Heme catabolism by tumor-associated macrophages controls metastasis formation.

Although the pathological significance of tumor-associated macrophage (TAM) heterogeneity is still poorly understood, TAM reprogramming is viewed as a promising anticancer therapy. Here we show that a distinct subset of TAMs (F4/80hiCD115hiC3aRhiCD88hi), endowed with high rates of heme catabolism by the stress-responsive enzyme heme oxygenase-1 (HO-1), plays a critical role in shaping a prometastatic tumor microenvironment favoring immunosuppression, angiogenesis and epithelial-to-mesenchymal transition. This population originates from F4/80+HO-1+ bone marrow (BM) precursors, accumulates in the blood of tumor bearers and preferentially localizes at the invasive margin through a mechanism dependent on the activation of Nrf2 and coordinated by the NF-κB1-CSF1R-C3aR axis. Inhibition of F4/80+HO-1+ TAM recruitment or myeloid-specific deletion of HO-1 blocks metastasis formation and improves anticancer immunotherapy. Relative expression of HO-1 in peripheral monocyte subsets, as well as in tumor lesions, discriminates survival among metastatic melanoma patients. Overall, these results identify a distinct cancer-induced HO-1+ myeloid subgroup as a new antimetastatic target and prognostic blood marker.

Gut Microbiota, Metabolome, and Body Composition Signatures of Response to Therapy in Patients with Advanced Melanoma.

Despite the recent breakthroughs in targeted and immunotherapy for melanoma, the overall survival rate remains low. In recent years, considerable attention has been paid to the gut microbiota and other modifiable patient factors (e.g., diet and body composition), though their role in influencing therapeutic responses has yet to be defined. Here, we characterized a cohort of 31 patients with unresectable IIIC-IV-stage cutaneous melanoma prior to initiation of targeted or first-line immunotherapy via the following methods: (i) fecal microbiome and metabolome via 16S rRNA amplicon sequencing and gas chromatography/mass spectrometry, respectively, and (ii) anthropometry, body composition, nutritional status, physical activity, biochemical parameters, and immunoprofiling. According to our data, patients subsequently classified as responders were obese (i.e., with high body mass index and high levels of total, visceral, subcutaneous, and intramuscular adipose tissue), non-sarcopenic, and enriched in certain fecal taxa (e.g., Phascolarctobacterium) and metabolites (e.g., anethole), which were potentially endowed with immunostimulatory and oncoprotective activities. On the other hand, non-response was associated with increased proportions of Streptococcus, Actinomyces, Veillonella, Dorea, Fusobacterium, higher neutrophil levels (and a higher neutrophil-to-lymphocyte ratio), and higher fecal levels of butyric acid and its esters, which also correlated with decreased survival. This exploratory study provides an integrated list of potential early prognostic biomarkers that could improve the clinical management of patients with advanced melanoma, in particular by guiding the design of adjuvant therapeutic strategies to improve treatment response and support long-term health improvement.

Blood to skin recirculation of CD4+ memory T cells associates with cutaneous and systemic manifestations of psoriatic disease.

Blood to skin recirculation could play a role in the pathogenesis of psoriasis. To investigate this possibility we dissected the phenotype of circulating T cells in psoriasis patients, calculated the correlation the clinical parameters of the disease and performed a parallel bioinformatics analysis of gene expression data in psoriatic skin. We found that circulating CCR6+ CD4+ TEM and TEFF cells significantly correlated with systemic inflammation. Conversely, the percentage of CXCR3+ CD4+ TEM cells negatively correlated with the severity of the cutaneous disease. Importantly CLA+ CD4+ TCM cells expressing CCR6+ or CCR4+CXCR3+ negatively correlated with psoriasis severity suggesting recruitment to the skin compartment. This assumption was reinforced by gene expression data showing marked increase of CCR7 and CLA-encoding gene SELPLG expression in psoriatic skin and strong association of their expression. The data enlightens a role for CD4+ T cells trafficking between blood and skin in cutaneous and systemic manifestations of psoriasis.

A role for CCR5(+)CD4 T cells in cutaneous psoriasis and for CD103(+) CCR4(+) CD8 Teff cells in the associated systemic inflammation.

Recent results have identified critical components of the T cell response involved in the initiation and amplification phases of psoriasis. However the link between T cell responses arising in the skin and the systemic inflammation associated with severe psoriasis is largely unknown. We hypothesized that specific subsets of memory T cells recirculating from the skin could play a role. We therefore dissected the circulating memory T cell compartment in patients by analyzing the TCM, TEM and Teff phenotype, the pattern of CCR4 and CCR5 chemokine receptor expression and the expression of the tissue homing molecule CD103. For each subset we calculated the correlation with the Psoriasis Area and Severity Index (PASI) and with the extent of systemic inflammation measured as serum level of the prototypic short pentraxin, C reactive protein (CRP). Validation was performed by comparison with gene expression data in psoriatic plaques. We found that circulating CD103(+)CCR4(+)CCR5(+) and CCR4(+)CCR6(-) CD8(+) Teff cells, were highly correlated with CRP levels as well as with the validated index PASI, reflecting a link between skin involvement and systemic inflammation in patients with severe psoriasis. In addition we observed a contraction of circulating CCR5(+) T cells in psoriasis patients, with a highly significant inverse correlation between CCR5(+)CD4 T cells and the PASI score. Increased expression of CCR5 and CCL5 genes in psoriatic skin lesions was consistent with an accumulation of CCR5(+) cells in psoriatic plaques indicating a role for CCR5/CCL5 axis in disease pathogenesis.

