High Levels Of PD-L1 Expression Research Articles

PD-L1 expression in tumor and inflammatory cells is associated with favorable tumor features and favorable prognosis in muscle-invasive urothelial carcinoma of the bladder not treated by immune checkpoint inhibitors

BackgroundA high level of PD-L1 expression is the most relevant predictive parameter for response to immune checkpoint inhibitor (CPI) therapy in urinary bladder cancer. Existing data on the relationship between PD-L1 expression and the natural course of disease are controversial and sparse.MethodsTo expand our understanding of the relationship between PD-L1 expression and parameters of cancer aggressiveness, PD-L1 was analyzed on tissue microarrays containing 2710 urothelial bladder carcinomas including 512 patients with follow-up data who underwent radical cystectomy and follow-up therapies in the pre-immune checkpoint inhibitor therapy era.ResultsTumor cell positivity in ≥10% of cells were seen in 513 (20%) and an immune cell positivity occurred in 872 (34%) of 2566 interpretable cancers. PD-L1 positivity in tumor cells increased from pTaG2 low grade (0.9% positive) to pTaG3 high grade (4.1%; p = 0.0255) and was even higher in muscle-invasive (pT2–4) carcinomas (29.3%; p < 0.0001). However, within pT2–4 carcinomas, PD-L1 positivity was linked to low pT stage (p = 0.0028), pN0 (p < 0.0001), L0 status (p = 0.0005), and a better prognosis within 512 patients with cystectomy who never received CPIs (p = 0.0073 for tumor cells and p = 0.0086 for inflammatory cells). PD-L1 staining in inflammatory cells was significantly linked to PD-L1 staining in tumor cells (p < 0.0001) and both were linked to a positive p53 immunostaining (p < 0.0001).ConclusionIt cannot be fully excluded that the strong statistical link between PD-L1 status and favorable histological tumor features as well as better prognosis could influence the outcome of studies evaluating CPIs in muscle-invasive urothelial carcinoma.

An exosome-derived lncRNA signature identified by machine learning associated with prognosis and biomarkers for immunotherapy in ovarian cancer.

Ovarian cancer (OC) has the highest mortality rate among gynecological malignancies. Current treatment options are limited and ineffective, prompting the discovery of reliable biomarkers. Exosome lncRNAs, carrying genetic information, are promising new markers. Previous studies only focused on exosome-related genes and employed the Lasso algorithm to construct prediction models, which are not robust. 420 OC patients from the TCGA datasets were divided into training and validation datasets. The GSE102037 dataset was used for external validation. LncRNAs associated with exosome-related genes were selected using Pearson analysis. Univariate COX regression analysis was used to filter prognosis-related lncRNAs. The overlapping lncRNAs were identified as candidate lncRNAs for machine learning. Based on 10 machine learning algorithms and 117 algorithm combinations, the optimal predictor combinations were selected according to the C index. The exosome-related LncRNA Signature (ERLS) model was constructed using multivariate COX regression. Based on the median risk score of the training datasets, the patients were divided into high- and low-risk groups. Kaplan-Meier survival analysis, the time-dependent ROC, immune cell infiltration, immunotherapy response, and immune checkpoints were analyzed. 64 lncRNAs were subjected to a machine-learning process. Based on the stepCox (forward) combined Ridge algorithm, 20 lncRNA were selected to construct the ERLS model. Kaplan-Meier survival analysis showed that the high-risk group had a lower survival rate. The area under the curve (AUC) in predicting OS at 1, 3, and 5 years were 0.758, 0.816, and 0.827 in the entire TCGA cohort. xCell and ssGSEA analysis showed that the low-risk group had higher immune cell infiltration, which may contribute to the activation of cytolytic activity, inflammation promotion, and T-cell co-stimulation pathways. The low-risk group had higher expression levels of PDL1, CTLA4, and higher TMB. The ERLS model can predict response to anti-PD1 and anti-CTLA4 therapy. Patients with low expression of PDL1 or high expression of CTLA4 and low ERLS exhibited significantly better survival prospects, whereas patients with high ERLS and low levels of PDL1 or CTLA4 exhibited the poorest outcomes. Our study constructed an ERLS model that can predict prognostic risk and immunotherapy response, optimizing clinical management for OC patients.

Polymorphisms of the PD-L1 gene 3'-untranslated region are associated with the expression of PD-L1 in non-small cell lung cancer.

