Mechanical force regulates the inhibitory function of PD-1.

PD-1 Imposes Qualitative Control of Cellular Transcriptomes in Response to T Cell Activation.

Programmed cell death 1 (PD-1) is a key immune checkpoint receptor that plays a pivotal role in maintaining immune tolerance by suppressing T cell activation. However, cancer cells frequently exploit the interaction between PD-1 and its ligand PD-L1, to evade immune surveillance. Although monoclonal antibodies targeting the PD-1/PD-L1 pathway have achieved considerable clinical success, their limitations such as high cost, immunogenicity, and limited tissue penetration highlight the need for alternative therapeutic strategies. In this study, a structure-based virtual screening strategy integrated with molecular docking and molecular dynamics (MD) simulations was employed to identify potential small-molecule inhibitors of human PD-1. An initial library of over 8.526 million compounds from the ZINC20 database was filtered for drug-likeness, toxicity, and pharmacokinetic properties, resulting in 18 lead compounds. Among these, four candidates, ZINC000009779862, ZINC000012821448, ZINC000095472904, and ZINC000102699623, exhibited superior binding affinities and interaction profiles with the PD-1 binding pocket. MD simulations confirmed the structural stability of PD-1/ligand complexes, while steered molecular dynamics (SMD) and umbrella sampling analyses revealed the mechanical and thermodynamic robustness of the interactions. Notably, ZINC000095472904 demonstrated the strongest binding strength and the highest binding free energy, making it the most promising candidate. Collectively, these findings suggest that the identified compounds, particularly ZINC000095472904, may interfere with PD-1/PD-L1 interaction and represent viable lead structures for the development of next-generation immune checkpoint inhibitors.

Programmed cell death-1 (PD-1) and programmed cell death-ligand 1 (PD-L1) are new therapeutic targets in cancer immunotherapy. The aim of this study was to investigate the clinicopathological characteristics of PD-1 and PD-L1 expression in extranodal natural killer/T‑cell lymphoma, nasal type (ENKTL). We performed PD-1 and PD-L1 immunostaining in 79 ENKTL biopsy samples and retrospectively analyzed medical records of all 79 patients from four tertiary referral hospitals. The expression of PD-1 and PD-L1 by tumor cells and/or infiltrating immune cells was evaluated. The expression rates of PD-L1 in tumor cells and infiltrating immune cells were 79.7 and 78.5%, respectively, whereas PD-1 in tumor cells and infiltrating immune cells were 1.3 and 11.4%. The PD-L1 positivity in tumor cells and infiltrating immune cells was significantly associated with low international prognostic index (IPI) (P = 0.044 and 0.037, respectively). Patients with normal range of serum lactate dehydrogenase demonstrated a significantly higher PD-L1 positivity in tumor cells (P = 0.020). PD-L1-positive patients had a trend toward better overall survival compared with that in patients with PD-L1-negative in tumor cells and infiltrating immune cells (P = 0.498 and 0.435, respectively). The expression rate of PD-L1 was up to 79.7% in ENKTL, while PD-1 expression rate was very low. This is the first report describing the clinicopathological features and survival outcome according to expression of PD-1 and PD-L1 in ENKTL.

Specific PD-1 Deletion on Regulatory T Cells Leads to Enhanced Anti-Tumor Responses

Fatal Autoimmune Hepatitis Induced by Concurrent Loss of Naturally Arising Regulatory T Cells and PD-1-Mediated Signaling

The programmed cell death 1 (PD-1) receptor on the surface of immune cells is an immune checkpoint molecule that mediates the immune escape of tumor cells. Consequently, antibodies targeting PD-1 have shown efficacy in enhancing the antitumor activity of T cells in some types of cancers. However, the potential effects of PD-1 on tumor cells remain largely unknown. Here, we show that PD-1 is expressed across a broad range of tumor cells. The silencing of PD-1 or its ligand, PD-1 ligand 1 (PD-L1), promotes cell proliferation and colony formation in vitro and tumor growth in vivo. Conversely, overexpression of PD-1 or PD-L1 inhibits tumor cell proliferation and colony formation. Moreover, blocking antibodies targeting PD-1 or PD-L1 promote tumor growth in cell cultures and xenografts. Mechanistically, the coordination of PD-1 and PD-L1 activates its major downstream signaling pathways including the AKT and ERK1/2 pathways, thus enhancing tumor cell growth. This study demonstrates that PD-1/PD-L1 is a potential tumor suppressor and potentially regulates the response to anti-PD-1/PD-L1 treatments, thus representing a potential biomarker for the optimal cancer immunotherapeutic treatment.

