Indoleamine 2,3-dioxygenase 1 activation in mature cDC1 promotes tolerogenic education of inflammatory cDC2 via metabolic communication.

Conventional dendritic cells (cDCs), cDC1 and cDC2, act both to initiate immunity and maintain self-tolerance. The tryptophan metabolic enzyme indoleamine 2,3-dioxygenase 1 (IDO1) is used by cDCs in maintaining tolerance, but its role in different subsets remains unclear. At homeostasis, only mature CCR7+ cDC1 expressed IDO1 that was dependent on IRF8. Lipopolysaccharide treatment induced maturation and IDO1-dependent tolerogenic activity in isolated immature cDC1, but not isolated cDC2. However, both human and mouse cDC2 could induce IDO1 and acquire tolerogenic function when co-cultured with mature cDC1 through the action of cDC1-derived l-kynurenine. Accordingly, cDC1-specific inactivation of IDO1 in vivo exacerbated disease in experimental autoimmune encephalomyelitis. This study identifies a previously unrecognized metabolic communication in which IDO1-expressing cDC1 cells extend their immunoregulatory capacity to the cDC2 subset through their production of tryptophan metabolite l-kynurenine. This metabolic axis represents a potential therapeutic target in treating autoimmune demyelinating diseases.

Impact of metronomic trabectedin combined with low-dose cyclophosphamide on sarcoma microenvironment and correlation with clinical outcome: results from the TARMIC study.

Soft tissue sarcomas (STS) are diverse mesenchymal tumors with few therapeutic options in advanced stages. Trabectedin has global approval for treating STS patients resistant to anthracycline-based regimens. Recent pre-clinical data suggest that trabectedin's antitumor activity extends beyond tumor cells to influencing the tumor microenvironment (TME), especially affecting tumor-associated macrophages and their pro-tumoral functions. We present the phase I/II results evaluating a combination of metronomic trabectedin and low-dose cyclophosphamide on the TME in patients with advanced sarcomas. 50 patients participated: 20 in phase I and 30 in phase II. Changes in the TME were assessed in 28 patients using sequential tumor samples at baseline and day two of the cycle. Treatment notably decreased CD68 + CD163 + macrophages in biopsies from tumor lesions compared to pre-treatment samples in 9 of the 28 patients after 4 weeks. Baseline CD8 + T cell presence increased in 11 of these patients. In summary, up to 57% of patients exhibited a positive immunological response marked by reduced M2 macrophages or increased CD8 + T cells post-treatment. This positive shift in the TME correlated with improved clinical benefit and progression-free survival. This study offers the first prospective evidence of trabectedin's immunological effect in advanced STS patients, highlighting a relationship between TME modulation and patient outcomes.This study was registered with ClinicalTrial.gov, number NCT02406781.

Reshaping the tumor microenvironment of cold soft-tissue sarcomas with oncolytic viral therapy: a phase 2 trial of intratumoral JX-594 combined with avelumab and low-dose cyclophosphamide.

Most soft-tissue sarcomas (STS) exhibit an immunosuppressive tumor microenvironment (TME), leading to resistance against immune checkpoint inhibitors (ICIs) and limited therapeutic response. Preclinical data suggest that oncolytic viral therapy can remodel the TME, facilitating T cell accumulation and enhancing the immunogenicity of these tumors.We conducted the METROMAJX, a phase II clinical trial, to investigate the combination of JX-594, an oncolytic vaccinia virus engineered for selective tumor cell replication, with metronomic cyclophosphamide and the PD-L1 inhibitor avelumab in patients with advanced, 'cold' STS, characterized by an absence of tertiary lymphoid structures. The trial employed a two-stage Simon design. JX-594 was administered intratumorally at a dose of 1.109 pfu every 2 weeks for up to 4 intra-tumoral administrations. Cyclophosphamide was given orally at 50 mg twice daily in a week-on, week-off schedule, and avelumab was administered at 10 mg/kg biweekly. The primary endpoint was the 6-month non-progression rate.Fifteen patients were enrolled, with the most frequent toxicities being grade 1 fatigue and fever. Fourteen patients were assessable for efficacy analysis. At 6 months, only one patient remained progression-free, indicating that the trial did not meet the first stage endpoint of Simon's design. Analysis of sequential tissue biopsies and plasma samples revealed an increase in CD8 density and upregulation of immune-related protein biomarkers, including CXCL10.Intra-tumoral administration of JX-594 in combination with cyclophosphamide and avelumab is safe and capable of modulating the TME in cold STS. However, the limited efficacy observed warrants further research to define the therapeutic potential of oncolytic viruses, particularly in relation to specific histological subtypes of STS.

