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Antitumor treatments based on the infusion of T cells expressing chimeric antigen receptors (CAR T cells) are still relatively ineffective for solid tumors, due to the presence of immunosuppressive mediators [such as prostaglandin E2 (PGE2) and adenosine] and poor T-cell trafficking. PGE2 and adenosine activate protein kinase A (PKA), which then inhibits T-cell receptor (TCR) activation. This inhibition process requires PKA to localize to the immune synapse via binding to the membrane protein ezrin. We generated CAR T cells that expressed a small peptide called the “regulatory subunit I anchoring disruptor” (RIAD) that inhibits the association of PKA with ezrin, thus blunting the negative effects of PKA on TCR activation. After exposure to PGE2 or adenosine in vitro, CAR-RIAD T cells showed increased TCR signaling, released more cytokines, and showed enhanced killing of tumor cells compared with CAR T cells. When injected into tumor-bearing mice, the antitumor efficacy of murine and human CAR-RIAD T cells was enhanced compared with that of CAR T cells, due to resistance to tumor-induced hypofunction and increased T-cell infiltration of established tumors. Subsequent in vitro assays showed that both mouse and human CAR-RIAD cells migrated more efficiently than CAR cells did in response to the chemokine CXCL10 and also had better adhesion to various matrices. Thus, the intracellular addition of the RIAD peptide to adoptively transferred CAR T cells augments their efficacy by increasing their effector function and by improving trafficking into tumor sites. This treatment strategy, therefore, shows potential clinical application for treating solid tumors. Cancer Immunol Res; 4(6); 541–51. ©2016 AACR.

Supplementary Figure Legends

Supplemental Figure 1. Schematic representations of viral-engineered constructs for transductions of T cells. Supplemental Figure 2. Successful transduction of primary T cells and detection of transgenes. Supplemental Figure 3. Phenotypic characterization of human CAR T cells at baseline. Supplemental Figure 4. Phenotypic characterization of murine CAR T cells at baseline. Supplemental Figure 5. Heightened cytokine production by CAR-RIAD T cells upon in vitro restimulation with mesothelin-coated beads. Supplemental Figure 6. Densitometry analyses of signaling proteins. Supplemental Figure 7. Expression of PGE2 in our tumor cells and the effect of mesoCAR-RIAD T cells on tumors that do not express the antigen mesothelin. Supplemental Figure 8. Greater influx of CD4+ CAR T cells in human and murine tumors.

Expression of inhibitory receptors on tumor vs. spleen infiltrating lymphocytes.

Purpose: Tumor-infiltrating lymphocytes (TILs) become hypofunctional, although the mechanisms are not clear. Our goal was to generate a model of human tumor-induced TIL hypofunction to study mechanisms and to test anti-human therapeutics.Experimental Design: We transduced human T cells with a published, optimized T-cell receptor (TCR) that is directed to a peptide within the cancer testis antigen, NY-ESO-1. After demonstrating antigen-specific in vitro activity, these cells were used to target a human lung cancer line that expressed NY-ESO-1 in the appropriate HLA context growing in immunodeficient mice. The ability of anti-PD1 antibody to augment efficacy was tested.Results: Injection of transgenic T cells had some antitumor activity, but did not eliminate the tumors. The injected T cells became profoundly hypofunctional accompanied by upregulation of PD1, Tim3, and Lag3 with coexpression of multiple inhibitory receptors in a high percentage of cells. This model allowed us to test reagents targeted specifically to human T cells. We found that injections of an anti-PD1 antibody in combination with T cells led to decreased TIL hypofunction and augmented the efficacy of the adoptively transferred T cells.Conclusions: This model offers a platform for preclinical testing of adjuvant immunotherapeutics targeted to human T cells prior to transition to the bedside. Because the model employs engineering of human T cells with a TCR clone instead of a CAR, it allows for study of the biology of tumor-reactive TILs that signal through an endogenous TCR. The lessons learned from TCR-engineered TILs can thus be applied to tumor-reactive TILs. Clin Cancer Res; 22(2); 436–47. ©2015 AACR.See related commentary by Yang, p. 275

Co-expression of inhibitory receptors on tumor infiltrating lymphocytes.

Description of cell culture conditions, isolation/activation/transduction/expansion of T cells, generation of target cell lines, flow cytometric conditions, ELISA measurements, statistical analyses, and animal use.

Comparison of inhibitory receptors on CD8 and CD4 tumor infiltrating lymphocytes.

