Immune-Desert Tumor Microenvironment in Thoracic SMARCA4-Deficient Undifferentiated Tumors with Limited Efficacy of Immune Checkpoint Inhibitors.

BACKGROUND: Thoracic SMARCA4-deficient undifferentiated tumors (SMARCA4-UT) are aggressive neoplasms. Data linking BAF alterations with tumor microenvironment (TME) and efficacy of immune checkpoint inhibitors (ICI) are contradictory. The TME of SMARCA4-UT and their response to ICI are unknown. MATERIALS AND METHODS: Patients diagnosed with SMARCA4-UT in our institution were included. Immunostainings for tertiary lymphoid structures (TLS), immune cell markers, and checkpoints were assessed. Validation was performed using an independent transcriptome dataset including SMARCA4-UT, non-small cell lung cancers (NSCLC) with/without SMARCA4 mutations, and unclassified thoracic sarcomas (UTS). CXCL9 and PD-L1 expressions were assessed in NSCLC and thoracic fibroblast cell lines, with/without SMARCA4 knockdown, treated with/without interferon gamma. RESULTS: Nine patients were identified. All samples but one showed no TLS, consistent with an immune desert TME phenotype. Four patients received ICI as part of their treatment, but the only one who responded, had a tumor with a TLS and immune-rich TME. Unsupervised clustering of the validation cohort using immune cell scores identified 2 clusters associated with cell ontogeny and immunity (cluster 1 enriched for NSCLC independently of SMARCA4 status (n = 9/10; P = .001); cluster 2 enriched for SMARCA4-UT (n = 11/12; P = .005) and UTS (n = 5/5; P = .0005). SMARCA4 loss-of-function experiments revealed interferon-induced upregulation of CXCL9 and PD-L1 expression in the NSCLC cell line with no effect on the thoracic fibroblast cell line. CONCLUSION: SMARCA4-UT mainly have an immune desert TME with limited efficacy to ICI. TME of SMARCA4-driven tumors varies according to the cell of origin questioning the interplay between BAF alterations, cell ontogeny and immunity.

Critical Roles of Tumor Extracellular Vesicles in the Microenvironment of Thoracic Cancers.

Extracellular vesicles (EVs), such as exosomes, are critical mediators of intercellular communication between tumor cells and other cells located in the microenvironment but also in more distant sites. Exosomes are small EVs that can carry a variety of molecules, such as lipids, proteins, and non-coding RNA, especially microRNAs (miRNAs). In thoracic cancers, including lung cancers and malignant pleural mesothelioma, EVs contribute to the immune-suppressive tumor microenvironment and to tumor growth and metastasis. In this review, we discuss the recent understanding of how exosomes behave in thoracic cancers and how and why they are promising liquid biomarkers for diagnosis, prognosis, and therapy, with a special focus on exosomal miRNAs.

Postoperative Inflammation Is an Independent Prognostic Factor in Patients With Thoracic Esophageal Squamous Cell Carcinoma.

BACKGROUND: The aim of this study was to elucidate the impact of postoperative inflammatory response on prognosis in patients with stage I thoracic esophageal squamous cell carcinoma (ESCC). MATERIALS AND METHODS: Seventy-five consecutive patients who underwent subtotal esophagectomy for clinical stage I thoracic ESCC were reviewed retrospectively. Maximum serum CRP level (CRPmax) and white blood celI count (WBCmax) were evaluated as postoperative inflammatory parameters. Prognostic factors were analyzed using Cox proportional hazards modeling. RESULTS: Optimal cut-off values were 10.7 mg/dl for CRPmax and 19,700/mm3 for WBCmax On univariate analyses, older age, worse performance status, higher WBCmax, and infectious complications were significantly associated with poorer overall survival. Multivariate analysis revealed WBCmax >19,700/mm3 to be an independent prognostic factor for poorer overall survival (hazard ratio=3.356; 95% confidence interval=1.221-9.220; p=0.019). CONCLUSION: A high WBCmax in the early postoperative phase, but not infectious complications, was an independent prognostic factor for poor overall survival in patients with clinical stage I thoracic ESCC.

Companion diagnostic tests for treatment of lung cancer patients: what are the current and future challenges?

