Phase II Study of Short-Term Cultured Anti-Tumor Autologous Lymphocytes After Lymphocyte-Depleting Chemotherapy in Metastatic Melanoma

Background:

• Tumor Infiltrating Lymphocytes (TIL) can mediate the regression of bulky metastatic melanoma when administered to an autologous patient with high dose (HD) IL-2 following a non-myeloablative (NMA) but lymphodepleting chemotherapy preparative regimen.
• Clinical investigations and preclinical animal models have demonstrated that less time in culture, longer telomeres, and a less differentiated lymphocyte phenotype are associated with TIL that are capable of mediating objective clinical responses and persisting long term in the host.
• Previous methods for generating TIL require screening for anti-tumor specificity using gamma-interferon (IFN) production by the TIL. However, in vitro screening depends on autologous tumor reagents that are often unavailable; and gamma-IFN release in vitro may not be the best correlate to in vivo efficacy. Additionally, this method necessitates long in vitro culture times (44 days), and therefore reduces the clonal heterogeneity of TIL cultures, and results in TIL cultures with shorter telomere lengths and phenotypes that are skewed toward a more differentiated phenotype.
• In Surgery Branch pre-clinical experiments, we evaluated a method for rapidly generating young TIL from melanoma tumors with optimal phenotypic characteristics.

Objectives:

• In cohort 1, to determine the ability of autologous TIL cells infused after minimal in vitro culture in conjunction with high dose aldesleukin (IL-2) following a non-myeloablative lymphodepleting preparative regimen to mediate tumor regression in patients with metastatic melanoma.
• In cohort 2, to determine the ability of autologous cluster of differentiation 4 (CD4+) cell depleted TIL cells infused after minimal in vitro culture in conjunction with high dose aldesleukin (IL-2) following a non-myeloablative lymphodepleting preparative regimen to mediate tumor regression in patients with metastatic melanoma.
• In cohort 3, to determine the ability of autologous CD4+ cell depleted TIL cells infused after minimal in vitro culture in conjunction with high dose aldesleukin following chemoradiation lymphoid depleting regimen to mediate complete tumor regression in patients with metastatic melanoma.
• In a prospective randomized fashion, to compare the ability of autologous TIL cells (cohort 4) and autologous CD4plus cell depleted TIL cells (cohort regimen, to mediate tumor regression, progression free survival, and overall survival in patients with metastatic melanoma.
• Evaluate the toxicity of these treatment regimens.
• Determine the rate of repopulation of the young TIL cells in treated patients and establish in vitro correlates of TIL cultures that mediate objective response and in vivo persistence.

Eligibility:

Patients who 18 years of age or older must have:

• Metastatic melanoma;
• Normal values for basic laboratory values.

Patients may not have:

• Received prior cell transfer therapy that included non-myeloablative or ablative chemotherapy;
• Concurrent major medical illnesses;
• Any form of immunodeficiency;
• Severe hypersensitivity to any of the agents used in this study;
• Contraindications for high dose IL-2 administration.

Design:

• Patients will undergo resection to obtain tumor for generation of autologous TIL cultures.
• Cohort 1:

• All patients will receive a non-myeloablative lymphocyte depleting preparative regimen of cyclophosphamide (60 mg/kg/day IV) on days -7 and -6 and fludarabine (25 mg/m^2/day IV) on days -5 through -1.
• On day 0 patients will receive the infusion of autologous TIL and then begin high-dose aldesleukin (720,000 IU/kg IV every 8 hours for up to 15 doses).
• Clinical and Immunologic response will be evaluated about 4-6 weeks after TIL infusion.
• Using a small optimal two-stage Phase II design, initially 21 patients will be enrolled, and if two or more of the first 21 patients has a clinical response (partial response (PR) or complete response (CR)), accrual will continue to 41 patients, targeting a 20% goal for objective response. Cohort 1 will be closed with amendment D.
• Cohort 2 will be initiated with amendment D whereby CD4+ cells will be eliminated from the cultures, using the Miltenyi Clinimacs apparatus, prior to performing the rapid expansion of the young TIL cells. Patients in cohort 2 will receive CD4+ cell depleted young unselected TIL. Patients will also receive high dose IL-2 after non-myeloablative but lymphodepleting chemotherapy preparative regimen as described above for cohort 1. Clinical and immunologic response will be evaluated about 4-6 weeks after TIL infusion. Using a small optimal two-stage Phase II design, initially 18 patients will be enrolled, and if three or more of the first 18 patients have a clinical response (PR or CR), accrual will continue to 35 patients, targeting a 30% goal for objective response. With the initiation of Cohort 3 with amendment H, patients will only be accrued to Cohort 2 if they are not eligible to receive 600 cGy due to prior radiation, or to inability to mobilize cluster of differentiation 34 (CD34+) cells. Also at this time, accrual will be expanded to a total of 50 patients in cohort 2. Cohort 2 will be closed with amendment K.
• Cohort 3 will be initiated with amendment H, whereby patients will receive a chemoradiation lymphocyte depleting preparative regimen consisting of cyclophosphamide, fludarabine, and 600 cGy total body irradiation followed by intravenous infusion of autologous CD4+ cell depleted young TIL plus IV high dose IL-2. Clinical and immunologic response will be evaluated about 4-6 weeks after TIL infusion. Using a small optimal two-stage Phase II design, initially 26 patients will be enrolled, and if one or more of the first 26 patients have a complete response (CR), accrual will continue to 51 patients, targeting a 10% goal for complete response. Cohort 3 will be closed with amendment K.

