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There are approximately 44,000 cases of human papillomavirus–associated (HPV-associated) cancer each year in the United States, most commonly caused by HPV types 16 and 18. Prophylactic vaccines successfully prevent healthy people from acquiring HPV infections via HPV-specific antibodies. In order to treat established HPV-associated malignancies, however, new therapies are necessary. Multiple recombinant gorilla adenovirus HPV vaccine constructs were evaluated in NSG-β2m–/– peripheral blood mononuclear cell–humanized mice bearing SiHa, a human HPV16+ cervical tumor, and/or in the syngeneic HPV16+ TC-1 model. PRGN-2009 is a therapeutic gorilla adenovirus HPV vaccine containing multiple cytotoxic T cell epitopes of the viral oncoproteins HPV 16/18 E6 and E7, including T cell enhancer agonist epitopes. PRGN-2009 treatment reduced tumor volume and increased CD8+ and CD4+ T cells in the tumor microenvironment of humanized mice bearing the human cervical tumor SiHa. PRGN-2009 monotherapy in the syngeneic TC-1 model also reduced tumor volumes and weights, generated high levels of HPV16 E6–specific T cells, and increased multifunctional CD8+ and CD4+ T cells in the tumor microenvironment. These studies provide the first evaluation to our knowledge of a therapeutic gorilla adenovirus HPV vaccine, PRGN-2009, showing promising preclinical antitumor efficacy and induction of HPV-specific T cells, along with the rationale for its evaluation in clinical trials.

Background The safety and efficacy of bintrafusp alfa, a first-in-class bifunctional fusion protein composed of the extracellular domain of the TGF-βRII receptor (a TGF-β ‘trap’) fused to a human IgG1 mAb blocking PD-L1, have been demonstrated in patients with human papillomavirus (HPV)-related cancers in an open label, multicenter phase 1 trial (NCT02517398), and an open-label, single center phase 2 trial (NCT03427411). The current study aimed to investigate whether HPV-16-specific T cells are expanded with therapy and associate with the clinical response of patients in these trials. We also present pre-clinical evidence from a mouse model of HPV-associated cancer supporting the combination of bintrafusp alfa with an HPV-16 targeted therapeutic vaccine and an immunostimulatory cytokine. Methods Peripheral blood mononuclear cells (PBMC) were obtained from 33 patients prior to and 2 weeks after 1 and/or 3 cycles of bintrafusp alfa and evaluated for HPV-16 specific CD4+ and CD8+ T cells. PBMCs were stimulated with 15-mer peptide pools of the HPV-16 E6 and E7 oncoproteins, and T cell responses were assessed for the production of cytokines (TNFa, IFNg, IL-2) and positivity for the degranulation marker CD107a. Multifunctional T cells, positive for >2 measures, were also enumerated. For pre-clinical studies, a syngeneic mouse model of TC-1 carcinoma was treated with bintrafusp alfa alone or in combination with a liposomal-based HPV-16 therapeutic vaccine (PDS 0101) and a tumor targeting immunocytokine (NHS-muIL12) and evaluated for anti-tumor activity and immune responses. Results HPV-16 specific T cells were increased after 1 cycle of bintrafusp alfa in a greater proportion of responders (9/14) than non-responders (4/17) (p=0.03). In addition, the magnitude of HPV-16 specific T cells was greater after 1 (p=0.04) and 3 (p Conclusions An early increase in HPV-16 specific T cells (after a single administration of bintrafusp alfa, prior to restaging) was associated with clinical activity in patients with HPV-related cancers undergoing bintrafusp alfa therapy. This evidence, and the pre-clinical finding of enhanced antitumor activity observed when combining bintrafusp alfa with an HPV-16 targeted vaccine and an immunostimulatory cytokine have provided the rational for an ongoing study evaluating this combination in patients with advanced HPV-associated malignancies (NCT04287868). Ethics Approval All patients provided written informed consent for participation in a clinical trial that was approved by the Institutional Review Board at the National Cancer Institute (NCT02517398, NCT03427411)

