Death Receptor Research Articles

Self-Illuminating Nanoagonist Simultaneously Induces Dual Cell Death Pathways via Death Receptor Clustering for Cancer Therapy.

Inducing death receptor 5 (DR5) clustering holds particular promise in tumor-specific therapeutics because it could trigger an apoptotic cascade in cancerous cells. Herein, we present a tumor microenvironment H2O2-responsive self-illuminating nanoagonist, which could induce dual tumor cell death pathways through enhancing DR5 clustering. By conjugating DR5 ligand peptides onto the surfaces of self-illuminating nanoparticles with cross-linking capacity, this strategy not only provides scaffolds for ligands to bind receptors but also cross-links them through photo-cross-linking. This strategy allows for efficient activation of DR5 downstream signaling, initiating the extrinsic apoptosis pathway and immunogenic cell death of tumor cells, and contributes to improved tumor-specific immune responses, resulting in enhanced antitumor efficacy and minimized systemic adverse effects.

Mechano-inhibition of endocytosis sensitizes cancer cells to Fas-induced Apoptosis

The transmembrane death receptor Fas transduces apoptotic signals upon binding its ligand, FasL. Although Fas is highly expressed in cancer cells, insufficient cell surface Fas expression desensitizes cancer cells to Fas-induced apoptosis. Here, we show that the increase in Fas microaggregate formation on the plasma membrane in response to the inhibition of endocytosis sensitizes cancer cells to Fas-induced apoptosis. We used a clinically accessible Rho-kinase inhibitor, fasudil, that reduces endocytosis dynamics by increasing plasma membrane tension. In combination with exogenous soluble FasL (sFasL), fasudil promoted cancer cell apoptosis, but this collaborative effect was substantially weaker in nonmalignant cells. The combination of sFasL and fasudil prevented glioblastoma cell growth in embryonic stem cell-derived brain organoids and induced tumor regression in a xenograft mouse model. Our results demonstrate that sFasL has strong potential for apoptosis-directed cancer therapy when Fas microaggregate formation is augmented by mechano-inhibition of endocytosis.

The impact of PD-1 inhibitors on prognosis in unresectable hepatocellular carcinoma treated with TACE and lenvatinib: a retrospective study

Our aim was to explore whether programmed death receptor-1 (PD-1) inhibitors would improve the prognosis of unresectable hepatocellular carcinoma (HCC) treated with transarterial chemoembolization (TACE) plus lenvatinib. In this single-center retrospective study, patients with unresectable HCC who underwent TACE and were administered lenvatinib with or without PD-1 inhibitors were enrolled and divided into the TACE + lenvatinib group and TACE + lenvatinib + PD-1 group. Overall survival (OS), progression-free survival (PFS) and tumor response were assessed by the Response Evaluation Criteria in Solid Tumors (RECIST v1.1 and mRECIST). Treatment-related adverse events (AEs) were evaluated according to the Common Terminology Criteria for Adverse Events (CTCAE, version 5.0). In total, 35 eligible patients with unresectable HCC were included; 82.9% of patients had Hepatitis B virus (HBV) infection, and 88.6% of patients had liver cirrhosis. A total of 88.6% of patients had multiple tumors, and the median diameter of the largest tumor was 10.1 cm. A total of 14.3% of patients had extrahepatic metastasis, and 51.4% of patients had portal vein tumor thrombus. The percentages of BCLC stages A, B and C were 5.7%, 28.6% and 65.7%, respectively. There were 16 patients in the TACE + lenvatinib group and 19 patients in the TACE + lenvatinib + PD-1 group. The median follow-up time was 7.7 months (ranging from 1.7 to 31.6 months). Neither group reached the median overall survival. Under RECIST v1.1 criteria, the median PFS was 10.4 and 7.9 months in the TACE + lenvatinib and TACE + lenvatinib + PD-1 groups (HR, 1.13; 95% CI 0.45–2.84; p = 0.80), the objective response rates (ORR) were 31.3% and 31.6% (p > 0.05), and the disease control rates (DCR) were 93.8% and 78.9% (p > 0.05), respectively. Under mRECIST criteria, the median PFS was 10.4 and 10.1 months (HR, 0.98; 95% CI 0.38–2.54, p = 0.97), the ORR was 62.5% and 63.2% (p > 0.05), and the DCR was 93.8% and 73.7% (p > 0.05), respectively. Overall, AEs were relatively similar between the two groups. PD-1 inhibitors did not improve the PFS and tumor response of unresectable HCC treated with TACE plus lenvatinib. Hepatitis B infection, liver cirrhosis, portal vein tumor thrombus, multiple tumors and large tumor diameter may be potential factors that affect the efficacy of PD-1 inhibitors but need further validation.

