An in vitro technique was developed to generate activated rat T cells, with antitumor activity. Splenic mononuclear cells (SMC) from outbred Wistar and inbred Wistar-Munich rats were stimulated with Concanavalin A and recombinant human interleukin-2 (rIL-2) in vitro for 48 h. After 2 days, the nonadherent cells began proliferating and were maintained in rIL-2 for up to 18 days in vitro. FACScan analysis revealed that SMC was a mixture of cell types; however, CD5+ T cells rapidly increased and became the predominant cell type after 5 days in culture. SMC induced cytolysis of YAC-1, but not C6 glioma cells in 4 h 51Cr release assays. In contrast, 5- and 9-day T cells lysed C6 glioma and YAC-1 cells. The C6 cells were admixed with cultured effector cells at various effector-to-target (E:T) ratios and were injected into the right cerebral hemisphere of Wistar and Wistar-Munich rats for a Winn assay. Histopathologic evaluations revealed that a) SMC had no effect; b) 2- and 5-day T cells, injected at E:T ratios greater than 5:1, caused significant reduction in tumor size; and c) 2- or 5-day T cells, at a 40:1 E:T ratio, resulted in little or no histologic evidence of tumor. Eighty-three percent of animals receiving C6 and 5-day mitogen-stimulated lymphokine activated killer cells at an E:T ratio of 40:1 were alive 120 days postinjection (p less than 0.05).

Peripheral blood mononuclear cells (PBM) from normal donors and patients with recurrent glioma were activated initially for 48-72 h with phytohemagglutinin-P (PHA) and recombinant human interleukin-2 (IL-2), and then proliferated in vitro for up to 5 months with IL-2. These cells are termed mitogen-stimulated lymphokine-activated T killer (T-LAK) cells. We measured patterns of T-LAK cell growth, in vitro cytolytic activity on a panel of continuous and primary tumor cells, and the phenotypes of the cells in these cultures. Lymphocyte viability declined dramatically over the first 3-5 days; and then the remaining cells in these cultures began to divide and maintained a constant 30-36 h doubling time for long periods in vitro. Phenotyping revealed that cells in the initial few days of culture were heterogeneous, but became almost totally CD3 T cells after 7-10 days in culture. The T-LAK cells from individual normal donors and cancer patients demonstrated a non-genetically restricted cytolytic ability against a panel of both continuous cell lines and primary autologous and allogeneic glioblastoma cells in vitro. This technique provides a method of generating large numbers of autologous cytolytic T cells with non-restricted anti-tumor activity that can be derived from peripheral blood mononuclear cells.

Associations between HLA class I antigen expression and the efficacy of a melanoma vaccine (Melacine; Corixa Corp.) were initially described in stage IV melanoma. Similar associations were observed in S9035, a phase III adjuvant trial evaluating Melacine for 2 years compared with observation in patients with stage II melanoma. This report provides long-term results. The effects of treatment on relapse-free survival (RFS) and overall survival (OS) were evaluated, and prespecified analyses investigated associations between treatment and HLA expression. Multivariable analyses were adjusted for tumor thickness, ulceration and site, method of nodal staging, and sex. P = 0.01 was considered statistically significant in subset analyses to account for multiple comparisons. For the entire study population of 689 patients, there were no significant differences in RFS or OS by treatment arm. HLA serotyping was performed on 553 (80%) patients (vaccine, 294; observation, 259). Among the subpopulation with HLA-A2 and/or HLA-Cw3 serotype, vaccine arm patients (n = 178) had marginally improved RFS (adjusted P = 0.02) and significantly improved OS compared with observation arm patients (n = 145), with 10-year OS of 75% and 63%, respectively [hazard ratio (HR), 0.62; 99% confidence interval (CI), 0.37-1.02; P = 0.01]. There was no impact of HLA-A2 and/or HLA-Cw3 expression on observation arm patients. An analysis of mature data from S9035 indicates a significant OS benefit from adjuvant vaccine therapy for patients with HLA-A2- and/or HLA-Cw3-expressing melanoma. The possibility of interactions between HLA type and outcome should be considered in future immunotherapy trials. Further investigations of melanoma-associated antigens present in Melacine and presented by HLA-A2 and HLA-Cw3 may be warranted.