Identification of microenvironment features associated with primary resistance to anti-PD-1/PD-L1 + antiangiogenesis in gastric cancer through spatial transcriptomics and plasma proteomics.

Anti-angiogenic agents elicit considerable immune modulatory effects within the tumor microenvironment, underscoring the rationale for synergistic clinical development of VEGF and immune checkpoint inhibitors in advanced gastric cancer (AGC). Early phase studies involving Asian patients demonstrated encouraging anti-tumor efficacies. We report the results of the REGOMUNE phase II study, in which Caucasian patients were administered regorafenib, a multi-tyrosine kinase inhibitor, in combination with avelumab, a PD-L1-targeting monoclonal antibody. This therapeutic regimen resulted in deep and durable responses in 19% of patients, with the median duration of response not yet reached. Notwithstanding, a significant proportion of AGC patients exhibited no therapeutic advantage, prompting investigations into mechanisms of inherent resistance. Comprehensive biomarker profiling elucidated that non-responders predominantly exhibited an augmented presence of M2 macrophages within the tumor microenvironment and a marked overexpression of S100A10 by neoplastic cells, a protein previously implicated in macrophage chemotaxis. Additionally, peripheral biomarker assessments identified elevated levels of cytokines, including CSF-1, IL-4, IL-8, and TWEAK, correlating with adverse clinical outcomes, thereby accentuating the role of macrophage infiltration in mediating resistance. These insights furnish an invaluable foundation for elucidating, and potentially circumventing, resistance mechanisms in current AGC therapeutic paradigms, emphasizing the integral role of tumor microenvironmental dynamics and immune modulation.

AT-0174, a novel dual IDO1/TDO2 enzyme inhibitor, synergises with temozolomide to improve survival in an orthotopic mouse model of glioblastoma.

BACKGROUND: Glioblastoma is an aggressive brain cancer, usually of unknown etiology, and with a very poor prognosis. Survival from diagnosis averages only 3 months if left untreated and this only increases to 12-15 months upon treatment. Treatment options are currently limited and typically comprise radiotherapy plus a course of the DNA-alkylating chemotherapeutic temozolomide. Unfortunately, the disease invariably relapses after several months of treatment with temozolomide, due to the development of resistance to the drug. Increased local tryptophan metabolism is a feature of many solid malignant tumours through increased expression of tryptophan metabolising enzymes. Glioblastomas are notable for featuring increased expression of the tryptophan catabolizing enzymes indole-2,3-dioxygenase-1 (IDO1), and especially tryptophan-2,3-dioxygenase-2 (TDO2). Increased IDO1 and TDO2 activity is known to suppress the cytotoxic T cell response to tumour cells, and this has led to the proposal that the IDO1 and TDO2 enzymes represent promising immuno-oncology targets. In addition to immune modulation, however, recent studies have also identified the activity of these enzymes is important in the development of resistance to chemotherapeutic agents. METHODS: In the current study, the efficacy of a novel dual inhibitor of IDO1 and TDO2, AT-0174, was assessed in an orthotopic mouse model of glioblastoma. C57BL/6J mice were stereotaxically implanted with GL261(luc2) cells into the striatum and then administered either vehicle control, temozolomide (8 mg/kg IP; five 8-day cycles of treatment every 2 days), AT-0174 (120 mg/kg/day PO) or both temozolomide + AT-0174, all given from day 7 after implantation. RESULTS: Temozolomide decreased tumour growth and improved median survival but increased the infiltration of CD4+ Tregs. AT-0174 had no significant effect on tumour growth or survival when given alone, but provided clear synergy in combination with temozolomide, further decreasing tumour growth and significantly improving survival, as well as elevating CD8+ T cell expression and decreasing CD4+ Treg infiltration. CONCLUSION: AT-0174 exhibited an ideal profile for adjunct treatment of glioblastomas with the first-line chemotherapeutic drug temozolomide to prevent development of CD4+ Treg-mediated chemoresistance.