Protein O-fucosyltransferase 1 promotes PD-L1 stability to drive immune evasion and directs liver cancer to immunotherapy

Background and aimsThe immunosuppressive tumor microenvironment (TME) plays an essential role in cancer progression and immunotherapy response. Despite the considerable advancements in cancer immunotherapy, the limited response to immune checkpoint...

Plasma circulating tumor DNA unveils the efficacy of PD-1 inhibitors and chemotherapy in advanced gastric cancer

Programmed Death Receptor 1 (PD-1) inhibitors, when combined with chemotherapy, have exhibited notable effectiveness in enhancing the survival outcomes of patients afflicted with advanced gastric cancer. However, it is important to acknowledge that not all patients derive substantial benefits from this therapeutic approach, highlighting the crucial necessity of identifying efficacious biomarkers to inform immunotherapy interventions. In this study, we sought to investigate the predictive utility of circulating tumor DNA (ctDNA) as a biomarker in a cohort of 30 patients diagnosed with advanced gastric cancer, all of whom underwent first-line treatment involving PD-1 inhibitor administration alongside chemotherapy. We procured peripheral blood samples both at baseline and following the completion of two treatment cycles. Additionally, baseline tissue specimens were collected for the purpose of genomic alteration assessment, employing both 47-gene and 737-gene next-generation sequencing panels for plasma and tumor tissue, respectively. We delineated a ctDNA response as the eradication of maximum variant allele frequencies relative to baseline levels. Notably, the objective response rate among individuals exhibiting a ctDNA response proved significantly superior in comparison to non-responders (P = 0.0073). Furthermore, patients who manifested a ctDNA response experienced markedly prolonged progression-free survival (PFS) and overall survival (OS) when juxtaposed with those devoid of a ctDNA response (median PFS: 15.6 vs. 6.0 months, P = 0.003; median OS: not reached [NR] vs. 9.0 months, P = 0.011). In summation, patients with advanced gastric cancer receiving first-line treatment with PD-1 inhibitors and chemotherapy, dynamic changes in ctDNA can serve as a potential biomarker for predicting treatment efficacy and long-term outcomes.

Evaluation of the Expression of Programmed Death-Ligand 1 and Its Role in Differentiating Low-Grade and High-Grade Urothelial Carcinoma.

Urothelial carcinoma (UC) is a common malignancy, predominantly affecting males. Many tumor cells use the interaction between programmed death-ligand 1 (PD-L1) and programmed death receptor (PD-1) to inactivate T-cells in the microenvironment and evade host immune response. Our study aims to evaluate the expression of PD-L1 in UC and correlate its expression with histomorphological parameters. After obtaining approval from the Institute Ethics Committee, we conducted a prospective observational study on transurethral resection of urinary bladder tumor (TURBT) and cystectomy specimens histopathologically diagnosed as UC between 2022 and 2023, comprising 50 cases. All standard protocol was followed and immunohistochemistry (IHC) was done using PD-L1 with rabbit anti-human PD-L1 monoclonal antibody (Clone: IHC411; Biogenics Inc., San Francisco, CA, USA). Results: Among the 50 cases of UC, the majority were papillary type (35 cases), high grade (28 cases), and non-muscle invasive (30 cases). Among the cases studied, 15 of them showed PD-L1 positivity; 55% of the cases of muscle-invasive bladder cancer were found to be positive for PD-L1 out of which the results were statistically significant. PD-L1 expression by IHC staining can differentiate between muscle-invasive and non-muscle-invasive UC cases. This observation allows for further exploring the potential role of immune checkpoint inhibitors in adjuvant and neoadjuvant therapy, especially in muscle-invasive cases of UC.

Development of an in vitro assay for screening programmed death receptor-1/programmed cell death ligand 1 monoclonal antibody therapy in dogs

