High Levels Of Programmed Death Ligand-1 Research Articles

Expression of alternatively spliced PD-L1 RNA with skipping of transmembrane domain.

e14596 Background: Programmed death-ligand 1 (PD-L1) is a transmembrane ligand for the programmed cell death protein 1 (PD-1), a receptor that inhibits T-cell activity. The PD-L1/PD-1 immune checkpoint axis has been successfully targeted to enhance antitumor immune responses. This gene is composed of seven exons. Exons 1-4 are responsible for the extracellular domain, exon 5 for the transmembrane domain, and exons 6-7 for the intracellular domain. Emerging data indicates that alternative splicing in genes is an important mechanism for expression of different isoforms that may influence antibody-based therapies. We explored the potential of the presence of alternative splicing in PD-L1 transcripts via sequencing of PD-L1 RNA in patients with various type of solid tumor cancers. Methods: RNA was extracted from 3443 FFPE solid tumor samples including lung, colon, pancreas, breast, and other cancer types. In addition, cfRNA was extracted from 532 peripheral blood plasma samples from patients with solid tumor cancers. RNA was sequenced using a hybrid capture targeted RNA panel on FFPE samples and peripheral blood with analysis focused on PD-L1 gene transcript. Quantification of RNA transcript was done using Salmon algorithm. Results: Of the 3443 FFPE samples, 38 (1.2%) samples showed alternative splicing involving deletion of exon 5 (PD-L1∆Ex5). The alternative splicing was in 2.10% of lung cancer, 2% of colon cancer, 0.70% of pancreatic cancer, 1.40% of gastric cancer, 0.38% of breast cancer, 0.30% of ovarian cancer, 1.10% of sarcoma cases, 1.50% of glioblastomas, 3.30% of thyroid cancer and 2% of urothelial cancer. Of the 532 cfRNA samples, 20 (3.7%) showed PD-L1∆Ex5. The median percentage of PD-L1∆Ex5 transcripts was 2.2% of total PD-L1 transcripts in tissue samples as compared with 6.6% in cfDNA samples. In tissue samples, there was a correlation between levels of PD-L1 and PD-L1∆Ex5 (r = .567, p < .001). In contrast, cfRNA showed no significant correlation between levels of PD-L1 and levels of PD-L1∆Ex5 (r = .268). Conclusions: PD-L1 exon 5 skipping was detected in a number of patients with different types of solid tumor cancers. The percentage of skipping transcripts is low in cells and relatively higher in cfRNA. Since exon 5 skipping deletes the transmembrane domain, the PD-L1 protein may remain in the cytoplasm or perhaps is secreted as cell-free PD-L1 protein. The demonstration of relatively higher levels of PD-L1∆Ex5 in cfRNA is likely reflecting higher turnover of cells and raises the possibility that cells with this isoform of PD-L1 are more aggressive than cells without the expression of this isoform. Further studies are needed to explore the clinical relevance of the expression of such abnormal PD-L1 protein on treatment with immune checkpoint inhibitors.

Intraoral salivary lymphoepithelial carcinoma: clinicopathologic and immunophenotypic characterization of 3 cases indicates elevated programmed death-ligand 1 expression

Intraoral salivary lymphoepithelial carcinoma (ISLEC) is a rare malignancy with programmed death-ligand 1 (PD-L1) expression levels that have been greatly understudied. We examined the clinicopathologic and immunophenotypic characteristics, including PD-L1 levels, of 3 cases of ISLEC. We searched the archives of 2 oral and maxillofacial pathology laboratories for specimens diagnosed as ISLEC between 1985 and 2022. We collected patient demographic and clinical data. Immunostaining for AE1/AE3, CK7, CD3, CD20, p16, p53, Ki67, and PD-L1 (SP263), as well as Epstein-Barr virus-encoded small RNAs (EBER) in situ hybridization (ISH) were performed. All 3 cases affected males aged 42 to 84 years (median=61y) and involved the floor of the mouth, soft palate/uvula, and tongue. The lesions showed diffuse infiltration by non-keratinizing sheets and islands of undifferentiated carcinoma cells with associated dense lymphoplasmacytic inflammation. Immunohistochemically, all tumors showed AE1/AE3 positivity, selective p53 staining, and negativity for CK7 and p16. Ki67 highlighted 20%-80% of lesional cells. The inflammatory infiltrate comprised a mixed population of T and B lymphocytes. EBER ISH was positive in one case. All ISLECs displayed membranous, focal-to-diffuse, PD-L1 staining with tumor proportion score > 95% in two and 40-50% in the third case. The clinicopathologic and immunophenotypic characteristics of the cases we examined highlight the rarity of ISLEC and indicate overall high PD-L1 levels in this type of malignancy, rendering patients with ISLEC potential candidates for targeted α-PD-1/PD-L1 immunotherapy.

Programmed death-ligand 1 and tumor-infiltrating lymphocytes (TILs) - low TIL density may predict poorer long-term prognosis in T1 laryngeal cancer.

