Radiation, Chemotherapy, Vaccine and Anti-MART-1 and Anti-gp100 Cells for Patients With Metastatic Melanoma

Background:

• We have engineered human peripheral blood lymphocyte (PBLs) to express a T-cell receptor (TCR) that recognizes human leukocyte antigens (HLAA*0201) restricted epitopes derived from the gp100 or the MART-1 melanoma antigen.
• We constructed single retroviral vectors that contain both Alpha and Beta T cell receptor (TCR) chains and can mediate genetic transfer of this TCR with high efficiency (greater than 30 percent) without the need to perform any selection.
• In co-cultures with HLA-A*0201 positive melanomas these TCR transduced T cells secreted significant amount of interferon-gamma (IFN-y) but no significant secretion was observed in control co-cultures.
• gp100:154-162 TCR or MART-1:27-35 TCR transduced T-cells could efficiently kill HLAA*0201 positive tumors. There was little or no recognition of normal fibroblasts.
• Adoptive transfer of either of these TCR transferred gene modified peripheral blood lymphocyte (PBL) following a nonmyeloablative lymphodepleting regimen could mediate tumor regression in from 13-30 percent of patients with metastatic melanoma though no complete responses were seen.

Objectives:

Primary objectives:

• In cohort 1 and 2, to determine if the administration of both the anti-gp100:154-162 TCR-engineered and anti- MART-1:27-35 TCR-engineered peripheral blood lymphocytes (PBL), aldesleukin and either the gp100:154-162 peptide or the MART-1:26-35(27L) peptide to patients following a chemoradiation lymphodepleting preparative regimen will result in complete tumor regression in patients with metastatic melanoma.
• In cohort 3 and 4, to determine if the administration of both the anti-gp100:154-162 TCR-engineered and anti-MART-1:27-35 TCR-engineered CD8+ peripheral blood lymphocytes (PBL), aldesleukin and either the gp100:154-162 peptide or the MART-1:26-35(27L) peptide to patients following a chemoradiation lymphodepleting preparative regimen will result in complete tumor regression in patients with metastatic melanoma.
• Determine whether the administration of the specific vaccine (the gp100:154-162 peptide or the MART-1:26-35(27L) peptide) can increase the persistence of the specific transferred cells (anti-gp100:154-162 TCR PBL, anti-gp100:154-162 TCR CD8+ PBL, the anti-MART- 1:27-35 TCR PBL, or the anti-MART-1:27-35 TCR CD8+ PBL), respectively.

Secondary objectives:

• Determine the toxicity profile of these treatment regimen.

Eligibility:

Patients who are HLA-A*0201 positive and 18 years of age or older must have:

• metastatic melanoma with measurable disease
• been previously treated with IL-2 for melanoma;
• normal values for basic laboratory values.

Patients may not have:

• concurrent major medical illnesses;
• any form of primary or secondary immunodeficiency;
• severe hypersensitivity to any of the agents used in this study;
• contraindications for high dose aldesleukin administration.

Design:

• In cohort 1 and 2:

• Peripheral blood mononuclear cells (PBMC) will be obtained by leukapheresis (approximately 5 times 10(9) cells) and cultured in the presence of anti-CD3 (OKT3) and aldesleukin and separate aliquots will be transduced with the anti-gp100:154-162 TCR and the anti-MART-1:27-35 TCR retroviral vector.
• Transduction is initiated by exposure of approximately 10(8) to 5 times 10(8) cells to supernatant containing the retroviral vectors. These gp100 and MART-1 TCR transduced cells will be separately expanded and tested for their anti-tumor activity.
• Once engineered lymphocytes are demonstrated to be biologically active according to the strict criteria outlined in the Certificate of Analysis, patients will receive a chemoradiation lymphocyte depleting preparative regimen consisting of cyclophosphamide and fludarabine and 600 cGy total body irradiation followed by intravenous infusion of ex vivo tumor reactive, gp100 and MART-1 TCR gene-transduced cells plus IV aldesleukin (720,000 IU/kg q8h for a maximum of 15 doses).
• Patients will be randomized to receive either the gp100:154-162 peptide or the MART-1:26-35(27L) peptide emulsified in incomplete Freund's adjuvant.
• Patients will undergo complete evaluation of tumor with physical examination, computed tomography (CT) of the chest, abdomen and pelvis and clinical laboratory evaluation four to six weeks after treatment and then monthly for approximately 3 to 4 months or until off study criteria are met.
• The study will be conducted using a phase II optimal design where initially 21 evaluable patients will be enrolled into each of two cohorts. If none of the 21 patients per cohort experiences a complete clinical response, then no further patients will be enrolled but if 1 or more of the first 21 evaluable patients enrolled in that cohort have a complete clinical response, then accrual to that cohort will continue until a total of 41 evaluable patients have been enrolled in that cohort.
• Cohort 1 and 2 will be closed with the approval of amendment D.
• Cohort 3 and 4 will be initiated with approval of amendment D.

