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

 -  Melanoma antigen recognized by T-cells (MART-1) and gp100 are two genes found in
 melanoma cells. An experimental procedure developed for treating patients with advanced
 melanoma uses these genes and a type of virus to make special cells called anti-MART-1
 and anti-gp100 cells, which are designed to destroy the patient's tumor. The cells are
 created in the laboratory using the patient's own tumor cells or blood cells.

 -  The procedure also uses one of two vaccines-the anti-MART-1 peptide or the anti-gp100
 peptide-to stimulate cells in the immune system that may increase the effectiveness of
 the anti-MART-1 and anti-gp100 cells. Both vaccines are made from a virus that is
 modified to carry a copy of the MART-1 gene or gp100 gene. The virus cannot cause
 disease in humans.

 Objectives:

 - To evaluate the safety and effectiveness of anti-MART-1 and anti-gp100 cells and peptide
 vaccines for treating patients with advanced melanoma.

 Eligibility:

 - Patients 18 years of age with metastatic melanoma for whom standard treatments, including
 aldesleukin (IL-2) therapy to boost immune response, have not been effective.

 Design:

 -  Participants have an initial evaluation with complete medical history, as well as scans,
 x-rays, and other tests as directed by researchers. Most of the treatments for this
 study will be given on an inpatient basis.

 -  Before the treatment begins, participants will undergo leukapheresis (removal of
 selected blood cells) to obtain cells for preparing the anti-MART-1 and anti-gp100
 cells, and for later stem cell transplantation.

 -  Preinfusion treatment: 5 days of chemotherapy and 2 days of total-body irradiation to
 prepare the immune system for receiving the anti-MART-1 and anti-gp100 cells.

 -  Infusion of cells, followed by IL-2 treatment to improve immune response. IL-2 is given
 as a 15-minute infusion through a vein every 8 hours for a maximum of 15 doses (over 5
 days).

 -  After the cell infusion, participants will be divided into two groups and will receive
 either the gp100 peptide or MART-1 vaccine, given once a week for 3 weeks. Participants
 will also have stem cell transplantation (from previously collected stem cells) to
 promote cell survival.

 -  Periodic follow-up clinic visits after hospital discharge for physical examination,
 review of treatment side effects, laboratory tests and scans every 1 to 6 months.
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:

 1. Metastatic melanoma with measurable disease.

 2. Previously received aldesleukin (IL-2) and have been either non-responders
 (progressive disease) or have recurred.

 3. 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).

 4. Greater than or equal to 18 years of age.

 5. Willing to sign a durable power of attorney.

 6. Able to understand and sign the Informed Consent Document.

 7. Clinical performance status of Eastern Cooperative Oncology Group (ECOG) 0 or 1.

 8. Life expectancy of greater than three months.

 9. 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.

 10. Must be human leukocyte antigens (HLA-A*0201) positive

 11. Serology:

 1. 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.)

 2. Seronegative for hepatitis B antigen and hepatitis C antibody unless antigen
 negative.

 l. Hematology:

 1. Absolute neutrophil count greater than 1000/m^3

 2. White blood cell (WBC) (greater than 3000/mm^3.

 3. Platelet count greater than 100,000/mm^3.

 4. Hemoglobin greater than 8.0 g/dl.

 m. Chemistry

 1. Serum alanine aminotransferase (ALT)/aspartate aminotransferase (AST) less than
 or equal to 2.5 times the upper limit of normal.

 2. Serum creatinine less than or equal to 1.6 mg/dl.

 3. 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:

 1. 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.

 2. 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.

 3. Any form of primary immunodeficiency (such as Severe Combined Immunodeficiency
 Disease).

 4. Systemic steroid therapy.

 5. History of severe immediate hypersensitivity reaction to any of the agents used in
 this study.

 6. History of coronary revascularization

 7. 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:

 1. A prolonged history of cigarette smoking (greater than 20 pack/year within the past 2
 years)

 2. Symptoms of respiratory distress