We evaluated the prognostic role of programmed death-ligand 1 (PD-L1) and tumor-infiltrating lymphocytes (TILs) in T1 glottic laryngeal squamous cell carcinoma (LSCC). T1 glottic LSCC patients (n = 174) treated at five Finnish university hospitals between 2003 and 2013 were included. Tissue microarray (TMA) blocks were used for PD-L1 immunohistochemistry. TILs were scored from intratumoral and stromal regions in whole tissue sections. Of 174 patients, 92 (53%) had negative, 66 (38%) intermediate, and 16 (9%) high PD-L1 levels. Of 80 patients whose TILs were analyzed, 50 (63%) had low and 30 (38%) high stromal TIL density. Patients with a local recurrence or a new primary tumor of the larynx had lower TIL density than had other patients (p = 0.047). High PD-L1 expression with low stromal TIL density was associated with inferior 5-year disease-specific survival (85% vs. 100%, p = 0.02). In conclusion, in patients treated for T1 glottic LSCC, low stromal TIL density was associated with local recurrences and new primary tumors of the larynx. High PD-L1 expression with low stromal TIL density may be associated with worse survival in T1 glottic LSCC.

Abstract 6543: Comparative analysis of immune landscape according to EGFR mutation and tumor PD-L1 proportion scores in non-small cell lung cancer at single cell resolution

Abstract Introduction: In EGFR-mutant non-small cell lung cancer (NSCLC), the efficacy of immune checkpoint blockade (ICB) is limited with higher incidence of toxicity profiles. EGFR-mutant NSCLC is associated with immune suppressive microenvironment, including lower programmed death-ligand 1 (PD-L1), tumor mutational burden (TMB), and tumor-infiltrating CD8+ T lymphocytes. However, tumors expressing high levels of PD-L1 show improved and durable clinical benefit in immunotherapy. In addition, Understanding the dynamics of tumor microenvironment may elucidate immune response in terms of immune composition, and help overcome resistance to immune checkpoint inhibitors. Here, we demonstrate that conventional type 1 dendritic cells (cDC1s) is one of the key tumor microenvironment, resulting in priming of CD8+ T cells and production of chemokines, such as CXCL9/CXCL10 and cytokines that augment anti-tumor immunity. Methods: A total of 31 surgical samples of NSCLC, including EGFR wild and mutant NSCLC of stages I-III, and 16 normal lung tissue samples were collected. PD-L1 expression in EGFR wild type was categorized into tumor proportion score (TPS) of < 50% and ≥ 50% using immunohistochemistry (IHC). Single cell RNA sequencing was performed with the Scanpy toolkit on the retrospective cohorts of normal (n=16), EGFR mutant (n=14, L858R=6, E19Del=8), EGFR wild TPS ≥ 50% (n=10), and TPS < 50% (n=7). Results: We observed that an overall tumor specific immune response occurred in EGFR wild-type TPS ≥ 50% compared with others. XCL1+ innate-like T cells, which act as an orchestra of tumor specific immune response, were found. These cells had high expression of FCER1G and IL2RB, a receptor for IL-15, However, expression of CD8A/B, CD4 and FCGR3A was low. Interestingly, in these innate-like T cells, the expression of ITGAE, which interacts with E-cadherin in cancer cells, and CD81, which interacts with CD3 and enhances immunological synapse, was observed in EGFR wild TPS ≥ 50%. According to these results, XCR1+ cDC1s were enriched in EGFR wild-type TPS ≥ 50%. CXCL9/CXCL10, which recruits CXCR3+ effector T cells which are primed at the lymph node, was highly expressed in activated DCs at EGFR wild TPS ≥ 50%. CD8+ T cells were significantly enriched in EGFR wild-type TPS ≥ 50%. These cells also expressed IFNG and FLT3LG, which maintain the function of intra-tumoral dendritic cells (DCs), and were also up-regulated in EGFR wild-type TPS ≥ 50%. Among them, exhausted CD8+ T cells expressing PDCD1 were observed in EGFR wild TPS ≥ 50%, and these cells also expressed ITGAE and CD81. In addition, these cells also showed high expression of CXCR3, a receptor for CXCL9/CXCL10 which were expressed in the activated DCs. Conclusion: cDC1s-mediated tumor specific immune response occurred in patients with EGFR wild TPS ≥ 50%, which could be an important mechanism to predict anti-PD-1 response. Citation Format: YoungJoon Park, Su-Jin Choi, Jii Bum Lee, Dong Kwon Kim, Sun Min Lim, Seong Gu Heo, Ke Jia Qi, Byoung Chul Cho, Kyoung-Ho Pyo. Comparative analysis of immune landscape according to EGFR mutation and tumor PD-L1 proportion scores in non-small cell lung cancer at single cell resolution [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 6543.

PD-L1: expression regulation.