PEOPLE (NTC03447678), a phase II trial to test pembrolizumab as first-line treatment in patients with advanced NSCLC with PD-L1 <50%: a multiomics analysis.

BACKGROUND: Chemoimmunotherapy represents the standard of care for patients with advanced non-small cell lung cancer (NSCLC) and programmed death-ligand 1 (PD-L1) <50%. Although single-agent pembrolizumab has also demonstrated some activity in this setting, no reliable biomarkers yet exist for selecting patients likely to respond to single-agent immunotherapy. The main purpose of the study was to identify potential new biomarkers associated with progression-free-survival (PFS) within a multiomics analysis. METHODS: PEOPLE (NTC03447678) was a prospective phase II trial evaluating first-line pembrolizumab in patients with advanced EGFR and ALK wild type treatment-naïve NSCLC with PD-L1 <50%. Circulating immune profiling was performed by determination of absolute cell counts with multiparametric flow cytometry on freshly isolated whole blood samples at baseline and at first radiological evaluation. Gene expression profiling was performed using nCounter PanCancer IO 360 Panel (NanoString) on baseline tissue. Gut bacterial taxonomic abundance was obtained by shotgun metagenomic sequencing of stool samples at baseline. Omics data were analyzed with sequential univariate Cox proportional hazards regression predicting PFS, with Benjamini-Hochberg multiple comparisons correction. Biological features significant with univariate analysis were analyzed with multivariate least absolute shrinkage and selection operator (LASSO). RESULTS: From May 2018 to October 2020, 65 patients were enrolled. Median follow-up and PFS were 26.4 and 2.9 months, respectively. LASSO integration analysis, with an optimal lambda of 0.28, showed that peripheral blood natural killer cells/CD56dimCD16+ (HR 0.56, 0.41-0.76, p=0.006) abundance at baseline and non-classical CD14dimCD16+monocytes (HR 0.52, 0.36-0.75, p=0.004), eosinophils (CD15+CD16-) (HR 0.62, 0.44-0.89, p=0.03) and lymphocytes (HR 0.32, 0.19-0.56, p=0.001) after first radiologic evaluation correlated with favorable PFS as well as high baseline expression levels of CD244 (HR 0.74, 0.62-0.87, p=0.05) protein tyrosine phosphatase receptor type C (HR 0.55, 0.38-0.81, p=0.098) and killer cell lectin like receptor B1 (HR 0.76, 0.66-0.89, p=0.05). Interferon-responsive factor 9 and cartilage oligomeric matrix protein genes correlated with unfavorable PFS (HR 3.03, 1.52-6.02, p 0.08 and HR 1.22, 1.08-1.37, p=0.06, corrected). No microbiome features were selected. CONCLUSIONS: This multiomics approach was able to identify immune cell subsets and expression levels of genes associated to PFS in patients with PD-L1 <50% NSCLC treated with first-line pembrolizumab. These preliminary data will be confirmed in the larger multicentric international I3LUNG trial (NCT05537922). TRIAL REGISTRATION NUMBER: 2017-002841-31.

Landscape of immune-related signatures induced by targeting of different epigenetic regulators in melanoma: implications for immunotherapy.

BACKGROUND: Improvement of efficacy of immune checkpoint blockade (ICB) remains a major clinical goal. Association of ICB with immunomodulatory epigenetic drugs is an option. However, epigenetic inhibitors show a heterogeneous landscape of activities. Analysis of transcriptional programs induced in neoplastic cells by distinct classes of epigenetic drugs may foster identification of the most promising agents. METHODS: Melanoma cell lines, characterized for mutational and differentiation profile, were treated with inhibitors of DNA methyltransferases (guadecitabine), histone deacetylases (givinostat), BET proteins (JQ1 and OTX-015), and enhancer of zeste homolog 2 (GSK126). Modulatory effects of epigenetic drugs were evaluated at the gene and protein levels. Master molecules explaining changes in gene expression were identified by Upstream Regulator (UR) analysis. Gene set enrichment and IPA were used respectively to test modulation of guadecitabine-specific gene and UR signatures in baseline and on-treatment tumor biopsies from melanoma patients in the Phase Ib NIBIT-M4 Guadecitabine + Ipilimumab Trial. Prognostic significance of drug-specific immune-related genes was tested with Timer 2.0 in TCGA tumor datasets. RESULTS: Epigenetic drugs induced different profiles of gene expression in melanoma cell lines. Immune-related genes were frequently upregulated by guadecitabine, irrespective of the mutational and differentiation profiles of the melanoma cell lines, to a lesser extent by givinostat, but mostly downregulated by JQ1 and OTX-015. GSK126 was the least active drug. Quantitative western blot analysis confirmed drug-specific modulatory profiles. Most of the guadecitabine-specific signature genes were upregulated in on-treatment NIBIT-M4 tumor biopsies, but not in on-treatment lesions of patients treated only with ipilimumab. A guadecitabine-specific UR signature, containing activated molecules of the TLR, NF-kB, and IFN innate immunity pathways, was induced in drug-treated melanoma, mesothelioma and hepatocarcinoma cell lines and in a human melanoma xenograft model. Activation of guadecitabine-specific UR signature molecules in on-treatment tumor biopsies discriminated responding from non-responding NIBIT-M4 patients. Sixty-five % of the immune-related genes upregulated by guadecitabine were prognostically significant and conferred a reduced risk in the TCGA cutaneous melanoma dataset. CONCLUSIONS: The DNMT inhibitor guadecitabine emerged as the most promising immunomodulatory agent among those tested, supporting the rationale for usage of this class of epigenetic drugs in combinatorial immunotherapy approaches.

