Tumor-induced Immune Suppression Research Articles

Progressive multifocal leukoencephalopathy, a possible complication in a patient with bladder cancer

Progressive multifocal leukoencephalopathy (PML) is a demyelinating disease of the brain, caused by reactivation of the polyomavirus JC (JCV). PML has classically been described in individuals with profound cellular immunosuppression such as patients with AIDS, hematological malignancies, organ transplant recipients or those treated with immunosuppressive or immunomodulatory medications. The occurrence of PML in patients without identifiable immunodeficiency represents a therapeutic challenge. Tumor-induced immune suppression is a fundamental problem in cancer biology. We present a case of 71-year-old man with bladder cancer. The patient was diagnosed with PML after an epileptic seizure followed by a coma. MRI images were compatible with PML. Presence of JC virus in the cerebrospinal fluid confirmed the diagnosis. Unfortunately the patient died shortly after the diagnosis. There is no specific treatment for JCV. The reconstitution of the immune system seems the most effective approach.

Myeloid‐Derived Suppressor Cells and Therapeutic Strategies in Cancer

Development of solid cancer depends on escape from host immunosurveillance. Various types of immune cells contribute to tumor-induced immune suppression, including tumor associated macrophages, regulatory T cells, type 2 NKT cells, and myeloid-derived suppressor cells (MDSCs). Growing body of evidences shows that MDSCs play pivotal roles among these immunosuppressive cells in multiple steps of cancer progression. MDSCs are immature myeloid cells that arise from myeloid progenitor cells and comprise a heterogeneous immune cell population. MDSCs are characterized by the ability to suppress both adaptive and innate immunities mainly through direct inhibition of the cytotoxic functions of T cells and NK cells. In clinical settings, the number of circulating MDSCs is associated with clinical stages and response to treatment in several cancers. Moreover, MDSCs are reported to contribute to chemoresistant phenotype. Collectively, targeting MDSCs could potentially provide a rationale for novel treatment strategies in cancer. This review summarizes recent understandings of MDSCs in cancer and discusses promissing clinical approaches in cancer patients.

Histone Deacetylase 11 (HDAC11) As a Novel Transcriptional Regulator of C/EBP-β, in Immature Myeloid Cell to Myeloid Derived Suppressor Cell Transition

In normal myelopoiesis, immature myeloid cells (IMCs) differentiate into macrophages, dendritic cells, and neutrophils. However, in pathological conditions such as in cancer, these immature cells differentiate into myeloid derived suppressor cells (MDSCs). Transcription factor CCAAT-enhancer-binding protein beta (C/EBP-β) plays a crucial role in the accumulation of MDSCs in several pathological conditions. Of note, mice lacking C/EBP-β in the bone marrow compartment lose the ability to differentiate IMCs into pathologically active MDSCs2.In contrast, up-regulation of C/EBP-β can partly induce MDSC expansion through a mechanism involving the STAT3 pathway3.Epigenetic changes, such as histone acetylation status, have been recently shown to modulate the regulatory function of MDSCs4. However, the epigenetic mechanism(s) involved in regulation of C/EBP-β gene expression in MDSCs are poorly understood. Here we show for the first time that among all the HDACs known, HDAC11, the newest member of this family of enzymes, is recruited to the C/EBP-β gene promoter region of primary myeloid cells isolated from C57BL/6 mice. Such a finding led us to explore the functional consequences of disrupting HDAC11 in myeloid cells. First, we observed a markedly higher expression of C/EBP-β mRNA (194 fold difference) in the CD11b+/Ly6G+ granulocytic compartment and a 6.7 fold difference in the CD11b+/Ly6C+monocytic compartments of HDAC11KO mice relative to control wild type mice. Second, MDSCs from HDAC11 KO mice display a higher suppressive phenotype and mechanistically we have found that they have increased expression of the immunosuppressive STAT3/IL10 axis. Third, inoculation of EL4 tumor cells into HDAC11KO mice and controls resulted in a higher expansion capacity of splenic MDSCs in mice devoid of HDAC11. Such an effect was associated with a more rapid tumor growth in HDAC11KO mice as compared to WT mice. Similar aggressive tumor growth was also observed in HDAC11 myeloid conditional KO mice (LyZ-Cre-HDAC11KO).Taken together, we have uncovered a previously unknown role for HDAC11 as a transcriptional regulator of C/EBP-β in MDSCs. Indeed, in the absence of this epigenetic checkpoint of C/EBP-β gene expression, the fully suppressive potential of MDSCs was fully unleashed. A better understanding of this novel role of HDAC11 in myeloid biology will ultimately lead to targeted epigenetic therapies to manipulate the suppressive abilities of these immunoregulatory cells.1. Hirai H, Zhang P, Dayaram T, et al. C/EBPbeta is required for 'emergency’ granulopoiesis. Nat Immunol 2006; 7(7): 732-9.2. Marigo I, Bosio E, Solito S, et al. Tumor-induced tolerance and immune suppression depend on the C/EBPbeta transcription factor. Immunity 2010; 32(6): 790-802.3. Condamine T, Gabrilovich DI. Molecular mechanisms regulating myeloid-derived suppressor cell differentiation and function. Trends Immunol 2011; 32(1): 19-25.4. Youn JI, Kumar V, Collazo M, et al. Epigenetic silencing of retinoblastoma gene regulates pathologic differentiation of myeloid cells in cancer. Nat Immunol 2013; 14(3): 211-20. DisclosuresNo relevant conflicts of interest to declare.

