NR5A2 is a key regulator of zygotic genome activation (ZGA), which primarily takes place at the 4-8-cell stage in porcine embryos; however, its associated metabolic programs remain largely uncharacterized. In this study, single-blastomere RNA sequencing was performed using Smart-seq2 in 8-cell-stage porcine embryos derived from in vitro fertilization (IVF) or somatic cell nuclear transfer (SCNT). Based on the bimodal expression pattern of NR5A2, individual blastomeres were categorized into NR5A2-active and NR5A2-inactive groups. Differential expression analysis identified 46 differentially expressed genes (DEGs) in IVF embryos, which were mainly enriched in steroid hormone and lipid metabolism. In SCNT embryos, the top-ranked DEGs were functionally associated with mitochondrial respiration and ribosome biogenesis. A total of 14 NR5A2-associated DEGs were consistently identified in both embryo types and clustered into three core metabolic modules: glycolysis and energy metabolism (represented by HK2), lipid and steroid metabolism (including STAR, LIPG, PLCD1 and DLC1), and inositol metabolism (represented by MIOX). Single-blastomere qRT-PCR in parthenogenetic embryos further validated the bimodal expression of NR5A2, HK2, STAR and MIOX. Collectively, this study reveals an NR5A2-associated metabolic transcriptional signature in blastomeres of both IVF and SCNT embryos. These findings identify candidate metabolic genes that may inform future optimization of in vitro culture media to enhance the developmental competence of porcine embryos and warrant subsequent functional validation.

Squamous cell carcinoma (SCC) of the bladder is a rare disease with poor prognosis and limited molecular profiling. Here we present a multi-layer, comparative investigation on bladder SCC and related cancers, namely pure urothelial carcinoma (UC), UC with squamous differentiation, and bladder adenocarcinoma. The mutational signatures of SCC, UC with squamous differentiation, and pure UC imply similar etiologies, with APOBEC-derived signatures found in around a third of samples. SCC and UC with squamous differentiation both highly express basal/squamous markers, different from the luminal profile in UC. We have also dissected the tumor microenvironment and cell type-specific expression in primary tumor and lymph node metastases of SCC at single-cell resolution, and SCC tumor cells also exhibit high expression of basal/squamous markers compared with the luminal feature in UC and adenocarcinoma. Similar mutational signatures present at different contributing fractions, combined with distinct transcriptomic features in various types of bladder cancer, provide an interesting perspective on etiological factors and suggest tumor initiation from potentially different urothelial cell types. The molecular landscape of SCC and related bladder cancers presented in this study improves our understanding of their etiologies and possible cellular origins, and may facilitate future prevention strategies and therapy development. © 2026 The Pathological Society of Great Britain and Ireland.

PURPOSE: This study aimed to characterize the clinicopathological, immunophenotypic, and molecular features of gastrointestinal stromal tumors (GISTs) harboring NTRK fusions and to evaluate their diagnostic, prognostic, and therapeutic implications. METHODS: Twenty-six cases of KIT/PDGFRA/SDH/BRAF wild-type GISTs were evaluated using pan-TRK immunohistochemistry (IHC), fluorescence in situ hybridization (FISH) for NTRK1/2/3, and next-generation sequencing (NGS). Transcriptome analysis was performed on all NTRK fusion-positive cases. Seven KIT-mutant GISTs served as controls. Clinicopathological parameters, IHC profiles, genetic alterations, and treatment responses were analyzed, supplemented by a literature review. RESULTS: Five of the 26 wild-type GISTs harbored NTRK fusions, all confirmed by NGS as ETV6::NTRK3. Pan-TRK IHC showed 100% sensitivity and 66.7% specificity. All five patients were male; four tumors were intestinal and one gastric. Four cases were high-risk and one very low-risk. Two cases recurred post-resection, showing additional mutations and copy number variations (CNVs). Preliminary evidence from transcriptome sequencing pointed to the possibility that NTRK fusion-positive GISTs represented a heterogeneous group and showed similarities in their molecular profiles to common KIT-mutant GISTs. Both recurrent patients received multi-line TKI therapy (imatinib, sunitinib, regorafenib, ripretinib) with disease progression; one subsequently achieved remission with larotrectinib. CONCLUSION: NTRK fusion-positive GISTs are rare and exhibit distinct clinicopathological characteristics. FISH and NGS are reliable detection methods, while pan-TRK IHC has limited specificity. Co-occurring genetic alterations may confer aggressive behavior. These tumors respond to TRK inhibition but are resistant to conventional TKIs, underscoring the need for molecularly guided therapy.

