Single-Cell Profiling of Tumor Immune Microenvironment Reveals Immune Irresponsiveness in Gastric Signet-Ring Cell Carcinoma.

BACKGROUND & AIMS: Gastric cancer (GC) is a major cancer type characterized by high heterogeneity in both tumor cells and the tumor immune microenvironment (TIME). One intractable GC subtype is gastric signet-ring cell carcinoma (GSRCC), which is associated with poor prognosis. However, it remains unclear what the GSRCC TIME characteristics are and how these characteristics may contribute to clinical outcomes. METHODS: We enrolled 32 patients with advanced GC of diverse subtypes and profiled their TIME using an immune-targeted single-cell profiling strategy, including (1) immune-targeted single-cell RNA sequencing (n = 20 patients) and (2) protein expression profiling by a targeted antibody panel for mass cytometry (n = 12 patients). We also generated matched V(D)J (variable, diversity, and joining gene segments) sequencing of T and B cells along CD45+ immunocytes. RESULTS: We found that compared to non-GSRCC, the GSRCC TIME appears to be quiescent, where both CD4+ and CD8+ T cells are difficult to be mobilized, which further impairs the proper functions of B cells. CXCL13, mainly produced by follicular helper T cells, T helper type 17, and exhausted CD8+ T cells, is a central coordinator of this transformation. We show that CXCL13 expression can predict the response to immune checkpoint blockade in GC patients, which may be related to its effects on tertiary lymphoid structures. CONCLUSIONS: Our study provides a comprehensive molecular portrait of immune cell compositions and cell states in advanced GC patients, highlighting adaptive immune irresponsiveness in GSRCC and a mediator role of CXCL13 in TIME. Our targeted single-cell transcriptomic and proteomic profiling represents a powerful approach for TIME-oriented translational research.

Efficacy of PD-1 Inhibitors in First-Line Treatment for Advanced Gastroesophageal Junction and Gastric Cancer by Subgroups: A Systematic Review and Meta-Analysis.

BACKGROUND: PD-1 inhibitors have been approved for the first-line treatment of patients with advanced gastric cancer, gastroesophageal junction cancer, or esophageal adenocarcinoma. However, the results of several clinical trials are not entirely consistent, and the dominant population of first-line immunotherapy for advanced gastric/gastroesophageal junction cancer still needs to be precisely determined. OBJECTIVE: This objective of this study is to evaluate the efficacy of anti-PD-1/PD-L1 therapy in advanced gastric/gastroesophageal junction adenocarcinoma patients through a systematic review and meta-analysis of relevant clinical trials. METHOD: The PubMed, Embase, and Cochrane Library electronic databases were searched up to August 1, 2022, for clinical trials of anti-PD-1/PD-L1 immunotherapy for the first-line treatment of advanced gastroesophageal cancer. Hazard ratios and 95% confidence intervals for overall survival, progression-free survival, and objective response rates were extracted and pooled for meta-analysis. Prespecified subgroups included the following: agent type, PD-L1 expression, and high microsatellite instability. RESULTS: This study analyzed 5 RCTs involving 3,355 patients. Compared with the chemotherapy group, the combined immunotherapy group had a significantly higher objective response rate (OR = 0.63, 95% CI: 0.55-0.72, p < 0.00001) and prolonged overall survival (HR = 0.82, 95% CI: 0.76-0.88, p < 0.00001) and progression-free survival (HR = 0.75, 95% CI: 0.69-0.82, p < 0.00001). The combination of immunotherapy and chemotherapy prolonged OS in both MSI-H (HR = 0.38, p = 0.002) and MSS (HR = 0.78, p < 0.00001) populations, but there was a significant difference between groups (p = 0.02). However, in improving ORR, the benefit of ICI combined with chemotherapy in the MSS group and MSI-H group was not significantly different between groups (p = 0.52). Combination therapy with ICIs was more effective than chemotherapy alone in prolonging OS in the subgroup with a high CPS, regardless of the CPS cutoff for PD-L1. However, when the cutoff of CPS was 1, the difference between subgroups did not reach statistical significance (p = 0.12), while the benefit ratio of the MSI-H group was higher when the cutoff was 10 (p = 0.004) than when the cutoff value was 5 (p = 0.002). CONCLUSIONS: For first-line treatment of advanced gastroesophageal cancer, an ICI combination strategy is more effective than chemotherapy. The subgroup of patients with a CPS ≥10 has a more significant benefit, and CPS ≥10 has the potential to be used as an accurate marker of the dominant population of immuno-combined therapy.

Development and validation of a novel prognostic lncRNA signature based on the APOBEC3 family genes in gastric cancer.