Combinations of immunotherapy and radiation in cancer therapy.

The immune system has the ability to recognize and specifically reject tumors, and tumors only become clinically apparent once they have evaded immune destruction by creating an immunosuppressive tumor microenvironment. Radiotherapy (RT) can cause immunogenic tumor cell death resulting in cross-priming of tumor-specific T-cells, acting as an in situ tumor vaccine; however, RT alone rarely induces effective anti-tumor immunity resulting in systemic tumor rejection. Immunotherapy can complement RT to help overcome tumor-induced immune suppression, as demonstrated in pre-clinical tumor models. Here, we provide the rationale for combinations of different immunotherapies and RT, and review the pre-clinical and emerging clinical evidence for these combinations in the treatment of cancer.

Abstract PL02-03: Immune modulation for cancer therapy: Assessing antagonists and agonists

Abstract PL02-03: Immune modulation for cancer therapy: Assessing antagonists and agonists

Abstract 5028: Metronomic cyclophosphamide enhances the immunogenicity and anti-tumor activity of a DepoVax based vaccine and may be further enhanced with inhibitors of CTLA-4 or PD-1

Abstract To counteract tumor-induced immune suppression, cancer vaccines are increasingly being combined with immune modulators that can not only reverse immune suppression but also enhance vaccine induced immune responses. We found that metronomic cyclophosphamide (50 mg BID) enhanced the immunogenicity of a DepoVaxTM (DPX) based cancer vaccine (DPX-Survivac) in ovarian cancer patients in a phase I clinical study. We emulated these results using transplantable tumor models which allowed us to study the underlying mechanisms of cyclophosphamide induced immune modulation. In several different models, mice were given mCPA on alternating weeks (20 mg/kg/day PO) in combination with a DPX vaccine containing relevant peptide antigens every three weeks. In these models, only the combination provided effective and significant long-term control of tumor growth. Notably, we found that the efficacy of the combination was comparable when vaccine was given at the beginning or end of a cycle of mCPA. Metronomic CPA had a pronounced lymphodepletive effect on the vaccine draining lymph node, yet did not reduce the development of antigen-specific CD8+ T cells induced by vaccination as detected by MHC-multimer flow cytometry. Combination treatment also increased cytotoxic T cell activity in the spleen measured by IFN-γ ELISPOT and in vivo cytotoxic T cell assay. Analysis of immune gene signatures in the tumor microenvironment by RT-qPCR detected elevated levels of cytotoxic markers, such as IFN-γ and granzyme B, as well as co-inhibitory markers, such as PD-1 and CTLA-4, in mice treated with combination therapy, the latter providing a strong rational for modulating these pathways with recently available monoclonal antibodies. Analysis of spleen cell populations by flow cytometry indicated that mCPA induced transient lymphodepletion that was marked by a selective expansion of myeloid-derived suppressor cells in the absence of vaccination. Selective depletion of regulatory T cells was not observed, in contrast to other regimens of low dose CPA. These results demonstrate that mCPA provides a complex form of immune modulation that is most effective when combined with active immunization. Using these models we can evaluate other immune modulator agents that may enhance vaccine activity, including checkpoint inhibitors or other immune based therapies. Citation Format: Genevieve Weir, Olga Hrytsenko, Marianne M. Stanford, Neil L. Berinstein, Mohan Karkada, Robert S. Liwski, Marc Mansour. Metronomic cyclophosphamide enhances the immunogenicity and anti-tumor activity of a DepoVax based vaccine and may be further enhanced with inhibitors of CTLA-4 or PD-1. [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 5028. doi:10.1158/1538-7445.AM2014-5028