Immunomodulatory antibody drugs that modulate the function of immune checkpoint molecules, such as programmed death receptor-1 (PD-1) and programmed cell death ligand 1 (PD-L1), have been established as new cancer treatments in human medicine. In recent years, there have also been reports on antibodies that inhibit immune checkpoint molecules in dogs, and clinical trials using such antibodies for canine cancer have been gradually increasing in number. Because inhibitory antibodies restore T-cell function by inhibiting the binding of PD-1 on T cells and its ligand PD-L1, the quality of antibody function has been evaluated using activated T cells or peripheral blood mononuclear cells isolated from healthy dogs; however, the assays and dogs used significantly vary. Therefore, in the present study, we developed a reporter gene assay using reporter cells (Jurkat/NFATluc/cPD1) and effector cells (CTAC/OKT3/cPDL1). Jurkat/NFATluc/cPD1 were generated by introducing both of the NFAT-responsive luciferase gene as a marker of T-cell signaling and canine PD-1, into a human T lymphoid cell line, Jurkat. CTAC/OKT3/cPDL1 were generated by introducing single-chain FV (scFV) of anti-human CD3 antibody (OKT3) and canine PD-L1 into a canine thyroid carcinoma cell line, CTAC. Ligation of PD-1 on Jurkat/NFATluc/cPD1 via binding of PD-L1 on CTAC/OKT3/cPDL1 suppressed NFAT luciferase activity induced by CD3 ligation by scFV of OKT3. The addition of anti-canine PD-1 and PD-L1 antibodies, both of which were previously developed in our laboratory, restored this suppression with high sensitivity, although the anti-human PD-L1 antibody atezolizumab induced a very weak restoration. This assay is an useful method for functionally evaluating the inhibition of canine PD-1 and PD-L1 binding.

Self-Assembled PD-L1 Downregulator to Boost Photodynamic Activated Tumor Immunotherapy Through CDK5 Inhibition.

The immunosuppressive characteristics and acquired immune resistance can restrain the therapy-initiated anti-tumor immunity. In this work, an antibody free programmed death receptor ligand 1 (PD-L1) downregulator (designated as CeSe) is fabricated to boost photodynamic activated immunotherapy through cyclin-dependent kinase 5 (CDK5) inhibition. Among which, FDA approved photosensitizer of chlorin e6 (Ce6) and preclinical available CDK5 inhibitor of seliciclib (Se) are utilized to prepare the nanomedicine of CeSe through self-assembly technique without drug excipient. Nanoscale CeSe exhibits an increased stability and drug delivery efficiency, contributing to intracellular production of reactive oxygen species (ROS) for robust photodynamic therapy (PDT). The PDT of CeSe can not only suppress the primary tumor growth, but also induce the immunogenic cell death (ICD) to release tumor associated antigens. More importantly, the CDK5 inhibition by CeSe can downregulate PD-L1 to re-activate the systemic anti-tumor immunity by decreasing the tumor immune escape and therapy-induced acquired immune resistance. This work provides an antibody free strategy to activate systemic immune response for metastatic tumor treatment, which may accelerate the development of translational nanomedicine with sophisticated mechanism.

Navigating the landscape of PD-1/PD-L1 imaging tracers: from challenges to opportunities.

Immunotherapy targeted to immune checkpoint inhibitors, such as the program cell death receptor (PD-1) and its ligand (PD-L1), has revolutionized cancer treatment. However, it is now well-known that PD-1/PD-L1 immunotherapy response is inconsistent among patients. The current challenge is to customize treatment regimens per patient, which could be possible if the PD-1/PD-L1 expression and dynamic landscape are known. With positron emission tomography (PET) imaging, it is possible to image these immune targets non-invasively and system-wide during therapy. A successful PET imaging tracer should meet specific criteria concerning target affinity, specificity, clearance rate and target-specific uptake, to name a few. The structural profile of such a tracer will define its properties and can be used to optimize tracers in development and design new ones. Currently, a range of PD-1/PD-L1-targeting PET tracers are available from different molecular categories that have shown impressive preclinical and clinical results, each with its own advantages and disadvantages. This review will provide an overview of current PET tracers targeting the PD-1/PD-L1 axis. Antibody, peptide, and antibody fragment tracers will be discussed with respect to their molecular characteristics and binding properties and ways to optimize them.

Evaluation of Immune Exhaustion and Co-Inhibitory Receptor Expression in Mycobacterium avium Subspecies paratuberculosis (MAP) Seropositive Diarrhoeic Bovines.

