Dendritic Cell Vaccination During Lymphoid Reconstruction

This is a dose ranging study of intranodal administration of autologous dendritic cells (DC) pulsed with tumor antigen class I peptides derived from MART-1 (26-35) (27L), gp100 (209-217 (210M), gp100 280-288 (288V), NY-ESO-1 157-165 (165V) and tyrosinase 207-215 as well as class II MART-1 (51-73), NY-ESO-1 (119-143), MAGE-3 (243-258) and tyrosinase (450-462) peptides preceded by Autologous Lymphocyte Infusion (ALI) and one of two doses of Fludarabine. The nine or ten amino acid peptides representing HLA-A2 restricted T cell epitopes of MART-1, gp100, NY-ESO-1 and tyrosinase will be pulsed onto autologous dendritic cells produced by incubation of peripheral blood mononuclear cells obtained by apheresis with interleukin-4 (IL-4) and GM-CSF and pulsed with four helper peptides then matured with a cytokine cocktail including TNF-a, IL-6, IL-1b and PGE2. Melanoma antigen peptide-pulsed dendritic cells will be administered at a total dose of 10 million cells each for four intranodal injections to patients with chemotherapy-naïve metastatic melanoma.

DC matured with a cytokine cocktail and pulsed with class I and II peptides will be injected intranodally, weekly for two doses, then every two weeks for two doses, for a total of four injections to each cohort.

Inclusion criteria

• Metastatic melanoma with measurable disease after attempted curative surgical therapy and without prior chemotherapy; adjuvant interferon or isolated limb perfusion is allowed.
• Tumor tissue must be available for immunohistochemical analysis, and specimens will stained for MART-1/tyrosinase/NY-ESO-1 by immunohistochemical staining and will also be stained for HMB-45 by immunohistochemistry, and positivity for at least one will be an entry requirement.
• Patients must be HLA-A *0201 positive by a DNA polymerase chain reaction (PCR) analysis.
• Serum creatinine of 2.0 mg/dl or less, total bilirubin of 2.0 mg/dl or less, and alanine transaminase/aspartic transaminase (ALT/AST) of less than 3X institutional upper limit of normal (ULN).
• Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1.
• Patients must be able to understand and sign an Institutional Review Board (IRB) approved informed consent form.
• Patients must have whit blood count (WBC) of 3000 or greater, platelets of 100,000 or greater, and hemoglobin of 9.0 gm/dl or more.
• Patients must be seropositive for Epstein-Barr virus (EBV).
• Patients with unresectable stages III/IV uveal melanoma and metastatic mucosal melanoma will be eligible for this trial.

Exclusion criteria

• Patients who are undergoing or have undergone in the past month any other therapy for their melanoma, including radiation therapy, chemotherapy and adjuvant therapy.
• Have major systemic infections, coagulation disorders, or other major medical illnesses (MI) of the cardiovascular or respiratory systems, or have had a documented MI in the last 6 months.
• Require steroid therapy.
• Are pregnant or lactating.
• Are known to be positive for hepatitis BsAg, Hepatitis C or human immunodeficiency virus (HIV) antibody, since cells for DC cannot be grown in the laboratory when virus contaminated.
• Have a prior history of uveitis or autoimmune inflammatory eye disease.
• Have previously received the gp100 209-217 (210M), MART-1 26-35 (27L), gp100 280-288 (288V), tyrosinase 207-215 or NY-ESO-1 157-165 (165V) peptides.
• Have had another malignancy other than cervical carcinoma-in-situ or basal cell

/squamous cancer of the skin, unless they have undergone curative therapy more than 5 years ago and are still free of detectable disease.

• Since this trial increase the risk of immunological impairment, patients with the following will be excluded from this trial: Hypogammaglobulinemia, Lymphocytopenia, History of impaired immune response, tuberculosis (TB) or positive purified protein derivative (PPD) unless they have received BCG vaccine.