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PD-L1 in pancreatic ductal adenocarcinoma: a retrospective analysis of 373 Chinese patients using an in vitro diagnostic assay.

ABSTRACT: BACKGROUND:Programmed death ligand 1 (PD-L1) has shown potential as a therapeutic target in numerous solid tumors. Its prognostic significance has also been established in pancreatic ductal adenocarcinoma (PDAC). The present study aimed to explore PD-L1 expression in PDAC cases in a large Chinese cohort using an in vitro diagnostic (IVD) assay to provide further insight into the potential value of programmed cell death protein 1 (PD-1) as a therapeutic target. METHODS:Three hundred seventy-three PDAC patients were retrospectively recruited in this study. Tissue microarray (TMA) blocks were made from available formalin-fixed and paraffin-embedded (FFPE) tumor and matched adjacent tissue specimens. We evaluated PD-L1 protein expression via immunohistochemistry (IHC) using a U.S. Food and Drug Administration (FDA)-approved IVD assay. The relationships between PD-L1 positivity and both clinicopathological characteristics and patient prognosis were analyzed. PD-1 expression and clinicopathological significance were also evaluated. RESULTS:PD-L1 and PD-1 positivity were observed in 3.2% and 7.5% of cases, respectively. PD-L1 showed a predominantly membranous pattern in tumor cells, while no positive PD-L1 staining was observed in normal regions. Statistical analyses revealed that PD-L1 expression was associated with lymph node metastasis. PD-L1 positivity was a prognostic indicator of progression-free survival (PFS) and overall survival (OS) in univariate analyses, but only PFS remained statistically significant in multivariate analysis. PD-1 expression was detected in lymphocytes and was not associated with any clinicopathological feature except a history of pancreatitis. CONCLUSIONS:The PD-L1 positivity rate is low in PDAC when evaluated using a companion diagnostic assay. It remains an independent prognostic factor for poor PFS.

SUBMITTER: Liang X

PROVIDER: S-EPMC6389094 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Immunotherapy targeting programmed cell death-1 (PD-1) has considerably improved the prognosis of patients with advanced cancers; however, its efficacy in the treatment of pancreatic ductal adenocarcinoma (PDAC) is unfavourable. To address the issue of PDAC immunotherapy, we investigated the expression of two PD-1 ligands, PD-L1 and PD-L2, in PDAC, analysed their role in survival, and explored their correlation with clinicopathological features, immune infiltration, and DNA damage response molecules. Immunohistochemistry was performed on 291 surgically resected PDAC samples. In tumour cells (TCs) and immune cells (ICs), the positivity of PD-L1 expression was 30 and 20% and that of PD-L2 expression was 40 and 20%, respectively. Moreover, PD-L1 expression on TCs correlated with its expression on ICs (p < 0.0001); a similar result was observed for PD-L2 (p < 0.0001). Nonetheless, no correlation was observed between PD-L1 and PD-L2 expression. Positive PD-L1 expression on TCs was related to N1 stage (p = 0.011) and AJCC II stage (p = 0.002), whereas positive PD-L2 expression on TCs was associated with high FOXP3+ cell infiltration (p = 0.001) and high BRCA2 expression (p < 0.0001). Survival analysis revealed that positive PD-L1 (p = 0.046) and PD-L2 (p = 0.028) expression on TCs was an independent risk factor for unfavourable disease-specific survival (DSS). Furthermore, positive PD-L2 expression on TCs was an independent risk factor for lower DSS in the pN0 (p = 0.023), moderate and well tumour differentiation (p = 0.004), low BRCA1 (p = 0.017), wild-type p53 (p = 0.034), and proficient mismatch repair (p = 0.004) subgroups. Moreover, post-operative adjuvant chemotherapy could significantly affect DSS, regardless of PD-L1/PD-L2 expression status (positive or negative) on TCs, while it only prolonged DSS in PDL1-ICs(-) (p < 0.0001) and PDL2-ICs(-) (p < 0.0001) subgroups. This study provides a comprehensive understanding of the roles of PD-L1 and PD-L2 in PDAC, supporting anti-PD-1 axis immunotherapy for PDAC.

Project description:IntroductionImmune checkpoint inhibitors (ICI), e.g., targeting programmed cell death protein 1-ligand 1 (PD-L1) or its receptor PD-1, have markedly improved the therapy of many cancers but so far failed in pancreatic ductal adenocarcinoma (PDAC). Macrophages represent one of the most abundant immune cell populations within the tumor microenvironment (TME) of PDAC being able to either support or restrain tumor progression depending on their phenotype. To better understand treatment failure of PD-L1/PD-1 inhibitors in PDAC, this study examined PD-L1 expression in the context of a dynamic TME in PDAC with a particular focus on the impact of macrophages.MethodsFormalin-fixed and paraffin embedded tissue samples of primary PDAC tissues and corresponding liver metastases were used for immunohistochemical analyses. Serial sections were stained with antibodies detecting Pan-Cytokeratin, CD68, CD163, CD8, and PD-L1.To investigate whether the PD-1/PD-L1 axis and macrophages contribute to immune escape of PDAC cells, a stroma enriched 3D spheroid coculture model was established in vitro, using different PDAC cell lines and macrophages subtypes as well as CD8+ T cells. Functional and flow cytometry analyses were conducted to characterize cell populations.ResultsImmunohistochemical analyses revealed that PD-L1 is mainly expressed by stroma cells, including macrophages and not PDAC cells in primary PDAC tissues and corresponding liver metastases. Notably, high local abundance of macrophages and strong PD-L1 staining were commonly found at invasion fronts of tumoral lesions between CD8+ T cells and tumor cells. In order to investigate whether PD-L1 expressing macrophages impact the response of PDAC cells to treatment with PD-L1/PD-1 inhibitors, we developed a spheroid model comprising two different PDAC cell lines and different ratios of in vitro differentiated primary M1- or M2-like polarized macrophages. In line with our in situ findings, high PD-L1 expression was observed in macrophages rather than PDAC cells, which was further increased by the presence of PDAC cells. The effector phenotype of co-cultured CD8+ T cells exemplified by expression of activation markers and release of effector molecules was rather enhanced by PDAC macrophage spheroids, particularly with M1-like macrophages compared to mono-culture spheroids. However, this was not associated with enhanced PDAC cell death. ICI treatment with either Durvalumab or Pembrolizumab alone or in combination with Gemcitabine hardly affected the effector phenotype of CD8+ T cells along with PDAC cell death. Thus, despite strong PD-L1 expression in macrophages, ICI treatment did not result in an enhanced activation and cytotoxic phenotype of CD8+ T cells.ConclusionOverall, our study revealed novel insights into the interplay of PDAC cells and macrophages in the presence of ICI.