BackgroundWhile prophylactic human papillomavirus (HPV) vaccines will certainly reduce the incidence of HPV-associated cancers, these malignancies remain a major health issue. PDS0101 is a liposomal-based HPV therapeutic vaccine consisting of the immune activating cationic lipid R-DOTAP and HLA-unrestricted HPV16 peptides that has shown in vivo CD8+ T cell induction and safety in a phase I study. In this report, we have employed the PDS0101 vaccine with two immune modulators previously characterized in preclinical studies and which are currently in phase II clinical trials. Bintrafusp alfa (M7824) is a first-in-class bifunctional fusion protein composed of the extracellular domains of the transforming growth factor-β receptor type II (TGFβRII) fused to a human IgG1 monoclonal antibody blocking programmed cell death protein-1 ligand (PDL1), designed both as a checkpoint inhibitor and to bring the TGFβRII ‘trap’ to the tumor microenvironment (TME). NHS-interleukin-12 (NHS-IL12) is a tumor targeting immunocytokine designed to bring IL-12 to the TME and thus enhance the inflammatory Th1 response.MethodsWe employed TC-1 carcinoma (expressing HPV16 E6 and E7 and devoid of PDL1 expression) in a syngeneic mouse model in monotherapy and combination therapy studies to analyze antitumor effects and changes in immune cell types in the spleen and the TME.ResultsAs a monotherapy, the PDS0101 vaccine generated HPV-specific T cells and antitumor activity in mice bearing HPV-expressing mEER oropharyngeal and TC-1 lung carcinomas. When used as a monotherapy in the TC-1 model, NHS-IL12 elicited antitumor effects as well as an increase in CD8+ T cells in the TME. When used as a monotherapy, bintrafusp alfa did not elicit antitumor effects or any increase in T cells in the TME. When all three agents were used in combination, maximum antitumor effects were observed, which correlated with increases in T cells and T-cell clonality in the TME.ConclusionThese studies provide the rationale for the potential clinical use of combinations of agents that can (1) induce tumor-associated T-cell responses, (2) potentiate immune responses in the TME and (3) reduce immunosuppressive entities in the TME.

The lack of serial biopsies in patients with a range of carcinomas has been one obstacle in our understanding of the mechanism of action of immuno-oncology agents as well as the elucidation of mechanisms of resistance to these novel therapeutics. While much information can be obtained from studies conducted with syngeneic mouse models, these models have limitations, including that both tumor and immune cells being targeted are murine and that many of the immuno-oncology agents being evaluated are human proteins, and thus multiple administrations are hampered by host xenogeneic responses. Some of these limitations are being overcome by the use of humanized mouse models where human peripheral blood mononuclear cells (PBMC) are engrafted into immunosuppressed mouse strains. Bintrafusp alfa (M7824) is an innovative first-in-class bifunctional fusion protein composed of the extracellular domain of the TGF-βRII to function as a TGF-β “trap” fused to a human IgG1 antibody blocking PD-L1. A phase I clinical trial of bintrafusp alfa showed promising anti-tumor efficacy in heavily pretreated advanced solid tumors, and multiple clinical studies are currently ongoing. There is still much to learn regarding the mechanism of action of bintrafusp alfa, including its effects on both human immune cells in the periphery and in the tumor microenvironment (TME), and any temporal effects upon multiple administrations. By using the NSG-β2m−/− mouse strain humanized with PBMC, we demonstrate here for the first time: (a) the effects of bintrafusp alfa administration on human immune cells in the periphery vs. the TME using three different human xenograft models; (b) temporal effects upon multiple administrations of bintrafusp alfa; (c) phenotypic changes induced in the TME, and (d) variations observed in the use of multiple different PBMC donors. Also discussed are the similarities and differences in the data thus far obtained employing murine syngeneic models, from clinical trials, and in the use of this humanized mouse model. The results described here may guide the future use of this agent or similar immunotherapy agents as monotherapies or in combination therapy studies.

