Figure S5 Blocking WNT7A synergizes with ICB.

2D interaction maps of small molecules obtained via protein screening of the ChemBridge library.

Figure S2 Blocking WNT7A leads to an increase in MHC-I expression.

Figure S4 Inhibition of WNT7A leads to the upregulation of MHC-I via the NF-κB pathway.

Figure S6 Discovery of small chemical inhibitors targeting WNT7A–FZD5.

Figure S3 WNT7A blockage inhibits tumor growth and enhances anti-tumor immunity.

Figure S1 High WNT7A expression correlates with low CTL cell infiltration, reduced MHC-I expression, and unfavorable patient survival.

Figure S7 1,365-0,109 improves ICB efficacy without significant toxic side effects.

The metabolic basis of tumor-associated macrophages (TAMs)-driven immune checkpoint blockade (ICB) resistance remains poorly understood. Here, in patients with immunotherapy-resistant cancer, we identify significant enrichment of TAMs marked by elevated aldehyde dehydrogenase 2 (ALDH2) expression. Myeloid-restricted ALDH2 ablation converts TAMs from a pro-tumorigenic phenotype to immunostimulatory regulators, concomitantly amplifying CD8+ T cell infiltration and cytotoxicity to improve ICB responsiveness. Mechanistically, ALDH2 deficiency induces the intracellular accumulation of reactive aldehydes, specifically 4-hydroxynonenal, which activates the PI3K-AKT signaling axis. This pathway phosphorylates and suppresses EZH2 methyltransferase activity, leading to the erosion of H3K27me3-mediated epigenetic silencing at CXCL9 promoter region. Subsequent CXCL9 derepression in TAMs facilitates persistent CD8+ T cell infiltration and enhances their cytotoxic effector functions. Clinical validation confirms that pronounced ALDH2 elevation in TAMs correlates with accelerated immunotherapy failure. Therapeutically, as a clinically approved ALDH2 inhibitor, disulfiram exerts its anti-tumor effect by selective reprogramming TAMs metabolism. Overall, our findings delineate a druggable ALDH2-metabolism-epigenetics axis in antitumor immunity, nominating ALDH2 inhibition for combination immunotherapy.

Abstract The limited infiltration of CD8+ T cells in tumors hampers the effectiveness of T cell–based immunotherapy, yet the mechanisms that limit tumor infiltration by CD8+ T cells remain unclear. Through bulk RNA sequencing of human tumors, we identified a strong correlation between WNT7A expression and reduced CD8+ T-cell infiltration. Further investigation demonstrated that inhibiting WNT7A substantially enhanced MHC-I expression on tumor cells. Mechanistically, WNT7A inhibition inactivated the Wnt/β-catenin signaling pathway and thus resulted in reduced physical interaction between β-catenin and p65 in the cytoplasm, which increased the nuclear translocation of p65 and activated the NF-κB pathway, ultimately promoting the transcription of genes encoding MHC-I molecules. We found that our lead compound, 1365-0109, disrupted the protein–protein interaction between WNT7A and its receptor FZD5, resulting in the upregulation of MHC-I expression. In murine tumor models, both genetic and pharmaceutical suppression of WNT7A led to increased MHC-I levels on tumor cells, and consequently enhanced the infiltration and functionality of CD8+ T cells, which bolstered antitumor immunity and improved the effectiveness of immune checkpoint blockade therapy. These findings have elucidated the intrinsic mechanisms of WNT7A-induced immune suppression, suggesting that therapeutic interventions targeting WNT7A hold promise for enhancing the efficacy of immunotherapy.