Recent results show that polymorphisms of programmed death ligand 1 (PD-L1, also known as CD274 or B7-H1) might be used as a possible marker for effectiveness of chemotherapy and cancer risk. However, the effect of PD-L1 gene variations on PD-L1 expression remain unclear. Given the post-transcriptional machinery in tumor PD-L1 expression, we investigated single nucleotide polymorphisms (SNPs) in the 3'-untranslated region (3'-UTR) of the PD-L1 gene, rs4143815 and rs4742098, using formalin-fixed paraffin-embedded sections of 154 patients with non-small cell lung cancers (NSCLCs). In rs4143815, the GG genotype showed significant association with PD-L1 expression (P = 0.032). In rs4742098, the AA genotype was significantly associated with histology and PD-L1 expression (P = 0.022 and P = 0.008, respectively). In multivariate logistic regression analysis, the AA genotype in rs4742098 was correlated with PD-L1 expression (odds ratio 0.408, P = 0.048). Interestingly, approximately 10% of the NSCLC cases showed somatic mutation when we compared genotypes of these SNPs between NSCLC tissues and non-tumor tissues from the same patients. In addition, cases with somatic mutation showed higher levels of PD-L1 expression than cases with germline mutation in rs4143815 GG. In conclusion, we demonstrated that the rs4143815 and rs4742098 SNPs in the 3'-UTR of PD-L1 were associated with tumor PD-L1 expression in NSCLCs.

The multiple arrays of a PD1-derived peptide on chromatin specifically bind to PD-L1 and induce doxorubicin resistance in cancer cell lines

Programmed cell death-1 (PD-1) and its ligand 1 (PD-L1) have been extensively studied to block the activation of the PD-1 axis immune checkpoint pathway on T cells. However, recent evidence suggests that PD-1-independent PD-L1 pathways in cancer cells and macrophages are important in cellular proliferation and survival of those cell types but the underlying mechanism is little understood. To recapitulate the binding of multiple PD-1 to the high levels of PD-L1 expression on cancer cell membranes, recombinant histones carrying a PD-1 receptor-derived peptide (PDR) were prepared and used to assemble a PDR-displayed nucleosome array. PDR-displayed chromatin showed physiologically spaced nucleosomes, unaffected by N-terminal histone tails and biotinylated DNA modifications. PDR-displayed chromatin exhibited selective binding to the breast cancer cell line with high PD-L1 expression, MDA-MB-231, where saturated binding occurred within 3 h, comparable to well-known antigen-antibody reactions. Notably, PDR-displayed chromatin enhanced resistance to doxorubicin in PD-L1-overexpressed cancer cells, revealing a PD-L1 level-dependent effect on cell survival upon chemotherapy. Our study sheds light on the potential of PDR-displayed chromatin as a tool to induce chemoresistance in cancer cells without employing anti-PD-L1 antibodies or soluble PD-1 receptors. Further investigation into the underlying signaling pathways and therapeutic applications is warranted, positioning PDR-displayed chromatin as a valuable tool for understanding PD-L1-mediated intracellular signaling pathways in cancer cells with high PD-L1 expression.

Tolerogenic phenotype of dendritic cells is induced after hapten sensitization followed by attenuated contact hypersensitivity response in atopic dermatitis model NC/Nga mice

Allergic contact dermatitis (ACD) and atopic dermatitis (AD) are common inflammatory diseases. We previously reported attenuated contact hypersensitivity (CHS) responses in AD model mice using 2,4-dinitrofluorobenzene, reflecting clinical experiments. However, previous studies have not addressed the commonality of findings across haptens and mechanisms focused on dendritic cells (DCs). Thus, this study evaluated CHS responses to fluorescein isothiocyanate (FITC) and DC migration and maturation in the sensitization phase of CHS in AD. CHS responses to FITC were compared between NC/Nga mice without and with AD induction (non-AD and AD mice, respectively). T-cell responses and DC migration and maturation after FITC-induced sensitization were examined in the draining lymph nodes of non-AD and AD mice. AD mice demonstrated reduced CHS responses to FITC under decreased T-cell proliferation following sensitization and interferon-γ production by hapten-specific T cells compared with non-AD mice. In addition, the number of FITC+CD11c+MHC class IIhigh migratory DCs 24 h after FITC sensitization was comparable between non-AD and AD mice. However, FITC+CD11c+MHC class IIhigh migratory DCs in AD mice exhibited lower expression levels of CD80 and CD86 and higher expression levels of PD-L1 and mRNA of transforming growth factor beta than non-AD mice. These findings suggest that attenuated CHS responses may be hapten-independent and the induction of the tolerogenic phenotype of hapten-bearing DCs can contribute to reduced T-cell proliferation after sensitization and CHS responses in AD.

High PD-L1 expression is associated with unfavorable clinical features in myelodysplastic neoplasms