Metronomic cyclophosphamide enhances HPV16E7 peptide vaccine induced antigen-specific and cytotoxic T-cell mediated antitumor immune response

In clinical trials, metronomic cyclophosphamide (CPA) is increasingly being combined with vaccines to reduce tumor-induced immune suppression. Previous strategies to modulate the immune system during vaccination have involved continuous administration of low dose chemotherapy, studies that have posed unique considerations for clinical trial design. Here, we evaluated metronomic CPA in combination with a peptide vaccine targeting HPV16E7 in an HPV16-induced tumor model, focusing on the cytotoxic T-cell response and timing of low dose metronomic CPA (mCPA) treatment relative to vaccination. Mice bearing C3 tumors were given metronomic CPA on alternating weeks in combination with immunization with a DepoVax vaccine containing HPV16E749–57 peptide antigen every 3 weeks. Only the combination therapy provided significant long-term control of tumor growth. The efficacy of the vaccine was uncompromised if given at the beginning or end of a cycle of metronomic CPA. Metronomic CPA had a pronounced lymphodepletive effect on the vaccine draining lymph node, yet did not reduce the development of antigen-specific CD8+ T cells induced by vaccination. This enrichment correlated with increased cytotoxic activity in the spleen and increased expression of cytotoxic gene signatures in the tumor. Immunity could be passively transferred through CD8+ T cells isolated from tumor-bearing mice treated with the combinatorial treatment regimen. A comprehensive survey of splenocytes indicated that metronomic CPA, in the absence of vaccination, induced transient lymphodepletion marked by a selective expansion of myeloid-derived suppressor cells. These results provide important insights into the multiple mechanisms of metronomic CPA induced immune modulation in the context of a peptide cancer vaccine that may be translated into more effective clinical trial designs.

PD-1 pathway inhibitors: changing the landscape of cancer immunotherapy.

Immunotherapeutic approaches to treating cancer have been evaluated during the last few decades with limited success. An understanding of the checkpoint signaling pathway involving the programmed death 1 (PD-1) receptor and its ligands (PD-L1/2) has clarified the role of these approaches in tumor-induced immune suppression and has been a critical advancement in immunotherapeutic drug development. A comprehensive literature review was performed to identify the available data on checkpoint inhibitors, with a focus on anti-PD-1 and anti-PD-L1 agents being tested in oncology. The search included Medline, PubMed, the ClinicalTrials.gov registry, and abstracts from the American Society of Clinical Oncology meetings through April 2014. The effectiveness and safety of the available anti-PD-1 and anti-PD-L1 drugs are reviewed. Tumors that express PD-L1 can often be aggressive and carry a poor prognosis. The anti-PD-1 and anti-PD-L1 agents have a good safety profile and have resulted in durable responses in a variety of cancers, including melanoma, kidney cancer, and lung cancer, even after stopping treatment. The scope of these agents is being evaluated in various other solid tumors and hematological malignancies, alone or in combination with other therapies, including other checkpoint inhibitors and targeted therapies, as well as cytotoxic chemotherapy. The PD-1/PD-L1 pathway in cancer is implicated in tumors escaping immune destruction and is a promising therapeutic target. The development of anti-PD-1 and anti-PD-L1 agents marks a new era in the treatment of cancer with immunotherapies. Early clinical experience has shown encouraging activity of these agents in a variety of tumors, and further results are eagerly awaited from completed and ongoing studies.