Programmed death-ligand 1 (PD-L1), expressed on the surface of tumor cells, can bind to programmed cell death-1 (PD-1) on T cells. The interaction of PD-1 and PD-L1 can inhibit T-cell responses by decreasing T-cell activity and accelerating their apoptosis. Various cancers express high levels of PD-L1 and exploit PD-L1/PD-1 signaling to evade T-cell immunity, and immunotherapies targeting the PD-1/PD-L1 axis have been shown to exert remarkable anti-tumor effects; however, not all tumor patients benefit from these therapies. Therefore, study of the mechanisms regulating PD-L1 expression are imperative. In this review, we explore regulation of PD-L1 expression in the contexts of gene transcription, signaling pathways, histone modification and remodeling, microRNAs, long noncoding RNAs, and post-translational modification. Current developments in studies of agents that block PD-L1 and correlations between immunotherapies targeting PD-1/PD-L1 and PD-L1 expression are also summarized. Our review will assist in understanding of PD-L1 expression regulation and discusses the implications of reported findings in cancer diagnosis and immunotherapy.

PD-L1 on Circulating Tumor Cells Indicates Poor Prognosis in Breast Cancer

Background: In breast cancer, research concerning programmed death-ligand 1 (PD-L1) on circulating tumor cells (CTCs) is limited. Besides, the best cell type for determining the prognostic and predictive values of PD-L1 has not been established. This study investigated associations between PD-L1 on tumor cells (TCs), CTCs, and tumor-infiltrating immune cells (TIICs), respectively, and prognosis and clinicopathological features in breast cancer. Methods: Twenty patients with breast cancer were recruited; one was excluded for confirmed lymphoma. The PD-L1 on TCs and TIICs was determined via immunohistochemistry; PD-L1 mRNA expression on CTCs was analyzed. The chi-squared test and Kaplan-Meier and Cox proportional hazards model analyses were applied. Results: The median follow-up time was 60 months. Of the 19 patients, 14 had >1 CTC/10 mL peripheral blood. Among these, each had ≥1 CTC showing PD-L1. At baseline, there was no difference between groups with or without metastasis regarding CTCs and PD-L1 expression. Patients with high PD-L1 levels on CTCs had poor overall survival (p = 0.034). In the multivariate analysis, PD-L1 levels on CTCs and T stage were independent prognostic factors (p = 0.029, 0.010, respectively). Conclusions: High levels of PD-L1 on CTCs may prognose shorter overall survival in breast cancer.

Does Elevated Pre-Treatment Plasma PD-L1 Level Indicate an Increased Tumor Burden and Worse Prognosis in Metastatic Colorectal Cancer?

Background: Programmed death-ligand 1 (PD-L1) and programmed cell death protein 1 (PD-1) have been reported as possibly favorable prognostic factors in colorectal cancer (CRC). However, their longitudinal effect is unknown. Methods: A pilot study was performed to investigate whether baseline PD-1/PD-L1 levels are associated with further laboratory changes and/or shorter survival. Results: A total of 506 laboratory measurements from 37 metastatic CRC patients were analyzed. The baseline plasma PD-1 and PD-L1 levels were 27.73 ± 1.20 pg/mL and 16.01 ± 1.09 pg/mL, respectively. Disease progression (p = 0.0443) and baseline high-sensitivity C-reactive protein (p = 0.0011), aspartate transaminase (p = 0.0253), alanine transaminase (p = 0.0386), and gamma-glutamyl transferase (p = 0.0103) were associated with higher PD-L1 levels. Based on the baseline PD-1/PD-L1 levels, low and high PD-1/PD-L1 groups were created. Constant, pathological levels of complete blood count values, high-sensitivity C-reactive protein, serum albumin, high-density lipoprotein cholesterol, and lactate dehydrogenase were characteristic for patients with high baseline PD-L1. High PD-L1 levels were significantly associated with increased tumor burden. Disease-specific survival and progression-free survival were significantly shorter in patients with high PD-L1. Conclusions: Abnormal levels of laboratory parameters and intensified tumor burden can be expected if elevated baseline plasma PD-1/PD-L1 levels are found.

Antitumor activity of bintrafusp alfa in previously treated patients with recurrent or metastatic nasopharyngeal cancer (NPC): A single arm, prospective phase II trial.