Introduction: Gastric Cancer (GC) refers to a prevalent malignant cancer accompanied by a weak prognosis. The APOBEC3 family genes and lncRNAs are linked with cancer progression. Nevertheless, there is still a scarcity of data concerning the prognostic value of APOBEC3-related lncRNAs in GC. Methods: We extracted the data from GC samples, including transcriptome as well as clinical data, obtained from the TCGA database. Then, we screened for lncRNAs that were correlated with the APOBEC3 family genes and constructed an APOBEC3-related lncRNA prognostic signature (LPS) by utilizing univariate Cox and lasso regression analysis. Furthermore, we validated our constructed signature and evaluated it thoroughly, including analysis of its function, immunity, mutations, and clinical applications via multiple methods, including Metascape, GSEA, and analyses including TIC and TME, immune checkpoints, CNV and SNPs, Kaplan-Meier survival curves, nomogram, decision tree and drug prediction analysis. Finally, we overexpressed LINC01094 to evaluate the impacts on the proliferation as well as migration with regards to KATO-2 cells. Results: We selected eight lncRNAs for our APOBEC3-related LPS, which is demonstrated as a valuable tool in predicting the individual GC patients' prognosis. Subsequently, we segregated the samples into subgroups of high-as well as low-risk relying on the risk score with regards to APOBEC3-related LPS. By performing functional analysis, we have shown that immune-as well as tumor-related pathways were enriched in high- and low-risk GC patients. Furthermore, immune analysis revealed a robust correlation between the APOBEC3-related LPS and immunity. We found that immune checkpoints were significantly associated with the APOBEC3-related LPS and were greatly exhibited in GC tumor and high-risk samples. Mutational analysis suggested that the mutational rate was greater in low-risk samples. Furthermore, we predicted small molecular drugs displayed greater sensitivity in patients categorized as high-risk. Moreover, the immune response was also better in high-risk patients. Of these drugs, dasatinib was significant in both methods and might be considered a potential novel drug for treating high-risk GC patients. Finally, we found that LINC01094 has the potential to enhance the migration, proliferation as well as inhibit apoptosis of KATO-2 in GC cells. And Dasatinib has an inhibitory effect on the migration as well as proliferation in GC cells. Conclusion: We created a novel APOBEC3-related LPS in predicting the prognosis with regards to individual GC patients. Importantly, this APOBEC3-related LPS was closely associated with immunity and might guide clinical treatment.

Distinct roles of miR-34 family members on suppression of lung squamous cell carcinoma.

miR-34, whose mimic was used on phase I clinical trial, has been extensively reported since its dysfunction in various cancers including non-small-cell lung cancer (NSCLC). However, the roles of miR-34 family members in the progression of lung squamous carcinoma (SCC) in patients who have occupational-exposure experience are unclear yet. Here, we comprehensively investigated the expression levels of miR-34 family members in SCC patients and compared the roles of them in SCC in vitro and vivo. The results showed that the average levels of miR-34a and miR-34b/c were decreased in patients. The analysis of miR-34a to miR-34b/c levels in patients graded different stages or metastases or recurrence showed that miR-34b/c was reduced earlier and more significantly than miR-34a. In vitro assays demonstrated that both miR-34a and miR-34b/c inhibits SCC cells proliferation, migration and invasion via Notch1 pathway, while miR-34b/c effects more than miR-34a does. As miR-34a was significantly decreased in cancer recurrence, the further analysis of relationship between miR-34a and stem cell adhesion molecular CD44 showed that miR-34a was significantly correlated with CD44 levels in patients. Knockdown of CD44 significantly blocked miR-34a mediated inhibition of cell migration and invasion. Treating the purified CD44hi cells with miR-34 overexpression lentivirus inhibited the tumor outgrowth. By contrast, anti-miR-34 facilitated tumor development of CD44low cells. Our study showed that miR-34 family members are negative regulator for SCC development, even though the inhibition is mediated by multiple and complicated signal pathways, which provides theoretical basis for SCC treatment and a biomarker candidate for SCC prognosis.

Dimerization-induced self-assembly of a redox-responsive prodrug into nanoparticles for improved therapeutic index.