Immune checkpoints are regulators of the immune system response that allow self-tolerance. Molecules such as Programmed Cell Death Protein 1 (PD-1) and its Ligand (PD-L1) participate in the immune checkpoint by signaling co-inhibition of lymphocyte responses. In cancers, PD-L1 expression is associated with the immune evasion mechanism, which favors tumor growth. The use of anti-PD-1/PD-L1 drugs is already well described in solid tumors, but still not fully understood in hematologic malignancies. Myelodysplastic neoplasms (MDSs) are heterogeneous bone marrow disorders with an increased risk of progression to Acute Myeloid Leukemia (AML). The MDS affects hematopoietic stem cells and its pathogenesis is linked to genetic and epigenetic defects, in addition to immune dysregulation. The influence of the PD-L1 on the MDS remains unknown. In this study, we evaluated the mRNA expression of the PD-L1 in 53 patients with MDS, classified according to the WHO 2016 Classification. Patients with dyserythropoiesis presented significantly higher PD-L1 expression than patients without dyserythropoiesis (p=0.050). Patients classified as having MDS with an excess of blasts 2 (MDS-EB2) presented a significant upregulation in the mRNA expression of the PD-L1 compared to the MDS with an excess of blasts 1 (MDS-EB1) (p=0.050). Furthermore, we detected three patients with very high levels of PD-L1 expression, being statistically classified as outliers. We suggested that the high expression of the PD-L1 is associated with a worse prognosis in the MDS and functional studies are necessary to evaluate the possible use of anti-PD-L1 therapies for high-risk MDS, such as the MDS-EBs.

Abstract 6647: Dostarlimab shows dose-dependent immune activation of the tumor microenvironment in a patient-derived NSCLC explant model similar to pembrolizumab

Abstract The ability to more faithfully mimic the tumor microenvironment (TME) of cancer patients offers a wide range of possibilities for non-clinical evaluation of anti-cancer therapeutics, including guidance for patient selection, dose, and combinations. We have established a patient-derived tumor explant (PDE) model to monitor immune activation within a structurally preserved TME, following ex vivo incubation with immune check point inhibitors (ICI). When testing anti-PD-1 agents, such as dostarlimab and pembrolizumab, in a cohort of NSCLC explants from stage I-III treatment-naïve patients, we observed that approximately half demonstrate release of inflammatory cytokines after two days incubation. Interestingly, responsive tumors presented with a high level of PD-L1 expression (high TPS), had elevated TMB, and were highly infiltrated by CD8+ T cells at baseline, in line with tumor baseline characteristics predictive of clinical response to anti-PD-1. When testing a dose-range of dostarlimab and pembrolizumab comparable potencies were observed when measuring cytokine release after treatment of the NSCLC explants. In conclusion, we describe here an in vitro patient-derived NSCLC model that mirrors the heterogeneity of patients’ response for anti-PD-1 agents observed in clinical settings, representing a valuable tool for comparative analysis of novel ICI assets and combinations, such as the CD226 axis members. Importantly, this system is also amenable to extensive multi-omics and genetics-based analyses, providing the opportunity to better identify predictive or pharmacodynamic characteristics within patient tumors. Citation Format: Jessica Perrin, Frank Wippich, Valeriia Sherina, Dmytro Dvornikov, Katharina Lupar, Murad M. Melhem, Patrick Hanafin, Jeremy Waight, Jong Yu, Sapna Yadavilli, Daniel Poeckel, Giovanna Bergamini. Dostarlimab shows dose-dependent immune activation of the tumor microenvironment in a patient-derived NSCLC explant model similar to pembrolizumab [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6647.

Abstract 4626: Spatial profiling of the tumor microenvironment in head and neck cancer identifies immune checkpoints and proteins involved in cell death signaling biomarkers of immunotherapy response

Abstract Immune checkpoint inhibitors (ICIs) Pembrolizumab and Nivolumab have shown promising results in patients with recurrent or metastatic (R/M) head and neck squamous cell carcinoma (HNSCC). Nonetheless, only up to 20% of patients may benefit from them. Next generation predictive biomarkers of response to ICI therapies are currently needed. This retrospective study profiled pre-treatment tissue samples collected from R/M HNSCC patients (n=30) treated with Pembrolizumab or Nivolumab. Targeted spatial proteomics profiling was performed using NanoString’s GeoMx® Digital Spatial Profiler (GeoMx). To develop an informed region-of-interest (ROI) selection strategy for the GeoMx, we analyzed H&amp;E images from serial sections and performed Oncotopix® Discovery software, which considered pathology inputs and machine learning to analyze H&amp;E imagery and 4-channel GeoMx visualization inputs. We profiled 80 proteins simultaneously using the GeoMx across modules in cell death, immune activation, immune cell typing, immuno-oncology drug target, immune profiling, PI3K/AKT signaling, and pan-tumor markers.. Next, we performed differential expression (DE) analysis of the proteins localized in the tumor and stromal compartments against response to therapy parameters of complete, partial, stable and progressive disease (according to RECIST criteria). Four-channel GeoMx immunofluorescence images provided per-ROI cell counts (total and phenotyped based on 3 markers), and these data were used to aid in area and cell count normalization within the tumor and stromal compartments. We found that patients responsive to immunotherapy had higher expression levels of PD-L1, Bcl-2, BCLX, and BIM in the tumor, whereas VISTA, FOXP3, and CD66b were downregulated. In the stromal compartment, it was found that responders had higher expression levels of B7-H3, CD40, and SMA, but lower expression of PARP, S100B, and NY-ESO-1 in comparison to non-responders. In terms of best response analysis, patients with PR (n=5) had higher levels of PD-L1, ER-alpha, and CD68 expression, but lower levels of VISTA, CD27, and CD95/Fas in tumor regions than patients with PD (n=8). In the stroma, PD-L1, CD68, and HLA-DR were upregulated, while VISTA, BIM, and BAD were downregulated in patients with PR versus those with PD. We found that informed ROI selection strategy aided in defining key features in the tumor microenvironment for comparative analysis across samples, and for data normalization methods. Moreover, we found that immune checkpoints and proteins involved in cell death signaling play important roles in the immune responsive tumor microenvironment of HNSCC based on response to immunotherapy. Citation Format: Habib Sadeghirad, Ning Liu, James Monkman, Chin Wee Tan, Caroline Cooper, Jeni Caldera, Sarah Church, Fabian Schneider, James Robert Mansfield, Ken O'Byrne, Melissa Davis, Brett Hughes, Arutha Kulasinghe. Spatial profiling of the tumor microenvironment in head and neck cancer identifies immune checkpoints and proteins involved in cell death signaling biomarkers of immunotherapy response. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4626.