The prognosis and progression mechanisms of bladder cancer (BLCA) are highly heterogeneous, driven by complex genetic and epigenetic alterations. This study aimed to construct a robust prognostic signature using epigenetic modification-related genes and to investigate the underlying molecular mechanisms driving its predictive power. We developed a prognostic signature by applying a machine learning-based approach to screen epigenetic genes in the TCGA (The Cancer Genome Atlas)-BLCA cohort. Its performance was rigorously evaluated against 101 other machine learning algorithms and 110 previously published signatures across four independent validation datasets (IMvigor210, E-MTAB-4321, GSE31684, GSE48075). Associations with clinical, genetic, and transcriptomic features were analyzed. Immune infiltration, cell-cell interactions, and drug responses were assessed using both bulk and single-cell RNA-seq data. The functional role of a key signature gene, YTHDC1 (YTH Domain-Containing 1), was investigated through in vitro assays. A six-gene epigenetic signature was constructed. It significantly stratified patients into high- and low-risk groups with distinct overall survival (median survival 20.5 vs. 86.8 months, HR = 2.12, p = 7.7e-7). Our signature demonstrated superior predictive accuracy (C-index and 1-year AUROC) compared to other models. High-risk scores correlated with adverse clinical features (e.g., advanced stage), elevated PD-1/PD-L1, higher genomic instability, and immunosuppressive microenvironments. Single-cell analysis revealed altered T-cell interactions in high-risk cases. Mechanistically, YTHDC1 was shown to bind and stabilize POU5F1 (OCT4) mRNA, thereby inhibiting proliferation and migration in BLCA cell lines (T24, 5637). This anti-tumor effect was dependent on POU5F1. The machine learning-derived epigenetic signature is a robust indicator of BLCA heterogeneity across multiple dimensions. YTHDC1, a core component, inhibits cancer progression by stabilizing POU5F1 mRNA, highlighting a novel regulatory axis.

BACKGROUND:Streptococcus anginosus has been linked with an increasing risk of gastric cancer (GC) and recognised as a signature for GC screening. OBJECTIVE:To investigate the promotional effect of S. anginosus in terms of its metabolic interactions with the host. DESIGN:We used the functional profiles of shotgun metagenomic sequencing from stools to detect bioactive molecules relevant to S. anginosus. In vivo and in vitro experiments were used to validate the facilitation of S. anginosus to GC progression. S. anginosus clinical strains were isolated and cultivated from cancerous tissues to verify its promotion of GC via methionine production. S. anginosus ΔmetE mutant strains were constructed to confirm the critical role of metE in methionine biosynthesis. RESULTS:We verified S. anginosus facilitated GC progression in vivo and in vitro. Our functional analysis of metagenomes revealed a significant enrichment of bacterial methionine biosynthesis pathways in GC patients with high S. anginosus abundance. Methionine, identified here as one of the primary microbial metabolites derived from S. anginosus, contributed to GC progression in humans and mice. S. anginosus strains from cancerous tissues were found to promote GC via methionine production. We further observed a higher abundance and prevalence of metE gene in cancer stool metagenomes. By constructing an S. anginosus ΔmetE mutant strain, we confirmed the critical role of metE in methionine biosynthesis. CONCLUSION:Our results elucidate the role of S. anginosus-derived methionine in GC progression, shedding light on intricate metabolic interplay between S. anginosus and host.

BACKGROUND:The ketogenic diet (KD) is an established therapy for drug-resistant epilepsy, yet its antiseizure mechanisms remain unclear. This study investigates whether KD modulates lipid metabolism-related genes to exert antiseizure effects in a mouse model of temporal lobe epilepsy. METHODS:Chronic epilepsy was induced in adult mice via systemic kainic acid (KA). Epileptic mice were fed either a standard chow diet (CD) or KD. Seizure activity was monitored via electroencephalography and behavioral scoring. Temporal lobe transcriptomic changes were analyzed using RNA sequencing. The role of the peroxisome proliferator-activated receptor gamma (PPARγ) pathway was investigated in vivo and in primary hippocampal neurons using pharmacological agonists/antagonists and siRNA-mediated knockdown. RESULTS:KD-fed mice exhibited significantly reduced seizure frequency and severity compared to CD-fed controls. Transcriptomic analysis identified 41 differentially expressed lipid metabolism-related genes in the KD group. Perilipin 1 (Plin1) emerged as a top hub gene and was markedly upregulated in the hippocampus of KD-fed mice, associated with a significant increase in neuronal lipid droplet formation. In vitro, KD's effects were recapitulated by a PPARγ agonist and blocked by an antagonist, demonstrating that PPARγ mediates this transcriptional regulation. Knockdown of Plin1 in KA-treated neurons reduced lipid droplet formation and concurrently increased neuronal excitability (c-Fos expression), establishing a direct mechanistic link. CONCLUSION:The ketogenic diet exerts antiseizure effects that may be related to activation of the PPARγ pathway, which upregulates Plin1 to promote lipid droplet formation in hippocampal neurons. This sequestration of fatty acids represents a novel mechanism for reducing neuronal excitability in chronic epilepsy.