Summary of publications describing tumor induced hypofunction of human tumor infiltrating lymphocytes.

Expression of PDL1 on tumors.

FAP-CAR T cells restrict AE17.OVA tumor growth through reduced proliferation, increased apoptosis and altered tumor stroma.

Alteration of tumor structure by FAP-CAR T cells in A549 tumors.

Supplementary materials and methods

Combination of FAP-CAR T cells and gemcitabine treatment augments anti-tumor response in 4662 tumors.

Reagents for immunohistochemistry studies.

Supplementary Figures 1-8. Supplementary Fig. 1. CD11b+Gr1+ populations were isolated by FACS from spleens of tumor-bearing p120-/- and control mice. Supplementary Fig. 2. CD38 is expressed by the CD11b+Gr1+ population in a murine model of Barrett's Esophagus. Supplementary Fig. 3. The HNM007 tumors induce CD38 expression on MDSC population similarly to the autochthonous p120-/- tumors. Supplementary Fig. 4. Immunohistochemical analysis of HNM007 tumors co-injected with CD38High or CD38Low MDSCs. Supplementary Fig. 5. CD38high CD11b+Gr1+ cells produce higher levels of iNOS and pNFκB. Supplementary Fig.6. Network analysis of differentially regulated genes in CD38high and CD38low MDSCs shows upregulation of RB1 pathway. Supplementary Fig.7. Distribution of G-MDSC, M-MDSC and mature monocytes is perturbed in tumor-bearing Cd38-/- mice. Supplementary Fig. 8. CD38 is expressed on human MDSC-like cell population that is expanded in the peripheral blood of advanced-stage cancer patients.

Analysis of immune infiltrating cells in 4662 tumors following FAP-CAR Therapy.

Alteration of tumor structure by FAP-CAR T cells in murine PDAs.

Supplementary materials and methods.

Supplementary Tables 1 and 2. Supplementary Table 1. List of the genes differentially expressed in CD11b+Gr1+ splenocytes from tumor-bearing, compared to control mice. Supplementary Table 2. List of the genes differentially expressed in CD38high, compared to CD38low, CD11b+Gr1+ splenocytes from tumor-bearing mice.

Supplementary Figure Legend 1-8.

Identification of FAP+ cancer-associated stromal cells in desmoplastic A549 tumors.

Malignant cells drive the generation of a desmoplastic and immunosuppressive tumor microenvironment. Cancer-associated stromal cells (CASC) are a heterogeneous population that provides both negative and positive signals for tumor cell growth and metastasis. Fibroblast activation protein (FAP) is a marker of a major subset of CASCs in virtually all carcinomas. Clinically, FAP expression serves as an independent negative prognostic factor for multiple types of human malignancies. Prior studies established that depletion of FAP+ cells inhibits tumor growth by augmenting antitumor immunity. However, the potential for immune-independent effects on tumor growth have not been defined. Herein, we demonstrate that FAP+ CASCs are required for maintenance of the provisional tumor stroma because depletion of these cells, by adoptive transfer of FAP-targeted chimeric antigen receptor (CAR) T cells, reduced extracellular matrix proteins and glycosaminoglycans. Adoptive transfer of FAP-CAR T cells also decreased tumor vascular density and restrained growth of desmoplastic human lung cancer xenografts and syngeneic murine pancreatic cancers in an immune-independent fashion. Adoptive transfer of FAP-CAR T cells also restrained autochthonous pancreatic cancer growth. These data distinguish the function of FAP+ CASCs from other CASC subsets and provide support for further development of FAP+ stromal cell-targeted therapies for the treatment of solid tumors. Cancer Res; 75(14); 2800–10. ©2015 AACR.

The adaptive immune response plays a critical role in FAP-CAR T cell-mediated anti-tumor activity in immunogenic AE17.OVA tumors.