e18029 Background: Patients with recurrent or metastatic nasopharyngeal cancer (R/M NPC) who failed platinum-based chemotherapy have poor prognoses. We report the clinical activity and safety of bintrafusp alfa, a first-in-class bifunctional fusion protein composed of the extracellular domain of the transforming growth factor (TGF)-βRII receptor fused to a human IgG1 antibody blocking programmed death-ligand 1 (PD-L1), in patients with heavily pretreated R/M NPC. Tumor and plasma-based biomarkers were investigated in an exploratory analysis. Methods: Eligible patients had histologically confirmed NPC that had recurred at distant sites and were not amenable to curative treatment. All patients received at least one prior line of platinum-based chemotherapy for recurrent disease. Patients were treated with bintrafusp alfa (1200mg every 2 weeks) until disease progression. The sample size was estimated to assume a 40% objective response rate (ORR) to bintrafusp alfa compared with 20% for checkpoint inhibitors. Modified Simon two-stage optimal design was used (power, 80%; a = 0.05; P0 = 0.20; P1 = 0.40; n1 = 18; n = 33 with an additional five patients to allow for ineligibility or other reasons). The primary endpoint was ORR and secondary endpoints included survival and toxicity. Expression of PD-L1 in archived tumors, plasma clearance of Epstein-Barr virus (EBV) DNA, plasma clearance of TGF-β, and exosomal PD-L1 were assessed for a potential correlation with ORR. (NCT 04396886). Results: Out of 43 patients screened, 38 patients were enrolled. After a median follow-up of 14.9 months (range: 1.6-23.3 months), the confirmed ORR was 23.7% (95% CI: 12.4-38.8%) (complete response, n = 1; partial response, n = 8). The median treatment duration was 1.8 months (range: 0.5-14.3 months). 8 patients (21.1%) and 2 patients (5.3%) received bintrafusp alfa for > 6 months and > 12 months respectively. The 1-year overall survival (OS) rate was 57.5% (95% CI, 40.2% to 71.5%) and 1-year progression-free survival rate was 23% (95% CI, 10.1% to 39.4%). ORR was higher in patients with a decreasing trend in EBV-DNA at week 4 (40% vs. 6.3%, p = 0.02), whereas high exosomal PD-L1 levels at week 4 were predictive of worse ORR (5.3% vs. 41.7%, p = 0.012). There were no associations between clinical outcome and tissue PD-L1 expression (p = 0.952) or plasma TGF-β clearance (p = 0.28). 16 patients (42.4%) experienced ≥ grade 3 treatment-related adverse events, most commonly anemia (n = 9, 23.7%) and secondary malignancies (n = 4, 10.5%). Conclusions: Bintrafusp alfa has promising activity in heavily pretreated R/M NPC and a favorable 1-year OS rate, though the observed activity was not as high as the study initially aimed. The biomarker results warrant validation in larger cohorts. Clinical trial information: NCT04396886.

NDRG2 Expression in Breast Cancer Cells Downregulates PD-L1 Expression and Restores T Cell Proliferation in Tumor-Coculture.

Simple SummaryN-myc downstream-regulated gene 2 (NDRG2) is a candidate tumor suppressor in various cancers, including breast cancer. Increased expression of programmed death ligand 1 (PD-L1) is frequently observed in human cancers. Despite its role in cancer cells, the effects of NDRG2 on PD-L1 expression and PD-L1-PD-1 pathway disruption have not been investigated. We demonstrated that NDRG2 overexpression inhibits PD-L1 expression in human breast cancer cells. Blocking T cell proliferation by coculture with 4T1 mouse tumor cells that express high levels of PD-L1 could be significantly reversed by NDRG2 overexpression in the same tumor cells. NDRG2 knockdown in NDRG2-transfected cells elicited the upregulation of PD-L1 expression and accelerated the inhibition of T cell proliferation. These findings were confirmed from The Cancer Genome Atlas (TCGA) data that PD-L1 expression in basal and triple-negative breast cancer (TNBC) patients, but not in luminal A or B cancer patients, was negatively correlated with the NDRG2 expression.(1) Background: The aim of the present study was to evaluate the effect of NDRG2 expression in regulating PD-L1 or PD-L2 on malignant breast cancer cells. (2) Methods: Overexpression and knockdown of the NDRG2 gene in human and mouse cancer cells were applied and quantitative real-time PCR and Western blot analysis were performed. T cell proliferation and TCGA analysis were conducted to validate negative correlation of the PD-L1 expression with the NDRG2 expression. (3) Results: We found that NDRG2 overexpression inhibits PD-L1 expression in human breast cancer cells through NF-κB signaling. NDRG2 overexpression in 4T1 mouse breast cancer cells followed by PD-L1 downregulation could block the suppressive activity of cancer cells on T cell proliferation and knockdown of NDRG2 expression enhanced the expression of PD-L1, leading to the inhibition of T cell proliferation by tumor cell coculture. Finally, we confirmed from TCGA data that PD-L1 expression in basal and triple-negative breast cancer patients was negatively correlated with the expression of NDRG2. Intriguingly, linear regression analysis using TNBC cell lines showed that the PD-L1 level was negatively associated with the NDRG2 expression level. (4) Conclusions: Our findings demonstrate that NDRG2 expression is instrumental in suppressing PD-L1 expression and restoring PD-L1-inhibited T cell proliferation activity in TNBC cells.

PD-L1 sustains chronic, cancer cell–intrinsic responses to type I interferon, enhancing resistance to DNA damage

Programmed death ligand 1 (PD-L1), an immune-checkpoint protein expressed on cancer cells, also functions independently of the immune system. We found that PD-L1 inhibits the killing of cancer cells in response to DNA damage in an immune-independent manner by suppressing their acute response to type I interferon (IFN; IFN-I). In addition, PD-L1 plays a critical role in sustaining high levels of constitutive expression in cancer cells of a subset of IFN-induced genes, the IFN-related DNA damage resistance signature (IRDS) which, paradoxically, protects cancer cells. The cyclic GMP-AMP synthase-stimulator of the IFN genes (cGAS-STING) pathway is constitutively activated in a subset of cancer cells in the presence of high levels of PD-L1, thus leading to a constitutive, low level of IFN-β expression, which in turn increases IRDS expression. The constitutive low level of IFN-β expression is critical for the survival of cancer cells addicted to self-produced IFN-β. Our study reveals immune-independent functions of PD-L1 that inhibit cytotoxic acute responses to IFN-I and promote protective IRDS expression by supporting protective chronic IFN-I responses, both of which enhance the resistance of cancer cells to DNA damage.