INTRODUCTION: Companion diagnostic tests (CDXs) are considered mandatory for decision-making for treatment with targeted therapies in thoracic oncology. The emergence of immunotherapy has also given rise to the development of CDXs. Some CDXs, in particular PD-L1 immunohistochemistry tests, have been questioned and re-examined for use with new combination therapies that are being evaluated in clinical trials. Current questions include: Can we establish therapeutic indications in thoracis oncology without CDXs? Would the addition of new tests benefit patient outcome? Areas covered: This review covers the use of CDXs for decision-making in the treatment of lung cancer but also covers the limits of certain tests. It discusses the major challenges for present and future development of CDXs in daily practice. Expert opinion: CDXs can predict the efficacy of drugs if crucial steps in development and validation are fully controlled. Future development of CDXs must consider the detection of biomarkers of resistance and toxicity that are complementary to CDXs predicting therapeutic drug efficacy. Certain CDXs that have already been developed may be of interest for new indications in the field of thoracic oncology.

Transcriptome profiling of caspase-2 deficient EµMyc and Th-MYCN mouse tumors identifies distinct putative roles for caspase-2 in neuronal differentiation and immune signaling.

Caspase-2 is a highly conserved cysteine protease with roles in apoptosis and tumor suppression. Our recent findings have also demonstrated that the tumor suppression function of caspase-2 is context specific. In particular, while caspase-2 deficiency augments lymphoma development in the EµMyc mouse model, it leads to delayed neuroblastoma development in Th-MYCN mice. However, it is unclear how caspase-2 mediates these differential outcomes. Here we utilized RNA sequencing to define the transcriptomic changes caused by caspase-2 (Casp2-/-) deficiency in tumors from EµMyc and Th-MYCN mice. We describe key changes in both lymphoma and neuroblastoma-associated genes and identified differential expression of the EGF-like domain-containing gene, Megf6, in the two tumor types that may contribute to tumor outcome following loss of Casp2. We identified a panel of genes with altered expression in Th-MYCN/Casp2-/- tumors that are strongly associated with neuroblastoma outcome, with roles in melanogenesis, Wnt and Hippo pathway signaling, that also contribute to neuronal differentiation. In contrast, we found that key changes in gene expression in the EµMyc/Casp2-/- tumors, are associated with increased immune signaling and T-cell infiltration previously associated with more aggressive lymphoma progression. In addition, Rap1 signaling pathway was uniquely enriched in Casp2 deficient EµMyc tumors. Our findings suggest that Casp2 deficiency augments immune signaling pathways that may be in turn, enhance lymphomagenesis. Overall, our study has identified new genes and pathways that contribute to the caspase-2 tumor suppressor function and highlight distinct roles for caspase-2 in different tissues.

Antibody-Drug Conjugates for the Therapy of Thoracic Malignancies.

Antibody-drug conjugates (ADCs) are a novel class of therapeutic agents incorporating both target-specific monoclonal antibodies and cytotoxic small molecules via a chemical linker. They were first introduced into the clinic for the treatment of advanced hematologic malignancies. The only approved ADC for solid tumors targets erb-b2 receptor tyrosine kinase (HER2), a validated antigen in breast cancer. Many ADCs are under active investigation for various types of solid tumors. In this article, we review the literature from several perspectives including the design, pharmacology, and mechanism-based toxicities of antibody-drug conjugates. We then discuss ADCs currently in clinical development for thoracic malignancies.

Update on International Cooperative Groups Studies in Thoracic Malignancies: The Emergence of Immunotherapy.

Cancer cooperative groups have historically played a critical role in the advancement of non-small-cell lung cancer therapy. Representatives from cooperative groups worldwide convene at the International Lung Cancer Congress annually. The International Lung Cancer Congress had its 17th anniversary in the summer of 2016. The present review highlights the thoracic malignancy studies discussed by presenters. The included studies are merely a sample of the trials of thoracic malignancies ongoing globally.

Immunotherapy: a new standard of care in thoracic malignancies? A summary of the European Respiratory Society research seminar of the Thoracic Oncology Assembly.

In May 2017, the second European Respiratory Society research seminar of the Thoracic Oncology Assembly entitled "Immunotherapy, a new standard of care in thoracic malignancies?" was held in Paris, France. This seminar provided an opportunity to review the basis of antitumour immunity and to explain how immune checkpoint inhibitors (ICIs) work. The main therapeutic trials that have resulted in marketing authorisations for use of ICIs in lung cancer were reported. A particular focus was on the toxicity of these new molecules in relation to their immune-related adverse events. The need for biological selection, currently based on immunohistochemistry testing to identify the tumour expression of programmed death ligand (PD-L)1, was stressed, as well as the need to harmonise PD-L1 testing and techniques. Finally, sessions were dedicated to the combination of ICIs and radiotherapy and the place of ICIs in nonsmall cell lung cancer with oncogenic addictions. Finally, an important presentation was dedicated to the future of antitumour vaccination and of all ongoing trials in thoracic oncology.