// Samuel T. Pellom Jr. 1, 2, 3 , Duafalia F. Dudimah 1 , Menaka C. Thounaojam 1 , Roman V. Uzhachenko 1 , Ashutosh Singhal 1 , Ann Richmond 4, 5, 6, 7, 8, 9 , Anil Shanker 1, 3, 7, 8, 9 1 Department of Biochemistry and Cancer Biology, School of Medicine, Meharry Medical College, Nashville, Tennessee, USA 2 Department of Microbiology and Immunology, School of Medicine, Meharry Medical College, Nashville, Tennessee, USA 3 School of Graduate Studies and Research, Meharry Medical College, Nashville, Tennessee, USA 4 Tennessee Valley Healthcare System, Nashville, Tennessee, USA 5 Department of Veterans Affairs, Nashville, Tennessee, USA 6 Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, Tennessee, USA 7 Host-Tumor Interactions Research Program, Vanderbilt-Ingram Comprehensive Cancer Center, Vanderbilt University, Nashville, Tennessee, USA 8 Vanderbilt Center for Immunobiology, Vanderbilt University, Nashville, Tennessee, USA 9 Vanderbilt Center for Translational and Clinical Immunology, Vanderbilt University, Nashville, Tennessee, USA Correspondence to: Anil Shanker, email: ashanker@mmc.edu Keywords: proteasome inhibition, CD8 + T cells, immunosuppression, cancer immunotherapy, adoptive cell therapy Received: November 07, 2016 Accepted: December 07, 2016 Published: December 29, 2016 ABSTRACT Tumor-induced immune tolerance poses a major challenge for therapeutic interventions aimed to manage cancer. We explored approaches to overcome T-cell suppression in murine breast and kidney adenocarcinomas, and lung fibrosarcoma expressing immunogenic antigens. We observed that treatment with a reversible proteasome inhibitor bortezomib (1 mg/kg body weight) in tumor-bearing mice significantly enhanced the expression of lymphocyte-stimulatory cytokines IL-2, IL-12, and IL-15. Notably, bortezomib administration reduced pulmonary nodules of mammary adenocarcinoma 4T1.2 expressing hemagglutinin (HA) model antigen (4T1HA) in mice. Neutralization of IL-12 and IL-15 cytokines with a regimen of blocking antibodies pre- and post-adoptive transfer of low-avidity HA 518-526 -specific CD8 + T-cells following intravenous injection of 4T1HA cells increased the number of pulmonary tumor nodules. This neutralization effect was counteracted by the tumor metastasis-suppressing action of bortezomib treatments. In bortezomib-treated 4T1HA tumor-bearing mice, CD4 + T-cells showed increased IL-2 production, CD11c + dendritic cells showed increased IL-12 and IL-15 production, and HA-specific activated CD8 + T-cells showed enhanced expression of IFNγ, granzyme-B and transcription factor eomesodermin. We also noted a trend of increased expression of IL-2, IL-12 and IL-15 receptors as well as increased phosphorylation of STAT5 in tumor-infiltrating CD8 + T-cells following bortezomib treatment. Furthermore, bortezomib-treated CD8 + T-cells showed increased phosphorylation of mitogen-activated protein kinase p38, and Akt, which was abrogated by phosphatidylinositide 3-kinase (PI3K) inhibitor. These data support the therapeutic potential of bortezomib in conjunction with other immunotherapies to augment the strength of convergent signals from CD8 + T-cell signaling molecules including TCR, cytokine receptors and downstream PI3K/Akt/STAT5 pathways to sustain CD8 + T-cell effector function in the tumor microenvironment.

Here we describe a novel bifunctional fusion protein, designated N-809. This molecule comprises the IL-15/IL15Rα superagonist complex containing the Fc-domain of IgG1 (N-803, formerly designated as ALT-803) fused to two single chain anti-PD-L1 domains. The fully human IgG1 portion of the N-809 molecule was designed to potentially mediate antibody dependent cellular cytotoxicity (ADCC). The studies reported here show that N-809 has the same ability to bind PD-L1 as an anti-PD-L1 monoclonal antibody. RNAseq studies show the ability of N-809 to alter the expression of an array of genes of both CD4+ and CD8+ human T cells, and to enhance their proliferation; CD8+ T cells exposed to N-809 also have enhanced ability to lyse human tumor cells. An array of genes was differentially expressed in human natural killer (NK) cells following N-809 treatment, and there was increased expression of several surface activating receptors; there was, however, no increase in the expression of inhibitory receptors known to be upregulated in exhausted NK cells. N-809 also increased the cytotoxic potential of NK cells, as shown by increased expression of granzyme B and perforin. The lysis of several tumor cell types was increased when either NK cells or tumor cells were exposed to N-809. Similarly, the highest level of ADCC was seen when both NK cells (from donors or cancer patients) and tumor cells were exposed to N-809. These studies thus demonstrate the multi-functionality of this novel agent.