Tumor necrosis factor-alpha (TNF-alpha) is an immunomodulatory cytokine that has exhibited anti-tumor activity in a variety of experimental systems. However, the toxicities associated with systemic administration of TNF-alpha have limited its clinical utility and have led to the investigation of targeted delivery techniques with the ability to present the TNF-alpha dose directly to the vascular bed of the tumor. The intra-arterial (IA) administration of TNF-alpha to patients with liver metastases represents one such approach, and recent work suggests that subsequent ablation of the tumor's arterial supply via embolization may enhance the efficacy of intra-arterial treatments (hepatic chemoembolization). The present study was undertaken to test the hypothesis that IA administration of TNF-alpha is superior to the intravenous (IV) route for inhibition of tumor growth in a regionally confined rat mammary adenocarcinoma model that provides for ablation of the arterial supply to the tumor following cytokine therapy. Rats bearing hind limb mammary adenocarcinomas received single IA or IV infusions of 8 x 10(5), 1 x 10(6), and 1.5 x 10(6) units of TNF-alpha via the common femoral artery (CFA) followed 1 h later by ligation of the artery. Control animals received either no treatment or IA infusion of 2% normal rat serum (NRS) followed by ipsilateral CFA ligation. Tumor size was measured every other day after treatment. Tumor growth inhibition occurred in the first 5 to 10 days after treatment. IV administration of TNF-alpha did not result in visual tumor necrosis or significant reduction in the rate of tumor growth. IA administration of TNF-alpha resulted in statistically significant diminution of tumor size as compared to untreated controls and animals receiving IA 2% normal rat serum (NRS; P<0.05 at days 6, 8 and 10), regardless of the dose employed. The maximum growth inhibition with IA TNF-alpha was a 91% reduction in tumor volume that was achieved with a dose of 1 x 10(6) U TNF-alpha. These results demonstrate improved anti-tumor activity with the IA administration of TNF-alpha over the IV route in a regionally confined mammary adenocarcinoma. IA administration of biologic response modifiers like TNF-alpha may therefore be a useful approach for the hepatic chemoembolization of breast adenocarcinomas metastatic to the liver.

Natural killer (NK) cells are large granular lymphocytes that promote the antitumor response via communication with other cell types in the tumor microenvironment. Previously, we have shown that NK cells secrete a profile of immune stimulatory factors (e.g., IFNγ, MIP-1α, and TNFα) in response to dual stimulation with the combination of antibody (Ab)-coated tumor cells and cytokines, such as IL12. We now demonstrate that this response is enhanced in the presence of autologous monocytes. Monocyte enhancement of NK cell activity was dependent on cell-to-cell contact as determined by a Transwell assay. It was hypothesized that NK cell effector functions against Ab-coated tumor cells were enhanced via binding of MICA on monocytes to NK cell NKG2D receptors. Strategies to block MICA-NKG2D interactions resulted in reductions in IFNγ production. Depletion of monocytes in vivo resulted in decreased IFNγ production by murine NK cells upon exposure to Ab-coated tumor cells. In mice receiving trastuzumab and IL12 therapy, monocyte depletion resulted in significantly greater tumor growth in comparison to mock-depleted controls (P < 0.05). These data suggest that NK cell-monocyte interactions enhance NK cell antitumor activity in the setting of monoclonal Ab therapy for cancer. Cancer Immunol Res; 5(9); 778-89. ©2017 AACR.

Recent advances have permitted successful therapeutic targeting of the immune system in head and neck squamous cell carcinoma (HNSCC). These new immunotherapeutic targets and agents are being rapidly adopted by the oncologic community and hold considerable promise. The National Cancer Institute sponsored a Clinical Trials Planning Meeting to address the issue of how to further investigate the use of immunotherapy in patients with HNSCC. The goals of the meeting were to consider phase 2 or 3 trial designs primarily in 3 different patient populations: those with previously untreated, human papillomavirus-initiated oropharyngeal cancers; those with previously untreated, human papillomavirus-negative HNSCC; and those with recurrent/metastatic HNSCC. In addition, a separate committee was formed to develop integrative biomarkers for the clinical trials. The meeting started with an overview of key immune components and principles related to HNSCC, including immunosurveillance and immune escape. Four clinical trial concepts were developed at the meeting integrating different immunotherapies with existing standards of care. These designs were presented for implementation by the head and neck committees of the National Cancer Institute-funded National Clinical Trials Network. This article summarizes the proceedings of this Clinical Trials Planning Meeting, the purpose of which was to facilitate the rigorous development and design of randomized phase 2 and 3 immunotherapeutic trials in patients with HNSCC. Although reviews usually are published immediately after the meeting is held, this report is unique because there are now tangible clinical trial designs that have been funded and put into practice and the studies are being activated to accrual. Cancer 2017;123:1259-1271. © 2016 American Cancer Society.

Abstract Scientific discoveries that provide strong evidence of antitumor effects in preclinical models often encounter significant delays before being tested in patients with cancer. While some of these delays have a scientific basis, others do not. We need to do better. Innovative strategies need to move into early stage clinical trials as quickly as it is safe, and if successful, these therapies should efficiently obtain regulatory approval and widespread clinical application. In late 2009 and 2010 the Society for Immunotherapy of Cancer (SITC), convened an "Immunotherapy Summit" with representatives from immunotherapy organizations representing Europe, Japan, China and North America to discuss collaborations to improve development and delivery of cancer immunotherapy. One of the concepts raised by SITC and defined as critical by all parties was the need to identify hurdles that impede effective translation of cancer immunotherapy. With consensus on these hurdles, international working groups could be developed to make recommendations vetted by the participating organizations. These recommendations could then be considered by regulatory bodies, governmental and private funding agencies, pharmaceutical companies and academic institutions to facilitate changes necessary to accelerate clinical translation of novel immune-based cancer therapies. The critical hurdles identified by representatives of the collaborating organizations, now organized as the World Immunotherapy Council, are presented and discussed in this report. Some of the identified hurdles impede all investigators; others hinder investigators only in certain regions or institutions or are more relevant to specific types of immunotherapy or first-in-humans studies. Each of these hurdles can significantly delay clinical translation of promising advances in immunotherapy yet if overcome, have the potential to improve outcomes of patients with cancer.