Background: Visualizations are used to communicate data about important political, social, environmental, and health topics to a wide range of audiences; however, perceptions of graphs as objective conduits of factual data make them an easy means for spreading misinformation. Research questions: 1. Are people deceived by common deceptive tactics or exaggerated titles used in data visualizations about non-controversial topics? 2. Does a person's previous data visualization coursework mitigate the extent to which they are deceived by deceptive tactics used in data visualizations? 3. What parts of data visualizations (title, shape, data labels) do people use to answer questions about the information being presented in data visualizations? Literature review: Although scholarship from psychology, human-computer interaction, and computer science has examined how data visualizations are processed by readers, scholars have not adequately researched how susceptible people are to a range of deceptive tactics used in data visualizations, especially when paired with textual content. Methodology: Participants (n = 329) were randomly assigned to view one of four treatments for four different graph types (bar, line, pie, and bubble) and then asked to answer a question about each graph. Participants were asked to rank the ease with which they read each graph and comment on what they used to respond to the question about each graph. Results/Discussion: Results show that deceptive tactics caused participants to misinterpret information in the deceptive versus control visualizations across all graph types. Neither graph titles nor previous coursework impacted responses for any of the graphs. Qualitative responses illuminate people's perceptions of graph readability and what information they use to read different types of graphs. Conclusions: Recommendations are made to improve data visualization instruction, including critically examining software defaults and the ease with which people give agency over to software when preparing data visualizations. Avenues of future research are discussed.

Gene-mediated cytotoxic immunotherapy (GMCI) is an immuno-oncology approach involving local delivery of a replication-deficient adenovirus expressing herpes simplex thymidine kinase (AdV-tk) followed by anti-herpetic prodrug activation that promotes immunogenic tumor cell death, antigen-presenting cell activation, and T cell stimulation. This phase I dose-escalation pilot trial assessed bronchoscopic delivery of AdV-tk in patients with suspected lung cancer who were candidates for surgery. A single intra-tumoral AdV-tk injection in three dose cohorts (maximum 1012 viral particles) was performed during diagnostic staging, followed by a 14-day course of the prodrug valacyclovir, and subsequent surgery 1 week later. Twelve patients participated after appropriate informed consent. Vector-related adverse events were minimal. Immune biomarkers were evaluated in tumor and blood before and after GMCI. Significantly increased infiltration of CD8+ T cells was found in resected tumors. Expression of activation, inhibitory, and proliferation markers, such as human leukocyte antigen (HLA)-DR, CD38, Ki67, PD-1, CD39, and CTLA-4, were significantly increased in both the tumor and peripheral CD8+ T cells. Thus, intratumoral AdV-tk injection into non-small-cell lung cancer (NSCLC) proved safe and feasible, and it effectively induced CD8+ T cell activation. These data provide a foundation for additional clinical trials of GMCI for lung cancer patients with potential benefit if combined with other immune therapies., Graphical Abstract, Predina and colleagues describe a phase I dose-escalation trial using bronchoscopic delivery of a replication-deficient adenovirus expressing herpes simplex thymidine kinase (AdV-tk) in lung cancer patients who underwent surgery. AdV-tk injection proved safe, feasible, and effectively induced CD8+ T cell activation in blood and in resected tumor tissues.

Hardware breakage during orthopaedic surgery especially closed intramedullary nailing is a nightmare for orthopaedic surgeons. During hip fracture surgery a mechanical failure of the guidewire or the reamer poses an additional risk of intrapelvic migration and neurovascular or visceral injury which can lead to devastating complications and litigation. We report a case of removal of the broken guidewire using a cannulated reamer & discectomy forceps and recommend some suggestions for prevention of this catastrophe.

Immune checkpoint inhibitors demonstrate clinical activity in many tumor types, however, only a fraction of patients benefit. Combining CD137 agonists with these inhibitors increases anti-tumor activity preclinically, but attempts to translate these observations to the clinic have been hampered by systemic toxicity. Here we describe a human CD137xPD-L1 bispecific antibody, MCLA-145, identified through functional screening of agonist- and immune checkpoint inhibitor arm combinations. MCLA-145 potently activates T cells at sub-nanomolar concentrations, even under suppressive conditions, and enhances T cell priming, differentiation and memory recall responses. In vivo, MCLA-145 anti-tumor activity is superior to immune checkpoint inhibitor comparators and linked to recruitment and intra-tumor expansion of CD8 + T cells. No graft-versus-host-disease is observed in contrast to other antibodies inhibiting the PD-1 and PD-L1 pathway. Non-human primates treated with 100 mg/kg/week of MCLA-145 show no adverse effects. The conditional activation of CD137 signaling by MCLA-145, triggered by neighboring cells expressing >5000 copies of PD-L1, may provide both safety and potency advantages., The anti-tumour effect of immune checkpoint inhibitors is potentiated by CD137 agonists in preclinical models, but translation of these results to the clinical practice is hampered by toxicity. Authors describe here a human CD137xPD-L1 bispecific antibody with improved anti-cancer activity whilst maintaining low toxicity in non-human primates.