Although some formats of nanomedicines are now available for clinical use, the translation of new nanoparticles to the clinic remains a considerable challenge. Here, we describe a simple yet cost-effective strategy that converts a toxic drug, cabazitaxel, into a safe and effective nanomedicine. The strategy involves the ligation of drug molecules via a self-immolating spacer, followed by dimerization-induced self-assembly to assemble stable nanoparticles. Self-assembled cabazitaxel dimers could be further refined by PEGylation with amphiphilic polymers suitable for preclinical studies. This protocol enables the formation of systemically injectable nanoparticles (termed SNPs) with nearly quantitative entrapment efficiencies and exceptionally high drug loading (> 86%). In healthy mice, PEGylated SNPs show a favorable safety profile, with reduced systemic toxicity and negligible immunotoxicity. In two separate mouse xenograft models of cancer, administration of SNPs produces efficient antitumor activity with durable tumor suppression during therapeutic studies. Overall, this methodology opens up a practical and expedient route for the fabrication of clinically useful nanomedicines, transforming a hydrophobic and highly toxic drug into a systemic self-deliverable nanotherapy. STATEMENT OF SIGNIFICANCE: Despite the great progress in cancer nanomedicines, clinical translation of nanomedicines still remains a considerable challenge. In this study, we designed a self-assembling nanoplatform based on cabazitaxel dimer reversibly ligated via a bioactivatable linker. This approach enabled the generation of systemically injectable nanomedicines with quantitative entrapment efficiencies and exceptionally high drug loading (> 86%), which greatly obviates concerns about excipient-associated side effects. Self-assembled dimeric cabazitaxel exhibited a higher safety profile than free cabazitaxel and negligible immunotoxicity in animals. This is a practical and expedient example how the chemical ligation of a hydrophobic and highly toxic anticancer drug can be leveraged to create a self-assembling delivery nanotherapy which preserves inherent pharmacologic efficacy while reduces in vivo systemic and immune toxicity.

A temperature-sensitive phase-change hydrogel of tamoxifen achieves the long-acting antitumor activation on breast cancer cells.

Background: Breast cancer is one of the foremost threats to female health nowadays. Tamoxifen, an antagonist of estrogen receptor-α (ERα), is the first choice for endocrine-dependent breast cancer (ERα-positive breast cancer) treatment. However, ERα has an important function in the normal physical regulation of estrogen, and current oral administration of tamoxifen has potential side effects on normal endocrine secretion. In the present work, we aim to develop novel approaches to increase the antitumor effect of tamoxifen on breast cancer cells and decrease the potential side effects in the human body during treatment. Methods: A temperature-sensitive phase-change hydrogel for tamoxifen (Tam-Gel) was generated. After establishing subcutaneous tumors formed by MCF-7, an ERα-positive breast cancer cell line, in nude mice, an intratumoral injection of Tam-Gel was performed to examine whether Tam-Gel facilitated the slow-release or antitumor effect of tamoxifen. A metastatic breast cancer model was established using the intrahepatic growth of MCF-7 cells in immunodeficient rats. Results: Tam-Gel can transform from liquid to hydrogel at room temperature. An intratumoral injection of Tam-Gel facilitated the slow-release or antitumor effect of tamoxifen. Once Tam-Gel, but not Tam-Sol, was administered by intratumoral injection, it significantly decreased the uptake of radionuclide probes (18F-fluoroestradiol or 18F-fluorodeoxyglucose) by cells in rats' livers and the intrahepatic growth of MCF-7 cells in rats' livers. Conclusion: A novel slow-release system was successfully prepared to facilitate the long-term release of tamoxifen in breast cancer tissues, and achieved an antitumor effect in the long term.

MicroRNA-645 targets urokinase plasminogen activator and decreases the invasive growth of MDA-MB-231 triple-negative breast cancer cells.

BACKGROUND: Urokinase plasminogen activator (uPA) promotes the in vivo invasive growth of HCC cells by cleaving and activating matrix metalloproteinases (MMPs) to induce the destruction of the extracellular matrix of triple-negative breast cancer (TNBC) cells. The identification of microRNAs that target uPA and decrease uPA expression would be useful for attenuating the in vivo invasive growth of TNBC cells. MATERIALS AND METHODS: MicroRNA-645 (miR-645) was identified using an online tool (miRDB) as potentially targeting uPA; miR-645 inhibition of uPA was confirmed by western blot experiments. The effects of miR-645 on the in vivo invasive growth of TNBC cells were examined using an intrahepatic tumor model in nude mice, and the miR-645 mechanism of action was explored with MMP cleaving experiments. RESULTS: Through virtual screening, we discovered that miR-645 potentially targeted the uPA 3' untranslated region. This targeting was confirmed by western blot experiments and miR-645 lentiviral particle (LV-645) transduction that inhibited uPA expression in MDA-MB-231 TNBC cells. The LV-645 inhibition of uPA led to the decreased invasive growth of TNBC cells in nude mice. The mechanism data indicated that the uPA inhibition resulted in a decreased cleaving of the pro-MMP-9 protein. CONCLUSION: Targeting uPA with miR-645 decreased the in vivo invasive growth of TNBC cells. These results suggest that miR-645 may represent a promising treatment strategy for TNBC.