Background: HER2-positive breast cancer patients with a high risk of recurrence are recommended extended treatment involving sequential therapy with anti-HER2 monoclonal antibody followed by maintenance therapy with HER2-targeted tyrosine kinase inhibitor (TKI). Pyrotinib, an oral irreversible pan-HER TKI, commonly induces diarrhea. This study aims to develop strategies to reduce the incidence and severity of diarrhea and evaluate the efficacy and safety of pyrotinib as an extended adjuvant treatment for early-stage HER2-positive breast cancer. Methods: In this multicenter, prospective, randomized cohort study (ChiCTR2200060339), stage II-III HER2-positive high-risk breast cancer patients who had completed adjuvant treatment post-surgery, stratified by disease stage (II vs. III) and hormone receptor (HR) status (positive vs. negative), were randomized 1:1:1 into three cohorts to receive extended adjuvant therapy with 12 months of pyrotinib. Cohort 1 received pyrotinib 240 mg daily from D1-14, 320 mg from D15-28, and 400 mg from D29-364. Cohort 2 received pyrotinib 320 mg daily from D1-28 and 400 mg from D29-364. Cohort 3 received pyrotinib 400 mg daily from D1-364, combined with loperamide 4 mg thrice daily from D1-7 and twice daily from D8-28. The primary endpoint was the incidence of grade ≥3 diarrhea. Secondary endpoints included serious adverse events (AEs), other AEs, incidence of diarrhea, invasive disease-free survival, and overall survival. Results: Between June 2022 and May 2024, 102 patients were enrolled, with 34 patients in each cohort. Twelve patients (35.3%) in Cohort 1, 13 (38.2%) in Cohort 2, and 13 (38.2%) in Cohort 3 had ≥4 lymph nodes involved post-surgery. Additionally, 19 patients (55.9%) in Cohort 1, 21 (61.8%) in Cohort 2, and 19 (55.9%) in Cohort 3 were HR positive. Grade ≥3 diarrhea was reported in 9 (26.5%) patients in Cohort 1, 9 (26.5%) in Cohort 2, and 5 (14.7%) in Cohort 3. The median time to the first occurrence of grade ≥3 diarrhea was 15 days (range: 8-296) in Cohort 1, 4 days (range: 1-43) in Cohort 2, and 3 days (range: 1-14) in Cohort 3. The median number of grade ≥3 diarrhea episodes was 1 (range: 1-3) in Cohort 1, 3 (range: 1-4) in Cohort 2, and 1 (range: 1-2) in Cohort 3. The median duration of each grade ≥3 diarrhea episode was 1 day (range: 1-2) in Cohort 1, 1 day (range: 1-4) in Cohort 2, and 1 day (range: 1-3) in Cohort 3. Dose reduction due to diarrhea occurred in 2 (5.9%) patients in Cohort 1, 7 (20.6%) in Cohort 2, and 10 (29.4%) in Cohort 3. Dose interruption for diarrhea was observed in 14 (41.2%) patients in Cohort 1, 15 (44.1%) in Cohort 2, and 16 (47.1%) in Cohort 3. Discontinuation of pyrotinib due to diarrhea was reported in 2 (5.9%) patients in Cohort 1, 4 (11.8%) in Cohort 2, and 2 (5.9%) in Cohort 3. The most common AEs were diarrhea (79.4% vs. 91.2% vs. 88.2%), nausea (17.6% vs. 20.6% vs. 44.1%), vomiting (11.8% vs. 8.8% vs. 35.3%), fatigue (14.7% vs. 17.6% vs. 26.5%), abdominal pain (8.8% vs. 11.8% vs. 20.6%), and abdominal distension (5.9% vs. 5.9% vs. 17.6%). Grade 1 constipation was observed in 1 patient (2.9%) in Cohort 1, 1 patient (2.9%) in Cohort 2, and 3 patients (8.8%) in Cohort 3. Conclusions: Loperamide prophylaxis showed a numerically lower incidence of grade ≥3 diarrhea. Diarrhea is a predictable and manageable side effect of pyrotinib treatment, occurring early after initiation and rarely in the later treatment course. It is advisable to combine loperamide with pyrotinib to reduce grade ≥3 pyrotinib-induced diarrhea. Additionally, timely adjustment of loperamide dose is necessary to prevent other gastrointestinal AEs, such as nausea and abdominal distension. Citation Format: Qiao Cheng, Hongyuan Li, Xiaowei Qi, Zhen Zhang, Fuyun Tong, Maoshan Chen, Hai Lei, Lei Xing, Xiang Zhang, Mengyuan Wang, Chao Zhang, Chunyan Li, Qian Wang, Yuxian Wei, Fan Li, Jinxiang Tan, Xuedong Yin, Yixiao Feng, Xiaoyi Wang, Man Huang. Optimizing Tolerability of Pyrotinib in HER2-Positive Early-Stage High-Risk Breast Cancer: A Multicenter, Randomized, Three-Cohort Study [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P1-04-12.