Standardized Pathology Screening of Mature Tertiary Lymphoid Structures in Cancers.

Mature tertiary lymphoid structures (mTLSs) are organized lymphoid structures containing B lymphocytes admixed to CD23+ follicular dendritic cells. Their presence has been linked to improved survival and sensitivity to immune checkpoint inhibitors in several cancers, emerging as a promising pancancer biomarker. However, the requirements for any biomarker are clear methodology, proven feasibility, and reliability. In 357 patients' samples, we studied tertiary lymphoid structures (TLSs) parameters using multiplex immunofluorescence (mIF), hematoxylin-eosin-saffron (HES) staining, double CD20/CD23 staining, and single CD23 immunohistochemistry. The cohort included carcinomas (n = 211) and sarcomas (n = 146), gathering biopsies (n = 170), and surgical specimens (n = 187). mTLSs were defined as TLSs containing either a visible germinal center on HES staining or CD23+ follicular dendritic cells. Focusing on 40 TLSs assessed using mIF, double CD20/CD23 staining was less sensitive than mIF to assess maturity in 27.5% (n = 11/40) but was rescued by single CD23 staining in 90.9% (n = 10/11). In 97 patients, several samples (n = 240) were reviewed to characterize TLS distribution. The likelihood of finding TLSs in surgical material was 6.1 higher than in biopsy and 2.0 higher in primary samples than in metastasis after adjustment with a type of sample. Interrater agreement rates over 4 examiners were 0.65 (Fleiss kappa, 95% CI [0.46, 0.90]) for the presence of TLS and 0.90 for maturity (95% CI [0.83, 0.99]). In this study, we propose a standardized method to screen mTLSs in cancer samples using HES staining and immunohistochemistry that can be applied to all specimens.

Cabozantinib in patients with advanced Ewing sarcoma or osteosarcoma (CABONE): a multicentre, single-arm, phase 2 trial.

BACKGROUND: Patients with Ewing sarcoma or osteosarcoma have a median overall survival of less than 12 months after diagnosis, and a standard treatment strategy has not yet been established. Pharmacological inhibition of MET signalling and aberrant angiogenesis has shown promising results in several preclinical models of Ewing sarcoma and osteosarcoma. We aimed to investigate the activity of cabozantinib, an inhibitor of MET and VEGFR2, in patients with advanced Ewing sarcoma and osteosarcoma. METHODS: We did a multicentre, single-arm, two-stage, phase 2 trial in patients with advanced Ewing sarcoma or osteosarcoma recruited from ten centres in the French Sarcoma Group. Key eligibility criteria were aged 12 years or older, Eastern Cooperative Oncology Group performance status of 0-1, and documented disease progression (according to Response Evaluation Criteria in Solid Tumors version 1.1) before study entry. The number of previous lines of treatment was not limited. Patients received cabozantinib (adults 60 mg, children [<16 years] 40 mg/m2) orally once daily in 28-day cycles until disease progression, unacceptable toxicity, the investigator's decision to discontinue, or participant withdrawal. The primary endpoint for Ewing sarcoma was best objective response within 6 months of treatment onset; for osteosarcoma, a dual primary endpoint of 6-month objective response and 6-month non-progression was assessed. All enrolled patients who received at least one dose of cabozantinib were included in the safety analysis, and all participants who received at least one complete or two incomplete treatment cycles were included in the efficacy population. This study was registered with ClinicalTrials.gov, number NCT02243605. FINDINGS: Between April 16, 2015, and July 12, 2018, 90 patients (45 with Ewing sarcoma 45 with osteosarcoma) were recruited to the study. Median follow-up was 31·3 months (95% CI 12·4-35·4) for patients with Ewing sarcoma and 31·1 months (24·4-31·7) for patients with osteosarcoma. 39 (87%) patients with Ewing sarcoma and 42 (93%) patients with osteosarcoma were assessable for efficacy after histological and radiological review. In patients with Ewing sarcoma, ten (26%; 95% CI 13-42) of 39 patients had an objective response (all partial responses) by 6 months; in patients with osteosarcoma, five (12%; 4-26) of 42 patients had an objective response (all partial responses) and 14 (33%; 20-50) had 6-month non-progression. The most common grade 3 or 4 adverse events were hypophosphataemia (five [11%] for Ewing sarcoma, three [7%] for osteosarcoma), aspartate aminotransferase increase (two [4%] for Ewing sarcoma, three [7%] for osteosarcoma), palmar-plantar syndrome (three [7%] for Ewing sarcoma, two [4%] for osteosarcoma), pneumothorax (one [2%] for Ewing sarcoma, four [9%] for osteosarcoma), and neutropenia (two [4%] for Ewing sarcoma, four [9%] for osteosarcoma). At least one serious adverse event was reported in 61 (68%) of 90 patients. No patients died from drug-related toxic effects. INTERPRETATION: Cabozantinib has antitumor activity in patients with advanced Ewing sarcoma and osteosarcoma and was generally well tolerated. Cabozantinib could represent a new therapeutic option in this setting, and deserves further investigation. FUNDING: Institut Bergonié; French National Cancer Institute; Association pour la Recherche contre le Cancer.