The Cre-lox site-specific recombinase system is one of the most powerful and versatile technology platforms for studying neural stem cells (NSCs) in adult brain, which is now challenged due to the complex and dynamic nature of in vivo gene expression. In this study, we develop an inducible dual recombinase-mediated intersectional genetics by combining Dre-rox and Cre-lox recombination technologies to specifically target two subpopulations of NSCs (α- and β-NSCs). By visiting their cell lineage and functionality, we find that α- and β-NSCs display distinct self-renewal and differentiation potential, as well as differential responses to external stimuli. Notably, in contrast to α-NSCs, the number of β-NSCs is not affected in aged mice and an APP/PS1 mouse model of Alzeimer’s disease. Single cell transcriptome analysis reveals divergent molecular signatures between type α- and β-NSCs and identifies PRMT1 as an important regulatory element to differentially regulate the neurogenic potential of α- and β-NSCs. Inhibition of PRMT1 specifically enhances the neurogenic capacity of β-NSCs and promotes the cognition functions in aged mice. Importantly, PRMT1 inhibition combined with increased BDNF levels pharmacologically ameliorates the cognitive impairments in APP/PS1 mice. Together, our study suggests that understanding the functional heterogeneity of NSCs might pave the way for harnessing the specific subpopulation of NSCs to treat brain disorders.

Background Anorexia nervosa (AN) has been characterised as a psychiatric disorder associated with increased control. Currently, it remains difficult to predict treatment response in patients with AN. Their cognitive abilities are known to be resistant to treatment. It has been established that the frontoparietal control network (FPCN) is the direct counterpart of the executive control network. Therefore, the resting-state brain activity of the FPCN may serve as a biomarker to predict treatment response in AN.Aims The study aimed to investigate the association between resting-state functional connectivity (RSFC) of the FPCN, clinical symptoms and treatment response in patients with AN.Methods In this case-control study, 79 female patients with AN and no prior treatment from the Shanghai Mental Health Center and 40 matched healthy controls (HCs) were recruited from January 2015 to March 2022. All participants completed the Questionnaire Version of the Eating Disorder Examination (version 6.0) to assess the severity of their eating disorder symptoms. Additionally, RSFC data were obtained from all participants at baseline by functional magnetic resonance imaging. Patients with AN underwent routine outpatient treatment at the 4th and 12th week, during which time their clinical symptoms were evaluated using the same measures as at baseline.Results Among the 79 patients, 40 completed the 4-week follow-up and 35 completed the 12-week follow-up. The RSFC from the right posterior parietal cortex (PPC) and dorsolateral prefrontal cortex (dlPFC) increased in 79 patients with AN vs 40 HCs after controlling for depression and anxiety symptoms. By multiple linear regression, the RSFC of the PPC to the inferior frontal gyrus was found to be a significant factor for self-reported eating disorder symptoms at baseline and the treatment response to cognitive preoccupations about eating and body image, after controlling for age, age of onset and body mass index. The RSFC in the dlPFC to the middle temporal gyrus and the superior frontal gyrus may be significant factors in the treatment response to binge eating and loss of control/overeating in patients with AN.Conclusions Alterations in RSFC in the FPCN appear to affect self-reported eating disorder symptoms and treatment response in patients with AN. Our findings offer new insight into the pathogenesis of AN and could promote early prevention and treatment.