Mycobacterium avium subspecies paratuberculosis (MAP) infection leads to chronic, persistent granulomatous enteritis, causing prolonged diarrhoea and emaciation. The disease is managed using medications such as antibiotics, live vaccines, mycobacteriophage therapies and other treatments; however, a notable proportion of affected animals do not show improvement with this approach. We hypothesise that immunoinhibitory receptors TIM-3 (T cell immunoglobulin mucin protein-3) and PD-1 (Programmed death receptor 1) may be upregulated on Peripheral blood mononuclear cells (PBMCs) of MAP-seropositive bovines, potentially contributing to immune exhaustion. Samples (blood and faeces) were collected from 32 diarrhoeic bovines suspected of MAP infection; eight apparently healthy buffaloes from the dairy farm at Hisar, Haryana and from 14 cows (suffering from chronic diarrhoea, weakness and emaciation) housed in stray cattle shed. MAP infection was estimated using indigenous ELISA (i-ELISA), faecal IS900 PCR, culture and acid-fast staining. TIM-3 and PD-1 gene expression on PBMCs were determined using qRT-PCR. TIM3 expression was relatively higher (~400-fold, 330-fold, 112-fold, 65-fold and 16-fold) in 5 chronically diarrhoeic PBMCs samples (MAP-seropositive), and higher PD-1 expression (around ~7-fold, 1.75-fold, 2.5-fold, 7.6-fold) was recorded in 4 diarrhoeic MAP-seropositive animals, compared to apparently healthy and other MAP-seronegative diarrhoeic animals. High co-expression of TIM-3 and PD-1 levels was also recorded in chronically diarrhoeic, emaciated stray cattle. Understanding immune responses in field conditions might aid in the therapeutic management of paratuberculosis.

Tabersonine Induces the Apoptosis of Human Hepatocellular Carcinoma In vitro and In vivo.

Tabersonine, a natural indole alkaloid derived from Apocynaceae plants, exhibits antiinflammatory and acetylcholinesterase inhibitory activities, among other pharmacological effects. However, its anti-tumor properties and the underlying molecular mechanisms remain underexplored. The present study aims to investigate the anti-tumor effects of tabersonine and its mechanisms in inducing apoptosis in hepatocellular carcinoma. The inhibitory effects of tabersonine on the viability and proliferation of liver cancer cells were evaluated using MTT assay and colony formation assay. AO/EB, Hoechst, and Annexin V-FITC/ PI staining techniques were employed to observe cell damage and apoptosis. JC-1 staining was used to detect changes in mitochondrial membrane potential. Western blot analysis was conducted to study the anti-tumor mechanism of tabersonine on liver cancer cells. Additionally, a xenograft model using mice hepatoma HepG2 cells was established to assess the anti-tumor potency of tabersonine in vivo. Our findings revealed that tabersonine significantly inhibited cell viability and proliferation, inducing apoptosis in liver cancer cells. Treatment with tabersonine inhibited Akt phosphorylation, reduced mitochondrial membrane potential, promoted cytochrome c release from mitochondria to the cytoplasm, and increased the ratio of Bax to Bcl-2. These findings suggested that tabersonine induces apoptosis in liver cancer cells through the mitochondrial pathway. Furthermore, tabersonine treatment activated the death receptor pathway of apoptosis. In vivo studies demonstrated that tabersonine significantly inhibited xenograft tumor growth. Our study is the first to demonstrate that tabersonine induces apoptosis in HepG2 cells through both mitochondrial and death receptor apoptotic pathways, suggesting its potential as a therapeutic agent candidate for hepatic cancer.

A phase 1/2 study of ubamatamab (REGN4018), a MUC16×CD3 bispecific antibody, administered alone or in combination with cemiplimab (anti–PD-1) in patients with recurrent ovarian cancer or MUC16+ endometrial cancer.