KRAS G12C mutated advanced non-small cell lung cancer (NSCLC): Characteristics, treatment patterns and overall survival from a Danish nationwide observational register study

ObjectivesWe aimed to characterize the advanced NSCLC population in terms of KRAS G12C prevalence, patient characteristics, and survival outcomes after the introduction of immunotherapies. Materials and MethodsWe identified adult patients diagnosed with advanced NSCLC between January 1, 2018 and June 30, 2021 using the Danish health registries. Patients were grouped by mutational status (any KRAS mutation, KRAS G12C, and KRAS/EGFR/ALK wildtype [Triple WT]). We analyzed KRAS G12C prevalence, patient and tumor characteristics, treatment history, time-to-next-treatment (TTNT), and overall survival (OS). ResultsWe identified 7,440 patients of whom 40% (n = 2,969) were KRAS tested prior to the first line of therapy (LOT1). Among the KRAS tested, 11% (n = 328) harbored KRAS G12C. More KRAS G12C patients were women (67%), smokers (86%), had a high (≥50%) level of PD-L1 expression (54%), and more frequently received anti-PD-L1 treatment than any other group. From the date of the mutational test result, OS (7.1–7.3 months) was similar between the groups. OS from LOT1 (14.0 months) and LOT2 (10.8 months), and TTNT from LOT1 (6.9 months) and LOT2 (6.3 months) was numerically longer for the KRAS G12C mutated group compared to any other group. However, from LOT1 and LOT2, the OS and TTNT were comparable when stratifying the groups by PD-L1 expression level. Regardless of the mutational group, OS was markedly longer for patients with high PD-L1 expression. ConclusionIn patients diagnosed with advanced NSCLC after the implementation of anti-PD-1/L1 therapies, the survival in KRAS G12C mutated patients is comparable to patients with any KRAS mutation, Triple WT, and all NSCLC patients.

The PD-L1 Expression and Tumor-Infiltrating Immune Cells Predict an Unfavorable Prognosis in Pancreatic Ductal Adenocarcinoma and Adenosquamous Carcinoma

The tumor microenvironment (TME) plays a vital role in the development, progression, and metastasis of pancreatic cancer (PC). The composition of the TME and its potential prognostic value remains to be fully understood, especially in adenosquamous carcinoma of pancreas (ASCP) patients. Immunohistochemistry was used to explore the clinical significance of CD3, CD4, CD8, FoxP3, and PD-L1 expression within the TME and to identify correlations with the prognosis of PC in a series of 29 patients with ASCP and 54 patients with pancreatic ductal adenocarcinoma (PDAC). Data from the Gene Expression Omnibus (GEO) and the Cancer Genome Atlas (TCGA) were accessed to obtain the scRNA-seq data and transcriptome profiles. Seurat was used to process the scRNA-seq data, and CellChat was used to analyze cell-cell communication. CIBERSORT was used to approximate the constitution of tumor-infiltrating immune cell (TICs) profiles. Higher levels of PD-L1 were linked with a shorter overall survival in ASCP (p = 0.0007) and PDAC (p = 0.0594). A higher expression of CD3+ and CD8+ T-cell infiltration was significantly correlated with a better prognosis in PC. By influencing the composition of tumor-infiltrating immune cells (TICs), high levels of PD-L1 expression are linked with a shorter overall survival in ASCP and PDAC.