Aryl hydrocarbon receptor control of a disease tolerance defence pathway.

Disease tolerance is the ability of the host to reduce the effect of infection on host fitness. Analysis of disease tolerance pathways could provide new approaches for treating infections and other inflammatory diseases. Typically, an initial exposure to bacterial lipopolysaccharide (LPS) induces a state of refractoriness to further LPS challenge (endotoxin tolerance). We found that a first exposure of mice to LPS activated the ligand-operated transcription factor aryl hydrocarbon receptor (AhR) and the hepatic enzyme tryptophan 2,3-dioxygenase, which provided an activating ligand to the former, to downregulate early inflammatory gene expression. However, on LPS rechallenge, AhR engaged in long-term regulation of systemic inflammation only in the presence of indoleamine 2,3-dioxygenase 1 (IDO1). AhR-complex-associated Src kinase activity promoted IDO1 phosphorylation and signalling ability. The resulting endotoxin-tolerant state was found to protect mice against immunopathology in Gram-negative and Gram-positive infections, pointing to a role for AhR in contributing to host fitness.

In vitro α-synuclein neurotoxicity and spreading among neurons and astrocytes using Lewy body extracts from Parkinson disease brains.

Synucleinopathies are a group of diseases characterized by the presence of intracellular protein aggregates containing α-synuclein (α-syn). While α-syn aggregates have been shown to induce multimodal cellular dysfunctions, uptake and transport mechanisms remain unclear. Using high-content imaging on cortical neurons and astrocytes, we here define the kinetics of neuronal and astrocytic abnormalities induced by human-derived α-syn aggregates grounding the use of such system to identify and test putative therapeutic compounds. We then aimed at characterizing uptake and transport mechanisms using primary cultures of cortical neurons and astrocytes either in single well or in microfluidic chambers allowing connection between cells and cell-types. We report that astrocytes take up α-syn-aggregates far more efficiently than neurons through an endocytic event. We also highlight that active α-syn transport occurs between cells and any cell-types. Of special interest regarding the disease, we also show that uptake and spreading of α-syn from astrocytes to neurons can lead to neuronal death. Altogether, we here show that patients-derived α-synuclein aggregates, which are taken up by neurons and astrocytes, induce a differential endogenous response in the two cell types including a peculiar astrocytic toxic gain-of-function that leads to neuronal death.

Deciphering the correlation between metabolic activity through 18F-FDG-PET/CT and immune landscape in soft-tissue sarcomas: an insight from the NEOSARCOMICS study.