TPS5632 Background: Mucin 16 (MUC16) is a cell surface glycoprotein that is overexpressed in ovarian cancer (OC) and endometrial cancer (EC). Ubamatamab is a MUC16 × cluster of differentiation 3 (MUC16×CD3) bispecific antibody that bridges MUC16 on tumor cells and CD3 on T cells to promote T-cell–mediated cytotoxicity. Cemiplimab, a monoclonal antibody targeting the programmed cell death-1 (PD-1) receptor, enhanced MUC16×CD3 activity in preclinical models. In Phase 1 of this first-in-human study (NCT03564340), ubamatamab alone and in combination with cemiplimab demonstrated an acceptable safety profile, durable clinical activity across a range of doses, and linear pharmacokinetics (PK) in patients (pts) with recurrent OC (Liu et al. IGCS 2023. Plenary 03). Phase 2 will assess the efficacy of ubamatamab alone and in combination with cemiplimab in pts with recurrent OC. Ubamatamab will also be evaluated for recurrent advanced or metastatic EC that is MUC16+. Methods: For individuals with recurrent OC, elevated serum cancer antigen (CA)-125 ≥2× the upper limit of normal, and up to four prior lines of cytotoxic therapy, weekly dosing of ubamatamab alone or in combination with cemiplimab will be evaluated in monotherapy and combination therapy expansion cohorts. A Phase 2 cohort will also be randomized 1:1:1 to receive ubamatamab 250 mg IV Q3W or 800 mg IV Q3W or ubamatamab 250 mg IV Q3W + cemiplimab 350 mg IV Q3W. For the EC cohort (≥25% of tumor cells MUC16+, as assessed by immunohistochemistry [IHC]) pts with advanced or metastatic EC that has progressed or is recurrent after prior anti–PD-1 therapy and prior platinum-based chemotherapy will be enrolled. The initial EC cohort will receive ubamatamab 250 mg IV Q3W. Based upon observed safety and activity, additional cohorts may be enrolled to receive ubamatamab 800 mg IV Q3W and/or ubamatamab 250 mg IV Q3W + cemiplimab 350 mg IV Q3W. To limit the risk of cytokine release syndrome, all pts will receive ubamatamab once-weekly step-up dosing prior to addition of cemiplimab and/or Q3W dosing. Expansion cohorts will use a Simon 2-stage study design, with an interim analysis after the first 20 pts. Any treatment arm with ≥3 objective responses may be expanded to 50 pts. In Phase 2, the primary endpoint will be the objective response rate for each arm (RECIST v1.1). Secondary endpoints include duration of response and progression-free survival, safety, PK, and change from baseline in quality of life. Exploratory objectives include clinical correlation with serum CA-125, MUC16 IHC, and immune markers. The study is currently enrolling to these OC and EC expansion cohorts. Clinical trial information: NCT03564340 .

Real-world treatment patterns and outcomes of patients with advanced melanoma treated with nivolumab plus relatimab.

e21501 Background: Programmed death-1 (PD-1) receptor inhibitors significantly increase survival rates for advanced melanoma, yet first-line PD-1 inhibitor therapy shows limited overall response rates (32.6%) and progression-free survival (4.6 months). Relatimab (RELA) was recently approved for the treatment of metastatic and unresectable stage III to IV melanoma in combination with nivolumab (NIVO). RELATIVITY-047 showed an ORR of 43% as a first-line treatment while RELATIVITY 020 showed an ORR of 9-12% in patients progressed on anti-PD-1 therapy. Although clinical trials have demonstrated NIVO-RELA provides benefit for patients with advanced melanoma, little is known about real-world treatment patterns and outcomes. Methods: We performed a retrospective review of all patients with stage III and IV melanoma treated with NIVO-RELA as first-(1L) or second-line or beyond (2L+) therapy at Inova Schar Cancer Institute between September 2021 to September 2023. Primary endpoints were overall response rate (ORR), complete response (CR), partial response (PR), stable disease (SD), and progression of disease (PD) which were evaluated using the RECIST v1.1 criteria. The Kaplan–Meier method was used to generate overall survival (OS) and progression free survival (PFS) median values. Results: Eighty-eight patients were followed for an average of 6.5 months following NIVO-RELA initiation. For 1L treated patients, ORR was 58% (21% PR, 37% CR). Median PFS was 11 months (95% CI 3.1-11). For 2L+ treated patients, ORR was 33% (14% PR, 19% CR). Median PFS was 9.8 months (95% CI 4.7-17.2). Subgroup analyses observed significantly different ORR between patients previously treated with versus without anti-CTLA4 therapy (29% v 44%; p = 0.017), patients treated with NIVO-RELA more than vs within 6 months from their last other therapy (50% v 27%; p = 0.004), and among patients with stage III vs stage IV disease (67% v 30%; p = 0.031) (Table 1). Conclusions: These real-world results support the use of NIVO + RELA in patients with advanced melanoma and emphasize the impressive efficacy of the drug combination in the IL and 2L+ settings compared to RELATIVITY-020 and 047. PFS and OS for 1L and 2L+ therapy with NIVO + RELA in this study aligned with or superposed results from previous trials. Further analyses with increased sample sizes and longer follow-up is warranted. [Table: see text]

Immune checkpoint inhibitors and immune mediated diabetes mellitus: Meta-review of case reports.