Tributyltin (TBT), a toxic environmental contaminant, has been widely utilized for various industrial, agricultural and household purposes. Its usage has led to a global contamination and its bioaccumulation in aquatic organisms and terrestrial mammals. Previous studies suggest that TBT has debilitating effects on the overall immune function of animals, rendering them more vulnerable to diseases. TBT (at concentrations that have been detected in human blood) alters secretion of inflammatory cytokines from human lymphocytes ex vivo. Thus, it is important to determine if specified levels of TBT can alter levels of cytokines in an in vivo system. Mice were exposed to biologically relevant concentrations of TBT (200, 100 or 25 nM final concentrations). The quantitative determination of interferon (IFN)-γ, tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL2, IL5, IL7, IL12βp40, IL13, IL15, keratinocyte chemoattractant (KC), macrophage inflammatory protein 1β (MIP), MIP2 and regulated on activation normal T-cell-expressed and secreted (RANTES) was performed in mouse sera by MAGPIX analysis and Western blot. Results indicated alterations (both decreases and increases) in several cytokines. The pro-inflammatory cytokines IFNγ, TNFα, IL-1β, IL-2, IL5, IL12βp40 and IL-15 were altered as were the chemokines MIP-1 and RANTES and the anti-inflammatory cytokine IL-13. Increases in IFNγ and TNFα were seen in the serum of mice exposed to TBT for less than 24 h. Levels of IL1β, IL-12 βp40, IL-5 and IL-15 were also modulated in mouse serum, depending on the specific experiment and exposure level. IL-2 was consistently decreased in mouse serum when animals were exposed to TBT. There were also TBT-induced increases in MIP-1β, RANTES and IL-13. These results from human and murine samples clearly suggest that TBT exposures modulate the secretion inflammatory cytokines.

Cancer immunotherapy shows great promise but many patients fail to show objective responses, including in cancers that can respond well, such as melanoma and renal adenocarcinoma. The proteasome inhibitor bortezomib sensitizes solid tumors to apoptosis in response to TNF-family death ligands. Because T cells provide multiple death ligands at the tumor site, we investigated the effects of bortezomib on T-cell responses in immunotherapy models involving low-avidity antigens. Bortezomib did not affect lymphocyte or tissue-resident CD11c+CD8+ dendritic cell counts in tumor-bearing mice, did not inhibit dendritic cell expression of costimulatory molecules, and did not decrease MHC class I/II-associated antigen presentation to cognate T cells. Rather, bortezomib activated NF-κB p65 in CD8+ T cells, stabilizing expression of T-cell receptor CD3ζ and IL2 receptor-α, while maintaining IFNγ secretion to improve FasL-mediated tumor lysis. Notably, bortezomib increased tumor cell surface expression of Fas in mice as well as human melanoma tissue from a responsive patient. In renal tumor-bearing immunodeficient Rag2−/− mice, bortezomib treatment after adoptive T-cell immunotherapy reduced lung metastases and enhanced host survival. Our findings highlight the potential of proteasome inhibitors to enhance antitumor T-cell function in the context of cancer immunotherapy. Cancer Res; 75(24); 5260–72. ©2015 AACR.

Bortezomib is an inhibitor of the ubiquitin-proteasome proteolytic pathway responsible for intracellular protein turnover. Cellular proteins controlled by this pathway represent a diverse group of potential therapeutic targets, particularly in cancer cells, which exploit this proteasomal pathway to promote their growth and diminish apoptosis. Along with inhibiting the proteasome and thus sensitizing tumor cells to apoptosis, bortezomib may also have multiple effects on the host immune responses. This review summarizes the effects that bortezomib may play on immune cell subsets in various disease states in modifying lymphocyte receptors, ligands, the expression of various cytokines and chemokines and their downstream signaling. We also propose steps that can be taken to refine combinatorial strategies that include bortezomib to improve current immunotherapeutic approaches.

Butyrate is a histone deacetylase inhibitor implicated in many studies as a potential therapy for various forms of cancer. High concentrations of butyrate (>1.5 mM) have been shown to activate apoptosis in several cancer cell lines including prostate, breast, and leukemia. Butyrate is also known to influence multiple signaling pathways that are mediators of cytokine production. The purpose of this study was to evaluate the impact of high concentrations of butyrate on the cancer microenvironment vis-à-vis apoptosis, cellular migration, and capacity to modulate cytokine expression in cancer cells. The results indicate that high concentrations of butyrate induced a 2-fold activation of caspase-3 and reduced cell viability by 60% in U937 leukemia cells. Within 24 hours, butyrate significantly decreased the levels of chemokines CCL2 and CCL5 in HL-60 and U937 cells, and decreased CCL5 in THP-1 leukemia cells. Differential effects were observed in treatments with valproic acid for CCL2 and CCL5 indicating butyrate-specificity. Many of the biological effects examined in this study are linked to activation of the AKT and MAPK signaling pathways; therefore, we investigated whether butyrate alters the levels of phosphorylated forms of these signaling proteins and how it correlated with the expression of chemokines. The results show that butyrate may partially regulate CCL5 production via p38 MAPK. The decrease in p-ERK1/2 and p-AKT levels correlated with the decrease in CCL2 production. These data suggest that while promoting apoptosis, butyrate has the potential to influence the cancer microenvironment by inducing differential expression of cytokines.