Metabolic elevation in soft-tissue sarcomas (STS), as documented with 18F-Fluorodeoxyglucose positron emission tomography (18F-FDG-PET/CT) has been linked with cell proliferation, higher grade, and lower survivals. However, the recent diagnostic innovations (CINSARC gene-expression signature and tertiary lymphoid structure [TLS]) and therapeutic innovations (immune checkpoint inhibitors [ICIs]) for STS patients underscore the need to re-assess the role of 18F-FDG-PET/CT. Thus, in this correspondence, our objective was to investigate the correlations between STS metabolism as assessed by nuclear imaging, and the immune landscape as estimated by transcriptomics analysis, immunohistochemistry panels, and TLS assessment. Based on a prospective cohort of 85 adult patients with high-grade STS recruited in the NEOSARCOMICS trial (NCT02789384), we identified 3 metabolic groups according to 18F-FDG-PET/CT metrics (metabolic-low [60%], -intermediate [15.3%] and high [24.7%]). We found that T-cells CD8 pathway was significantly enriched in metabolic-high STS. Conversely, several pathways involved in antitumor immune response, cell differentiation and cell cycle, were downregulated in extreme metabolic-low STS. Next, multiplex immunofluorescence showed that densities of CD8+, CD14+, CD45+, CD68+, and c-MAF cells were significantly higher in the metabolic-high group compared to the metabolic-low group. Lastly, no association was found between metabolic group and TLS status. Overall, these results suggest that (i) rapidly proliferating and metabolically active STS can instigate a more robust immune response, thereby attracting immune cells such as T cells and macrophages, and (ii) metabolic activity and TLS could independently influence immune responses.

TROP2 Is Associated with Primary Resistance to Immune Checkpoint Inhibition in Patients with Advanced Non-Small Cell Lung Cancer.

PURPOSE: Mechanisms of primary resistance to inhibitors of the programmed cell death-1 (PD-1)/programmed death-ligand 1 (PD-L1) signaling axis in non-small cell lung cancer (NSCLC) are still poorly understood. While some studies suggest the involvement of trophoblast cell surface antigen 2 (TROP2) in modulating tumor cell resistance to therapeutic drugs, its specific role in the context of PD-1/PD-L1 axis blockade is not definitively established. EXPERIMENTAL DESIGN: We performed high-throughput analysis of transcriptomic data from 891 NSCLC tumors from patients treated with either the PD-L1 inhibitor atezolizumab or chemotherapy in two large randomized clinical trials. To confirm our results at the protein level, we complemented this transcriptional approach by performing a multiplex immunofluorescence analysis of tumor tissue samples as well as a proteomic profiling of plasma. RESULTS: We observed a significant association of TROP2 overexpression with worse progression-free survival and overall survival on PD-L1 blockade, independent of other prognostic factors. Importantly, we found increased TROP2 expression to be predictive of survival in patients treated with atezolizumab but not chemotherapy. TROP2 overexpression was associated with decreased T-cell infiltration. We confirmed these results at the proteomic level both on tumor tissue and in plasma. CONCLUSIONS: Our results suggest an important contribution of TROP2 expression to the primary resistance to PD-L1 blockade in NSCLC. TROP2-biomarker-based strategy may be relevant in selecting patients with NSCLC who are more likely to benefit from a combination of immunotherapy and an anti-TROP2 agent.

Predictive value of tumor microenvironment on pathologic response to neoadjuvant chemotherapy in patients with undifferentiated pleomorphic sarcomas.

Undifferentiated pleomorphic sarcomas (UPS) represent a prevalent and aggressive subtype of soft tissue sarcomas (STS) in adults. Despite advancements in loco regional treatments, many patients with high grade STS, including UPS, develop metastatic disease. Neoadjuvant chemotherapy is a standard approach to mitigate this risk, but response variability necessitates refined patient selection strategies. This study investigated the correlation between UPS microenvironment and neoadjuvant chemotherapy response in resectable UPS. The NEOSARCOMICS study (NCT02789384) enrolled patients with resectable STS from six sarcoma centers in France. Patients received anthracycline based chemotherapy, followed by surgery. Histological response, gene expression profiling, and multiplex immunohistofluorescence were performed on baseline and post treatment tumor samples. Plasma proteomics was analyzed to identify biomarkers. Good responders to neoadjuvant chemotherapy showed enrichment in genes related to stemness and cell cycle regulation, while poor responders exhibited immune related gene enrichment. Proteomic profiling revealed immune pathway activation and downregulation of cell cycle pathways in non responders. Despite being associated with a good prognosis, high immune infiltration, particularly of CD8 + T cells and CD20 + B cells, predicts a poor response to neoadjuvant chemotherapy in UPS, suggesting the need for alternative therapeutic strategies for patients with inflamed UPS.Ongoing clinical trials are exploring the efficacy of combining chemotherapy with immune checkpoint inhibitors to improve outcomes.