Abstract 4-1BB (CD137, TNFRSF9) is a co-stimulatory receptor initially identified on T cells, belonging to the tumor necrosis factor receptor superfamily. Primarily expressed on activated CD8+ T cells, it plays a crucial role in immune activation. Moreover, the intracellular domain of 4-1BB is leveraged to enhance the proliferation and cytotoxicity of Chimeric Antigen Receptor-T (CAR-T) cells. However, the development of 4-1BB agonist antibodies has encountered challenges, with limited clinical efficacy (e.g., utomilumab) or dose-dependent liver toxicity (e.g., urelumab). Consequently, there is a significant demand for the development of next-generation 4-1BB agonist antibodies, leading to numerous ongoing preclinical R&D efforts and clinical trials. Here, we introduce a novel fully humanized anti-4-1BB antibody, HLX25, and discuss the development of HLX34, a bispecific antibody targeting both 4-1BB and Her2. HLX25 boasts a unique binding epitope distinct from first-generation 4-1BB antibodies and demonstrates multiple cross-species reactivities. Additionally, HLX25 incorporates an engineered Fc region to enhance intratumoral clustering while reducing toxicity. In vitro studies reveal that HLX25 exhibits activity similar to natural 4-1BB ligands. In contrast, utolimumab shows inadequate activity, and urelumab displays superior activity, suggesting a correlation between efficacy and safety. As reported previously, (As been profiled in our Lab,) HLX25 exhibits robust dose-dependent anti-tumor activity in the MC38/h4-1BB KI mouse model. Noteworthy is the superior safety profile of HLX25 in the h4-1BB KI model, where both HLX25 and utolimumab induce no significant AST and ALT upregulation, unlike urelumab, which demonstrates such upregulation in vivo. A comparative analysis of HLX25 with other second-generation anti-4-1BB antibodies in clinical trials in vivo reveals that HLX25 provides superior tumor inhibition without observed toxicity within the effective dose range. Furthermore, we've developed HLX34, a Her2x4-1BB bispecific antibody, derived from Trastuzumab and HLX25. Its format underwent optimization using a reporter assay, indicating superior in vitro activity. Validation through a cytokine release assay with primary PBMCs in vitro and confirmed in vivo activity in animal models, without observed toxicity, strongly supports the efficacy and safety of HLX34 in Her2-positive tumors. These results provide evidence that our novel anti-4-1BB antibody activates proper 4-1BB signaling through tumor-enriched FcgRIIB or tumor-associated antigen-mediated clustering, inhibiting tumor growth with good safety in both in vitro and in vivo settings. This underscores HLX34 as a promising alternative therapeutic strategy for next-generation cancer immunotherapy. Citation Format: Jen-Kuan Chang, Jie Xue, Chen Dong, An Ju, Toya Baral, Julie Yoo, Yanling Wang, Marco Muda, Qiang Liu, Wenfeng Xu, Weidong Jiang, Lixin Feng. A novel anti-4-1BB antibody with no liver toxicity and its application in a bi-specific antibody [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5299.

PurposeHead and neck cancer cells commonly express programmed death ligand 1 (PD-L1) and epidermal growth factor receptor (EGFR), both of which play pivotal roles in the antitumor cellular immune response. Pembrolizumab, a PD-1 inhibitor, and cetuximab, an EGFR inhibitor, are typically effective agents combined with neoadjuvant platinum-based chemotherapy for the treatment of head and neck squamous cell carcinoma (HNSCC). This study aims to evaluate the efficacy and safety of neoadjuvant immunochemotherapy in patients with HNSCC.MethodsPatients with HNSCC underwent radical surgery and complete cervical lymph node dissection following neoadjuvant immunochemotherapy at RenJi Hospital from January 2021 to June 2024 were retrospectively analyzed. The primary endpoint was major pathological response (MPR). We further explored the relationship between the efficacy and immune estimators.FindingsTwenty-one patients were enrolled in this retrospective study. The MPR was 66.7%, including 11 patients who achieved a pathological complete response (pCR). The overall response rate (ORR) was 90.5%, and the complete response (CR) rate was 28.6%. The oropharynx, as the primary site, was the sensitive tumor type to neoadjuvant immunochemotherapy. The most common adverse event (AEs) was anemia (61.9%). No grade 4 AE or delayed surgery was reported. Laryngeal preservation rates were 90.9% (10/11), and pathological findings confirmed negative surgical margins for all patients. Moreover, pre-treatment peripheral lymphocyte count, monocyte count, and platelet to lymphocyte ratio (PLR) displayed a significant correlation with the treatment response.ConclusionPembrolizumab plus cetuximab with chemotherapy for patients with HNSCC is a feasible and safe clinical protocol fulfilling organ preservation and life quality improvement. Pre-treatment peripheral immune estimators could help to screen patients who may respond to the neoadjuvant immunochemotherapy.