Project description:Pancreatic ductal adenocarcinoma (PDA) is an extremely lethal malignancy arising from the pancreas. The treatment of PDA is complicated by ineffective treatments and a lack of biomarkers predictive of treatment success. We have designed a patient-derived organoid (PDO) based high-throughput drug screening assay to model treatment response to a variety of conventional and investigational treatments for PDA. Consecutive patients undergoing endoscopic ultrasound-guided fine-needle biopsy for tissue diagnosis of PDA at Rush University Medical Center were offered to participate in the study. Biopsies were immediately processed to develop organoids. Fifteen PDOs were screened for sensitivity to 18 compounds, including conventional PDA chemotherapies and FDA-approved investigational targeted therapies in cancer using Cell-titer GLO 3D (Promega) cell viability assay. The area under the curve (AUC) was calculated and normalized to the maximum area under the curve to generate a normalized AUC between 0 and 1. Molecular profiling of PDOs was conducted using RNA-seq. Human PDA transcriptomic was extracted from The Cancer Genome Atlas (TCGA). The drug response curves were reproducible. We observed variation in response to conventional therapies overall as well as among individual patients. There were distinct transcriptome signatures associated with response to the conventional chemotherapeutics in PDA. The transcriptomic profile of overall resistance to conventional therapies in our study was associated with poor survival in PDA patients in TCGA. Our pathway analysis for targeted drugs revealed a number of predictors of response associated with the mechanism of action of the tested drug. The multiplex organoid-based drug assay could be used in preclinical to inform patient stratification and therapeutic selection in PDA. When combined with omics data, ex vivo response to treatment could help identify gene signatures associated with response to novel therapies.

Project description:IntroductionPancreatic ductal adenocarcinoma (PDAC) represents the 4th most common cause of cancer-related deaths in Western countries. Most patients are diagnosed at advanced stages, often already with metastases. The main site of metastasis is the liver and hepatic myofibroblasts (HMF) play a pivotal role in metastatic outgrowth. Immune checkpoint inhibitors (ICI) targeting programmed death ligand 1 (PD-L1) or programmed cell death protein 1 (PD-1) improved treatment of several cancers but not of PDAC. Therefore, this study aimed to better understand the impact of HMF on PD-L1 expression and immune evasion of PDAC cells during liver metastasis.MethodsFormalin-fixed and paraffin embedded biopsy samples or diagnostic resection specimens from liver metastases of 15 PDAC patients were used for immunohistochemical analyses. Serial sections were stained with antibodies directed against Pan-Cytokeratin, αSMA, CD8, and PD-L1. To investigate whether the PD-1/PD-L1 axis and HMF contribute to immune escape of PDAC liver metastases, a stroma enriched 3D spheroid coculture model was established in vitro, using two different PDAC cell lines, HMF, and CD8+ T cells. Here, functional and flow cytometry analyses were conducted.ResultsImmunohistochemical analysis of liver tissue sections of PDAC patients revealed that HMF represent an abundant stroma population in liver metastases, with clear differences in the spatial distribution in small (1500 µm) and large (> 1500 μm) metastases. In the latter, PD-L1 expression was mainly located at the invasion front or evenly distributed, while small metastases either lacked PD-L1 expression or showed mostly weak expression in the center. Double stainings revealed that PD-L1 is predominantly expressed by stromal cells, especially HMF. Small liver metastases with no or low PD-L1 expression comprised more CD8+ T cells in the tumor center, while large metastases exhibiting stronger PD-L1 expression comprised less CD8+ T cells being mostly located at the invasion front. HMF-enriched spheroid cocultures with different ratios of PDAC cells and HMF well mimicking conditions of hepatic metastases in situ. Here, HMF impaired the release of effector molecules by CD8+ T cells and the induction of PDAC cell death, an effect that was dependent on the amount of HMF but also of PDAC cells. ICI treatment led to elevated secretion of distinct CD8+ T cell effector molecules but did not increase PDAC cell death under either spheroid condition.ConclusionOur findings indicate a spatial reorganization of HMF, CD8+ T cells, and PD-L1 expression during progression of PDAC liver metastases. Furthermore, HMF potently impair the effector phenotype of CD8+ T cells but the PD-L1/PD-1 axis apparently plays a minor role in this scenario suggesting that immune evasion of PDAC liver metastases relies on other immunosuppressive mechanisms.

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PD-L1 in pancreatic ductal adenocarcinoma: a retrospective analysis of 373 Chinese patients using an in vitro diagnostic assay.

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