Human hepatocellular carcinoma-infiltrating CD4+CD69+Foxp3− regulatory T cell suppresses T cell response via membrane-bound TGF-β1

Tumors can recruit, induce, and expand regulatory T cells (Tregs) to suppress antitumor immune responses for survival and progression. The complicated tumor-related Treg subsets and their functional mechanisms are not fully addressed yet. We have previously identified a novel CD4⁺CD69⁺Foxp3⁻ Treg subset in tumor-bearing mice, which suppresses CD4 T cell response via membrane-bound transforming growth factor beta 1 (mTGF-β1) and then promotes tumor progression. In hepatocellular carcinoma patients, here, we identified tumor-infiltrating human CD4⁺CD69⁺ Tregs which represent ~67.2 % of tumor-infiltrating CD4 T cells that is significantly higher than conventional CD4⁺CD25⁺Foxp3⁺ Tregs. They expressed mTGF-β1, PD-1, and CTLA-4, but not CD25 or Foxp3, and only produced a little interleukin (IL)-10 and TGF-β1. More importantly, they significantly suppressed CD4 T cell response via mTGF-β1 in vitro. Furthermore, the percentage of these CD4⁺CD69⁺ Tregs in tumor tissue was significantly correlated with tumor progression, which is more pronounced at the late stage of cancer patients. Thus, we have identified a tumor-induced new population of human CD4⁺CD69⁺ Tregs in cancer patients with phenotype of CD25⁻Foxp3⁻mTGF-β1⁺CTLA-4⁺PD-1⁺, and these Tregs can suppress antitumor immune response via mTGF-β1. Our results not only enrich the family of Treg subsets, providing new mechanistic insight to tumor-induced immune suppression in human, but also suggest a potential target for cancer immunotherapy. CD4⁺CD69⁺Foxp3⁻ regulatory T cells were identified in hepatocellular carcinoma patients. These Treg cells inhibit T cell response via membrane-bound TGF-β. The percentage of these cells was significantly correlated with tumor progression. The percentage of these cells was higher than conventional CD4⁺CD25⁺Foxp3⁺ Tregs. These Treg cells not only exist in tumor-bearing mice, but also in cancer patients.

Victory and defeat in the induction of a therapeutic response through vaccine therapy for human and canine brain tumors: a review of the state of the art.

Anti-tumor immunotherapy using tumor lysate-based vaccines has made great advances over recent decades. Cancer vaccines aim to elicit adaptive immune responses through various pathways by providing tumor and tumor-associated antigens with an immune stimulant or adjuvant. These anti-tumor vaccines are therefore developed as personalized treatments. Utilizing tumors as a source of vaccine antigens in immunotherapy has demonstrated promising results with minimal toxicity. However, to date, researchers have failed to overcome the overpowering immune suppressive effects within the tumor microenvironment. Immune suppression occurs naturally via multiple mechanisms. These mechanisms serve an important homeostatic role restoring a normal tissue microenvironment following an inflammatory response. Due to these suppressive mechanisms and the inherent heterogeneity of tumors, it is imperative to then elicit and maintain a specific tumoricidal response if vaccine therapy or some other combination of reagents is chosen. In this review, we focus on the historical use of tumors as a source of antigens to elicit a tumoricidal response and the limitations encountered that prevent greater success in immunotherapy. We describe the advantages and disadvantages of various vaccines and their ineffectiveness due to tumor-induced immune suppression.