Gene mutation profiles and clinicopathological features of patients with non-small cell lung cancer harboring KRAS G12C mutation: a single-center retrospective study

Objective: To accurately screen non-small cell lung cancer (NSCLC) patients with KRAS G12C mutation and to evaluate their clinicopathological features, prognostic factors and current treatment status. Methods: A total of 19 410 NSCLC cases diagnosed at the Department of Pathology of Shanghai Chest Hospital, Shanghai, China from January 2018 to September 2021 were retrospectively reviewed, and the cases with KRAS gene mutation detected by next-generation sequencing were included. The clinicopathological and genetic mutation data of these cases were collected and analyzed. Results: A total of 1 633 (8.4%) NSCLC patients carried a KRAS gene mutation, among whom G12C was the most frequent (468 cases, 28.7%) mutant subtype. The mutation was more commonly found in males (414/468, 88.5%), patients with a history of smoking (308/468, 65.8%), and patients with a pathological type of invasive adenocarcinoma (231/468, 49.4%). The most common co-mutated genes in KRAS G12C mutant NSCLC were TP53 (52.4%, 245/468), STK11 (18.6%, 87/468) and ATM (13.2%, 62/468). The proportion of PD-L1 expression (≥1%) in KRAS G12C mutant NSCLC was significantly higher than that in patients without G12C mutation [64.3% (90/140) vs. 56.1% (193/344), P=0.014]. Immune checkpoint inhibitors (ICIs) treatment significantly prolonged progression-free survival (PFS) in NSCLC patients (10.0 months vs. 5.0 months, P=0.011). However, combination of chemotherapy and ICIs with anti-angiogenesis inhibitors or multi-target inhibitors did not significantly improve PFS in patients with KRAS G12C mutant NSCLC (P>0.05). Patients with KRAS G12C mutation NSCLC treated with ICIs and KRAS G12C patients with TP53 mutation had significantly longer median PFS than those with STK11 mutation (9.0 months vs. 4.3 months, P=0.012). Conclusions: Patients with KRAS G12C mutant NSCLC have relatively higher levels of PD-L1 expression and can benefit from ICIs treatment. The feasibility of chemotherapy, ICIs therapy and their combination needs further investigation.

Investigation of the mutational status of the FGFR3 gene in urothelial bladder carcinoma

To study the somatic mutational status of the FGFR3 gene in urothelial bladder cancer (BC) and evaluate its relationship with the clinical and morphological characteristics of the tumor, deficiency of the DNA mismatch repair (dMMR), PD-L1 tumor status, and immunohistochemical (IHC) expression of the p16 protein. Surgical material of 40 patients with BC, on which the mutational status of the FGFR3 gene was studied using the molecular genetic method, as well as the MMR status, PD-L1 and p16 expression by the IHC method. FGFR3 mutations, such as G370C, S249C, S371C/Y373C, R248C, were detected in 35.0% of the studied BC samples. FGFR3 status did not depend on the gender and age of patients, as well as on the degree of tumor lymphoid infiltration (TILs). Statistically significant differences were found in the analysis of FGFR3 status depending on the histological structure and degree of tumor differentiation, as well as on the pT stage. The FGFR3 status of BC was not associated with the IHC expression of the studied proteins of the MMR system, as well as with the PD-L1 status. Higher levels of PD-L1 expression were demonstrated by BC tumor cells, in which no aberrations in FGFR3 were detected. There was no significant association between p16 status and the presence of FGFR3 mutations, but for FGFR3-positive carcinomas, the basal pattern of p16 staining by IHC was noted. A positive somatic mutational status of the FGFR3 gene was statistically significantly more common in the group of papillary low-grade non-muscle-invasive BC, demonstrating basal p16 IHC staining. In the study sample, there was no statistically significant relationship between the FGFR3 status of BC and gender and age differences, TILs, MMR status, PD-L1 status (SP142 and 22C3), and p16 status. The results of the study indicate the need to determine the FGFR3 status in patients with BC for further prescription of personalized therapy.

The association between the tumor immune microenvironments and clinical outcome in low-grade, early-stage endometrial cancer patients.

Endometrial tumors show substantial heterogeneity in their immune microenvironment. This heterogeneity could be used to improve the accuracy of current outcome prediction tools. We assessed the immune microenvironment of 235 patients diagnosed with low-grade, early-stage endometrial cancer. Multiplex quantitative immunofluorescence was carried out to measure CD8, CD68, FOXP3, PD-1, and PD-L1 markers, as well as cytokeratin (CK), on tissue microarrays. Clustering results revealed five robust immune response patterns, each associated with specific immune populations, cell phenotypes, and cell spatial clustering. Most samples (69%) belonged to the immune-desert subtype, characterized by low immune cell densities. Tumor-infiltrating lymphocyte (TIL)-rich samples (4%) displayed high CD8+ T-cell infiltration, as well as a high percentage of CD8/PD-1+ cells. Immune-exclusion samples (19%) displayed the lowest CD8+ infiltration combined with high PD-L1 expression levels in CK+ tumor cells. In addition, they demonstrated high tumor cell spatial clustering as well as increased spatial proximity of CD8+ /PD-1+ and CK/PD-L1+ cells. FOXP3 and macrophage-rich phenotypes (3% and 4% of total samples) displayed relatively high levels of FOXP3+ regulatory T-cells and CD68+ macrophages, respectively. These phenotypes correlated with clinical outcomes, with immune-exclusion tumors showing an association with tumor relapse. When compared with prediction models built using routine pathological variables, models optimized with immune variables showed increased outcome prediction capacity (AUC = 0.89 versus 0.78) and stratification potential. The improved prediction capacity was independent of mismatch repair protein status and adjuvant radiotherapy treatment. Further, immunofluorescence results could be partially recapitulated using single-marker immunohistochemistry (IHC) performed on whole tissue sections. TIL-rich tumors demonstrated increased CD8+ T-cells by IHC, while immune-exclusion tumors displayed a lack of CD8+ T-cells and frequent expression of PD-L1 in tumor cells. Our results demonstrate the capability of the immune microenvironment to improve standard prediction tools in low-grade, early-stage endometrial carcinomas. © 2022 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