Pretreatment Glasgow prognostic score predicts survival among patients with high PD‐L1 expression administered first‐line pembrolizumab monotherapy for non‐small cell lung cancer

BackgroundThere are no established biomarkers for predicting the efficacy of first‐line pembrolizumab monotherapy in patients with high programmed death‐ligand 1 (PD‐L1) expression. In this study, we investigated whether the Glasgow prognostic score (GPS), neutrophil‐to‐lymphocyte ratio (NLR), and body mass index (BMI) can be used to evaluate the effect of first‐line pembrolizumab monotherapy in patients with advanced non‐small cell lung cancer (NSCLC) who express high levels of PD‐L1.MethodsWe reviewed data from 142 patients with high PD‐L1 expression who underwent first‐line pembrolizumab monotherapy for NSCLC at six Japanese institutions between February 2017 and June 2019 and assessed the prognostic value of the GPS, NLR, and BMI. The Kaplan–Meier method and Cox proportional hazard models were used to examine differences in progression‐free survival (PFS) and overall survival (OS). The GPS, NLR, and BMI were calculated using C‐reactive protein and albumin concentrations, neutrophil and lymphocyte counts, and body weight and height, respectively.ResultsThe GPS independently predicted the first‐line pembrolizumab monotherapy efficacy, as a good GPS (GPS 0–1) was associated with a significantly better PFS and OS compared to a poor GPS (GPS 2) (PFS: 11.8 vs. 2.9 months, p < 0.0001; OS: not reached vs. 8.3 months, p < 0.0001). Furthermore, BMI independently predicted efficacy, as patients with high BMI (BMI ≥21.4) exhibited significantly better OS compared to those with low BMI (BMI <21.4) (OS: not reached vs. 14.1 months, p = 0.006).ConclusionsAmong patients with high PD‐L1 expression undergoing first‐line pembrolizumab monotherapy for NSCLC, the GPS is significantly correlated with both PFS and OS, and BMI with OS, indicating that they could be used to predict treatment outcome in these patients. To the best of our knowledge, this is the first study to assess the relationship among the GPS, NLR, and BMI and survival among patients with high PD‐L1 expression undergoing first‐line pembrolizumab monotherapy for NSCLC.

Nested and adjacent subgroups in cancer clinical trials: When the best interests of companies and patients diverge

The design of clinical trials with outcomes reported in cohorts including nested subgroups is common in novel agents seeking new indications for approval. This structure represents a tension between drug companies that have an incentive to pursue broad biomarker-agnostic approvals and patients whose best interest is to identify the subgroup(s) most likely to benefit from the drug. Programmed death ligand 1 (PD-L1) and checkpoint inhibitors are a prominent example with early trials reporting efficacy of checkpoint inhibitors in cohorts with high levels of PD-L1. Subsequent analyses incrementally report outcomes in broader patient cohorts that include the nested subgroup of high PD-L1 expression which drives the positive outcome in the entire cohort. Comparing aggregate outcomes between groups of patients with known heterogeneous outcomes deters the effective analysis of all available data. Exploring the optimal treatment for individual patients with different levels of PD-L1 expression, whether it is checkpoint inhibitors only, checkpoint inhibitors combined with chemotherapy or chemotherapy only, requires a granular approach to trial design and reporting. Such grouping of patients with different biomarker findings is increasingly seen in the setting of adjuvant therapy, as well as in targeted therapies that show efficacy in a single gene mutation which however are studied in the setting of panels of mutations. Here we discuss the difference between nested and adjacent subgroups in oncology.

Impact of tumor programmed death ligand-1 expression on osimertinib efficacy in untreated EGFR-mutated advanced non-small cell lung cancer: a prospective observational study.

BackgroundOsimertinib monotherapy is currently the standard of care as a first-line treatment for patients harboring epidermal growth factor receptor (EGFR) mutations; however, some EGFR-mutated non-small cell lung cancer (NSCLC) patients exhibit primary resistance and an insufficient response to EGFR-tyrosine kinase inhibitors (EGFR-TKIs). Elevated programmed death-ligand 1 (PD-L1) expression in tumors was reported as a negative predictive factor for outcomes of first- or second-generation EGFR-TKIs.MethodsWe prospectively assessed advanced NSCLC patients with EGFR mutations who were treated with osimertinib at 14 institutions in Japan between September 2019 and December 2020. Relationships between outcomes of osimertinib monotherapy and patients’ characteristics were reviewed.ResultsSeventy-one patients who underwent the tumor PD-L1 test were enrolled. Multivariate analysis identified tumor PD-L1 expression as an independent predictor for progression-free survival (PFS) with osimertinib treatment (P=0.029). The objective-response and disease-control rates for osimertinib treatment were significantly lower in patients demonstrating elevated PD-L1 levels relative to those with low or negative PD-L1 level (P=0.043 and P=0.007, respectively). Furthermore, among patients treated with osimertinib, those with high PD-L1 levels exhibited shorter PFS relative to those with low plus negative PD-L1 level (median PFS: 5.0 vs. 17.4 months; P<0.001).ConclusionsElevated tumor PD-L1 expression is associated with poor outcomes of osimertinib monotherapy in previously untreated advanced NSCLC patients with EGFR mutation. Further clinical trials are warranted to accumulate evidence demonstrating the effectiveness of combination therapy with osimertinib for EGFR-mutated advanced NSCLC patients with elevated tumor PD-L1 expression.Trial RegistrationUMIN000043942.