Resistance to Checkpoint Inhibition in Cancer Immunotherapy

Formation of eruptive cutaneous squamous cell carcinomas after programmed cell death protein-1 blockade

Programmed cell death4 (PDCD4) has been identified as a tumor‑suppressor gene that inhibits neoplastic transformation, tumor progression, and protein translation. It has been reported that multiple factors participate in the regulation of PDCD4 mRNA and protein. The endometrium is regulated by changing concentrations of ovarian hormones, such as estrogen and progesterone, and shows periodical changes. However, whether ovarian hormones regulate PDCD4 expression remains unclear. This study aimed to explore the effect and mechanism of estrogen or progesterone on PDCD4 mRNA and protein expression in human endometrial cancer cells. The expression of PDCD4 mRNA and protein in Ishikawa and HEC‑1‑A cells was detected by quantitative real‑time PCR and western blot analysis. The signaling pathway‑related proteins were detected by western blot analysis. The results showed that PDCD4 mRNA levels exhibited no significant changes after treatment with estrogen or progesterone in both Ishikawa and HEC‑1‑A cell lines. Estrogen also had no obvious effect on PDCD4 protein expression. However, progesterone effectively decreased the expression of PDCD4 protein and the PI3K/AKT pathway may be involved in the downregulation of PDCD4 protein induced by progesterone. These results suggest that the downregulation of PDCD4 induced by progesterone affects the therapeutic efficacy of progesterone in human endometrial cancer or endometriosis, which may have important implications for progesterone treatment in the clinic.

Lymphocyte-activation gene 3 (LAG-3) receptor is expressed on activated CD4 and CD8 T cells, γδ T cells, Treg, NK, NKT, B and plasmacytoid dendritic cells. LAG-3 binds to major histocompatibility complex class II (MHC II) and delivers inhibitory signals that regulate T cell proliferation and cytokine production. LAG-3 blockade augments T cell proliferation and activation. Programmed cell death 1 (PD-1) receptor upon binding to its ligands PD-L1 (B7-H1; CD274) and PD-L2 (B7-DC; CD273) also delivers an inhibitory checkpoint signal that is critical for the establishment and maintenance of peripheral T cell tolerance. PD-1 signaling plays a critical role in the tumor microenvironment by allowing tumors to escape immune surveillance. We tested in vivo activity of anti-mouse PD-1 and anti-mouse LAG-3 antibodies in several preclinical syngeneic tumor models and showed that combination treatment with both antibodies resulted in an additive anti-tumor effect compared to either single antibody treatment. TaqMan analysis in the MC38 tumor model demonstrated CD8+ T cell expansion in both the draining lymph nodes and spleens of mice in the combination treatment group. Anti-human LAG-3 antibody is a fully human monoclonal antibody developed for cancer immunotherapy. It binds with high affinity to human LAG-3 and blocks LAG-3/MHC II interaction. Fully human monoclonal antibody REGN2810 binds with high affinity to human PD-1 and blocks PD-1 interaction with PD-L1 and PD-L2. Double humanized LAG-3/PD-1 mice were engineered using VelociGene® technology to replace the extracellular domains of mouse Pdcd1 and Lag3 genes with the corresponding regions of human PD-1 and human LAG-3 genes. To validate humanized protein expression, we examined PD-1 and LAG-3 protein expression on T cells after anti-CD3/anti-CD28 antibody stimulation. We confirmed binding of human LAG-3 and PD-1 to the corresponding mouse ligands by cell adhesion assay for human LAG-3 and mouse MHC II interactions and by SPR-Biacore for human PD-1 and mouse PD-L1 interactions, respectively. Combination of REGN2810 and anti-hLAG-3 antibodies in MC38.ova tumor model in double humanized LAG-3/PD-1 mice, which allows testing of clinical antibodies that do not cross to mouse receptors, demonstrated improved efficacy, including reduced tumor growth and improved survival, compared to REGN2810 and anti-hLAG-3 monotherapies. Robust anti-tumor efficacy of REGN2810 and anti-hLAG-3 combination in preclinical setting supports their clinical development as a combination cancer immunotherapy. Citation Format: Elena Burova, Omaira Allbritton, Chandrika Taduriyasas, Venus Lai, William Poueymirou, Nicholas Papadopoulos, Douglas MacDonald, William Olson, Markus Mohrs, Ella Ioffe, Gavin Thurston. Combined treatment with anti-LAG-3 and anti-PD-1 fully human monoclonal antibodies inhibits tumor growth in immunocompetent double-humanized LAG-3/PD-1 mice. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1484.

