This is a phase 2 study to find the best way to give this new experimental regimen and determine if it can treat metastatic melanoma in humans. In this phase 2 study, the experimental products are given initially to a group of 8 people, and if safe and found to have a significant antitumor activity, it will be given to up to 14 other people, for a total of 22 people in this study.

The study investigators' main aim is to obtain sufficient information about the effects of gene-modified cells in humans. The gene modification of cells is an attempt to direct the patient's own cells to kill melanoma cancer cells. The gene-modified cells studied in this experimental protocol are cells from the patient's blood modified in the laboratory using genetic techniques to express a specific receptor against melanoma cells.

Gene modification of cells involves the transfer of foreign genetic material (DNA) into a cell, in this case immune system cells, using a form of virus that has been modified to express the melanoma specific receptor called MART-1 T cell receptor (or TCR).

This clinical trial has two main study drugs:

• The first study drug is called gene modified MART-1 TCR T cells. T cells are a special type of white blood cells that have the ability to kill cancer cells. They are also called a cytotoxic T lymphocytes (or CTL), which means that they can kill other cells. The CTLs will be obtained from body and taken to the laboratory to grow them and modify their genes. The CTL cells will be gene modified with a virus vector similar to the HIV virus called a retrovirus vector. This retrovirus is called a vector because it has been extensively modified to make sure that it cannot make copies of itself or induce AIDS in patients. The retrovirus vector will allow expression of a protein called the T cell receptor (or TCR). This TCR is specific for a marker on the surface of melanoma cells called the MART-1 tumor antigen. The investigators expect that the insertion of the TCR specific for the MART-1 melanoma marker will allow the CTLs to be redirected to recognize and attack melanoma cancer cells.
• The second study drug is called a dendritic cell vaccine loaded with the MART-1 melanoma protein. Dendritic cells are a type of blood cells that specialize in stimulating the immune system, and will be grown in the laboratory from the patient's own blood cells. The dendritic cells do not directly fight cancer cells. Instead, they teach other cells in the body to look for cancer cells with the MART-1 protein and destroy them, therefore they function as an "on switch" for the immune system. MART-1 is a protein produced by melanoma, which may be recognized by cells of the immune system. The goal of giving the dendritic cell vaccines to the patient will be to further help the ability of the gene modified MART-1 TCR CTLs to attack melanoma lesions in the body.

These two study drugs are given after administering chemotherapy to clear up the patient's immune system before administering the gene modified MART-1 TCR T cells, and the infusion of these cells is followed by 5 days of high dose Interleukin-2, which is an FDA-approved therapy for melanoma that boosts immune system cells.

This is a two-stage phase II clinical trial with the combined primary endpoints to determine the safety, feasibility and antitumor activity of adoptive transfer of peripheral blood mononuclear cells (PBMC) genetically engineered to express the alpha and beta chains of a high affinity T cell receptor (TCR) specific for the HLA-A*0201-restricted MART-1 melanoma tumor antigen to patients with locally advanced or metastatic melanoma.

Patients with MART-1-positive locally advanced or metastatic melanoma who are HLA-A*0201-positive, and HIV, hepatitis B and C seronegative, will receive a non-myeloablative but lymphocyte depleting chemotherapy conditioning regimen consisting of cyclophosphamide and fludarabine, and then receive the adoptive transfer of autologous PBMC transduced with the MSGV1-F5AfT2AB retroviral vector, which expresses a high affinity TCR for the MART-1 melanoma antigen (MART-1 F5 TCR). Following adoptive cell transfer, patients will receive MART-126-35 peptide-pulsed dendritic cell (DC) vaccines and high dose interleukin-2 (IL-2).