INCREASED PROGRAMMED DEATH LIGAND-1 EXPRESSION LEADS TO WORSE CLINICAL FEATURES IN PATIENTS WITH MYELODYSPLASTIC SYNDROME

Immune checkpoints are regulators of the immune system response that allow self-tolerance. Molecules such as Programmed Cell Death Protein 1 (PD-1) and its Ligand (PD-L1) participate in the immune checkpoint by signaling co-inhibition of lymphocyte responses. In cancers, PD-L1 expression is associated with the immune evasion mechanism, which favors tumor growth. The role of PD-L1 is already known for several solid tumors and anti-PD-L1 drugs are used for treatment. Myelodysplastic Syndrome (MDS) is a heterogeneous bone marrow disease with an increased risk of progression to Acute Myeloid Leukemia (AML). The pathogenesis of MDS involves several factors, including alterations in immune pathways. However, the role of PD-L1 still lacks understanding in this disease. The aim of this study was to evaluate if PD-L1 gene expression would be linked to clinical and prognostic features in patients with myelodysplastic syndrome. In this study, we evaluated PD-L1 mRNA expression by real-time PCR using TaqMan ® probes. Fifty-three patients classified according to WHO 2016 were analyzed. We observed that patients with dyserythropoiesis have higher PD-L1 expression than patients without dyserythropoiesis (p = 0.050). Furthermore, patients classified as excess of blasts 2 (MDS-EB2) presented an upregulation in mRNA expression of PD-L1 compared to excess of blasts 1 (MDS-EB1) (p = 0.050). Throughout the statistical analysis, we detected three patients with very high levels of PD-L1 expression, being classified as outliers. Analyzing these three patients specifically, all of them had hemoglobin lower than 8g/dL, blast count higher than 8%, dysplasia in at least two sectors and had died by the end of the study. The increase of PD-L1 in MDS may be considered a marker of poor prognosis.

Immune Checkpoint Inhibitors' Associated Renal Toxicity: A Series of 12 Cases.

We present a series of twelve patients, bearing a wide range of solid malignancies, who received either PD-L1 or a combination of PD-L1 and CTLA-4 inhibitors. Following immunotherapy administration, they exhibited the clinical signs indicative of renal toxicity, including increased serum creatinine levels, proteinuria, nephrotic syndrome and/or hematuria. All patients underwent renal biopsy. Results: All cases demonstrated some degree of interstitial inflammation and tubular injury, while in five patients, glomerular alterations consistent with a specific glomerulopathy were also observed: secondary “lupus-like” membranous glomerulopathy in two cases and membranoproliferative glomerulonephritis, IgA glomerulonephritis and secondary AA amyloidosis in each of the remaining three patients. The two patients with “lupus-like” nephritis and the one with amyloidosis experienced nephrotic syndrome, while their creatinine was within normal range. In the remaining nine cases, deterioration of renal function was the main manifestation. Conclusion: Our findings harmonize with bibliographical data that identify tubulointerstitial nephritis as the most frequent histological lesion related to ICIs administration. The preferential involvement of tubulointerstitial tissue could be associated with the reported higher expression levels of PD-L1 on tubular epithelial cells, compared to glomeruli. On the other hand, glomerular involvement is probably a consequence of a systemic immune system reconstruction, induced by immune-checkpoints inhibition.

FoRT 05-BEAT: A phase II randomized trial comparing atezolizumab versus atezolizumab + bevacizumab as first-line treatment in patients with PD-L1 high advanced/metastatic NSCLC.