• Peripheral blood mononuclear cells (PBMC) will be obtained by leukapheresis (approximately 5 times 10(9) cells) and CD8+ cells will be selected from the cultures, using the Miltenyi CliniMACS apparatus, prior to being stimulated and transduced. Separate aliquots of CD8+ cells will be transduced with the anti-gp100:154-162 TCR and the anti-MART-1:27-35 TCR retroviral vectors.
• Transduction is initiated by exposure of approximately 10(8) to 5 times 10(8) CD8+ cells to supernatant containing the retroviral vectors. These gp100 and MART-1 TCR transduced cells will be separately expanded and tested for their anti-tumor activity.
• Once engineered lymphocytes are demonstrated to be biologically active according to the strict criteria outlined in the Certificate of Analysis, patients will receive a chemoradiation lymphocyte depleting preparative regimen consisting of cyclophosphamide and fludarabine and 600 cGy total body irradiation followed by intravenous infusion of ex vivo tumor reactive, gp100 and MART-1 TCR gene-transduced CD8+ cells plus IV aldesleukin (720,000 IU/kg q8h for a maximum of 15 doses).
• Patients will be randomized to receive either the gp100:154-162 peptide (cohort 3) or the MART-1:26-35(27L) peptide (cohort 4) emulsified in incomplete Freund's adjuvant.
• Patients will undergo complete evaluation of tumor with physical examination, CT of the chest, abdomen and pelvis and clinical laboratory evaluation four to six weeks after treatment and then monthly for approximately 3 to 4 months or until off study criteria are met.
• The study will be conducted using a phase II optimal design where initially 21 evaluable patients will be enrolled into each of cohort 3 and 4. If none of the 21 patients per cohort experiences a complete clinical response, then no further patients will be enrolled but if 1 or more of the first 21 evaluable patients enrolled in that cohort have a complete clinical response, then accrual to that cohort will continue until a total of 41 evaluable patients have been enrolled in that cohort.

• INCLUSION CRITERIA:

• Metastatic melanoma with measurable disease.
• Previously received aldesleukin (IL-2) and have been either non-responders (progressive disease) or have recurred.
• Positive for gp100 and melanoma antigen recognized by T-cells (MART-1) (at least 1 plus and greater than 5 percent) as assessed by immunohistochemistry (IHC) in the Clinical Laboratory Improvement Amendments (CLIA) approved test in the Laboratory of Pathology, Center for cancer Research (CCR), National Cancer Institute (NCI), National Institutes of Health (NIH).
• Greater than or equal to 18 years of age.
• Willing to sign a durable power of attorney.
• Able to understand and sign the Informed Consent Document.
• Clinical performance status of Eastern Cooperative Oncology Group (ECOG) 0 or 1.
• Life expectancy of greater than three months.
• Patients of both genders must be willing to practice birth control from the time of enrollment on this study and for up to four months after receiving the preparative regimen.
• Must be human leukocyte antigens (HLA-A*0201) positive
• Serology:

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• Seronegative for human immunodeficiency virus (HIV) antibody. (The experimental treatment being evaluated in this protocol depends on an intact immune system. Patients who are HIV seropositive can have decreased immune-competence and thus be less responsive to the experimental treatment and more susceptible to its toxicities.)
• Seronegative for hepatitis B antigen and hepatitis C antibody unless antigen negative.

l. Hematology:

• Absolute neutrophil count greater than 1000/m^3
• White blood cell (WBC) (greater than 3000/mm^3.
• Platelet count greater than 100,000/mm^3.
• Hemoglobin greater than 8.0 g/dl.

m. Chemistry

• Serum alanine aminotransferase (ALT)/aspartate aminotransferase (AST) less than or equal to 2.5 times the upper limit of normal.
• Serum creatinine less than or equal to 1.6 mg/dl.
• Total bilirubin less than or equal to 1.5 mg/dl, except in patients with Gilbert's Syndrome who must have a total bilirubin less than 3.0 mg/dl.

n. Women of child-bearing potential must have a negative pregnancy test because of the potentially dangerous effects of the preparative chemotherapy on the fetus.

o. More than four weeks must have elapsed since any prior systemic therapy at the time the patient receives the preparative regimen, and patients' toxicities must have recovered to a grade 1 or less (except for toxicities such as alopecia or vitiligo).

p. Six weeks must have elapsed since prior anti-CTLA4 (cytotoxic T-lymphocyte antigen 4) antibody therapy to allow antibody levels to decline, and patients who have previously received anti-CTLA4 antibody must have a normal colonoscopy with normal colonic biopsies.

Exclusion criteria

• Women of child-bearing potential who are pregnant or breastfeeding because of the potentially dangerous effects of the preparative chemotherapy on the fetus or infant.
• Active systemic infections; coagulation disorders or other major medical illnesses of the cardiovascular, respiratory or immune system; myocardial infarction; cardiac arrhythmias; obstructive or restrictive pulmonary disease.
• Any form of primary immunodeficiency (such as Severe Combined Immunodeficiency Disease).
• Systemic steroid therapy.
• History of severe immediate hypersensitivity reaction to any of the agents used in this study.
• History of coronary revascularization
• Documented left ventricular ejection fraction (LVEF) of less than 45 percent in patients with:

a) Clinically significant atrial and/or ventricular arrhythmias including but not limited to: atrial fibrillation, ventricular tachycardia, 2 degree or 3 degree heart block

b) Age greater than or equal to 60 years old

h. Documented forced expiratory volume 1 (FEV1) less than or equal to 60 percent predicted for patients with:

• A prolonged history of cigarette smoking (greater than 20 pack/year within the past 2 years)
• Symptoms of respiratory distress