Peripheral blood mononuclear cells of patients with breast cancer can be reprogrammed to enhance anti-HER-2/neu reactivity and overcome myeloid-derived suppressor cells

Two major barriers in the immunotherapy of breast cancer include tumor-induced immune suppression and the establishment of long-lasting immune responses against the tumor. Recently, we demonstrated in an animal model of breast carcinoma that expanding and reprogramming tumor-sensitized lymphocytes, ex vivo, yielded T memory (Tm) cells as well as activated CD25+ NKT cells and NK cells. The presence of activated CD25+ NKT and NK cells rendered reprogrammed T cells resistant to MDSC-mediated suppression, and adoptive cellular therapy (ACT) of reprogrammed lymphocytes protected the host from tumor development and relapse. Here, we performed a pilot study to determine the clinical applicability of our protocol using peripheral blood mononuclear cells (PBMCs) of breast cancer patients, ex vivo. We show that bryostatin 1 and ionomycin combined with IL-2, IL-7, and IL-15 can expand and reprogram tumor-sensitized PBMCs. Reprogrammed lymphocytes contained activated CD25+ NKT and NK cells as well as Tm cells and displayed enhanced reactivity against HER-2/neu in the presence of MDSCs. The presence of activated NKT cells was highly correlated with the rescue of anti-HER-2/neu immune responses from MDSC suppression. Ex vivo blockade experiments suggest that the NKG2D pathway may play an important role in overcoming MDSC suppression. Our results show the feasibility of reprogramming tumor-sensitized immune cells, ex vivo, and provide rationale for ACT of breast cancer patients.

Combined lenalidomide, low‐dose dexamethasone, and rituximab achieves durable responses in rituximab‐resistant indolent and mantle cell lymphomas

Lenalidomide is an immunomodulatory drug with effects on the immune system that may enhance antibody-dependent cell-mediated cytotoxicity and reverse tumor-induced immune suppression. Furthermore, single-agent lenalidomide has therapeutic activity in relapsed/refractory B-cell lymphomas. These immunologic effects potentially may enhance the action of rituximab. To test the efficacy of lenalidomide combined with rituximab, the authors conducted a phase 2 trial of lenalidomide, low-dose dexamethasone, and rituximab in patients who had rituximab-resistant, relapsed/refractory, indolent B-cell or mantle cell lymphomas. Patients received two 28-day treatment cycles of lenalidomide 10 mg daily and dexamethasone 8 mg once weekly (part I). During cycle 3, 4 weekly doses of rituximab 375 mg/m2 were administered with lenalidomide-dexamethasone (part II). After the part II response assessment, stable or responding patients continued to receive lenalidomide-dexamethasone. Twenty-seven patients with follicular (n=18), mantle cell (n=5), small lymphocytic (n=3), and marginal zone (n=1) lymphomas started therapy; 3 of 27 patients discontinued therapy because of adverse events and were not evaluable for response. For 24 patients, the overall response rate after part I was 29% (4 patients had a complete response [CR] or CR unconfirmed, and 3 patients had a partial response), and the overall response rate after part II was 58% (8 patients had a CR, and 6 patients had a partial response). For 27 patients, at a median follow-up of 12.2 months, the median progression-free survival was 23.7 months. The combination of lenalidomide, low-dose dexamethasone, and rituximab achieved high response rates with durable responses in patients with rituximab-resistant, indolent B-cell and mantle cell lymphomas. Overall response rate increased from 29% after two 28-day cycles of lenalidomide and low-dose dexamethasone to 58% after the addition of rituximab, suggesting that lenalidomide can overcome resistance to rituximab.