TPS9145 Background: Several ongoing phase III studies are evaluating the efficacy of first-line atezolizumab in combination with chemotherapy in patients with non small-cell lung cancer (NSCLC). Among angiogenesis inhibitors, bevacizumab is approved as first-line therapy in combination with chemotherapy or in combination with erlotinib in patients with NSCLC harboring activating EGFR mutations. Recent evidence (Wallin 2016) suggests that the combination of atezolizumab and bevacizumab increases intra-tumoral CD8+T cells, suggesting that dual VEGF and PD-L1 inhibition improves antigen-specific T-cell migration. In addition, preliminary clinical data suggested a strong synergistic effects of bevacizumab with immune checkpoint inhibitors. There is therefore a strong rationale for investigating the combination of atezolizumab and bevacizumab in patients with advanced/metastatic NSCLC. The FoRT 05-BEAT is a multicenter, Italian, phase II, randomized study comparing atezolizumab monotherapy versus the combination of atezolizumab and bevacizumab in patients with chemo-naive metastatic NSCLC and high levels of PD-L1 expression. Methods: The trial was conducted in 35 Italian centers: chemotherapy naive metastatic NSCLC patients, with high levels of PD-L1 expression (PD-L1 TPS ≥50% or TC/IC 3 scoring) were randomly assigned to atezolizumab monotherapy (1200 mg every 3 weeks) or to the combination of atezolizumab (1200 mg every 3 weeks) and bevacizumab (15 mg/kg every 3 weeks).The primary endpoint is overall survival (OS) rate at 18 months. Secondary endpoints include response rate (RR), PFS, OS according to presence of bone and/or hepatic metastases. Safety considerations will be considered. Exploratory analysis of predictive biomarker on tumor tissue and blood samples has been planned. Sample size has been calculated assuming a 18 months OS of 50% in the atezolizumab arm. Therefore, a total of 186 patients is needed to detect an absolute improvement of 20%, thus obtaining a 18mOS of 70% in the combination arm, with a power of 80% at a significance level of 5%. Taking into account the percentage of patients lost-to-follow-up, the sample size has been increased by 10% (N = 206 patients, 103 per arm). At the drafting of this abstract, 57 patients have already been enrolled (47 randomized). Clinical trial information: NCT03896074.

A phase 1b/2 study of nanatinostat and valganciclovir in patients with advanced Epstein-Barr virus positive (EBV+) solid tumors and in combination with pembrolizumab in patients with recurrent/metastatic nasopharyngeal carcinoma (RM-NPC).

TPS6107 Background: Epstein-Barr virus (EBV) is linked to the development and pathogenesis of nasopharyngeal carcinoma (NPC). Most patients present with advanced stage disease at diagnosis (Li 2014); first-line chemoradiation is commonly followed by recurrence and poor prognosis emphasizing the need for new treatment options. Targeting EBV in NPC represents a novel therapeutic approach. EBV is predominantly latent in NPC; pre-clinical studies demonstrated that induction of the viral lytic phase by histone deacetylase inhibitors (HDACi) renders EBV+ tumor cells susceptible to the cytotoxic activity of ganciclovir (GCV) (Hui 2016). Nanatinostat (Nstat), a potent Class-I HDACi, induces the expression of the lytic BGLF4 protein kinase in EBV+ tumor cells, which activates the nucleoside analog GCV via phosphorylation. Phosphorylated GCV becomes incorporated into the cellular DNA causing chain termination and apoptosis. The all-oral combination of Nstat and valganciclovir (VGCV), a pro-drug of GCV, has demonstrated favorable safety and preliminary efficacy in a phase 1/2 study in patients with recurrent EBV+ lymphoma (NCT03397706). This phase 1b/2, open-label, multicenter study will evaluate the safety, tolerability, pharmacokinetics, and preliminary activity of Nstat + VGCV in patients with advanced EBV+ solid tumors. Additionally, the combination of pembrolizumab together with Nstat + VGCV will be evaluated in recurrent/metastatic NPC (RM-NPC) patients. NPC frequently exhibits high PD-L1 expression levels; however, PD-1 inhibitors resulted in limited response rates ranging from 20-30% in the RM-NPC setting. Methods: Phase 1b utilizes a 3+3 dose escalation design to determine the recommended Phase 2 dose (RP2D) of Nstat + VGCV in patients with EBV+ RM-NPC. In Phase 2, up to 60 patients with EBV+ RM-NPC will be randomized 1:1 to receive Nstat + VGCV at the RP2D with or without pembrolizumab to evaluate safety, tolerability, overall response rate, and potential pharmacodynamic markers of drug activity, including plasma EBV DNA levels. Additionally, patients with EBV+ solid tumors other than RM-NPC will receive Nstat + VGCV at the RP2D in a separate Phase 1b dose expansion cohort. Patients eligible for the phase 1b dose escalation and phase 2 will have EBV+ RM-NPC with 1-3 prior lines of platinum-based chemotherapy and no available curative therapies. Patients with advanced EBV+ non-NPC solid tumors (gastric cancer, lymphoepithelioma-like carcinoma, leiomyosarcoma) and no curative therapies are eligible for the phase 1b expansion cohort. All patients must have measurable disease per RECIST v1.1 and adequate bone marrow, liver, and renal function. Enrollment began in January 2022. Clinical trial information: NCT05166577.