Faecalibaterium prausnitzii strain EXL01 boosts efficacy of immune checkpoint inhibitors.

Gut microbiota impacts responses to immune checkpoint inhibitors (ICI). A high level of Faecalibacterium prausnitzii have been associated with a positive response to ICI in multiple cancer types. Here, based on fecal shotgun metagenomics data, we show in two independent cohorts of patients with non-small cell lung cancer and advanced melanoma that a high level of F. prausnitzii at baseline is positively associated with a better clinical response to ICI. In MCA205 tumor-bearing mice, administration of F. prausnitzii strain EXL01, already in clinical development for Inflammatory Bowel Disease, restores the anti-tumor response to ICI in the context of antibiotic-induced microbiota perturbation at clinical and tumor transcriptomics level. In vitro, EXL01 strain enhances T cell activation in the presence of ICI. Interestingly, oral administration of EXL01 strain did not induce any change in fecal microbiota diversity or composition, suggesting a direct effect on immune cells in the small intestine. F. prausnitzii strain EXL01 will be evaluated as an adjuvant to ICI in multiple cancers in the near future.

Antitumor Effects of PRMT5 Inhibition in Sarcomas.

Patients with advanced soft-tissue sarcomas (STS) have few therapeutic options. Protein arginine methyltransferase 5 (PRMT5), an anticancer target, has been extensively investigated in recent years in epithelial tumors. To date, no data related to the biological role of PRMT5 inhibition and its potential effect as a treatment in STS have been reported.To investigate the therapeutic potential of PRMT5 targeting in STS, we first evaluated the prognostic value of PRMT5 expression in two different cohorts of patients with STS. We then used the potent and selective GSK3326595 (GSK595) compound to investigate the antitumor effect of the pharmacologic inhibition of PRMT5 in vitro via MTT, apoptosis, cell cycle, clonogenicity, and proliferation assays. In vivo studies were performed with two animal models to evaluate the effects of GSK595 on tumor growth. The mechanisms of action were investigated by RNA sequencing, metabolic pathway analysis, Western blotting, and glucose uptake/lactate production assays.High PRMT5 gene expression levels were significantly associated with worsened metastasis-free survival of patients with STS. GSK595 decreased the global symmetric dimethylarginine level, the proliferation rate and clonogenicity of STS cell lines in vitro and tumor growth in vivo. Moreover, PRMT5 inhibition regulated aerobic glycolysis through downregulation of key enzymes of glycolysis as well as glucose uptake and lactate production.The current study demonstrated that PRMT5 regulates STS cell metabolism and thus represents a potential therapeutic target for STS. Additional studies in diverse sarcoma subtypes will be essential to confirm and expand upon these findings. SIGNIFICANCE: STSs have limited therapeutic options. We show here the poor prognostic value of high PRMT5 expression in STS. Moreover, we demonstrate that the pharmacologic inhibition of PRMT5 has significant antitumor activity through the downregulation of glycolysis. Our findings support the clinical investigation of PRMT5 inhibition in STSs.

Upregulation of Indoleamine 2,3-Dioxygenase 1 in Tumor Cells and Tertiary Lymphoid Structures is a Hallmark of Inflamed Non-Small Cell Lung Cancer.

PURPOSE: Overexpression of the tryptophan-catabolizing enzyme indoleamine 2,3-dioxygenase 1 (IDO1) has been reported in several tumor types, including non-small cell lung cancer (NSCLC), and has been shown to promote tumor-immune evasion and inhibit T-cell activation through increased tryptophan degradation and the production of several immunosuppressive metabolites collectively known as kynurenines. However, it remains unclear whether IDO1 expression by tumor cells is detrimental specifically in the context of programmed cell death protein 1 (PD-1)/programmed death ligand 1 (PD-L1) axis blockade. EXPERIMENTAL DESIGN: We analyzed the transcriptome of 891 NSCLC tumor samples from patients enrolled in two large randomized clinical trials investigating the safety and activity of atezolizumab, a humanized IgG1 mAb that targets PD-L1, versus docetaxel in patients with advanced NSCLC. We complemented these transcriptomics results at the protein level by using multiplex immunofluorescence and at the functional level with in vitro experiments. RESULTS: The increased expression of the tryptophan-catabolizing enzyme IDO1 was significantly associated with improved objective response, progression-free survival, and overall survival in patients treated with PD-L1 inhibitors, but not in those treated with chemotherapy. Strikingly, inflamed tumors had higher levels of IDO1, and IDO1 was also expressed in tertiary lymphoid structures (TLS) by mature follicular dendritic cells. L-kynurenine impaired the differentiation of antibody-producing B cells induced by follicular helper T (Tfh)/B-cell interactions, a hallmark process within TLS. CONCLUSIONS: IDO1 pathway in NSCLC is driven by the immune system rather than by tumor cells. Targeting IDO1 in combination with anti-PD-1/PD-L1 might be beneficial only in patients with inflamed tumors and particularly in those bearing TLS.

