Kathleen E. Morton

Cancer regression and autoimmunity induced by cytotoxic T lymphocyte-associated antigen 4 blockade in patients with metastatic melanoma

Cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) is a critical immunoregulatory molecule (expressed on activated T cells and a subset of regulatory T cells) capable of down-regulating T cell activation. Blockade of CTLA-4 has been shown in animal models to improve the effectiveness of cancer immunotherapy. We thus treated 14 patients with metastatic melanoma by using serial i.v. administration of a fully human anti-CTLA-4 antibody (MDX-010) in conjunction with s.c. vaccination with two modified HLA-A*0201-restricted peptides from the gp100 melanoma-associated antigen, gp100:209–217(210M) and gp100:280–288(288V). This blockade of CTLA-4 induced grade III/IV autoimmune manifestations in six patients (43%), including dermatitis, enterocolitis, hepatitis, and hypophysitis, and mediated objective cancer regression in three patients (21%; two complete and one partial responses). This study establishes CTLA-4 as an important molecule regulating tolerance to “self” antigens in humans and suggests a role for CTLA-4 blockade in breaking tolerance to human cancer antigens for cancer immunotherapy.

Durable Complete Responses in Heavily Pretreated Patients with Metastatic Melanoma Using T-Cell Transfer Immunotherapy

Purpose: Most treatments for patients with metastatic melanoma have a low rate of complete regression and thus overall survival in these patients is poor. We investigated the ability of adoptive cell transfer utilizing autologous tumor-infiltrating lymphocytes (TIL) to mediate durable complete regressions in heavily pretreated patients with metastatic melanoma. Experimental Design: Ninety-three patients with measurable metastatic melanoma were treated with the adoptive transfer of autologous TILs administered in conjunction with interleukin-2 following a lymphodepleting preparative regimen on three sequential clinical trials. Ninety-five percent of these patients had progressive disease following a prior systemic treatment. Median potential follow-up was 62 months. Results: Objective response rates by Response Evaluation Criteria in Solid Tumors (RECIST) in the 3 trials using lymphodepleting preparative regimens (chemotherapy alone or with 2 or 12 Gy irradiation) were 49%, 52%, and 72%, respectively. Twenty of the 93 patients (22%) achieved a complete tumor regression, and 19 have ongoing complete regressions beyond 3 years. The actuarial 3- and 5-year survival rates for the entire group were 36% and 29%, respectively, but for the 20 complete responders were 100% and 93%. The likelihood of achieving a complete response was similar regardless of prior therapy. Factors associated with objective response included longer telomeres of the infused cells, the number of CD8+CD27+ cells infused, and the persistence of the infused cells in the circulation at 1 month (all P2 < 0.001). Conclusions: Cell transfer therapy with autologous TILs can mediate durable complete responses in patients with metastatic melanoma and has similar efficacy irrespective of prior treatment. Clin Cancer Res; 17(13); 4550–7. ©2011 AACR.

Adoptive Cell Therapy for Patients With Metastatic Melanoma: Evaluation of Intensive Myeloablative Chemoradiation Preparative Regimens

PURPOSE The two approved treatments for patients with metastatic melanoma, interleukin (IL)-2 and dacarbazine, mediate objective response rates of 12% to 15%. We previously reported that adoptive cell therapy (ACT) with autologous antitumor lymphocytes in lymphodepleted hosts mediated objective responses in 51% of 35 patients. Here, we update that study and evaluate the safety and efficacy of two increased-intensity myeloablative lymphodepleting regimens. PATIENTS AND METHODS We performed two additional sequential trials of ACT with autologous tumor-infiltrating lymphocytes (TIL) in patients with metastatic melanoma. Increasing intensity of host preparative lymphodepletion consisting of cyclophosphamide and fludarabine with either 2 (25 patients) or 12 Gy (25 patients) of total-body irradiation (TBI) was administered before cell transfer. Objective response rates by Response Evaluation Criteria in Solid Tumors (RECIST) and survival were evaluated. Immunologic correlates of effective treatment were studied. RESULTS Although nonmyeloablative chemotherapy alone showed an objective response rate of 49%, when 2 or 12 Gy of TBI was added, the response rates were 52% and 72% respectively. Responses were seen in all visceral sites including brain. There was one treatment-related death in the 93 patients. Host lymphodepletion was associated with increased serum levels of the lymphocyte homeostatic cytokines IL-7 and IL-15. Objective responses were correlated with the telomere length of the transferred cells. CONCLUSION Host lymphodepletion followed by autologous TIL transfer and IL-2 results in objective response rates of 50% to 70% in patients with metastatic melanoma refractory to standard therapies.