OBJECTIVE:Branched-chain amino acid (BCAA) metabolism is involved in the development of colorectal cancer (CRC); however, the underlying mechanism remains unclear. Therefore, this study investigates the role of BCAA metabolism in CRC progression.METHODS:Dietary BCAA was administered to both azoxymethane-induced and azoxymethane/dextran sodium sulfate-induced CRC mouse models. The expression of genes related to BCAA metabolism was determined using RNA sequencing. Adjacent tissue samples, obtained from 58 patients with CRC, were subjected to quantitative real-time PCR and immunohistochemical analysis. Moreover, the suppressive role of branched-chain aminotransferase 2 (BCAT2) in cell proliferation, apoptosis, and xenograft mouse models was investigated. Alterations in BCAAs and activation of downstream pathways were also assessed using metabolic analysis and western blotting.RESULTS:High levels of dietary BCAA intake promoted CRC tumorigenesis in chemical-induced CRC and xenograft mouse models. Both the mRNA and protein levels of BCAT2 were decreased in tumor tissues of patients with CRC compared to those in normal tissues. Proliferation assays and xenograft models confirmed the suppressive role of BCAT2 in CRC progression. Furthermore, the accumulation of BCAAs caused by BCAT2 deficiency facilitated the chronic activation of mTORC1, thereby mediating the oncogenic effect of BCAAs.CONCLUSION:BCAT2 deficiency promotes CRC progression through inhibition of BCAAs metabolism and chronic activation of mTORC1.

BACKGROUND:Historically, Asia had a lower prostate cancer (PCa) incidence and mortality compared with Western countries, but the gap is narrowing. Paradoxically, Asians have been reported to present with more advanced disease though more favorable outcomes. Despite PCa becoming an emerging health priority in East Asia, our knowledge remains limited. We compared the prevalence of high-grade PCa on biopsy and disease progression after radical prostatectomy (RP) in East Asian men from Asia and non-East Asian men from Western countries. METHODS:This retrospective cohort study included men who underwent prostate biopsy and RP at academic centers in Shanghai, China, and Toronto, Canada (2014-2019). The expanded RP cohort included East Asian men from Singapore (n=282) and non-East Asians from Paris (n=192). Primary endpoints included the proportion of men with Gleason score (GS) ≥8 on biopsy and metastasis-free survival (MFS) after RP for GS ≥8. Multivariable logistic regression and Cox proportional hazard models were performed. Propensity score matching was used to reduce imbalances between cohorts. RESULTS:PCa was found on biopsy in 2,343 of 4,905 (48%) East Asians and 2,317 of 3,482 (67%) non-East Asians (P<.001). Prostate-specific antigen (PSA) levels at presentation and the proportion of men with GS ≥8 were higher in East Asians than non-East Asians (12.4 vs 6.6 ng/mL and 15.0% vs 8.8%, respectively; both P<.001). On multivariable analysis, there was no difference in the proportion of men with GS ≥8 between matched cohorts with PSA <20 ng/mL (n=3,572; odds ratio, 1.05 [95% CI, 0.77-1.43]; P=.76). No difference in MFS was found after RP between matched cohorts (hazard ratio, 0.97 [95% CI, 0.55-1.70]; P=.92). CONCLUSIONS:This contemporary study demonstrates that East Asian men are equally as likely to harbor aggressive PCa on biopsy as non-East Asian men at PSA levels observed in screening programs, with no difference in disease aggressiveness after RP. The assumption that unfavorable PCa at diagnosis is more common but less aggressive in East Asians should be revisited and viewed in the context of the expected increase in the PCa burden worldwide.

Large cohort studies examining trends in cancer-related suicide are lacking. We analyzed data from the Surveillance, Epidemiology, and End Results (SEER) database, encompassing a total of 4,870,410 patients diagnosed with cancer from 1975 to 2017 in the United States. Joinpoint regression was used to estimate the annual percent change (APC) and average annual percentage change (AAPC) of age-adjusted rates of suicide. In the past 40 years, we revealed a gradual increase in cancer-related suicide rates from 1975 to 1989, followed by a gradual decrease from 1989 to 2013, and a marked decrease from 2013 to 2017. These trends suggested the potential impact of advancements in psychosocial care for patients with cancer in contributing to the observed decrease in suicide rates.