Abstract A79: The adenosine pathway in ovarian carcinoma: Tumor cells and tumor-derived exosomes express CD39 and CD73 ectonucleotidases, produce adenosine and mediate immune suppression

Abstract Background: Ectonucleotidases CD39/CD73 have been reported to play an important role in functional supression of various immune cells via adenosine that is generated locally in the tumor microenvironment. In patients with ovarian cancer (OvCa) exosomes released by tumor cells (TEX) are abundant in body fluids, including the plasma or ascites and may be involved in tumor progression. Based on the observations suggesting that TEX carry proteins that are expressed on tumor cells from which TEX originate, we hypothesized that CD39+ and CD73+ TEX could deliver these enzymes to distant immune cells. Adenosine produced fom ATP in the presence of TEX could suppress functions of these cells or elevate suppressor activity of regulatory T cells (Treg) , The aim of the study was to: (1) investigate the expression and clinical significance of CD39, CD73, adenosine deaminase (ADA) and CD26 in OvCa tissues and in TEX isolated from OvCa body fluids; (2) determine whether OvCa TEX metabolize exogeneous ATP to adenosine. (3) characterize the molecular profile of TEX; and (4) test whether OvCa TEX can suppress activities of NK cells and up-regulate suppressive activity of Treg. Methods: The expression of CD39, CD73, ADA and CD26 in OvCa tissues was determined by immunohistochemistry (IHC). The relationship between expression of these enzymes and clinicopathological characteristics was analyzed. TEX were isolated from supernatant of two OvCa cell lines (A2780, SKOV-3) and from the patients' plasma by exclusion chromatpgraphy followed by ultracentrifugation, as previously described.. Western blots were used for molecular characterization of TEX. NK cells and Treg were separated from the PMBC of normal donors and co-cultured with TEX. The phenotype of NK cells and Treg was evaluated by flow cytometry. ATP hydrolysis was measured using a luciferase detection assay. Results: By IHC in tissue sections, 70% of tumor cells were CD39+, 77% were CD73+ and 100% were CD26+ADA+.Expression levels of the ectonucleotidases varied from strong to moderate, and patients with a more advanced disease stage had tumors showing strongest CD73 expression (p<0,05). Exosomes isolated from plasma of OvCa patients were enriched in TEX which carried LAMP-1, CD63, TGF-β1, MAGE3/6, CD39, CD73, ADA and Ep-CAM. In contrast, exosomes isolated from the plasma of healthy donors carried LAMP-1 and CD63. TEX obtained from OvCa patients hydrolyzed more exogeneous ATP than did TEX from OvCa cell line supernatants (p<0.05) and produced more adenosine (p<0,05). After co-incubation with TEX, normal NK cells downregulated expression of NKG2D, NKp44 and NKp46 (p<0,05) and Treg up-regulated expression of Perforin, FasL, CCR7 (p<0,05). Co-incubation of Treg with TEX resulted by increased suppression of responder cells (p<0,01). Conclusion: Similar to OvCa tumor cells in tissue exosomes isolated from the plasma of OvCa patients were found to carry enzymatically-active ectonucleotidases and to produce extracellular adenosine. These exosomes were capable of down-regulating NK cell functions and up-regulating Treg activity in vitro. The same TEX-mesiated mechanisms could contribute to tumor-induced immune suppression characteristic of OvCa and resulting in tumor immune escape and OvCa progression. Note: This abstract was not presented at the conference. Citation Format: Marta E. Szajnik-Szczepanski, Magdalena Derbis, Michal Lach, Paulina Patalas, Marcin Michalak, Hanna Drzewiecka, Marta Glura, Ewa Nowak-Markwitz, Marek Spaczynski, Theresa L. Whiteside. The adenosine pathway in ovarian carcinoma: Tumor cells and tumor-derived exosomes express CD39 and CD73 ectonucleotidases, produce adenosine and mediate immune suppression. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: From Concept to Clinic; Sep 18-21, 2013; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2013;19(19 Suppl):Abstract nr A79.

CpG oligodeoxynucleotide as immune adjuvant enhances photodynamic therapy response in murine metastatic breast cancer.