Prospective randomization between cohorts 4 and 5:

• Cohort 4:

• All patients will receive a non-myeloablative lymphocyte depleting preparative regimen of cyclophosphamide (60 mg/kg/day IV) on days -7 and -6 and fludarabine (25 mg/m^2/day IV) on days -5 through -1.
• On day 0 patients will receive the infusion of autologous TIL and then begin high-dose aldesleukin (720,000 IU/kg IV every 8 hours for up to 15 doses).
• Clinical and immunologic response will be evaluated about 4-6 weeks after TIL infusion.
• Cohort 5

• All patients will receive a non-myeloablative lymphocyte depleting preparative regimen of cyclophosphamide (60 mg/kg/day IV) on days -7 and -6 and fludarabine (25 mg/m^2/day IV) on days -5 through -1.
• On day 0 patients will receive the infusion of autologous CD4+ depleted TIL and then begin high-dose aldesleukin (720,000 IU/kg IV every 8 hours for up to 15 doses).
• Clinical and immunologic response will be evaluated about 4-6 weeks after TIL infusion.

-INCLUSION CRITERIA:

• Measurable metastatic melanoma with at least one lesion that is resectable for tumor infiltrating lymphocytes (TIL) generation.
• Patients with one to three brain metastases are eligible (lesions greater than or equal to 1 cm each, or symptomatic lesions must have been treated and stable for 3 months).
• Greater than or equal to 18 years of age .
• Willing to practice birth control during treatment and for four months after receiving the preparative regimen.
• Life expectancy of greater than three months.
• 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.
• Hematology:

• Absolute neutrophil count greater than 1000/mm^3 without support of filgrastim.
• Normal white blood cell (WBC) (greater than 3000/ mm^3).
• Hemoglobin greater than 8.0 g/dl.
• Platelet count greater than 100,000/ mm^3.
• Serology:

• 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 or hepatitis C.
• Chemistry: . Serum alanine aminotransferase (ALT)/aspartate aminotransferase (AST) less than three times the upper limit of normal. Serum creatinine less than or equal to 1.6 mg/dl. Total bilirubin less than or equal to 2 mg/dl, except in patients with Gilbert's Syndrome who must have a total bilirubin less than 3 mg/dl.
• 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). Patients may have undergone minor surgical procedures with the past 3 weeks, as long as all toxicities have recovered to grade 1 or less or as specified in the eligibility criteria in Section 2.1.1.
• Six weeks must have elapsed since prior MDX-010 (Ipilimumab) therapy to allow antibody levels to decline.
• Patients who have previously received any anti-CTLA4 (cytotoxic T-lymphocyte antigen 4) antibody and experienced treatment related colitis must have a normal colonoscopy with normal colonic biopsies.

Exclusion criteria

• Prior cell transfer therapy that included non-myeloablative or ablative chemotherapy (for cohorts 4 and 5).
• 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.
• Systemic steroid therapy required.
• Active systemic infections, coagulation disorders or other active major medical illnesses of the cardiovascular, respiratory or immune system, as evidenced by a positive stress thallium or comparable test, myocardial infarction, cardiac arrhythmias, obstructive or restrictive pulmonary disease.
• Any form of primary immunodeficiency (such as Severe Combined Immunodeficiency Disease and Acquired Immune Deficiency Syndrome (AIDS)).
• Opportunistic infections (The experimental treatment being evaluated in this protocol depends on an intact immune system. Patients who have decreased immune competence may be less responsive to the experimental treatment and more susceptible to its toxicities.)
• History of severe immediate hypersensitivity reaction to any of the agents used in this study.
• History of coronary revascularization or ischemic symptoms.
• Any patient known to have an left ventricular ejection fraction (LVEF) less than or equal to 45%.
• Documented LVEF of less than or equal to 45% tested in patients with:

• Clinically significant atrial and/or ventricular arrhythmias including but not limited to:

atrial fibrillation, ventricular tachycardia, second or third degree heart block.

• Age greater than or equal to 60 years old.
• Documented forced expiratory volume 1 (FEV1) less than or equal to 60% predicted tested in patients with:

• A prolonged history of cigarette smoking
• Symptoms of respiratory dysfunction