Identification of potential bacterial players in colorectal tumorigenesis has been a focus of intense research. Herein, we find that Clostridium symbiosum (C. symbiosum) is selectively enriched in tumor tissues of patients with colorectal cancer (CRC) and associated with higher colorectal adenoma recurrence after endoscopic polypectomy. The tumorigenic effect of C. symbiosum is observed in multiple murine models. Single-cell transcriptome profiling along with functional assays demonstrates that C. symbiosum promotes the proliferation of colonic stem cells and enhances cancer stemness. Mechanistically, C. symbiosum intensifies cellular cholesterol synthesis by producing branched-chain amino acids (BCAAs), which sequentially activates Sonic hedgehog signaling. Low dietary BCAA intake or blockade of cholesterol synthesis by statins could partially abrogate the C. symbiosum-induced cell proliferation in vivo and in vitro. Collectively, we reveal C. symbiosum as a bacterial driver of colorectal tumorigenesis, thus identifying a potential target in CRC prediction, prevention, and treatment.

Abstract Background: KRAS is the best-known oncogene but had long been considered ‘undruggable’ until the approval of the first KRAS G12C inhibitor sotorasib in 2021. However, most current clinical-stage KRAS inhibitors target the KRAS G12C mutation. Inhibitors of other KRAS mutants such as KRAS G12D, the most frequent KRAS mutation in human cancer, are needed for patients. We have developed GFH375 (VS-7375), an oral, selective KRAS G12D inhibitor targeting both “ON” (GTP-bound) and “OFF” (GDP-bound) states of KRAS proteins. The in vitro potency, selectivity, and in vivo efficacy of monotherapy and combination therapy with avutometinib, a unique RAF/MEK clamp, were evaluated in preclinical studies. Methods: Biochemical and cellular assays were used to investigate inhibition to KRAS cycling and signaling. CellTiter-Glo assay was performed to determine the effects on proliferation of tumor cell lines. Several KRAS G12D CDX tumor models were employed to study the in vivo pharmacodynamic and anti-tumor effects. Results: GFH375 inhibited both nucleotide exchange on GDP-bound KRAS G12D and interaction between GMPPNP-bound KRAS G12D with RAF1 with single-digit nanomolar IC50 values. GFH375 suppressed phosho-ERK1/2 (p-ERK) level with sub-nanomolar IC50 values and potently inhibited cell proliferation across a panel of KRAS G12D tumor cell lines. GFH375 showed high selectivity for KRAS G12D relative to non-G12D KRAS variants, KRAS wild type cells, and NRAS, HRAS, or BRAF mutated cells. Following a single oral dose, GFH375 produced deep and durable inhibition of p-ERK in KRAS G12D CDX tumors. GFH375 demonstrated dose-dependent anti-tumor activity with tumor regressions at 10 or 30 mg/kg given orally twice daily in multiple KRAS G12D PDAC and CRC CDX tumor models. GFH375 also showed strong anti-tumor efficacy in an intracranial KRAS G12D tumor model starting as low as 10 mg/kg orally twice daily. Strong synergy between GFH375 and avutometinib was observed in vitro and was validated in vivo as anti-tumor activity of GFH375 was further enhanced by avutometinib. Conclusions: GFH375 is a highly potent and selective orally active inhibitor of KRAS G12D (ON/OFF) and demonstrated promising anti-tumor activity in multiple KRAS G12D tumor models. GFH375 was also effective in an intracranial tumor model. GFH375 showed strong synergy with avutometinib in vitro and in vivo. GFH375 is currently in IND-enabling development in preparation for clinical studies of monotherapy and potentially in combination with other agents for patients with KRAS G12D mutant tumors. Citation Format: Feng Yan, Tao Jiang, Tao Liang, Lijian Cai, Leitao Zhang, Xiaoming Xu, Yanhui Zhao, Xiaoling Lan, Xiaohui Zhang, Meng Liu, Qiang Liu, Jinting Gao, Fubo Xie, Xueyan Gao, Li Wang, Jingyang Zhang, Hongcan Ren, Dong Liu, Siyuan Le, Lili Tang, Silvia Coma, Yaofeng Cheng, Nathan Sanburn, Jonathan A. Pachter, Fusheng Zhou, Jiong Lan, Qiang Lu. GFH375 (VS-7375): An oral, selective KRAS G12D (ON/OFF) inhibitor with potent anti-tumor efficacy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3318.