Cancer progression is marked by dysfunctional tumor-infiltrating lymphocytes (TIL) with high inhibitory receptor (IR) expression. Because IR blockade has led to clinical responses in some patients with non–small cell lung cancer (NSCLC), we investigated how IRs influenced CD8+ TIL function from freshly digested early-stage NSCLC tissues using a killing assay and intracellular cytokine staining after in vitro T-cell restimulation. Early-stage lung cancer TIL function was heterogeneous with only about one third of patients showing decrements in cytokine production and lytic function. TIL hypofunction did not correlate with clinical factors, coexisting immune cells (macrophages, neutrophils, or CD4+ T regulatory cells), nor with PD-1, TIGIT, TIM-3, CD39, or CTLA-4 expression. Instead, we found that the presence of the integrin αeβ7 (CD103), characteristic of tissue-resident memory cells (TRM), was positively associated with cytokine production, whereas expression of the transcription factor Eomesodermin (Eomes) was negatively associated with TIL function. These data suggest that the functionality of CD8+ TILs from early-stage NSCLCs may be influenced by competition between an antitumor CD103+ TRM program and an exhaustion program marked by Eomes expression. Understanding the mechanisms of T-cell function in the progression of lung cancer may have clinical implications for immunotherapy.

PD1 blockade to reinvigorate T cells has become part of standard of care for patients with NSCLC across disease stages. However, the majority of patients still do not respond. One potential mechanism of resistance is increased expression of other checkpoint inhibitory molecules on T cells leading to their suppression; however, this phenomenon has not been well studied in tumor-reactive, human T cells. The purpose of this study was to evaluate this compensatory mechanism in a novel model using human effector T cells infiltrating and reactive against human lung cancer. Immunodeficient mice with flank tumors established from a human lung cancer cell line expressing the NYESO1 antigen were treated with activated human T cells expressing a TCR reactive to NYESO1 (Ly95) with or without anti-PD1 alone and with combinations of anti-PD1 plus anti-TIM3 or anti-TIGIT. A month later, the effect on tumor growth and the phenotype and ex vivo function of the TILs were analyzed. Anti-PD1 and Ly95 T cells led to greater tumor control than Ly95 T cells alone; however, tumors continued to grow. The ex-vivo function of PD1-blocked Ly95 TILs was suppressed and was associated with increased T cell expression of TIM3/TIGIT. Administering combinatorial blockade of PD1+ TIM3 or PD1+ TIGIT with Ly95 T cells led to greater tumor control than blocking PD1 alone. In our model, PD1 blockade was suboptimally therapeutic alone. The effect of TIM3 and TIGIT was upregulated on T cells in response to PD1 blockade and anti-tumor activity could be enhanced when these inhibitory receptors were also blocked with antibodies in combination with anti-PD1 therapy.

Visualizations effectively communicate data about important political, social, environmental, and health topics to a wide range of audiences; however, longstanding trust of graphs as conveyors of factual data makes them an easy means for spreading misinformation. Scholars in technical and professional communication have not yet conducted needed empirical research into people's perception and comprehension of data visualizations, especially when part of larger information texts [1]. Our study investigated the extent to which people exaggerated the differences between data points when reading graphs about non-controversial topics that used deceptive techniques and/or exaggerated titles. Participants (n=329) were randomly assigned to view one of four treatments for four different graph types (bar, line, pie, and bubble) and then asked to answer a question about each graph. Results show that deceptive techniques used in the graphs (including truncated axes, 3-D exaggeration, and arbitrary sizing), caused participants to misinterpret information in the deceptive vs. control visualizations for all of the graphs regardless of graph type, previous visualization coursework or comfort level with reading graphs. Results also showed that the presence of exaggerated vs. control titles that accompanied each graph did not significantly influence the extent of the misinterpretation. We will discuss the implication of these findings for technical communication as well as avenues of future research already underway that investigate these topics further.