INTRODUCTION:Although autologous fat grafting has been widely adopted globally to improve poor cosmetic outcomes following breast cancer surgery, oncologic concerns persist regarding the potential risk of cancer recurrence associated with fat transfer performed near the tumor bed. We sought to prospectively evaluate the oncologic safety and clinical benefits for breast cancer women undergoing breast-conserving surgery (BCS) with immediate autologous fat grafting (IAFG). METHODS:This multicenter, prospective, randomized controlled clinical trial enrolled 360 women diagnosed with breast cancer between 3 March 2017 and 31 May 2021. Participants were randomly assigned in a 1:1 ratio to either the BCS with IAFG (IAFG group) or the BCS without IAFG (control group). The primary outcomes were the cumulative incidence rates of locoregional and systemic recurrence. The secondary outcomes included adverse events, patient's satisfaction, and psychosocial well-being. RESULTS:The average volume resected and volume grafted were 45.5 g and 66.5 mL, respectively, in the IAFG group. At a median follow-up period of 62.8 months (range: 4.3-86 months), the proportions of local relapse were 0.6% and 2.4% in the IAFG group and the control group, respectively (P = 0.65). There was no increased risk of distant recurrence (3.6% vs. 3.5%) or breast cancer-specific mortality (0.6% vs. 0.6%) in the IAFG group. Among the secondary outcomes, the occurrence rates of complications were similar; however, the IAFG group showed significantly higher scores of satisfaction with breast appearance, psychosocial well-being, and sexual well-being than the controls (78 vs. 65, P < 0.001; 83 vs. 71, P < 0.001; 74 vs. 66, P < 0.001, respectively). CONCLUSION:Our results provide clear evidence that IAFG is a safe and effective surgical technique that does not increase the risk of local or distant recurrence in breast cancer patients and yields higher satisfaction with postoperative breast appearance and psychosocial outcomes.

T cell dysfunction enables tumor immune evasion, understanding its mechanism is crucial for improving immunotherapy. Here we show, by RNA-sequencing analysis of human colon adenocarcinoma and triple-negative breast cancer tissues, that expression of Adipocyte Enhancer-Binding Protein 1 (AEBP1) positively correlates with T cell dysfunction and indicative of unfavorable patient outcomes. Subsequent single-cell RNA sequencing identifies cancer-associated fibroblasts (CAF) as the primary AEBP1 source. Fibroblast-specific AEBP1 deletion in mice enhances T cell cytotoxicity and suppresses tumor growth. Mechanistically, autocrine AEBP1 binds CKAP4 on CAFs, activating AKT/PD-L1 signaling to drive T cell dysfunction. By molecular-docking-based virtual screening we identify Chem-0199, a drug that disrupts the interaction between AEBP1 and CKAP4, thereby enhancing antitumor immunity. Both genetic and pharmacological AEBP1 inhibition synergize with immune checkpoint blockade in syngeneic models. Our study establishes AEBP1 as a key regulator of CAF-mediated T cell dysfunction and a therapeutic target.

AIMS:To identify the multiple mediating effects of resilience and depression between social support and self-care ability among patients with breast cancer during rehabilitation to provide reference for developing and implementing targeted interventions. DESIGN:A cross-sectional study reported according to the STROBE checklist. METHODS:A convenience sample of 320 patients with breast cancer during rehabilitation was recruited from one hospital in China. Data were collected from April to August 2022 using a self-report questionnaire, including the demographic and clinical information, Appraisal of Self-Care Agency Scale-Revised, Multidimensional Scale of Perceived Social Support, Connor-Davidson Resilience Scale-10 item, and Patient Health Questionnaire-9. The mediation analysis was conducted using the SPSS Process macro. RESULTS:Self-care ability was positively associated with social support (β = .229) and resilience (β = .290), and negatively associated with depression (β = -.208). The relationship between social support and self-care ability was mediated by resilience and depression, respectively, and together in serial. The multiple mediating effects accounted for 34.0% of the total effect of social support on self-care ability. CONCLUSION:Our findings identify resilience and depression as multiple mediators between social support and self-care ability and highlight the important roles of social support, resilience and depression in improving self-care ability. RELEVANCE TO CLINICAL PRACTICE:Healthcare providers should pay great attention to the underlying mechanisms of how social support affects patients' self-care ability during breast cancer rehabilitation. Integrated intervention programmes targeted at enhancing social support, building resilience and alleviating depression might be beneficial to the improvement of self-care ability. PATIENT OR PUBLIC CONTRIBUTION:No patient or public contribution. REPORTING METHOD:The Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) checklist for cross-sectional studies was applied to report the results.