Cell-Surface Programmed Death Ligand-1 Expression Identifies a Sub-Population of Distal Epithelial Cells Enriched in Idiopathic Pulmonary Fibrosis

Idiopathic lung fibrosis (IPF) is a fatal lung disease characterized by chronic epithelial injury and exhausted repair capacity of the alveolar compartment, associated with the expansion of cells with intermediate alveolar epithelial cell (AT2) characteristics. Using SftpcCreERT2/+: tdTomatoflox/flox mice, we previously identified a lung population of quiescent injury-activated alveolar epithelial progenitors (IAAPs), marked by low expression of the AT2 lineage trace marker tdTomato (Tomlow) and characterized by high levels of Pd-l1 (Cd274) expression. This led us to hypothesize that a population with similar properties exists in the human lung. To that end, we used flow cytometry to characterize the CD274 cell-surface expression in lung epithelial cells isolated from donor and end-stage IPF lungs. The identity and functional behavior of these cells were further characterized by qPCR analysis, in vitro organoid formation, and ex vivo precision-cut lung slices (PCLSs). Our analysis led to the identification of a population of CD274pos cells expressing intermediate levels of SFTPC, which was expanded in IPF lungs. While donor CD274pos cells initiated clone formation, they did not expand significantly in 3D organoids in AT2-supportive conditions. However, an increased number of CD274pos cells was found in cultured PCLS. In conclusion, we demonstrate that, similar to IAAPs in the mouse lung, a population of CD274-expressing cells exists in the normal human lung, and this population is expanded in the IPF lung and in an ex vivo PCLS assay, suggestive of progenitor cell behavior. CD274 function in these cells as a checkpoint inhibitor may be crucial for their progenitor function, suggesting that CD274 inhibition, unless specifically targeted, might further injure the already precarious lung epithelial compartment in IPF.

Identification of lncRNAs based on different patterns of immune infiltration in gastric cancer.

Gastric cancer is one of the most common malignant tumors in the world, which brings great challenges to people's life and health. The purpose of this study was to investigate immune related-lncRNAs and identify new biomarkers for the prognosis of gastric cancer (GC). We downloaded data from The Cancer Genome Atlas (TCGA) and used R software to determine the ESTIMATEScore, ImmuneScore, and StromalScore of each tumor sample. We performed prognostic analysis and identified the differentially expressed lnRNAs, which were then used to construct a prognostic model. Among the 44 hub genes in the competitive endogenous RNA (ceRNA) network, 3 differentially expressed genes were verified by qPCR. Based on the degree of immune infiltration, cluster A had a higher ESTIMATEScore, ImmuneScore, and StromalScore and higher expression levels of PD-L1 (CD274) and CTLA4 than cluster B. Univariate Cox analysis was conducted for these differential lncRNAs, and 57 lncRNAs were found to have prognostic value (P<0.05). gene cluster A had a worse prognosis than gene cluster B (P=0.021). Then, a prognostic model was constructed. The low-risk group had a significantly higher survival rate. Finally, the qPCR results showed that the expression levels of BMPER, PRUNE2, and RBPMS2 were low in GC cell lines. We identified a risk score of 19 lncRNAs as a prognostic marker of GC. There was a relationship between these 19 prognostic-related lncRNAs and the subtypes of infiltrating immune cells. An approach for predicting the prognosis of GC was therefore provided in this study.

Association of a Novel Prognosis Model with Tumor Mutation Burden and Tumor-Infiltrating Immune Cells in Thyroid Carcinoma.

Although immunotherapy has recently demonstrated a substantial promise in treating advanced thyroid carcinoma (THCA), it is not appropriate for all THCA patients. As a result, this study aims to identify biomarkers for predicting immunotherapy efficacy and prognosis in THCA patients based on a constructed prognostic model. The transcriptomic and corresponding clinical data of THCA patients were obtained from the Cancer Genome Atlas (TCGA) database. We identified differentially expressed genes (DEGs) between THCA and normal samples and performed an intersection analysis of DEGs with immune-related genes (IRGs) downloaded from the ImmPort database. Functional enrichment analysis was performed on the chosen immune-related DEGs. Subsequently, Cox and LASSO regression analyses were conducted to obtain three hub immune-related DEGs, including PPBP, SEMA6B, and GCGR. Following that, a prognostic risk model was established and validated based on PPBP, SEMA6B, and GCGR genes to predict immunotherapy efficacy and THCA prognosis. Finally, we investigated the association between the constructed risk model and tumor mutational burden (TMB), abundance of tumor-infiltrating immune cells (TICs) as well as immunotherapeutic targets (PDL-1, PD-1, and CTLA4) in THCA. THCA patients in the high-risk score (RS) group showed higher TMB levels and worse prognosis than the low RS group. Patients in the high-RS group had higher proportions of monocytes, M2 macrophages, and activated dendritic cells, whereas those in the low-RS group exhibited higher numbers of M1 macrophages and dendritic resting cells. Our data implied that the constructed THCA prognostic model was sound and we concluded that the THCA patients having high TMB and low PD-L1 expression levels might respond poorly to immunotherapy. Taken together, we constructed a novel prognostic model for THCA patients to predict their prognosis and immunotherapy efficacy, providing a viable option for the future management of THCA patients in the clinic.