Tryptophan metabolism activation by indoleamine 2,3-dioxygenase in adipose tissue of obese women: an attempt to maintain immune homeostasis and vascular tone.

Human obesity is characterized by chronic low-grade inflammation in white adipose tissue and is often associated with hypertension. The potential induction of indoleamine 2,3-dioxygenase-1 (IDO1), the rate-limiting enzyme in tryptophan/kynurenine degradation pathway, by proinflammatory cytokines, could be associated with these disorders but has remained unexplored in obesity. Using immunohistochemistry, we detected IDO1 expression in white adipose tissue of obese patients, and we focused on its contribution in the regulation of vascular tone and on its immunoregulatory effects. Concentrations of tryptophan and kynurenine were measured in sera of 36 obese and 15 lean women. The expression of IDO1 in corresponding omental and subcutaneous adipose tissues and liver was evaluated. Proinflammatory markers and T-cell subsets were analyzed in adipose tissue via the expression of CD14, IL-18, CD68, TNFα, CD3ε, FOXP3 [a regulatory T-cell (Treg) marker] and RORC (a Th17 marker). In obese subjects, the ratio of kynurenine to tryptophan, which reflects IDO1 activation, is higher than in lean subjects. Furthermore, IDO1 expression in both adipose tissues and liver is increased and is inversely correlated with arterial blood pressure. Inflammation is associated with a T-cell infiltration in obese adipose tissue, with predominance of Th17 in the omental compartment and of Treg in the subcutaneous depot. The Th17/Treg balance is decreased in subcutaneous fat and correlates with IDO1 activation. In contrast, in the omental compartment, despite IDO1 activation, the Th17/Treg balance control is impaired. Taken together, our results suggest that IDO1 activation represents a local compensatory mechanism to limit obesity-induced inflammation and hypertension.

Targeting Tryptophan Catabolism in Cancer Immunotherapy Era: Challenges and Perspectives.

Metabolism of tryptophan (Trp), an essential amino acid, represent a major metabolic pathway that both promotes tumor cell intrinsic malignant properties as well as restricts antitumour immunity, thus emerging as a drug development target for cancer immunotherapy. Three cytosolic enzymes, namely indoleamine 2,3-dioxygenase 1 (IDO1), IDO2 and tryptophan 2,3-dioxygenase (TDO2), catalyzes the first-rate limiting step of the degradation of Trp to kynurenine (Kyn) and modulates immunity toward immunosuppression mainly through the aryl hydrocarbon receptor (AhR) activation in numerous types of cancer. By restoring antitumor immune responses and synergizing with other immunotherapies, the encouraging preclinical data of IDO1 inhibitors has dramatically failed to translate into clinical success when combined with immune checkpoints inhibitors, reigniting the debate of combinatorial approach. In this review, we i) provide comprehensive evidences on immunomodulatory role of the Trp catabolism metabolites that highlight this pathway as relevant target in immuno-oncology, ii)ii) discuss underwhelming results from clinical trials investigating efficacy of IDO1 inhibitors and underlying mechanisms that might have contributed to this failure, and finally, iii) discuss the current state-of-art surrounding alternative approaches of innovative antitumor immunotherapies that target molecules of Trp catabolism as well as challenges and perspectives in the era of immunotherapy.

Prophylactic and Therapeutic Effect of Kynurenine for Experimental Autoimmune Encephalomyelitis (EAE) Disease.