BackgroundGlioma, an aggressive brain tumor, poses a challenge in understanding the mechanisms of treatment resistance, despite promising results from immunotherapy.MethodsWe identified genes associated with immunotherapy resistance through an analysis of The Cancer Genome Atlas (TCGA), Chinese Glioma Genome Atlas (CGGA), and Gene Expression Omnibus (GEO) databases. Subsequently, qRT-PCR and western blot analyses were conducted to measure the mRNA and protein levels of TBC1 Domain Family Member 1 (TBC1D1), respectively. Additionally, Gene Set Enrichment Analysis (GSEA) was employed to reveal relevant signaling pathways, and the expression of TBC1D1 in immune cells was analyzed using single-cell RNA sequencing (scRNA-seq) data from GEO database. Tumor Immune Dysfunction and Exclusion (TIDE) database was utilized to assess T-cell function, while Tumor Immunotherapy Gene Expression Resource (TIGER) database was employed to evaluate immunotherapy resistance in relation to TBC1D1. Furthermore, the predictive performance of molecules on prognosis was assessed using Kaplan-Meier plots, nomograms, and ROC curves.ResultsThe levels of TBC1D1 were significantly elevated in tumor tissue from glioma patients. Furthermore, high TBC1D1 expression was observed in macrophages compared to other cells, which negatively impacted T cell function, impaired immunotherapy response, promoted treatment tolerance, and led to poor prognosis. Inhibition of TBC1D1 was found to potentially synergistically enhance the efficacy of immunotherapy and prolong the survival of cancer patients with gliomas.ConclusionHeightened expression of TBC1D1 may facilitate an immunosuppressive microenvironment and predict a poor prognosis. Blocking TBC1D1 could minimize immunotherapy resistance in cancer patients with gliomas.

Traditional treatments against advanced non-small cell lung cancer (NSCLC) with high morbidity and mortality continue to be dissatisfactory. Given this situation, there is an urgent requirement for alternative modalities that provide lower invasiveness, superior clinical effectiveness, and minimal adverse effects. The combination of photodynamic therapy (PDT) and immunotherapy gradually become a promising approach for high-grade malignant NSCLC. Nevertheless, owing to the absence of precise drug delivery techniques as well as the hypoxic and immunosuppressive characteristics of the tumor microenvironment (TME), the efficacy of this combination therapy approach is less than ideal. In this study, we construct a novel nanoplatform that indocyanine green (ICG), a photosensitizer, loads into hollow manganese dioxide (MnO2) nanospheres (NPs) (ICG@MnO2), and then encapsulated in PD-L1 monoclonal antibodies (anti-PD-L1) reprogrammed exosomes (named ICG@MnO2@Exo-anti-PD-L1), to effectively modulate the TME to oppose NSCLC by the synergy of PDT and immunotherapy modalities. The ICG@MnO2@Exo-anti-PD-L1 NPs are precisely delivered to the tumor sites by targeting specially PD-L1 highly expressed cancer cells to controllably release anti-PD-L1 in the acidic TME, thereby activating T cell response. Subsequently, upon endocytic uptake by cancer cells, MnO2 catalyzes the conversion of H2O2 to O2, thereby alleviating tumor hypoxia. Meanwhile, ICG further utilizes O2 to produce singlet oxygen (1O2) to kill tumor cells under 808 nm near-infrared (NIR) irradiation. Furthermore, a high level of intratumoral H2O2 reduces MnO2 to Mn2+, which remodels the immune microenvironment by polarizing macrophages from M2 to M1, further driving T cells. Taken together, the current study suggests that the ICG@MnO2@Exo-anti-PD-L1 NPs could act as a novel drug delivery platform for achieving multimodal therapy in treating NSCLC.

T cell exclusion is crucial in enabling tumor immune evasion and immunotherapy resistance. However, the key genes driving this process remain unclear. We uncovered a notable increase of insulin-like growth factor 2 (IGF2) in immune- excluded tumors, predominantly secreted by cancer-associated fibroblasts (CAFs). Using mice with systemic or fibroblast- specific deletion of IGF2, we demonstrated that IGF2 deficiency enhanced the infiltration and cytotoxic activity of CD8+ T cells, leading to a reduction in tumor burden. Integration of spatial and single-cell transcriptomics revealed that IGF2 promoted interaction between CAFs and T cells via CXCL12 and programmed death ligand 1 (PD-L1). Mechanistically, autocrine IGF2 activated PI3K/AKT signaling by binding to the IGF1 receptor (IGF1R) on CAFs, which was required for the immunosuppressive functions of CAFs. Furthermore, genetic ablation of IGF2 or targeted inhibition of the IGF2/IGF1R axis with the inhibitor linsitinib markedly boosted the response to immune checkpoint blockade. Clinically, elevated levels of IGF2 in tumors or plasma correlated with an adverse prognosis and reduced efficacy of anti-programmed death 1 treatment. Together, these results highlight the pivotal role of IGF2 in promoting CAF-mediated immunoevasion, indicating its potential as a biomarker and therapeutic target in immunotherapy.