Breast cancer is the most common cause of cancer death in women. The side effects and complications following current breast cancer therapy can be devastating. Moreover, the prognosis in late stages of the diseases is usually poor. Photodynamic therapy (PDT) is a promising cancer treatment modality that is capable of both local tumor destruction and immune stimulation. However, treatment with PDT alone is often non-curative due to tumor-induced immune cell dysfunction and immune suppression. This phenomenon has motivated a new approach by combining immunostimulants with PDT to enhance anti-tumor immunity. In the present study, we investigated PDT mediated by verteporfin and 690 nm light delivered 15 min later, in combination with an immunomodulation approach using CpG oligodeoxynucleotide for the treatment of 4T1 metastatic breast cancer in a BALB/c immunocompetent mouse model. In vitro, CpG primed immature dendritic cells (DC) via toll like receptor 9 to phagocytose PDT killed tumor cells leading to DC maturation and activation. Peritumoral injection of CpG after PDT in mice gave improved local tumor control and a survival advantage compared to either treatment alone (p < 0.05). CpG may be a valuable dendritic cell targeted immunoadjuvant to combine with PDT.

Myeloid TGF-β Responsiveness Promotes Metastases

Tumor-induced immune suppression is a major impediment to many potentially effective cancer therapies. TGF-β has previously been described as having both tumor-promoting and tumor-suppressive characteristics. In this issue of Cancer Discovery, Pang and colleagues show that myeloid-specific TGF-β signaling is a critical mediator in tumor metastasis. These findings point to a more specific means to reduce cancer immunosuppression, prevent metastasis, and minimize treatment-related adverse events.

PD‐1 as a potential target in cancer therapy

Recently, an improved understanding of the molecular mechanisms governing the host response to tumors has led to the identification of checkpoint signaling pathways involved in limiting the anticancer immune response. One of the most critical checkpoint pathways responsible for mediating tumor-induced immune suppression is the programmed death-1 (PD-1) pathway, normally involved in promoting tolerance and preventing tissue damage in settings of chronic inflammation. Many human solid tumors express PD ligand 1 (PD-L1), and this is often associated with a worse prognosis. Tumor-infiltrating lymphocytes from patients with cancer typically express PD-1 and have impaired antitumor functionality. Proof-of-concept has come from several preclinical studies in which blockade of PD-1 or PD-L1 enhanced T-cell function and tumor cell lysis. Three monoclonal antibodies against PD-1, and one against PD-L1, have reported phase 1 data. All four agents have shown encouraging preliminary activity, and those that have been evaluated in larger patient populations appear to have encouraging safety profiles. Additional data are eagerly awaited. This review summarizes emerging clinical data and potential of PD-1 pathway–targeted antibodies in development. If subsequent investigations confirm the initial results, it is conceivable that agents blocking the PD-1/PD-L1 pathway will prove valuable additions to the growing armamentarium of targeted immunotherapeutic agents.Next-generation immunotherapy agents that target the PD-1 checkpoint pathway are demonstrating antitumor activity and encouraging safety profiles in early clinical trials. Current and future clinical trials will provide new insights, and the evaluation of biomarkers and rational combination therapies is ongoing.

Lenalidomide enhances the protective effect of a therapeutic vaccine and reverses immune suppression in mice bearing established lymphomas

Immunomodulatory drugs (IMiDs) are effective therapeutic agents with direct inhibitory effects on malignant B- and plasma-cells and immunomodulatory effects on the T-cell activation. This dual function of IMiDs makes them appealing candidates for combination with a cancer vaccine. We investigated the immune stimulatory effects of lenalidomide, administrated to mice in doses, which provided comparable pharmacokinetics to human patients, on the potency of a novel fusion DNA lymphoma vaccine. The combination was curative in the majority of mice with 8d pre-established syngeneic A20 lymphomas compared with vaccine or lenalidomide alone and induced immune memory. In vivo depletion experiments established the requirement for effector CD8(+) and CD4(+) T cells in protective immunity. Unexpectedly, lenalidomide alone was also associated with reduced numbers of systemic myeloid-derived suppressor cell (MDSC) and regulatory T cell (Treg) in tumor-bearing but not naïve mice, an effect that was independent of simple tumor burden reduction. These results confirm and extend results from other models describing the effect of lenalidomide on enhancing T-cell immunity, highlight the potency of this effect, and provide a rationale for clinical application. Independently, a novel mechanism of action reversing tumor-induced immune suppression by MDSC is suggested.