e14606 Background: Use of Immune checkpoint inhibitors (ICIs) is an increasingly popular antitumor immunotherapy targeting immunoreceptors on T lymphocytes, enhancing the anti-neoplastic immune response. ICIs are becoming first line therapy options for non-small cell lung cancer and gastric adenocarcinoma and can be used as second line for variety of others. Various endocrinologic adverse events have been reported with use of ICIs. Of these, new onset Type-1 diabetes mellitus (T1DM) is of particular interest given its significant morbidity and noted frequency of 1-5% of patients who receive ICIs. There is limited consolidated information about symptomatic presentations, risk factors, and demographics of patients developing Type 1 DM in relation to ICI use. We performed a systematic review of published case reports to identify commonalities between patient disease presentation, ICI use, and development of immune-mediated diabetes mellitus. Methods: A comprehensive systemic literature search from PubMed Web Science, CINAHL, and Google Scholars was performed identifying case reports showing development of new onset diabetes mellitus in patients using immune checkpoint inhibitors. Descriptive analysis was performed to better identify characteristics and presentation of patients. Results: Based on our parameters, 130 reports with 149 cases were included. Primary tumors among the patients comprised lung cancer (34.9%) and melanoma (31.5%). Amongst all the cases, about 88% of the patients were treated with anti-PD-1 (program cell death) receptor, with nivolumab alone (34.9%) and pembrolizumab alone (26.2%) being the most common agents. Cases were more commonly found to be males (61.7%) and of Asian (36.6%) or North American (35.6%) background. Patient ages ranged from 14-95 with a mean of 62.3. Presentation most commonly started after the third cycle (13.4%) and was noted to occur between 1-50 doses. Diabetic Ketoacidosis was diagnosed in 69.8% of individuals, and the average presenting blood glucose was 611 mg/dl with an HbA1c of 8.2%. Autoantibody data was largely missing; however, anti-GAD antibodies were noted to be present in 29.5% of cases. 34.2% of patients had coexisting endocrinopathies with thyroiditis being the most common, occurring in 20% of patients. 1.3% of patients did not survive the initial hospitalizations, with the vast majority (95.3%) remaining on insulin and a select few (2%) were eventually able to discontinue insulin. Conclusions: New onset diabetes is a rare but a potential life-threatening metabolic urgency. Thus, it is paramount that clinicians maintain close monitor of blood glucose and endocrine function when a cancer patient who recently started ICIs gets admitted for hyperglycemia. Further research is warranted to understand onset of diabetes mellitus at molecular level.

Application of Traditional Chinese Medicine in Inhibiting the PD-1/PD-L1 Pathway in the Treatment of Gastric and Colon Cancers

Immune checkpoint regulation is a negative feedback regulatory mechanism in the body, and sequential death receptor-1 (PD-1) and programmed death receptor ligand-1 (PD-L1) are known as immune checkpoints. The PD-1/PD-L1 pathway inhibits the activity of effector T cells through a negative regulatory mechanism to avoid excessive response-induced body damage. PD-L1 is highly expressed in many tumor tissues, and high PD-L1 expression can ultimately lead to tumor immune escape. Therefore, immune checkpoint blockade with inhibition of negative immune regulation therapy has become a cutting-edge hot spot for antitumor therapy, with the main target molecules being PD-1 and PD-L1. Recent years have seen promising progress in the study of traditional Chinese medicines and their effects on gastric and colon cancers, particularly in relation to the PD-1/PD-L1 pathway mechanisms. This review specifically examines the modulation of the PD-L1 pathway by certain traditional Chinese medicines in gastric and colon cancers, aiming to provide insights for the development of innovative drugs for these types of digestive cancers.

Maintenance treatment with toripalimab and anlotinib after anthracycline-based chemotherapy in patients with advanced soft tissue sarcoma (TORAN): A single-arm, phase 2 trial.

11572 Background: In patients with advanced soft tissue sarcoma (STS) who have achieved disease control after anthracycline-based chemotherapy, maintenance therapy is being explored. This study evaluated the efficacy and safety of toripalimab (a monoclonal antibody against the programmed cell death-1 receptor) combined with anlotinib (a multitargeted tyrosine kinase inhibitor) as a maintenance treatment. Methods: Patients with advanced STS who have achieved disease control after at least four cycles of first-line anthracycline-based chemotherapy were enrolled in this single-arm, phase 2 trial. Fifty patients are planned to receive treatment with toripalimab and anlotinib. The primary endpoint was progression-free survival rate at 24 weeks (PFR24w) during maintenance therapy. Secondary endpoints were median progression-free survival (PFS), median overall survival (OS), objective response rate (ORR), disease control rate (DCR) and safety. Results: By the cut-off date of Jan 25, 2024, 22 patients were enrolled, including patients with leiomyosarcoma (n =5), synovial sarcoma (n =3), epithelioid sarcoma (n =3), dedifferentiated liposarcoma (n =2) and other different subtypes of sarcomas (n =9). The median follow-up time was 11 months. Response has been evaluated in 18 patients. The ORR and DCR were 22.2% and 100%. The PFR24w was 67%, the median PFS was 11.8 months, and the median OS was not reached. The grade 3 or higher adverse events were hypertension (n =4), hemorrhage (n =1), adrenal insufficiency (n =1) and hypertriglyceridemia (n =1). Conclusions: Maintenance therapy with toripalimab and anlotinib is a promising treatment option for patients with advanced STS after first-line anthracycline-based chemotherapy. Clinical trial information: ChiCTR2100054901.