The effects of Tributyltin (TBT), a widespread environmental contaminant, on the induction of in vivo and ex vivo cytokine production and secretion were investigated. Human monocyte-depleted peripheral blood mononuclear cells and different groups of Balb/C mice were stimulated with desired concentrations of TBT (200-2.5 nM for human studies and 200-25 nM for mouse studies). The quantitative determination of IFNγ, TNFα and IL-1β was performed in mouse sera by MagPix analysis and western blot. The levels of these cytokines were measured at different time points: 6, 12, 24 and 48 h after injection. Analysis of IFNγ and TNFα in human cells were conducted by ELISA and western blot. MAPK and NF-κB pathways were inhibited to determine TBT-induced dysregulation of IFNγ and TNFα. Inhibition of the p38 and p44/42 pathways blocked the TBT-induced elevation of IFNγ, while the inhibition of the p38 pathway blocked the TBT-induced elevation of TNFα. Results showed that TBT increased the levels of IFNγ, TNFα and IL-1β in the sera of mice. IFNγ and TNFα secretion from human monocyte-depleted peripheral blood mononuclear cells also increased, showing striking agreement in the response to TBT between the mouse and human systems. These data suggest that TBT exposure can result in an inflammatory environment, which could predispose the host to various immunopathologies. Grant U54CA163066 from the National Institute of Health Citation Format: Shanieek Lawrence, Samuel T. Pellom, Anil Shanker, Margaret Whalen. Tributyltin-induced dysregulation of inflammatory cytokine levels in human and mouse immune cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 934.

Tributyltin (TBT) is an environmental contaminant that has been used for several industrial, agricultural and household purposes. Despite limited bans on the usage of Tributyltin, the stability and toxicity of the compound may prove to have negative effects on immune function in terrestrial mammals. Studies show that there is an increase in the incidences of tumors and decreased immune cell function in TBT-exposed mammals. TBT interferes with the ability of human natural killer (NK) cells to lyse target cells; it also alters the secretion of the pro-inflammatory cytokines Interferon gamma (IFNγ) and Tumor necrosis factor alpha (TNFα) from human immune cells ex vivo. Any dysregulation of the cytokine communication network may have detrimental effects on immune function, as cytokines play a key role in the regulation of immune responsiveness. There have been no in vivo studies on the effects of exposure to TBT on the cytokine communication network. The effects of 24 hour exposure to TBT on the production of IFNγ and TNFα from human immune cells were analyzed in an ex vivo system using western blotting. The serum of TBT-exposed mice was analyzed for changes in the levels of cytokine secretion and production using a Milliplex mouse cytokine/chemokine magnetic bead premixed 32 plex kit and western blotting; the effects of TBT-exposure was assessed using a time curve of 6h, 12h, 24h, 48h and control. TBT increased the levels of the cytokines IFNγ, TNFα, IL1β, IL5, IL7, IL12βp40, IL13, IL15, MIP1β, MIP2 and RANTES in the serum at a minimum of one time point. IFNγ and TNFα secretion and production from human cells also increased, showing striking agreement in the response to TBT between the human and mouse systems. Supported by: NIH grant 5U54CA163066-03 Citation Format: Shanieek T. Lawrence, Margaret Whalen, Samuel Pellom, Anil Shanker. Tributyltin alters the production and secretion of inflammatory cytokines from human and mouse immune cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2735. doi:10.1158/1538-7445.AM2015-2735