Histone/protein deacetylases (HDACs) are frequently upregulated in human malignancies and have therefore become therapeutic targets in cancer therapy. However, inhibiting certain HDAC isoforms can have protolerogenic effects on the immune system, which could make it easier for tumor cells to evade the host immune system. Therefore, a better understanding of how each HDAC isoform affects immune biology is needed to develop targeted cancer therapy. Here, we studied the immune phenotype of HDAC5(-/-) mice on a C57BL/6 background. While HDAC5(-/-) mice replicate at expected Mendelian ratios and do not develop overt autoimmune disease, their T-regulatory (Treg) cells show reduced suppressive function in vitro and in vivo. Likewise, CD4(+) T-cells lacking HDAC5 convert poorly to Tregs under appropriately polarizing conditions. To test if this attenuated Treg formation and suppressive function translated into improved anticancer immunity, we inoculated HDAC5(-/-) mice and littermate controls with a lung adenocarcinoma cell line. Cumulatively, lack of HDAC5 did not lead to better anticancer immunity. We found that CD8(+) T cells missing HDAC5 had a reduced ability to produce the cytokine, IFN-γ, in vitro and in vivo, which may offset the benefit of weakened Treg function and formation. Taken together, targeting HDAC5 weakens suppressive function and de-novo induction of Tregs, but also reduces the ability of CD8(+) T cells to produce IFN-γ.

Purpose: Tumor-infiltrating lymphocytes (TILs) become hypofunctional, although the mechanisms are not clear. Our goal was to generate a model of human tumor-induced TIL hypofunction to study mechanisms and to test anti-human therapeutics. Experimental Design: We transduced human T cells with a published, optimized T-cell receptor (TCR) that is directed to a peptide within the cancer testis antigen, NY-ESO-1. After demonstrating antigen-specific in vitro activity, these cells were used to target a human lung cancer line that expressed NY-ESO-1 in the appropriate HLA context growing in immunodeficient mice. The ability of anti-PD1 antibody to augment efficacy was tested. Results: Injection of transgenic T cells had some antitumor activity, but did not eliminate the tumors. The injected T cells became profoundly hypofunctional accompanied by upregulation of PD1, Tim3, and Lag3 with coexpression of multiple inhibitory receptors in a high percentage of cells. This model allowed us to test reagents targeted specifically to human T cells. We found that injections of an anti-PD1 antibody in combination with T cells led to decreased TIL hypofunction and augmented the efficacy of the adoptively transferred T cells. Conclusions: This model offers a platform for preclinical testing of adjuvant immunotherapeutics targeted to human T cells prior to transition to the bedside. Because the model employs engineering of human T cells with a TCR clone instead of a CAR, it allows for study of the biology of tumor-reactive TILs that signal through an endogenous TCR. The lessons learned from TCR-engineered TILs can thus be applied to tumor-reactive TILs. Clin Cancer Res; 22(2); 436–47. ©2015 AACR. See related commentary by Yang, p. 275

9050 Background: GMCI is a tumor-specific immuno-oncology approach implemented through local delivery of aglatimagene besadenovec(AdV-tk) followed by anti-herpetic prodrug. This leads to immunogenic tumor cell death, antigen presenting cell activation, and T cell stimulation resulting in CD8+ T cell dependent protection, as demonstrated in preclinical models and clinical trials in other tumor types. This is the first study to assess endobronchial delivery of AdV-tk for NSCLC. Methods: This Phase I dose escalation trial enrolled patients with suspected NSCLC who were candidates for surgery. A single AdV-tk injection was performed by endobronchial ultrasound (n = 11) or mediastinoscopy (n = 1) during the diagnostic staging procedure 3 weeks prior to surgery. Three dose levels were evaluated: 2.5x 1011, 5x1011, and 1x1012 vector particles (vp) in a 3+3 design. Valacyclovir was administered for 14 days, starting the day after AdV-tk injection. To assess the local and systemic effects of GMCI, immune biomarkers were evaluated in blood and tumor samples before and after GMCI. Results: From 2017-2019, 12 patients (9 men, 3 women, median age 65 [range 55-80]) received GMCI followed by surgery. Average tumor size was 5.1 cm (largest diameter) and final pathologic stage was I (n = 4), II (n = 3), and III (n = 5). Treatment-related adverse events were CTC grade 1 fever (n = 1), flu-like symptoms (n = 1) and nausea/vomiting/diarrhea (n = 1). The only > grade 2 lab abnormality was transient grade 3 lymphopenia (n = 2). A measurable reduction in tumor size was observed in one patient. The average amount of tumor necrosis was 29.4%. Significant infiltration of CD8+T cells (5.2-fold compared to baseline, p = 0.001) was found in tumor 19-22 days after AdV-tk injection. Within the CD8+tumor infiltrating lymphocytes, there was increased expression of CD38 (2.5-fold, p = 0.002), Ki67 (4.8-fold, p = 0.02), PD1 (1.9-fold, p = 0.002), CD39 (2.9-fold, p = 0.04) and CTLA-4 (4.8-fold, p < 0.001), without significant detected differences in Tim3 or TIGIT. Simultaneously, peripheral blood CD8+ cells displayed significant increases in CD38 (3.4-fold, p = 0.006), HLA-DR (4.2-fold, p = 0.002), and Ki67 (5.8-fold, p = 0.017). Conclusions: Intratumoral injection of AdV-tk into lung tumors was safe and feasible. Further, AdV-tk effectively induced peripheral blood and intra-tumoral CD8 T cell activation. Consequent upregulation of inhibitory receptors suggests a potential benefit for combination therapies. Clinical trial information: NCT03131037.