Chimeric Antigen Receptor (CAR) T-Cell Therapy for Thoracic Malignancies.

Chimeric antigen receptor (CAR) T cells are patient T cells that are transduced with genetically engineered synthetic receptors to target a cancer cell surface antigen. The remarkable clinical response rates achieved by adoptive transfer of T cells that target CD19 in patients with leukemia and lymphoma have led to a growing number of clinical trials exploring CAR T-cell therapy for solid tumors. Herein, we review the evolution of adoptive T-cell therapy; highlight advances in CAR T-cell therapy for thoracic malignancies; and summarize the targets being investigated in clinical trials for patients with lung cancer, malignant pleural mesothelioma, and esophageal cancer. We further discuss the barriers to successfully translating CAR T-cell therapy for solid tumors and present strategies that have been investigated to overcome these hurdles.

Fine needle aspirate flow cytometric phenotyping characterizes immunosuppressive nature of the mesothelioma microenvironment.

With the emergence of checkpoint blockade and other immunotherapeutic drugs, and the growing adoption of smaller, more flexible adaptive clinical trial designs, there is an unmet need to develop diagnostics that can rapidly immunophenotype patient tumors. The ability to longitudinally profile the tumor immune infiltrate in response to immunotherapy also presents a window of opportunity to illuminate mechanisms of resistance. We have developed a fine needle aspirate biopsy (FNA) platform to perform immune profiling on thoracic malignancies. Matching peripheral blood, bulk resected tumor, and FNA were analyzed from 13 mesothelioma patients. FNA samples yielded greater numbers of viable cells when compared to core needle biopsies. Cell numbers were adequate to perform flow cytometric analyses on T cell lineage, T cell activation and inhibitory receptor expression, and myeloid immunosuppressive checkpoint markers. FNA samples were representative of the tumor as a whole as assessed by head-to-head comparison to single cell suspensions of dissociated whole tumor. Parallel analysis of matched patient blood enabled us to establish quality assurance criteria to determine the accuracy of FNA procedures to sample tumor tissue. FNA biopsies provide a diagnostic to rapidly phenotype the tumor immune microenvironment that may be of great relevance to clinical trials.

Moving Immune Checkpoint Blockade in Thoracic Tumors beyond NSCLC.

SCLC and malignant pleural mesothelioma (MPM) are historically characterized by a disappointing lack of significant therapeutic breakthroughs for novel agents, and both malignancies represent true unmet medical needs. Given the promising results of anti-cytotoxic T-lymphocyte associated protein-4 and anti-programmed cell death-1/programmed death ligand-1 antibodies in the treatment of advanced NSCLCs, these immune checkpoint inhibitors are now also under investigation in SCLC and MPM, as well as in thymic epithelial tumors (TETs). Here, we review the biological and clinical rationale for immune checkpoint inhibition in SCLC, MPM, and TETs and present preliminary clinical results with available antibodies. Immunotherapeutic perspectives for these malignancies in terms of novel agents currently under evaluation or anticipated in the near future are also discussed. Current immune checkpoint blockers targeting cytotoxic T-lymphocyte associated protein-4 and the programmed cell death-1/programmed death ligand-1 axis, administered alone or in combination and as multimodality treatment, are likely to be a valuable addition to the therapeutic array for managing SCLC and MPM; studies in TETs, which are currently in their infancy, are merited. Close attention to potential toxicities will be important to the success of such strategies in these settings.

Modulated electro-hyperthermia enhances dendritic cell therapy through an abscopal effect in mice.