Background:Available evidence shows that the incidence of toxicities associated with cancer immunotherapy, such as programmed cell death 1 (PD-1) and programmed cell death 1 ligand 1 (PD-L1)-related toxicities, is estimated to be between 0.3 and 1.3%.Objective:This systematic review aimed to investigate cancer patients’ susceptibility to toxicities associated with PD-1/PD-L1 inhibitors and establish a clinically relevant landscape of side effects of PD-1/PD-L1 inhibitors.Data sources:Relevant publications from PubMed, Embase, Cochrane Library, Web of Science, and China National Knowledge Infrastructure (CNKI) between 2014 and 2019.Study eligibility criteria, participants, and interventions:We searched randomized controlled trials (RCTs) reporting treatment-related toxicities associated with PD-1 and PD-L1 inhibitors in the treatment of cancers. The primary endpoint was to assess the difference in the incidences of toxicities between cancer patients who did and did not receive PD-1/PD-L1 inhibitors. A total of 29 RCTs, incorporating 8576 patients, met the eligibility criteria.Study appraisal and synthesis methods:We calculated the pooled relative risks and corresponding 95% CIs using a random-effects model and assessed the heterogeneity between different groups. The subgroup analyses were conducted based on cancer type, toxicity grade (severity), system and organ, treatment regimens in the intervention arm and the control arm, PD-1/PD-L1 inhibitor drug type, and cancer type.Results:A total of 11 categories (e.g. endocrine toxicity), and 39 toxicity types (e.g. hyperthyroidism) were identified. For toxicities at any grade, those treated with PD-1/PD-L1 inhibitors were at lower risks for gastrointestinal toxicity, hematologic toxicity, and treatment event leading to discontinuation; and were at higher risks for respiratory toxicity (all P<0.05). Those treated with PD-1/PD-L1 inhibitors were at lower risks for fatigue, asthenia, and peripheral edema and were at higher risks for pyrexia, cough, dyspnea, pneumonitis, and pruritus.Limitations:The present research is a meta-analysis at the study level rather than at the patient level; insights on risk factors associated with the development of toxicities cannot be found in our study. There was a possible overlap in Common Terminology Criteria for Adverse Events (CTCAE) definitions which prevents understanding the true rates of specific toxicities.Conclusions and implications of key findings:For most toxicity types based on system and organ, the incidence proportions for patients in the intervention arm were lower than those in the control arm, which suggested the general safety of PD-1/PD-L1 inhibitors against conventional chemotherapy and cytotoxic t-lymphocyte-associated protein 4 (CTLA-4) inhibitors. Future research should focus on taking effective targeted measures to decrease the risks of different toxicities for different patient populations.Systematic review registration number:We registered the research protocol with PROSPERO (registration number CRD42019135113).

The immune checkpoint molecule lymphocyte activation gene 3 (LAG-3) is currently being investigated as a possible target for immunotherapy in triple-negative breast cancer (TNBC), frequently as an addition to treatment with programmed cell death protein 1/programmed death ligand 1 (PD-L1) inhibition. However, expression of LAG-3, the frequency of coexpression with PD-L1, and the prognostic significance of this marker have not been studied extensively in TNBC. For this study, tissue microarrays (TMAs) were constructed from surgical specimens of 514 patients with TNBC. TMAs were stained immunohistochemically for LAG-3 and PD-L1 expression. Tumor-infiltrating lymphocytes (TILs) were evaluated on full glass slides. LAG-3 expression was significantly associated with improved overall survival and relapse-free survival. When adjusted for clinicopathologic factors, each increment of 10 LAG-3-positive intratumoral lymphocytes per TMA core was associated with improved overall survival (hazard ratio=0.93, 95% confidence interval: 0.89-0.97, P=0.002), and recurrence-free survival (hazard ratio=0.91, 95% confidence interval: 0.85-0.97, P=0.002). PD-L1 expression on immune cells and PD-L1 expression evaluated with the combined positive score and TILs were also associated with improved survival in both univariate and multivariate analyses. PD-L1 expression on tumor cells was only associated with improved survival in univariate analysis. LAG-3 expression was associated with both TILs and PD-L1 expression. Coexpression of LAG-3 and PD-L1 did not confer additional survival benefits. In conclusion, LAG-3 expression is associated with improved survival in TNBC. LAG-3 is often coexpressed with PD-L1, confirming that TNBC is likely a suitable candidate for cotreatment with LAG-3 and programmed cell death protein 1/PD-L1 inhibitors. However, coexpression does not confer additional survival benefits.