In this paper we present the results of an empirical study that analyzed how people understand the data presented to them in deceptive data visualizations when those visualizations are paired with non-deceptive text. This study was administered as an online user survey and was designed to test the extent to which deceptive data visualizations can fool users, even when they are accompanied by a paragraph of accurate text. The study consisted of a basic demographic questionnaire, chart familiarity assessment, and data visualization survey. A total of 256 participants completed the survey and were evenly distributed between a control (non-deceptive) survey and a test (deceptive) survey in which participants were asked to observe a paragraph of text and a data visualization. Participants then answered a question relevant to the observed information to measure how they perceived the information. The results of the study confirmed that deceptive techniques in data visualizations caused participants to misinterpret the information in the deceptive data visualizations even when they were accompanied by accurate explanatory text. Furthermore, certain demographics and comfort levels with chart types were more susceptible to certain types of deceptive techniques. These results highlight the importance of education and awareness in the area of data visualizations to ensure deceptive practices are not utilized on the part of developers and to avoid misinformation on the part of users.

Exhausted CD8 T (Tex) cells are immunotherapy targets in chronic infection and cancer, but a comprehensive assessment of Tex cell diversity in human disease is lacking. Here, we developed a transcriptomic- and epigenetic-guided mass cytometry approach to define core exhaustion-specific genes and disease-induced changes in Tex cells in HIV and human cancer. Single-cell proteomic profiling identified 9 distinct Tex cell clusters using phenotypic, functional, transcription factor, and inhibitory receptor co-expression patterns. An exhaustion severity metric was developed and integrated with high-dimensional phenotypes to define Tex cell clusters that were present in healthy subjects, common across chronic infection and cancer or enriched in either disease, linked to disease severity, and changed with HIV therapy. Combinatorial patterns of immunotherapy targets on different Tex cell clusters were also defined. This approach and associated datasets present a resource for investigating human Tex cell biology, with implications for immune monitoring and immunomodulation in chronic infections, autoimmunity, and cancer. Exhausted T (Tex) cells have poor function in chronic infections and cancer but can be therapeutically re-invigorated. Bengsch et al. use genes modified epigenetically during exhaustion and high-dimensional CyTOF profiling to define Tex cell heterogeneity in humans with HIV or lung cancer and link Tex cell features to disease progression and response to immunotherapy.

Data from mouse tumor models suggest that tumor-associated monocyte/macrophage lineage cells (MMLCs) dampen antitumor immune responses. However, given the fundamental differences between mice and humans in tumor evolution, genetic heterogeneity, and immunity, the function of MMLCs might be different in human tumors, especially during early stages of disease. Here, we studied MMLCs in early-stage human lung tumors and found that they consist of a mixture of classical tissue monocytes and tumor-associated macrophages (TAMs). The TAMs coexpressed M1/M2 markers, as well as T cell coinhibitory and costimulatory receptors. Functionally, TAMs did not primarily suppress tumor-specific effector T cell responses, whereas tumor monocytes tended to be more T cell inhibitory. TAMs expressing relevant MHC class I/tumor peptide complexes were able to activate cognate effector T cells. Mechanistically, programmed death-ligand 1 (PD-L1) expressed on bystander TAMs, as opposed to PD-L1 expressed on tumor cells, did not inhibit interactions between tumor-specific T cells and tumor targets. TAM-derived PD-L1 exerted a regulatory role only during the interaction of TAMs presenting relevant peptides with cognate effector T cells and thus may limit excessive activation of T cells and protect TAMs from killing by these T cells. These results suggest that the function of TAMs as primarily immunosuppressive cells might not fully apply to early-stage human lung cancer and might explain why some patients with strong PD-L1 positivity fail to respond to PD-L1 therapy.

