina; coronary angioplasty within the past 6 months; uncontrolled atrial or ventricular cardiac arrhythmias. Active immunodeficiency or autoimmune disease. Vitiligo was not an exclusion criterion. Other serious acute or chronic illnesses, e.g. active infections requiring antibiotics, bleeding disorders, or other conditions requiring concurrent medications not allowed during this study. Other malignancy within 3 years prior to entry into the study, except for treated non-melanoma skin cancer and cervical carcinoma in situ. Lack of availability for immunological and clinical follow-up assessments. Participation in any other clinical trial involving another investigational agent within 4 weeks prior to enrollment. Pregnancy or breastfeeding. Women of childbearing potential: Refusal or inability to use effective means of contraception.

Ages Eligible for Study
-----------------
Minimum Age: 18 Years

Sexes Eligible for Study
-----------------
All

Accepts Healthy Volunteers
-----------------
No

Study Plan
=================
How is the study designed?
-----------------

Design Details

Primary Purpose: Treatment
Allocation: Non-Randomized
Intervention Model: Sequential Assignment
Interventional Model Description: The protocol planned the inclusion of 2 cohorts, each of them including 14 patients, with metastatic cutaneous melanoma, with at least one detectable metastasis (grades AJCC 2002 III N2b to N3, grades IV M1a, M1b, and M1c without elevation of the LDH rate and without central nervous system toxicity).
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Experimental: Cohort 1: 8 HLA-A2-restricted peptides and CpG 7909<br>Patients were immunized with a combination of 8 peptides corresponding to defined tumor antigens (MAGE-1.A2, MAGE-3.A2, MAGE-4.A2, MAGE-10.A2, MAGE-C2.A2, NY-ESO-1.A2, NA17.A2 and Tyrosinase.A2), mixed with CpG 7909. Patients received six sequential injections at 2-week intervals. | Biological: 8 HLA-A2-restricted peptides and CpG 7909<br> <br> |
| Experimental: Cohort 2: 8 HLA-A2-Restricted Peptides and Montanide ISA51<br>Patients were immunized with a combination of 8 peptides corresponding to defined tumor antigens (MAGE-1.A2, MAGE-3.A2, MAGE-4.A2, MAGE-10.A2, MAGE-C2.A2, NY-ESO-1.A2, and NA17.A2), mixed with Montanide ISA 51. Tyrosinase.A2 was administered without Montanide ISA51. Patients received six sequential injections at 2-week intervals. | Biological: 8 HLA-A2-restricted peptides and Montanide ISA51<br> <br> |

What is the study measuring?
-----------------
Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Number of Patients With Cytotoxic T-lymphocyte (CTL) Response to Individual Peptides After Immunization With a Combination of 8 Peptides and CpG 7909 or Montanide ISA51. | Peripheral blood lymphocytes (PBL) were collected prior to the first dose of vaccine and after the completion of the six vaccinations in Week 13. Specific CTL directed against the 8 vaccine antigens ( NA17.A2, MAGE-1.A2, MAGE-3.A2, MAGE-4.A2, MAGE-10.A2, MAGE-C2.A2 and Tyrosinase.A2) was assessed by using re-stimulation in vitro, followed by staining with the corresponding tetramer (MLPC/tetramer). A patient was considered to have a positive CTL response when the post-vaccine CTL response against at least one of the vaccine antigens was ten times higher than the corresponding pre-treatment value. | Week 13 | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Number of Patients With Dose Limiting Toxicities (DLT). | Toxicity was evaluated according to the National Cancer Institute (CTC Scale Version 3.0, published December 12, 2003). Dose limiting toxicity (DLT) is defined as: Any Grade 3 hematological or non-hematological toxicity other than skin or flu-like symptoms Any Grade 4 toxicity To be dose-limiting, an adverse event must be definitely, probably, or possibly related to the administration of the investigational agent. | up to Week 13 | 
| Number of Patients With Tumor Responses After Immunization With a Combination of 8 Peptides and CpG 7909 or Montanide ISA51 as Measured by RECIST. | Tumor response was assessed according to the Response Evaluation Criteria in Solid Tumors (RECIST). Tumor measurements were taken at screening and at the end of Cycle 1 in Week 13. Per RECIST, target lesions are categorized as follows: Complete Response (CR): Disappearance of all target lesions; Partial Response (PR): ≥ 30% decrease in the sum of the longest diameter of target lesions; Progressive Disease (PD): ≥ 20% increase in the sum of the longest diameter of target lesions; Stable Disease (SD): small changes that do not meet above criteria. | Week 13 | 
| Number of Patients With Cytotoxic T-lymphocyte (CTL) Responses and Tumor Expression of the Corresponding Genes. | Gene expression was determined by RT-PCR on a pre-treatment tumor sample. The correlation of the induction of a CTL response against a defined antigen to the prior expression of the gene coding for this antigen by the tumor removed before vaccination was assessed. | Week 13 | 

 Terms related to the study
=================
Keywords Provided by Centre Hospitalier Valida
-----------------
Melanoma, Vaccine, Peptide, Montanide