Cytokines are important regulators of immune responsiveness. Alterations of cytokine secretion from immune cells would have the potential to disrupt both immune destruction of cancer cells and to alter the proliferation and invasiveness of tumor cells due to their capacity to act as growth factors, angiogenic factors, and inducers of epithelial-mesenchymal transition. Tributyltin (TBT) is a widespread environmental contaminant having been used in a variety of industrial applications and is found in human blood at levels ranging as high as 261 nM. Studies in human immune cells have shown that secretion of tumor necrosis factor alpha (TNFα) is altered by exposures to TBT. There have been no in vivo studies of the effects of exposure to TBT on the cytokine communication network. As it is not possible to examine alterations of this network in exposed humans, we examined the effects of TBT exposure in a mouse model using Milliplex mouse cytokine/chemokine magnetic bead premixed 32 plex kit. Serum and spleens of TBT-exposed mice were analyzed for changes in cytokine secretion/levels. TBT increased the levels of IL13 in the serum of non-tumor-bearing mice and increased IL1β, IL6, KC, and M-CSF in tumor-bearing mice. The spleens of both non-tumor-bearing and tumor-bearing mice showed increases in Eotaxin when exposed to TBT. Of significant interest is the fact that the presence of tumor causes increases in Eotaxin, IL12p40, M-CSF, and TNFα and exposure to TBT alone can cause increases in these same cytokines. Tumors are known to increase levels of cytokines that enhance their ability to survive. The results showing that TBT can amplify the levels of several cytokines that are increased by tumor suggest that TBT may have tumor-promoting abilities by modulating tumor growth and/or invasiveness. Experiments are under way to dissect cytokine signaling pathways that are affected by TBT. The findings will allow identification of molecular targets that are necessary to intercept the tumor promoting signaling following TBT exposure. Supported by: NIH grant 5U54CA163066-03 Citation Format: Shanieek Lawrence, Samuel Troy Pellom, Tasia Hurd-Brown, Anil Shanker, Margaret Whalen. Elevation of tumor-promoting cytokines in mice exposed to the environmental contaminant tributyltin. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3191. doi:10.1158/1538-7445.AM2014-3191

Kidney cancer is a malignant tumor that accounts for about two percent of all cancers in the United States. African American men and women account for a higher incidence rate of kidney cancer compared to other races. The higher rate of failure of current immunotherapeutic strategies is attributed to the immunosuppressive effects as well as modification of the immune-regulatory circuits brought about by cancers. With the aim of enhancing immune mechanisms against kidney cancer, we undertook a combinatorial approach to reveal the therapeutic potential of adoptive cell transfer regimens using mouse models in conjunction with molecular targeting drugs that sensitize kidney tumor cell-death. The proteasome inhibitor, bortezomib, selectively sensitizes kidney tumor cells to apoptosis by activating caspase-8 in the extrinsic apoptosis pathway. Moreover, our studies thus far have revealed that bortezomib can also enhance natural killer (NK) cell antitumor function. We observed that bortezomib upregulates Fas and NK cell activating receptor NKG2D ligand Rae1-on renal cancer Renca cells in vivo. Moreover, preliminary observations suggest that bortezomib can be combined with adoptive NK cell transfer to provide greater therapeutic benefits with little adverse effects in vivo. These new data predict the potential of combining adoptive NK cell transfer regimens with bortezomib-induced sensitization of kidney tumor cells to induce apoptosis. Such a combinatorial approach of NK cell transfer coupled with bortezomib treatment would have dual effects by sensitizing tumor cells to cell death and improve NK cell effector functions. This could prevent tumor metastasis by eliminating tumor escape variants and significantly benefit tumor immunotherapy efforts in kidney cancer. Citation Information: Cancer Epidemiol Biomarkers Prev 2011;20(10 Suppl):B107.

Breast cancer mortality occurs primarily due to invasive metastasis and takes a disproportionately heavy toll in African American women. Clinical immunotherapy efforts prove largely ineffective, partly due to the numerous immune-regulatory circuits influenced by cancers, which alter the differentiation and function of immune cells, including T cells. The long-term objective of this study is to enhance immune mechanisms of cancer rejection to provide durable clinical benefits against breast cancer. Our recent work using mouse models has revealed the therapeutic potential of combinatorial adoptive cell transfer regimens along with immunostimulatory and tumor cell-death-sensitizing treatments. We found that the proteasome inhibitor bortezomib selectively sensitized mammary tumor cells to apoptosis by amplifying caspase-8 activation in the death-inducing signaling complex. This resulted in reduced lung metastases in mice. In addition, bortezomib could be combined with adoptive T cell transfer with no adverse effects on the antigen-specific T cell proliferative or cytolytic functions in vivo. However, no long-term survival benefits could be observed, likely due to immunosuppressive effects of tumor. More recently, we observed that Delta-like ligand (DLL1)-specific immune Notch activation could enhance antitumor T cell immunity by overcoming tumor-associated immunosuppression. These data predict the potential of combining immunostimulatory Notch activation by soluble clustered DLL1 ligand treatment and bortezomib-induced sensitization of breast tumor cells to apoptosis with T cell adoptive transfers. This could prevent tumor metastasis and significantly improve breast cancer therapy. Citation Information: Cancer Epidemiol Biomarkers Prev 2011;20(10 Suppl):B108.