Myeloid-derived suppressor cells (MDSC) are an immunosuppressive population of immature myeloid cells found in advanced-stage cancer patients and mouse tumor models. Production of inducible nitric oxide synthase (iNOS) and arginase, as well as other suppressive mechanisms, allows MDSCs to suppress T-cell–mediated tumor clearance and foster tumor progression. Using an unbiased global gene expression approach in conditional p120-catenin knockout mice (L2-cre;p120ctnf/f), a model of oral–esophageal cancer, we have identified CD38 as playing a vital role in MDSC biology, previously unknown. CD38 belongs to the ADP-ribosyl cyclase family and possesses both ectoenzyme and receptor functions. It has been described to function in lymphoid and early myeloid cell differentiation, cell activation, and neutrophil chemotaxis. We find that CD38 expression in MDSCs is evident in other mouse tumor models of esophageal carcinogenesis, and CD38high MDSCs are more immature than MDSCs lacking CD38 expression, suggesting a potential role for CD38 in the maturation halt found in MDSC populations. CD38high MDSCs also possess a greater capacity to suppress activated T cells, and promote tumor growth to a greater degree than CD38low MDSCs, likely as a result of increased iNOS production. In addition, we have identified novel tumor–derived factors, specifically IL6, IGFBP3, and CXCL16, which induce CD38 expression by MDSCs ex vivo. Finally, we have detected an expansion of CD38+ MDSCs in peripheral blood of advanced-stage cancer patients and validated targeting CD38 in vivo as a novel approach to cancer therapy. Cancer Res; 75(19); 4074–85. ©2015 AACR.

The role of psychologists and other mental health professionals in long-term care settings is undefined in Australia. Graduate psychology students receive little training in clinical geropsychology, and residential aged care providers do not routinely employ psychologists within such settings. Further, despite high rates of depression, neurocognitive problems, and other mental health problems, residents are rarely referred for evidence-based psychological treatment. This article presents four case studies showing how psychology services may be employed in such settings within the context of a postgraduate psychology placement programme. These case studies emphasise the importance of engagement, the use of flexible and individualised treatment approaches, and the involvement of family and professional carers in the provision of psychological services. Psychology services in residential settings can have a positive impact on the care of older adults and their families.

Naive CD4+ T cells require signals from the TCR and CD28 to produce IL-2, expand, and differentiate. However, these same signals are not sufficient to induce autocrine IL-2 production by naive CD8+ T cells, which require cytokines provided by other cell types to drive their differentiation. The basis for failed autocrine IL-2 production by activated CD8+ cells is unclear. We find that Ikaros, a transcriptional repressor that silences IL-2 in anergic CD4+ T cells, also restricts autocrine IL-2 production by CD8+ T cells. We find that CD8+ T cell activation in vitro in the absence of exogenous cytokines and CD4 help leads to marked induction of Ikaros, a known repressor of the Il2 gene. Naive murine CD8 T cells haplo-insufficient for Ikzf1 failed to upregulate Ikaros, produced autocrine IL-2, and differentiated in an IL-2–dependent manner into IFN-γ–producing CTLs in response to TCR/CD28 stimulation alone. Furthermore, Ikzf1 haplo-insufficient CD8+ T cells were more effective at controlling Listeria infection and B16 melanoma growth in vivo, and they could provide help to neighboring, non-IL-2–producing cells to differentiate into IFN-γ–producing effectors. Therefore, by repressing autocrine IL-2 production, Ikaros ensures that naive CD8+ T cells remain dependent on licensing by APCs and CD4+ T cells, and it may therefore act as a cell-intrinsic safeguard against inappropriate CTL differentiation and immunopathology.