Background: The essential amino acid, tryptophan, is predominantly metabolised through the kynurenine pathway (KP) to generate kynurenine, an aryl-hydrocarbon receptor (AhR) pro-ligand that exerts its effects in a ligand-dependent manner. Interaction between kynurenine and the AhR is an effector mechanism of immunosuppression. We previously found that the KP is involved in multiple sclerosis (MS) disease progression. We postulated that AhR activation by kynurenine might be neuroprotective by encouraging differentiation of Tregs. In this study, we assess both the prophylactic and therapeutic efficiency of kynurenine on disease severity and progression in mice with experimental autoimmune encephalomyelitis (EAE), an MS model. Methods: Myelin oligodendrocyte glycoprotein induced EAE mice (n = 6 per group) were treated with 200 mg/kg L-kynurenine once daily for 10 days beginning on either day 1 of EAE induction (prophylactic) or once they demonstrated motor weakness (therapeutic). Clinical disease severity measured by disease score, time on rotarod, and body weight. Results: The prophylactic kynurenine treatment significantly (P < .0001) prevented the development of a more severe disease course with mice demonstrating diminished relapse rate and improved clinical and behavioural outcomes. However, therapeutic kynurenine did not significantly (P = .4463) decrease the clinical signs until 36 days following induction of disease; after 36 days, it also significantly (P = .0479) reduced disease relapse. Mean body weight measurements only correlated with time on rotarod (r = -.6410; P = .0007) but not clinical scores (r = .1925; P = .3674). Conclusions: Kynurenine ameliorates EAE disease progression prophylactically and reduces relapses therapeutically. Further investigations are needed to elucidate the molecular mechanism explaining the therapeutic role of kynurenine for MS.

Phase 2 trial of intravenous oncolytic virus JX-594 combined with low-dose cyclophosphamide in patients with advanced breast cancer.

Breast cancer is one the most common cause of cancer death in women worldwide. We report here the first phase II study investigating a virus genetically engineered for tumor-selective replication in patients with breast cancer. Ten patients were treated with a combination of low-dose oral cyclophosphamide and intra-venous JX-594, a thymidine kinase gene-inactivated oncolytic vaccinia virus engineered for the expression of transgenes encoding human granulocyte-macrophage colony-stimulating factor (GM-CSF) and ß-galactosidase. Best response as per RECIST criteria was stable disease for 2 patients and progressive disease for 8 patients. Median progression-free and overall survival were 1.6 months (95% CI: [1.1-1.9]) and 14.4 months (95% CI: [2.0 - NA]) respectively. High throughput analysis of sequential plasma samples revealed an upregulation of protein biomarkers reflecting immune induction such as IFN gamma. Whether the combination of JX-594 with an immune checkpoint inhibitor is associated with meaningful clinical activity is therefore worth to investigate.

Randomized phase 2 trial of intravenous oncolytic virus JX-594 combined with low-dose cyclophosphamide in patients with advanced soft-tissue sarcoma.

JX-594 is an oncolytic vaccinia virus genetically modified to replicate selectively in tumor cells. Metronomic chemotherapy has shown preclinical synergy with oncolytic viruses. We report here the results of the METROMAJX which is a randomized phase II clinical trial investigating the combination of JX-594 combined with metronomic cyclophosphamide (arm 1) or metronomic cyclophosphamide (arm 2) in patients with advanced STS. A two-stage Simon design was used. JX-594 was administered intra-venously at the dose 1.109 every 2 weeks for the first 3 injections and then every 3 weeks. Cyclophosphamide was given orally at the dose of 50 mg BID 1 week on 1 week off. The primary endpoint was the 6-month non progression rate. 20 patients were included (arm 1:15, arm 2:5). The two most frequent toxicities were grade 1 fatigue and fever and grade 2 fatigue and grade 2 lymphopenia in arms 1 and 2, respectively. In arm 1, 12 patients were assessable for the efficacy analysis. None of them were progression-free at 6 months indicating that the first stage of the Simon's design was not satisfied. One patient out 4 assessable for efficacy was progression-free at 6 months in arm 2. High throughput analysis of sequential plasma samples revealed an upregulation of protein biomarkers reflecting immune induction such as CXCL10 and soluble CD8 antigen in arm 1. Systemic treatment with JX-594 is safe in patients with advanced STS. Further investigations are needed to improve immune response to oncolytic viruses and define their therapeutic potential in patients with STS.