Immunotherapy in preneoplastic disease: targeting early procarcinogenic inflammatory changes that lead to immune suppression and tumor tolerance

Recent advances in immunotherapy have demonstrated that single agent vaccines can be effective when given as primary prevention before exposure to the causative agent, and partially effective in some patients with existing cancer. However, as tumors develop and progress, tumor-induced immune suppression and tolerance present the greatest barrier to therapeutic success. Preneoplastic disease represents an important opportunity to intervene with tumor antigen-targeted vaccines before these mechanisms of immune evasion outpace efforts by the immune system to destroy precancerous cells. However, as we discuss in this review, emerging evidence suggests that procarcinogenic inflammatory changes occur early in cancer development, in both patients and mouse models of cancer progression. Defining early inhibitory signals within tumor microenvironments will yield insights that can eventually be used in the clinic to target these events and deliver treatments that can be used in addition to cancer vaccines to prevent premalignant and early invasive cancers.

Aging subverts immune function by dictating alternative STAT responses to cytokine signaling (P5078)

Abstract Chronic inflammation in the aged closely resembles tumor-induced immune suppression, manifested as a progressive T1→T2 shift and an increased presence of MDSCs. We hypothesize that reversing these age-related phenomena should reduce the elderly’s heightened susceptibility to malignancy. We have now optimized real-time analyses of phosphorylated STATs in murine T cells and MDSCs in order to correlate STAT activation with immune suppression. Both young and old T cells responded to IL-4 stimulation with STAT6 activation and to IFN-γ stimulation with STAT1 activation. Uniquely, however, young T cells also phosphorylated STAT1, STAT3 and STAT5 in response to IL-4, and responded to IL-6 with STAT3 activation, all consistent with a greater susceptibility to T1/T17 differentiation. Regarding MDSCs, both old and young displayed constitutive STAT3 activation which could be enhanced by G-CSF or IL-6 treatment. Both young and old MDSCs activated STAT1 briskly in response to IFN-γ, STAT5 in response to GM-CSF and STAT6 in response to IL-4. However, younger MDSCs, like younger T cells, displayed more diverse responsiveness to IL-4, activating not only STAT6 but also STAT1 and STAT3. While chronic in vivo exposure to T2 cytokines may explain the decreased responsiveness of older MDSCs and T cells to IL-4, paradoxically IL-4 still robustly activated STAT6 in these cells. Such selective responsiveness may be required to preserve the immune suppressive potential of older MDSCs.

Collagen matrix deposition by hepatic stellate cells protects hepatocellular carcinoma from NK-mediated cytotoxicity (P2013)

Abstract Liver fibrosis (LF), a leading risk factor for hepatocellular carcinoma (HCC), is caused by collagen-producing hepatic stellate cells (HSC) activated during chronic inflammation secondary to hepatitis-C infection, or alcohol liver disease (ALD). NK cells promote disease resolution via RAE1-dependant HSC clearance. A defect in NK function by an ill-defined mechanism contributes to a pro-fibrotic response. Using an engineered bioartifical collagen matrix, the expression of inhibitory leukocyte associated IgG-like receptor-1 (LAIR-1) was increased on dividing NK cells and IL-2-induced NK proliferation was inhibited. Thus, we hypothesized that the accumulation of HSC-derived collagen directly inhibits NK activity and protects both HSC and HCC from innate clearance. We found that deposition of native collagen matrix for 72h protected HSCs from NK lysis in Cr51 cytotoxicity assays. A HCC cell line (HEPG2), which produced no collagen when cultured in the absence of HSCs, was susceptible to activated NK killing and was unaffected by collagenase. To test if HSC-derived collagen protects HEPG2 cells, the populations were differentially labeled with ipophilic dyes and co-cultured for 72h for use in flow-based assays. Collagen matrix protected both targets, while collagenase pretreatment restored NK sensitivity. These results suggest that HSC-derived collagen may contribute to NK dysfunction in LF and HCC, and identifies LAIR-1 as a potential mediator of tumor-induced immune suppression.