Analysis of regulation of cell death pathway genes in CSC-enriched TNBC after treatment with doxorubicin: A potencial biomarker.

e13154 Background: Breast cancer is one of the most common cancers in women worldwide, and triple-negative breast cancer (TNBC) accounts for approximately 10-20% of all cases globally. TNBC has the worst prognosis among other types of breast cancer due to the lack of targeted therapy. In addition, TNBC tumors exhibit significant cellular and molecular heterogeneity, along with the presence of cells with cancer stem cell (CSC) characteristics, which contribute to the low response to available chemotherapy treatments, leading to chemoresistance, recurrence and metastasis. objective of this study is to evaluate the expression of genes related to cell death and survival pathways in CSCs-enriched spheroids DOX-treated obtained of BT-549 cells, a human TNBC cell line. The death receptor genes include TNFRSF10B, TNFRSF10C, CFLAR, RIPIK1, CASP8, NFKB1, RELA, and MLKL. Methods: To achieve this, gene expression was analyzed through RT-qPCR, comparing the expression in DOX-treated CSCs-enriched spheroids (IC50 55.7 μM/ 48h incubation) and untreated CSCs-enriched spheroids. Preliminary results indicated that, among the evaluated genes, only CFLAR, NFKB1, and RELA were expressed in the untreated CSCs-enriched spheroids, while the others were not expressed under any condition. Results: The genes CFLAR, NF- κ B and RELA showed considerable Ct values, while the remaining genes did not exhibit a Ct value in one or more samples. The gene CFLAR showed an overexpression, both in monolayers vs. spheroids and in treated spheroids vs. untreated spheroids, which can be related to the inhibition of Caspase 8 and consequently, the apoptotic activity of cells. The overexpression of the RELA, same as CFLAR, suggests increased inflammation pathway activation in the tumor microenvironment. However, the lower expression of the NF-κB1 gene in treated spheroids vs. untreated spheroids may be related to the production of ROS, suppressing its activation. Conclusions: The expression of NFKB1 and RELA genes is associated with the cellular survival pathway, which may be linked to the resistance to the drug observed in many patients. Previous studies of our group have shown that achieving a satisfactory result of DOX action on CSCs-enriched spheroids requires using a dose 100 times higher than that used in monolayer cells. This may be related to the non-expression of the TNFRSF10B, TNFRSF10C, RIPIK1, CASP8, and MLKL genes in CSCs-enriched spheroid DOX-treated. Thus, the results obtained so far indicate that DOX may have an impact on the expressed genes in CSCs-enriched spheroids and demonstrate the expression of apoptosis inhibitors in treated breast spheres. Consequently, the expression of these genes in BT-549 cells TNBC indicates a pro-inflammatory, anti-apoptotic, and pro-cell survival profile of CSC-enriched spheroids.

Human 3D Ovarian Cancer Models Reveal Malignant Cell-Intrinsic and -Extrinsic Factors That Influence CAR T-cell Activity.

In vitro preclinical testing of chimeric antigen receptor (CAR) T cells is mostly carried out in monolayer cell cultures. However, alternative strategies are needed to take into account the complexity and the effects of the tumor microenvironment. Here, we describe the modulation of CAR T-cell activity by malignant cells and fibroblasts in human three-dimensional (3D) in vitro cell models of increasing complexity. In models combining mucin-1 (MUC1) and TnMUC1 CAR T cells with human high-grade serous ovarian cancer cell spheroids, malignant cell-intrinsic resistance to CAR T-cell killing was due to defective death receptor signaling involving TNFα. Adding primary human fibroblasts to spheroids unexpectedly increased the ability of CAR T cells to kill resistant malignant cells as CCL2 produced by fibroblasts activated CCR2/4+ CAR T cells. However, culturing malignant cells and fibroblasts in collagen gels engendered production of a dense extracellular matrix that impeded CAR T-cell activity in a TGFβ-dependent manner. A vascularized microfluidic device was developed that allowed CAR T cells to flow through the vessels and penetrate the gels in a more physiological way, killing malignant cells in a TNFα-dependent manner. Complex 3D human cell models may provide an efficient way of screening multiple cytotoxic human immune cell constructs while also enabling evaluation of mechanisms of resistance involving cell-cell and cell-matrix interactions, thus accelerating preclinical research on cytotoxic immune cell therapies in solid tumors. Significance: Three-dimensional in vitro models of increasing complexity uncover mechanisms of resistance to CAR T cells in solid tumors, which could help accelerate development of improved CAR T-cell constructs.