• Biological: F5 TCR transgenic cells and MART-1 peptide pulsed dendritic cells
  After chemotherapy, patients receive up to 1 x 10(9) MART-1 F5 TCR transgenic T cells infused i.v., 1 x 10(7) MART-1 peptide pulsed dendritic cells intradermally, and high dose IL-2 at 600,000 IU/kg every 8 hours for up to 14 doses.
  Other Names:

  • F5 TCR transgenic cells
  • MART-1 peptide pulsed dendritic cells
  • Interleukin-2 (aldesleukin)
• Drug: non-myeloablative conditioning chemotherapy
  Patients receive non-myeloablative conditioning chemotherapy with 5 days of cyclophosphamide (60 mg/kg/day) and 2 days of fludarabine (25 mg/m2/day)

Inclusion Criteria:

• Histologically confirmed melanoma that is considered surgically incurable with either:

  • Stage IIIc melanoma including locally relapsed, satellite, in-transit lesions or bulky draining node metastasis.
  • Stage IV melanoma (M1a, M1b or M1c).
• At least 1 lesion amenable for outpatient biopsies; this should be a cutaneous or palpable metastatic site.
• MART-1 positive melanoma by RT-PCR or IHC.
• HLA-A*0201 (HLA-A2.1) positivity by molecular subtyping.
• Age greater than or equal to 18 years old.
• Life expectancy greater than 3 months assessed by a study physician.

• A minimum of one measurable lesion defined as:

  • Meeting the criteria for measurable disease according to Response Evaluation Criteria in Solid Tumors (RECIST).
  • Skin lesion(s) selected as non-completely biopsied target lesion(s) that can be accurately measured and recorded by color photography with a ruler to document the size of the target lesion(s).
• No restriction based on prior treatments.
• ECOG performance status (PS) 0 or 1.

• Adequate bone marrow and hepatic function determined within 30-60 days prior to enrollment, defined as:

  • Absolute neutrophil count more than 1.5 x 109 cells/L.
  • Platelets more than 100 x 109/L.
  • Hemoglobin more than 10 g/dL.
  • Aspartate and alanine aminotransferases (AST, ALT) less than 2.5 x ULN (less than 5 x ULN, if documented liver metastases are present).
  • Total bilirubin less than 2 x ULN (except patients with documented Gilbert's syndrome).
  • Creatinine less than 2 mg/dl (or a glomerular filtration rate greater than 60).
• Must be willing and able to accept at least two tumor biopsies.
• Must be willing and able to accept at least two leukapheresis procedures.
• Must be willing and able to provide written informed consent.

Exclusion Criteria:

• Previously known hypersensitivity to any of the agents used in this study.
• Received systemic treatment for cancer, including immunotherapy, within one month prior to dosing.
• Potential requirement for systemic corticosteroids or concurrent immunosuppressive drugs based on prior history or received systemic steroids within the last 4 weeks prior to enrollment (inhaled or topical steroids at standard doses are allowed).
• HIV seropositivity or other congenital or acquired immune deficiency state, which would increase the risk of opportunistic infections and other complications during chemotherapy-induced lymphodepletion. If there is a positive result in the infectious disease testing that was not previously known, the patient will be referred to their primary physician and/or infectious disease specialist.
• Hepatitis B or C seropositivity, which would increase the likelihood of hepatic toxicities from the chemotherapy conditioning regimen and supportive treatments. If there is a positive result in the infectious disease testing that was not previously known, the patient will be referred to their primary physician and/or infectious disease specialist.
• Dementia or significantly altered mental status that would prohibit the understanding or rendering of informed consent and compliance with the requirements of this protocol.
• Clinically active brain metastases. Radiological documentation of absence of active brain metastases at screening is required for all patients.
• Pregnancy or breast-feeding. Female patients must be surgically sterile or be postmenopausal for two years, or must agree to use effective contraception during the period of treatment and 6 months after. All female patients with reproductive potential must have a negative pregnancy test (serum/urine) within 14 days from starting the conditioning chemotherapy. The definition of effective contraception will be based on the judgment of the study investigators.

• Since IL-2 is administered following cell infusion:

  • Patients will be excluded if they have a history of clinically significant EKG abnormalities, symptoms of cardiac ischemia or arrhythmias and have a left ventricular ejection fraction (LVEF) < 45% on a cardiac stress test (stress thallium, stress MUGA, dobutamine echocardiogram, or other stress test)
  • Similarly, patients who are 50 years old with a baseline LVEF < 45% will be excluded.
  • Patients with a prolonged history of cigarette smoking or symptoms of respiratory dysfunction who do not have a normal pulmonary function test as evidenced by a FEV1 < 60% predicted will be excluded.

• California Institute of Technology
• University of Southern California
• University of Connecticut
• National Cancer Institute (NCI)