Randomized Study of High-Dose and Low-Dose Interleukin-2 in Patients With Metastatic Renal Cancer

PURPOSE This three-arm randomized study compares response rates and overall survival of patients with metastatic renal cell cancer (RCC) receiving high-dose or one of two low-dose interleukin-2 (IL-2) regimens. PATIENTS AND METHODS Patients with measurable metastatic RCC and a good performance status were randomized to receive either 720,000 U/kg (high-dose [HD]) or 72,000 U/kg (low-dose [LD]), both given by intravenous (IV) bolus every 8 hours. After randomly assigning 117 patients, a third arm of low-dose daily subcutaneous IL-2 was added, and an additional 283 patients were randomly assigned. RESULTS A total of 156 patients were randomly assigned to HD IV IL-2, and 150 patients to LD IV IL-2. Toxicities were less frequent with LD IV IL-2 (especially hypotension), but there were no IL-2-related deaths in any arm. There was a higher response proportion with HD IV IL-2 (21%) versus LD IV IL-2 (13%; P =.048) but no overall survival difference. The response rate of subcutaneous IL-2 (10%, partial response and complete response) was similar to that of LD IV IL-2, differing from HD IV (P =.033). Response durability and survival in completely responding patients was superior with HD IV compared with LD IV therapy (P =.04). CONCLUSION Major tumor regressions, as well as complete responses, were seen with all regimens tested. IL-2 was more clinically active at maximal doses, although this did not produce an overall survival benefit. The immunological factors which constrain the curative potential of IL-2 to only a small percentage of patients need to be further elucidated.

Chemotherapy-Refractory Diffuse Large B-Cell Lymphoma and Indolent B-Cell Malignancies Can Be Effectively Treated With Autologous T Cells Expressing an Anti-CD19 Chimeric Antigen Receptor

PURPOSE T cells can be genetically modified to express an anti-CD19 chimeric antigen receptor (CAR). We assessed the safety and efficacy of administering autologous anti-CD19 CAR T cells to patients with advanced CD19(+) B-cell malignancies. PATIENTS AND METHODS We treated 15 patients with advanced B-cell malignancies. Nine patients had diffuse large B-cell lymphoma (DLBCL), two had indolent lymphomas, and four had chronic lymphocytic leukemia. Patients received a conditioning chemotherapy regimen of cyclophosphamide and fludarabine followed by a single infusion of anti-CD19 CAR T cells. RESULTS Of 15 patients, eight achieved complete remissions (CRs), four achieved partial remissions, one had stable lymphoma, and two were not evaluable for response. CRs were obtained by four of seven evaluable patients with chemotherapy-refractory DLBCL; three of these four CRs are ongoing, with durations ranging from 9 to 22 months. Acute toxicities including fever, hypotension, delirium, and other neurologic toxicities occurred in some patients after infusion of anti-CD19 CAR T cells; these toxicities resolved within 3 weeks after cell infusion. One patient died suddenly as a result of an unknown cause 16 days after cell infusion. CAR T cells were detected in the blood of patients at peak levels, ranging from nine to 777 CAR-positive T cells/μL. CONCLUSION This is the first report to our knowledge of successful treatment of DLBCL with anti-CD19 CAR T cells. These results demonstrate the feasibility and effectiveness of treating chemotherapy-refractory B-cell malignancies with anti-CD19 CAR T cells. The numerous remissions obtained provide strong support for further development of this approach.

Tumor Progression Can Occur despite the Induction of Very High Levels of Self/Tumor Antigen-Specific CD8+ T Cells in Patients with Melanoma

The identification of many tumor-associated epitopes as nonmutated “self” Ags led to the hypothesis that the induction of large numbers of self/tumor Ag-specific T cells would be prevented because of central and peripheral tolerance. We report in this study on vaccination efforts in 95 HLA-A*0201 patients at high risk for recurrence of malignant melanoma who received prolonged immunization with the “anchor-modified” synthetic peptide, gp100209–217(210M). Vaccination using this altered peptide immunogen was highly effective at inducing large numbers of self/tumor-Ag reactive T cells in virtually every patient tested, with levels as high as 42% of all CD8+ T cells assessed by tetramer analysis. From 1 to 10% of all CD8+ cells were tumor-Ag reactive in 44% of patients and levels >10% were generated in 17% of patients. These studies were substantiated using the ELISPOT assay and a bulk cytokine release assay. Although our data regarding “tumor escape” were inconclusive, some patients had growing tumors that expressed Ag and HLA-A*0201 in the presence of high levels of antitumor T cells. There was no difference in the levels of antitumor Ag-specific T cells in patients who recurred compared with those that remained disease-free. Thus, the mere presence of profoundly expanded numbers of vaccine-induced, self/tumor Ag-specific T cells cannot by themselves be used as a “surrogate marker” for vaccine efficacy. Further, the induction of even high levels of antitumor T cells may be insufficient to alter tumor progression.