Pulsed Electromagnetic Field Enhances Caffeic Acid Phenethyl Ester-induced Death of MCF-7 Breast Cancer Cells.

Caffeic acid phenethyl ester (CAPE) exerts anticancer effects against several cancer types, including breast cancer. Pulsed electromagnetic field (PEMF) improves the efficiency of some chemotherapeutic drugs. In this study, we examined the effects of PEMF stimulation on the anticancer activity of CAPE in MCF-7 breast cancer cells and the underlying signal transduction pathways. MCF-7 cells were seeded and incubated for 24 h. Each of the drugs (5-fluorouracil, paclitaxel, gefitinib, or CAPE) was added to the cells on day 0. Then, cells were immediately stimulated with a 60-min PEMF session thrice a day (with 4-h interval between sessions) for 1-3 days. Cell death and viability were assessed by flow cytometry and trypan blue dye exclusion assay. Molecular mechanisms involved in cell death were confirmed by western blot assay. Compared with treatment with CAPE alone, co-treatment with CAPE and PEMF more strongly reduced the viability of MCF-7 cells, further increased the percentage of the sub-G1 population, poly (ADP-ribose) polymerase (PARP) cleavage, activation of apoptotic caspases, up-regulation of pro-apoptotic proteins, such as Fas cell surface death receptor (FAS) and BCL2 associated X, apoptosis regulator (BAX), and reduced the expression of anti-apoptotic proteins, such as BCL-2 apoptosis regulator (BCL-2), MCL-1 apoptosis regulator, BCL-2 family member (MCL-1), and survivin. PEMF stimulation also increased CAPE-induced phosphorylation of p53, and inhibition of p53 partially restored the PEMF-reduced viability of CAPE-treated MCF-7 cells. PEMF stimulation enhanced CAPE-induced cell death by activating p53, which regulates the expression of apoptosis-related molecules, subsequently activating the caspase-dependent apoptotic pathway in MCF-7 cells, suggesting that PEMF can be utilized as an adjuvant to enhance the effect of CAPE on breast cancer cells.

Enhancement of anti-PD-L1 antibody plus anlotinib efficacy due to downregulation of PD-L1 in the micro-conduit endothelium within the tumor: a randomized double-blind trial.

The possible enhancing effect of anlotinib on programmed death receptor ligand (PD-L1) antibody and the efficacy-predicting power of PD-L1 in micro-conduit endothelium, including lymphatic endothelial cells (LECs) and blood endothelial cells (BECs), were determined to identify patients who would benefit from this treatment. PD-L1 positivity in LECs, BECs, and tumor cells (TCs) was assessed using paraffin sections with multicolor immunofluorescence in an investigator's brochure clinical trial of TQB2450 (PD-L1 antibody) alone or in combination with anlotinib in patients with non-small cell lung cancer. Progression-free survival (PFS) with different levels of PD-L1 expression was compared between the two groups. Among 75 patients, the median PFS (mPFS) was longer in patients who received TQB2450 with anlotinib [10 and 12 mg (161 and 194 days, respectively)] than patients receiving TQB2450 alone (61 days) [hazard ratio (HR)10 mg = 0.390 (95% confidence interval {CI}, 0.201-0.756), P = 0.005; HR12 mg = 0.397 (0.208-0.756), P = 0.005]. The results were similar among 58 patients with high PD-L1 expression in LECs and TCs [159 and 209 vs. 82 days, HR10 mg = 0.445 (0.210-0.939), P = 0.034; HR12 mg = 0.369 (0.174-0.784), P = 0.009], and 53 patients with high PD-L1 expression in BECs and TCs [161 and 209 vs. 41 days, HR10 mg = 0.340 (0.156-0.742), P = 0.007; HR12 mg = 0.340 (0.159-0.727), P = 0.005]. No differences were detected in the mPFS between the TQB2450 and combination therapy groups in 13 low/no LEC-expressing and 18 low/no BEC-expressing PD-L1 cases. Mono-immunotherapy is not effective in patients with high PD-L1 expression in LECs and/or BECs. Anlotinib may increase efficacy by downregulating PD-L1 expression in LECs and/or BECs, which is presumed to be a feasible marker for screening the optimal immune patient population undergoing anti-angiogenic therapy.