The aim of this study was to assess whether modulated electro-hyperthermia (mEHT) can induce an abscopal effect and thereby enhance the antitumor effects of immunotherapy. We used an intratumoral dendritic cell (DC) injection and mEHT to treat C3H/He mice inoculated with squamous cell carcinoma SCCVII cells in the left leg, and we assessed the whole body antitumor effects. Tumors were examined every two or three days in order to assess growth inhibition. The tumor-draining lymph nodes were removed to enable flow cytometric analysis of CD3+ and CD8+ cells, whereas immunohistochemistry was used to assess CD8, S100 and Foxp3 expression in the tumors. Additionally, GP96 expression in the tumors from the different treatment groups was measured. In the control group, the mean tumor volume was larger than that in other groups. These results indicated that the combination therapy of an intratumoral DC injection and mEHT evoked systemic antitumor activity. A larger number of CD3+ and CD8+ cells were detected by flow cytometric analysis in the DC plus mEHT treatment group. Tumor tissue immunostaining showed that CD8 and S100 were more strongly expressed in the DC plus mEHT treatment group, although Foxp3 expression was much higher in the control group. The GP96 gene expression level in the mEHT group was significantly different from the expression level in the control group. An abscopal effect may be induced by mEHT, and the effect of immunotherapy with DCs was strongly enhanced by the overexpression of GP96. GP96 is thought to be one of the molecules explaining the abscopal effect. Direct intratumoral administration of DCs and mEHT may be a feasible future treatment strategy.

Carnosine may reduce lung injury caused by radiation therapy.

Ionising radiation is known one of the most effective tools in the therapy of cancer but in many thoracic cancers, the total prescribed dose of radiation that can be safely administered to the target volume is limited by the risk of complications arising in the normal lung tissue. One of the major reasons for cellular injury after radiation is the formation of reactive oxygen species (ROS). Radiation pneumonitis is an acute phase side-effect which generally subsides after a few weeks and is followed by a chronic phase characterized by inflammation and fibrosis, that can develop months or years after irradiation. Carnosine is a dipeptide composed by the amino acids beta-histidine and l-alanine. The exact biological role of carnosine is not totally understood, but several studies have demonstrated that it possesses strong and specific antioxidant properties, protects against radiation damage,and promotes wound healing. The antioxidant mechanism of carnosine is attributed to its chelating effect against metal ions, superoxide dismutase (SOD)-like activity, ROS and free radicals scavenging ability . Either its antioxidant or anti-inflammatuar properties, we propose that carnosine ameliorates irradiation-induced lung injury. Thus, supplementing cancer patients to whom applied radiation therapy with carnosine, may provide an alleviation of the symptoms due to radiation-induced lung injury. This issue warrants further studies.

Clinical and immunologic results of a randomized phase II trial of vaccination using four melanoma peptides either administered in granulocyte-macrophage colony-stimulating factor in adjuvant or pulsed on dendritic cells.

PURPOSE: To determine clinical and immunologic responses to a multipeptide melanoma vaccine regimen, a randomized phase II trial was performed. PATIENTS AND METHODS: Twenty-six patients with advanced melanoma were randomly assigned to vaccination with a mixture of four gp100 and tyrosinase peptides restricted by HLA-A1, HLA-A2, and HLA-A3, plus a tetanus helper peptide, either in an emulsion with granulocyte-macrophage colony-stimulating factor (GM-CSF) and Montanide ISA-51 adjuvant (Seppic Inc, Fairfield, NJ), or pulsed on monocyte-derived dendritic cells (DCs). Systemic low-dose interleukin-2 (Chiron, Emeryville, CA) was given to both groups. T-lymphocyte responses were assessed, by interferon gamma ELIspot assay (Chiron, Emeryville, CA), in peripheral-blood lymphocytes (PBLs) and in a lymph node draining a vaccine site (sentinel immunized node [SIN]). RESULTS: In patients vaccinated with GM-CSF in adjuvant, T-cell responses to melanoma peptides were observed in 42% of PBLs and 80% of SINs, but in patients vaccinated with DCs, they were observed in only 11% and 13%, respectively. The overall immune response was greater in the GM-CSF arm (P <.02). Vitiligo developed in two of 13 patients in the GM-CSF arm but in no patients in the DC arm. Helper T-cell responses to the tetanus peptide were detected in PBLs after vaccination and correlated with T-cell reactivity to the melanoma peptides. Objective clinical responses were observed in two patients in the GM-CSF arm and one patient in the DC arm. Stable disease was observed in two patients in the GM-CSF arm and one patient in the DC arm. CONCLUSION: The high frequency of cytotoxic T-lymphocyte responses and the occurrence of clinical tumor regressions support continued investigation of multipeptide vaccines administered with GM-CSF in adjuvant.