Novel Tumor Suppressor Protein Programmed Cell Death 4 (PDCD4) Suppresses Activity of PI3K/Akt Pathway and Regulates Expression of p27 (Kip1) and c-myc, DAP5 and Willm's Tumor (WT1) in Acute Myeloid Leukemia.

Programmed cell death protein-1/programmed cell death ligand-1 (PD-1/PD-L1) pathway has emerged as a critical target for cancer immunotherapy, and monoclonal antibodies that block either side of this inhibitory interaction have demonstrated impressive activity across a broad set of cancer subtypes. Clinical studies have shown that tumor PD-L1 expression is associated with clinical stages and poor prognosis. Tumor PD-L1 expression detection may help to screen the patients who are most likely to be benefit from anti-PD-1/PD-L1 therapy and evaluate the prognosis. Molecular imaging technology such as PET, SPECT and optical imaging may be the modalities that allowing for noninvasively dynamic detection and overall evaluation of multiple biomarkers. This review focuses on the state of the art of tumor molecular imaging targeting PD-1/PD-L1 pathway. Key words: Neoplasms; Programmed cell death 1 receptor; Antigens, CD274; Molecular imaging; Trends

Immune checkpoint mechanisms are important molecular cell systems that maintain tolerance toward autoantigens in order to prevent immunity-mediated accidental damage. It is well known that cancer cells may exploit these molecular and cellular mechanisms to escape recognition and elimination by immune cells. Programmed cell death protein-1 (PD-1) and its natural ligand programmed cell death ligand-1 (PD-L1) form the PD-L1/PD-1 axis, a well-known immune checkpoint mechanism, which is considered an interesting target in cancer immunotherapy. In fact, the expression of PD-L1 was found in various solid malignancies and the overactivation of PD-L1/PD-1 axis results in a poor patient survival rate. Breaking PD-L1/PD-1 axis, by blocking either the cancer side or the immune side of the axis, is currently used as anti-cancer strategy to re-establish a tumor-specific immune response. For this purpose, several blocking antibodies are now available. To date, three anti-PD-L1 antibodies have been approved by the FDA, namely atezolizumab, durvalumab and avelumab. The main advantages of anti-PD-L1 antibodies arise from the overexpression of PD-L1 antigen by a high number of tumor cells, also deriving from different tissues; this makes anti-PD-L1 antibodies potential pan-specific anti-cancer molecules. Despite the good results reported in clinical trials with anti-PD-L1 antibodies, there is a significant number of patients that do not respond to the therapy. In fact, it should be considered that, in some neoplastic patients, reduced or absent infiltration of cytotoxic T cells and natural killer cells in the tumor microenvironment or presence of other immunosuppressive molecules make immunotherapy with anti-PD-L1 blocking antibodies less effective. A strategy to improve the efficacy of antibodies is to use them as carriers for toxic payloads (toxins, drugs, enzymes, radionuclides, etc.) to form immunoconjugates. Several immunoconjugates have been already approved by FDA for treatment of malignancies. In this review, we focused on PD-L1 targeting antibodies utilized as carrier to construct immunoconjugates for the potential elimination of neoplastic cells, expressing PD-L1. A complete examination of the literature regarding anti-PD-L1 immunoconjugates is here reported, describing the results obtained in vitro and in vivo. The real potential of anti-PD-L1 antibodies as carriers for toxic payload delivery is considered and extensively discussed.