PD-L1 combined with HDAC9 is a useful prognostic predictor in hepatocellular carcinoma.

BackgroundThe expression of programmed death-ligand 1 (PD-L1) is associated with the response of patients to PD-1/PD-L1 blockade immunotherapy. It has been demonstrated that histone deacetylase (HDAC) inhibitors may alter the expression of PD-L1/PD-L2 and enhance the antitumor immune responses. However, the profile of PD-L1 expression and its association with HDACs in hepatocellular carcinoma (HCC) has not been accurately investigated.MethodsThe expression of PD-L1 and HDACs were examined by immunohistochemical (IHC) staining using tissue microarray (TMA) of 109 HCC specimens. Expression data from TCGA database and IHC staining of TMA were used for correlation analysis. Survival rates were analyzed based on data from 109 HCC patients.ResultsWe found that PD-L1 was upregulated in the majority of HCC samples. Furthermore, correlation analysis revealed that PD-L1 expression was positively correlated with HDAC9 and HDAC2 expression in HCC. Survival analysis showed that high levels of PD-L1 combined with increased expression of HDAC9 decreased overall survival (OS) in patients with HCC. In addition, univariate and multivariate analysis further suggested that the increased PD-L1/HDAC9 expression was an independent prognostic biomarker in HCC. HDAC9 overexpression promoted HCC growth and PD-L1 expression.ConclusionsThe results of the current study highlighted the strong association between HDACs with immunotherapy, thus providing a rational basis for combining HDAC9 specific inhibitors and PD-1 blockade in a future clinical approach for HCC.

Overexpression of PVR and PD-L1 and its association with prognosis in surgically resected squamous cell lung carcinoma

Targeting T-Cell Immunoreceptor with Ig and ITIM domain-poliovirus receptor (PVR) pathway is a potential therapeutic strategy in lung cancer. We analyzed the expression of PVR and programmed death ligand-1 (PD-L1) in surgically resected squamous cell lung carcinoma (SQCC) and determined its prognostic significance. We collected archival surgical specimens and data of 259 patients with SQCC at Yonsei Cancer Center (1998–2020). Analysis of variance was used to analyze the correlations between PVR and PD-L1 expression and patient characteristics. Kaplan–Meier curves were used to estimate recurrence-free survival (RFS) and overall survival (OS). Most patients were male (93%); the majority were diagnosed with stage 1 (47%), followed by stage 2 (29%) and stage 3 (21%). Overexpression of PVR resulted in a significantly shorter median RFS and OS (P = 0.01). PD-L1 expression was not significant in terms of prognosis. Patients were subdivided into four groups based on low and high PVR and PD-L1 expression. Those expressing high levels of PVR and PD-L1 had the shortest RFS (P = 0.03). PVR overexpression is associated with a poor prognosis in surgically resected SQCC. Inhibition of PVR as well as PD-L1 may help overcome the lack of response to immune checkpoint monotherapy.

Diagnostic Value of Vaginal Microecology, Serum miR-18a, and PD-L1 for Identifying HPV-Positive Cervical Cancer.

Objective:We aimed to investigate the diagnostic value of the vaginal microecology, serum miR-18a, and programmed death ligand-1 (PD-L1) for human papillomavirus (HPV)-positive cervical cancer.Methods:Eighty-four patients with HPV-positive cervical cancer were assigned to the observation group, 107 HPV-positive patients without cervical cancer were assigned to the positive group, and 191 healthy women were assigned to the control group. Vaginal microecology and serum levels of miR-18a and PD-L1 on the surface of CD4+ and CD8+ T cells were compared among the 3 groups. The observation group was further divided into subgroups according to patients’ characteristics for comparison. The diagnostic value of miR-18a and PD-L1 for HPV-positive cervical cancer was investigated.Results:Women in the control group had better vaginal microecology and lower levels of miR-18a and PD-L1 than those in the observation and the positive groups (all P < 0.05). Compared with the positive group, the observation group had similar vaginal microecology (all P > 0.05) but higher levels of miR-18a and PD-L1 (all P < 0.05). Moreover, the patients at stage III had higher levels of miR-18a and PD-L1 than those at stage I and II (all P < 0.05). The values of area under the curve for miR-18a and PD-L1 in the diagnosis of HPV-positive cervical cancer were over 0.8 (all P < 0.001).Conclusion:Patients with HPV-positive cervical cancer have vaginal microbial dysbiosis and high serum levels of miR-18a and PD-L1. miR-18a and PD-L1 have diagnostic value for identifying HPV-positive cervical cancer.

Effect of PD-L1 and tumor regression rate on prognosis of nasopharyngeal carcinoma.