Given the growing interest and promising preliminary results of immunotherapy in malignant pleural mesothelioma (MPM), it has become important to more fully understand the immune landscape in this tumor. This may be especially relevant in deciding who might benefit most from checkpoint blockade or agonist antibody therapy. Since the phenotype of tumor infiltrating lymphocytes (TILs) in MPM has not been fully described and their function has not been carefully assessed, we collected fresh tumor and blood from 22 patients undergoing surgical resection and analysed single cell suspensions by flow cytometry. The functionality of TILs was assessed by measurement of cytokine expression (IFN-γ) following overnight stimulation ex vivo. Results showed low numbers of CD8+ TILs whose function was either moderately or severely suppressed. The degree of TIL hypofunction did not correlate with the presence of co-existing macrophages or neutrophils, nor with expression of the inhibitory receptors PD-1, CD39 and CTLA-4. Hypofunction was associated with higher numbers of CD4 regulatory T cells (Tregs) and with expression of the inhibitory receptor TIGIT. On the other hand, presence of tissue-resident memory (Trm) cells and expression of TIM-3 on CD8+ cells were positively associated with cytokine production. However, Trm function was partially suppressed when the transcription factor Eomesodermin (Eomes) was co-expressed. Understanding the function of TILs in malignant mesothelioma may have clinical implications for immunotherapy, especially in choosing the best immunotherapy targets. Our data suggests that Treg cell blocking agents or TIGIT inhibitor antibodies might be especially valuable in these patients.

CD137 (4-1BB) is a transmembrane costimulatory receptor on T and NK cells that enhances adaptive immune responses and is a critical mediator of antitumor immunity. The development of CD137 targeted agents for cancer therapy has been hampered by on-target off-tumor toxicity in the case of agonist monospecific, bivalent mAbs or limited antitumor activity in the case of crosslinking mAbs. Here we have developed an Fc-silenced bispecific IgG1 antibody to CD137 and PD-L1 with monovalent binding specificity to each target. MCLA-145 drives transactivation of CD137 in the vicinity of cells expressing PD-L1, such as in the immunosuppressive tumor microenvironment. The degree of CD137 agonistic activity in T cells correlated with the expression level of PD-L1 on neighboring cells, as demonstrated in transactivation assays whereby reporter T cells were co-cultured with cells expressing different levels of PD-L1. PD-L1 expression as low as 6000 receptors per cell was sufficient to activate CD137 in neighboring T cells. In contrast, MCLA-145 blocked PD-1 signaling without requirement for CD137 binding in a PD-1/PD-L1 reporter assay. CD137 signaling was induced by MCLA-145 in multiple primary human immune cell assays including the mixed lymphocyte reaction, human PBMC, and whole blood SEB stimulation assays. MCLA-145 reversed T cell suppression mediated by M2 macrophages or Tregs, in vitro. In addition, MCLA-145 enhanced Ag-specific expansion and differentiation of human naïve CD8+ T cells in vitro. In vivo, MCLA-145 treatment resulted in significant tumor immune activation and antitumor responses in two separate humanized mouse tumor models. In one model, human T cells expressing NY-ESO specific TCR were adoptively transferred to mice bearing A549 tumors which expressed NY-ESO antigen and human PD-L1. MCLA-145 treatment at 5 mg/kg resulted in 54% tumor growth inhibition (TGI) as compared to T cell only treated mice. In the tumors of MCLA-145 treated mice, the percentage of NY-ESO specific CD8+ T cells were significantly increased compared to controls. In a second model, mice engrafted with human CD34+ cells were implanted with the breast tumor cell line MDA-MB-231. MCLA-145 at 0.5 mg/kg and 5 mg/kg induced significant tumor growth inhibition (55 and 57% respectively) as compared to vehicle control or Fc-silenced huIgG1 controls. Additionally, two out of nine animals in the 5 mg/kg MCLA-145-treated group had complete tumor regression. MCLA-145 increased the number of infiltrating CD8+ T cells, as well as the percentage of central memory CD8+ T cells. The cured animals were then re-challenged with MDA-MB-231 tumor cells, and tumors of previously cured mice were rejected as compared to no growth inhibition in treatment-naïve CD34+ NSG mice. In conclusion, these data support the clinical evaluation of MCLA-145 as a novel, PD-L1 dependent CD137 agonist immune therapy. Citation Format: Patrick Mayes, Paul Tacken, Steve Wang, Pieter-Fokko van Loo, Thomas Condamine, Hans van der Maaden, Eric Rovers, Steef Engels, Floris Fransen, Ashwini Kulkarni, Yao-bin Liu, Arpita Mondal, Leslie Hall, Soyeon Kim, Marina Martinez, Shaun O'Brien, Edmund Moon, Steven Albelda, Peggy Scherle, Gregory Hollis, Reid Huber, Mark Throsby, Cecile A. Geuijen. A bispecific Fc-silenced IgG1 antibody (MCLA-145) requires PD-L1 binding to activate CD137 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 539.