Fifteen-year follow-up of relapsed indolent non-Hodgkin lymphoma patients vaccinated with tumor-loaded dendritic cells.

We previously published the results of a pilot study showing that vaccination with tumor-loaded dendritic cells (DCs) induced both T and B cell response and produced clinical benefit in the absence of toxicity in patients with relapsed, indolent non-Hodgkin lymphoma (iNHL). The purpose of the present short report is to provide a 15-year follow-up of our study and to expand the biomarker analysis previously performed. The long-term follow-up highlighted the absence of particular or delayed toxicity and the benefit of active immunization with DCs loaded with autologous, heat-shocked and UV-C treated tumor cells in relapsed iNHL (5-year and 10-year progression-free survival (PFS) rates: 55.6% and 33.3%, respectively; 10-year overall survival (OS) rate: 83.3%). Female patients experienced a better PFS (p=0.016) and a trend towards a better OS (p=0.185) compared with male patients. Of note, we observed a non-negligible fraction of patients (22%) who experienced a long-lasting complete response. In a targeted gene expression profiling of pre-treatment tumor biopsies in 11 patients with available formalin-fixed, paraffin-embedded tissue, we observed that KIT, ATG12, TNFRSF10C, PBK, ITGA2, GATA3, CLU, NCAM1, SYT17 and LTK were differentially expressed in patients with responder versus non-responder tumors. The characterization of peripheral monocytic cells in a subgroup of 14 patients with available baseline blood samples showed a higher frequency of the subset of CD14++CD16+ cells (intermediate monocytes) in patients with responding tumors. Since in patients with relapsed iNHL the available therapeutic options are often incapable of inducing a long-lasting complete remission and can be sometimes characterized by intolerable toxicity, we think that the encouraging results of our long-term follow-up analysis represent a stimulus to further investigate the role of active vaccination in this specific setting and in earlier lines of therapy and to explore novel combinatorial strategies encompassing other innovative immunotherapy agents, such as immune-checkpoint inhibitors.

Immune Escape Mechanisms in Non Small Cell Lung Cancer.

Development of strong immune evasion has been traditionally associated with the late stages of solid tumor progression, since advanced cancers are more likely to have reached the third phase of the immunoediting process. However, by integrating a variety of approaches, evidence for active immune escape mechanisms has been found even in the pre-invasive lesions that later progress to the main NSCLC histotypes. Pre-invasive lesions of adenocarcinoma (LUAD) and of squamous cell carcinoma (LUSC) can show impaired antigen presentation, loss of heterozygosity at the Human Leukocyte Antigen (HLA) region, neoantigen silencing, activation of immune checkpoints, altered TH1/TH2 cytokine ratios, and immune contexture evolution. Analysis of large panels of LUAD vs. LUSC, of early stage NSCLC vs. normal lung tissue, of specific molecular subsets of NSCLC, and of distinct regions within the same tumor, indicates that all these processes of immune escape continue to evolve in the invasive stage of NSCLC, are associated with inter- and intra-tumor heterogeneity, and contribute to resistance to therapy by immune checkpoint blockade (ICB). In this review, we will discuss the most recent evidence on immune escape mechanisms developing from the precursor to invasive stage in NSCLC, and the contribution of immune evasion to resistance to ICB in lung cancer.

Vascular complications of pancreatectomy.

Despite decreased postoperative mortality, pancreatic resections continue to be associated with high morbidity rates. Vascular complications and, in particular, erosive bleeding from the large retroperitoneal vasculature are particularly difficult to treat and account for a large percentage of the residual postoperative mortality of pancreatic resections. We herein analyze the pathogenesis, diagnosis, preventive measures and possible remedies of either hemorrhagic or occlusive complications of pancreatic resections through a review of the literature and of our institutional experience consisting of 818 pancreatectomies.