To explore the expression level of programmed death ligand-1 (PD-L1) and the tumor regression rate after chemoradiotherapy, and to investigate their influence on the prognosis of patients with nasopharyngeal carcinoma. Eighty-six patients, who were initially diagnosed with non-metastatic nasopharyngeal carcinoma in the Second Xiangya Hospital of Central South University from January 2014 to June 2014, were recruited. Positive expression of PD-L1 in prepared sections of nasopharyngeal carcinoma tissues was detected by immunohistochemical staining. The clinical data of recruited patients and the tumor regression rate were calculated before chemoradiotherapy and 1 month after chemoradiotherapy. In addition, the potential influence of PD-L1 level and the tumor regression rate after chemoradiotherapy on therapeutic efficacy and the prognosis of nasopharyngeal carcinoma were assessed. Among the 86 patients with nasopharyngeal carcinoma, there were 30 patients with high expression of PD-L1 in nasopharyngeal carcinoma sections (34.88%), while there were 56 patients with low expression (65.12%). Expression levels of PD-L1 in nasopharyngeal carcinoma tissues were not related with sex, age, smoking history, concurrent chemoradiotherapy, pathological type, N staging, tumor staging, and pre-treatment EBV-DNA load in peripheral blood in the affected patients (all P>0.05). But the expression level of PD-L1 was related to the tumor regression rate and T staging in the patients with nasopharyngeal carcinoma (both P<0.05). The expression levels of PD-L1 in nasopharyngeal carcinoma tissues was a risk factor for tumor regression rate (OR=0.226, P<0.05). Compared with the patients with low expression levels of PD-L1, there was shorter disease-free survival in the patients with high expression levels of PD-L1 (P<0.05). Moreover, compared with the patients with low tumor regression rate, the nasopharyngeal carcinoma patients with high tumor regression rate showed a favorable disease-free survival (P<0.05). A high expression level of PD-L1 and a low tumor regression rate after chemoradiotherapy are unfavorable to the prognosis of nasopharyngeal carcinoma.

Local Activator and T Cell Engager (αBCMA x αPD-L1 x αCD3) with Enhanced Tumor Killing and Minimal Cytokine Release for the Treatment of Multiple Myeloma

BCMA-targeting bispecific T-cell engagers in clinical development have demonstrated encouraging preclinical efficacy. The most advanced of these is AMG 420, which showed significantly improved response rates in relapsed/refractory multiple myeloma (MM) patients. Nevertheless, median duration until relapse is currently limited to approximately 12 months, highlighting the need for new drugs with novel MoA. Recently, we reported on a Local Activator and T cell Engager (LocATE) antibody that targets BCMA and selectively blocks programmed death-ligand 1 (PD-L1) on malignant cells (ASCO, June 2019). LocATE induced superior T cell activation and cancer cell killing, in vitro and ex vivo, compared to a BCMAxCD3 BiTE alone or in combination with a PD-L1 inhibitor. Here, we sought to further characterize the novel MoA of our LocATE. To assess the therapeutic potential of the LocATE, we first investigated whether potent cytotoxicity is uncoupled from high levels of cytokine release. We evaluated three LocATE molecules with different PD-L1 affinities (low, medium, high). Using BCMA-expressing MM cell lines (U-266, MM.1S, RPMI-8226 and H929) with distinct PD-L1 surface expression levels (3 - 53%), we determined the cytokine profile (IL-2, IL-6, IFN-γ, TNF-α) and target cell lysis induced by each candidate in the presence of CD3-positive human T cells. All three candidates exhibited comparable killing potency, however, low-affinity PD-L1 LocATE antibodies induced significantly less cytokine release (up to 10-fold) than its higher PD-L1 affinity counterparts across all cell lines investigated. Notably, using the low-affinity PD-L1 LocATE, we observed a 2-fold increase in tumor cell killing compared to bispecific BCMAxCD3 targeting controls in cell lines expressing high PD-L1 levels (53%), underlining the contribution of PD-L1 inhibition. Accordingly, phenotypic profiling of effector cells showed that the LocATE more potently induced dose-dependent upregulation of the activation markers CD69, CD25 and HLA-DR compared to bispecific controls. Importantly, cytotoxic activity, T cell activation and cytokine release were not induced when BCMA-negative cells expressing high levels of PD-L1 were treated with LocATE, underlining the BCMA-selective killing mechanism. Since the superior efficacy of LocATE was found to correlate with the expression level of PD-L1 on MM cell lines and upregulation of PD-L1/PD-1 has been reported as one of the major myeloma cell escape mechanisms during treatment with BiTEs, we subsequently investigated the efficacy of LocATE using primary bone marrow samples and peripheral blood mononuclear cell (PBMCs) obtained from MM patients. Six bone marrow mononuclear cell (BMMC) and eight PBMC samples from MM donors of different disease stages were characterized for PD-1/PD-L1 expression levels; analysis of T cell frequency and level of activation/exhaustion was performed based on CD4, CD8, CD25, CD69, Tim-3, Lag-3 and PD-1 marker expression. Using patient samples with high frequencies of PD-1 expressing T cells prior to treatment, LocATE induced superior MM tumor cell lysis and T cell activation compared to BCMAxCD3 bispecific antibodies. No activity was observed on healthy cells, underlining the safe and selective killing mechanism through tumor-local reactivation of exhausted T cells. Collectively, these findings demonstrate that the simultaneous T cell redirection and tumor-specific checkpoint inhibition with the LocATE leads to an improved therapeutic index with robust tumor cell killing and low levels of cytokine release. Capable of counteracting adaptive immune resistance caused by increased PD-1/PD-L1 signaling, the LocATE antibody has the prospect to significantly improve survival for multiple myeloma patients. Disclosures Vrohlings: CDR-Life: Current Employment. Jungmichel:CDR-Life: Current Employment, Other: current option holder. Senn:CDR-Life: Current Employment. Schleier:CDR-Life: Current Employment, Current equity holder in private company. Scheifele:CDR-Life: Current Employment, Current equity holder in private company. Wendelspiess:CDR-Life: Current Employment. Leisner:CDR-Life: Current Employment, Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees. Jaeger:CDR Life AG: Consultancy, Research Funding; Miltenyi: Consultancy, Honoraria; Karyopharm: Honoraria; BMS/Celgene: Consultancy, Honoraria, Research Funding; Gilead: Honoraria, Research Funding; Novartis: Consultancy, Honoraria, Research Funding; AbbVie: Honoraria; F. Hoffmann-La Roche: Honoraria, Research Funding. Borras:CDR-Life: Current Employment, Current equity holder in private company.