Phase I Study of the Intravenous Administration of Attenuated Salmonella typhimurium to Patients With Metastatic Melanoma

PURPOSE A strain of Salmonella typhimurium (VNP20009), attenuated by chromosomal deletion of the purI and msbB genes, was found to target to tumor and inhibit tumor growth in mice. These findings led to the present phase I study of the intravenous infusion of VNP20009 to patients with metastatic cancer. PATIENTS AND METHODS In cohorts consisting of three to six patients, 24 patients with metastatic melanoma and one patient with metastatic renal cell carcinoma received 30-minute intravenous bolus infusions containing 10(6) to 10(9) cfu/m(2) of VNP20009. Patients were evaluated for dose-related toxicities, selective replication within tumors, and antitumor effects. RESULTS The maximum-tolerated dose was 3 x 10(8) cfu/m(2). Dose-limiting toxicity was observed in patients receiving 1 x 10(9) cfu/m(2), which included thrombocytopenia, anemia, persistent bacteremia, hyperbilirubinemia, diarrhea, vomiting, nausea, elevated alkaline phosphatase, and hypophosphatemia. VNP20009 induced a dose-related increase in the circulation of proinflammatory cytokines, such as interleukin (IL)-1beta, tumor necrosis factor alpha, IL-6, and IL-12. Focal tumor colonization was observed in two patients receiving 1 x 10(9) cfu/m(2) and in one patient receiving 3 x 10(8) cfu/m(2). None of the patients experienced objective tumor regression, including those patients with colonized tumors. CONCLUSION The VNP20009 strain of Salmonella typhimurium can be safely administered to patients, and at the highest tolerated dose, some tumor colonization was observed. No antitumor effects were seen, and additional studies are required to reduce dose-related toxicity and improve tumor localization.

IL-7 administration to humans leads to expansion of CD8+ and CD4+ cells but a relative decrease of CD4+ T-regulatory cells

Lymphopenia is a serious consequence of HIV infection and the administration of cancer chemotherapeutic agents. Although growth factors can be administered to patients to increase circulating neutrophils, there is no effective method to stimulate CD8+ lymphocyte production in humans, in vivo. This report is the first to describe the administration of recombinant interleukin-7 to humans and demonstrates the ability of this cytokine to mediate selective increases in CD4+ and CD8+ lymphocytes along with a decrease in the percentage of CD4+ T-regulatory cells. These studies suggest an important role for interleukin-7 in the treatment of patients with lymphopenia.

A Phase I Study of Nonmyeloablative Chemotherapy and Adoptive Transfer of Autologous Tumor Antigen-Specific T Lymphocytes in Patients With Metastatic Melanoma

This report describes a phase I clinical trial using nonmyeloablative, lympho-depleting chemotherapy in combination with adoptive immunotherapy in patients with metastatic melanoma. The chemotherapy-conditioning schedule that induced transient lymphopenia consisted of cyclophosphamide (30 or 60 mg/kg per day for 2 days) followed by fludarabine (25 mg/m2 per day for 5 days). Immunotherapy for all patients consisted of in vitro expanded, tumor-reactive, autologous T-cell clones selected for high avidity recognition of melanoma antigens. Cohorts of three to six patients each received either no interleukin (IL)-2, low-dose IL-2 (72,000 IU/kg intravenously three times a day to a maximum of 15 doses), or high-dose IL-2 (720,000 IU/kg intravenously three times a day for a maximum of 12 doses). The toxicities associated with this treatment were transient and included neutropenia and thrombocytopenia that resolved in all patients. High dose intravenous IL-2 was better tolerated by patients after chemotherapy than during previous immunotherapy cycles without chemotherapy. No patient exhibited an objective clinical response to treatment, although five patients demonstrated mixed responses or transient shrinkage of metastatic deposits. This study established a nonmyeloablative-conditioning regimen that could be safely administered in conjunction with adoptive T-cell transfer and IL-2 in patients with metastatic melanoma.

CD8+ Enriched “Young” Tumor Infiltrating Lymphocytes Can Mediate Regression of Metastatic Melanoma

Purpose: Tumor‐infiltrating lymphocytes (TIL) and interleukin (IL)-2 administered following lymphodepletion can cause the durable complete regression of bulky metastatic melanoma in patients refractory to approved treatments. However, the generation of a unique tumor-reactive TIL culture for each patient may be prohibitively difficult. We therefore investigated the clinical and immunologic impact of unscreened, CD8+ enriched “young” TIL. Experimental Design: Methods were developed for generating TIL that minimized the time in culture and eliminated the individualized tumor-reactivity screening step. Thirty-three patients were treated with these CD8+ enriched young TIL and IL-2 following nonmyeloablative lymphodepletion (NMA). Twenty-three additional patients were treated with CD8+ enriched young TIL and IL-2 after lymphodepletion with NMA and 6 Gy of total body irradiation. Results: Young TIL cultures for therapy were successfully established from 83% of 122 consecutive melanoma patients. Nineteen of 33 patients (58%) treated with CD8+ enriched young TIL and NMA had an objective response (Response Evaluation Criteria in Solid Tumors) including 3 complete responders. Eleven of 23 patients (48%) treated with TIL and 6 Gy total body irradiation had an objective response including 2 complete responders. At 1 month after TIL infusion the absolute CD8+ cell numbers in the periphery were highly correlated with response. Conclusions: This study shows that a rapid and simplified method can be used to reliably generate CD8+ enriched young TIL for administration as an individualized therapy for advanced melanoma, and may allow this potentially effective treatment to be applied at other institutions and to reach additional patients. Clin Cancer Res; 16(24); 6122–31. ©2010 AACR.