Role of PD-L1 Expression during the Progression of Submucosal Gastric Cancer

Introduction: Programmed death-ligand 1 (PD-L1) expression is a prognostic marker for gastric cancer that correlates with tumor diameter and depth of penetration. But the role of PD-L1 and mechanism(s) employed in the initial phase of invasion in early gastric cancer is yet to be understood. Objective: This study aims to elucidate the role of PD-L1 during the progression of gastric cancer, specifically invading the submucosa beyond the lamina muscularis mucosa. Methods: Using 107 patients with pathological submucosal gastric cancer, we determined the expression of PD-L1 based on the staining of the cell membrane or cytoplasm of tumor cells in the central and invasive front of the tumor. Samples were categorized into 3 groups based on the intensity of PD-L1 expression. CD8⁺ lymphocytes expressing PD-1 and CD163⁺ macrophages were used to determine the number of cell nuclei at the invasive front, similar to PD-L1. CMTM6 levels were determined and used to stratify samples into 3 groups. Results: PD-L1 expression was higher in the invasive front (26.2%) than in the central portion of the tumors (7.4%; p < 0.001). Moreover, lymphatic and vascular invasion were more frequently observed in samples with high levels of PD-L1 (lymphatic invasion: 60.7 vs. 35.4%, p = 0.0026, and vascular invasion: 39.3 vs. 16.5%, p = 0.0018). There was no correlation between PD-L1 expression and the levels of PD-1, CD8, CD163, and CMTM6. Conclusions: PD-L1-expressing cancer cells at the invasive front of gastric cancer influence the initial stages of tumor invasion and lymphovascular permeation in early-stage gastric cancers. Immune checkpoint signaling may be the driving force in the invasive front during the invasion of the submucosa beyond the lamina muscularis mucosa.

Prognosis significance of indoleamine 2, 3-dioxygenase, programmed death ligand-1 and tumor-infiltrating immune cells in microenvironment of breast cancer

IntroductionThe immune microenvironment plays an increasingly important role in predicting the prognosis of multiple tumors and selecting patients for immunotherapy trials. We studied the expression of indoleamine 2, 3-dioxygenase (IDO) and programmed death ligand-1 (PD-L1), detected the proportion of tumor-infiltrating immune cells (TIIs), and further analyzed the association of these immunological characteristics with the clinicopathological parameters and prognosis of breast cancer patients. MethodsImmunohistochemical staining for IDO, PD-L1, CD4, CD8, Foxp3, CD20, CD56 and CD68 expression in breast cancer tissues was carried out. IDO and PD-L1 expression were scored by extent in tumor cells. TIIs expressing CD4, CD8, Foxp3, CD20, CD56 or CD68 were evaluated by positive count. Clinicopathological characteristics and follow-up were recorded. ResultsThe frequencies of IDO-high-expressing and PD-L1-expressing tissue were 33.77% and 24.68%, respectively. The co-expression of IDO and PD-L1 was identified in 16/77 (20.78%) of cases. IDO high expression, CD4+ T cells and CD56+ cells were most frequently observed in patients with tumor-draining lymph nodes(TDLNs) metastasis. Immune cells were more common in non-luminal breast cancer than in luminal breast cancer. In survival analysis, PFS were not associated with high levels of IDO and PD-L1, nor were TIIs. However, CD20 and CD68 were significant risk factors for prognostic after adjusting covariates by COX regression. IDOhighFoxp3highT patients had a tendency with shorter progression-free survival. ConclusionsAlthough we found a limited prognostic effect of TIIs on survival in breast cancer patients, IDO combined with TIIs can help to evaluate the prognosis of patients.