mTOR inhibitors (mTORis) are Food and Drug Administration (FDA)-approved therapies for advanced gastroenteropancreatic neuroendocrine tumors (GEP-NETs), yet their clinical efficacy is often limited by transient responses and acquired resistance. To uncover sensitizing co-targets, we conduct a kinome-wide CRISPR-Cas9 screen, identifying the lipid kinase PIKfyve as a key vulnerability in GEP-NETs. PIKfyve is overexpressed and functionally linked to the regulation of lipid biosynthesis through the mTOR-SREBP1 axis. Mechanistically, PIKfyve inhibition impairs lysosome-mediated ferritin degradation, amplifying metabolic stress triggered by mTORi-induced ferritinophagy. Co-inhibition of mTOR and PIKfyve synergistically disrupts lipid and iron metabolism, leading to enhanced tumor suppression and improved survival in preclinical GEP-NET models. These findings nominate PIKfyve as a metabolic co-target to overcome mTORi resistance, offering a rationale for combination therapies in mTOR-driven malignancies.

Supplemental Figure 3. Compensatory mechanisms in CCR1 ablated tumor microenvironment elicits immune infiltration

Hypomorphic variants in the SEL1L-HRD1 ER-associated degradation (ERAD) complex have been linked to severe neurological syndromes in children, including neurodevelopmental delay, intellectual disability, motor dysfunction, and early death. Despite this association, its physiological importance and underlying mechanisms in neurons remain poorly understood. Here, we show that neuronal SEL1L-HRD1 ERAD is essential for maintaining one-carbon metabolism, motor function, and overall viability. Neuron-specific deletion of Sel1L in mice (Sel1LSynCre) resulted in growth retardation, severe motor impairments, and early mortality by 9 weeks of age - mirroring core clinical features observed in affected patients - despite preserved neuronal numbers and only modest ER stress. Multiomics analyses, including single-nucleus RNA sequencing and metabolomics, revealed significant dysregulation of one-carbon metabolism in ERAD-deficient brains. This included activation of the serine, folate, and methionine pathways, accompanied by elevated levels of S-adenosylmethionine and related metabolites, likely resulting from induction of the integrated stress response. Together, these findings uncover a previously unappreciated role for neuronal SEL1L-HRD1 ERAD in coordinating ER protein quality control with metabolic adaptation, providing insight into the molecular basis of ERAD-related neurodevelopmental disease.

Sex differences in immune responses impact cancer outcomes and treatment response, including in glioblastoma (GBM). However, host factors underlying distinct immune-cancer interactions are poorly understood. Here we identify γ-aminobutyric acid (GABA) as a female-specific driver of GBM-promoting immune response. We demonstrated that GABA receptor B (GABBR) signaling enhances the T cell suppressive function of granulocytic myeloid-derived suppressor cells (gMDSCs) from female mice by upregulating the cationic amino acid transporter 2-L-arginine-nitric oxide synthase 2 (NOS2) pathway. GABBR agonism promotes GBM growth in female preclinical models through gMDSCs, while GABBR antagonism extends survival and reduces NOS2 in tumor-infiltrating gMDSCs only in female mice. Immune cells from female participants with GBM have enriched GABA transcriptional signatures and a higher GABA concentration compared to male counterparts. Collectively, these results highlight the sex-specific immunomodulatory role of GABA in tumorigenesis, supporting future assessment of GABA pathway inhibitors for cancer immunotherapy.

In Type 1 Diabetes (T1D), disease onset and response to immunotherapy vary widely among individuals, reflecting heterogeneous stage-specific immune dysregulation that remains undefined. To investigate this heterogeneity, we engineered a microporous polycaprolactone scaffold that forms a synthetic immunological niche (IN) upon subcutaneous implantation, enabling in vivo capture of systemic immune dysregulation. In non-obese diabetic (NOD) mice, longitudinal transcriptomic profiling of IN-infiltrating cells identified early-stage genes relatively enriched for myeloid cells, followed by progressive increases in T cell-associated dysregulation at later stages that distinguished T1D progressors from non-progressors. We derived an early-stage IN-based T1D gene signature capturing immune alterations. The signature stratified NOD progressors from non-progressors as early as 6 weeks of age and was conserved across human T1D datasets, distinguishing T1D from non-diabetic individuals in spleen and pancreatic lymph node samples, but not peripheral blood. Signature-based stratification further revealed enrichment of macrophage-associated TNF-α pathways in NOD progressors, validated in human T1D islets. Given heterogeneous response to anti-TNF-α therapy, IN profiling identified resistance-associated mechanisms and enabled derivation of a pathway score that prospectively distinguished treatment-sensitive from resistant mice prior to therapy, establishing the IN as a minimally invasive platform for detecting stage-wise immune dysregulation and stratifying immunotherapy response in T1D.

Chromosome instability is highly prevalent in cancer and drives large-scale chromosomal imbalances, known as aneuploidies1-4. How aneuploidy contributes to tumorigenesis remains difficult to study due to the vast numbers of genes affected. Here we established a CRISPR knockout- and activation-linked assay (CRISPR-KOALA), enabling high-throughput bidirectional genetic screens in immunocompetent mouse models of cancer. We developed a compendium of the ten most frequent human chromosome-arm-level alterations in basal-like breast cancer (BLBC), a disease type that is driven by large copy-number alterations (CNAs)5-8. Using CRISPR-KOALA, we screened the mouse orthologues of 3,752 genes on these arms and identified 90 cancer driver genes, the function of the vast majority of which is unknown. These genes drive distinct signalling pathways including MAPK, HIPPO and WNT, reflecting the high degree of BLBC heterogeneity. Manipulating the identified cancer driver genes overcomes the need for CNAs in Trp53-mutant BLBC mouse models. Mechanistically, we identify that PLGRKT is a potent oncogene that lies on chromosome 9p and show that its tumour-promoting activity is associated with highly stress-resistant mitochondria and an increased ability to detoxify reactive oxygen species. Together, our findings reveal that arm-level CNAs can function to select specific driver genes to promote heterogeneous biological processes.

Colorectal carcinoma (CRC) remains a major cause of cancer-related mortality with rising incidence in individuals under 55, highlighting the need for novel therapeutic strategies. Hypoxia-inducible factor 2 alpha (HIF-2α) has been genetically validated as a critical driver of colorectal tumorigenesis, with intestinal epithelium–specific deletion in mice markedly reducing tumor formation. PT2385, a selective small-molecule HIF-2α inhibitor applied in renal cell carcinoma treatment, has not been evaluated in CRC. Here, we demonstrate that HIF-2α inhibition with PT2385 alone fails to suppress CRC growth in vitro under normoxic or hypoxic conditions and in xenograft models in vivo. To identify vulnerabilities induced by HIF-2α blockade, we performed an unbiased CRISPR metabolic screen. This revealed cholesterol biosynthesis as a critical dependency. Targeting this pathway with clinically approved statins (atorvastatin, pitavastatin, simvastatin) synergized with PT2385 to suppress CRC cell growth, reduce colony formation, and enhance cell death. Genetic knockdown of HIF-2α Mechanistic studies show that combined HIF-2α and HMG-CoA reductase inhibition with statins promotes ferroptosis, characterized by increased lipid peroxidation and depletion of antioxidant metabolites. These effects are fully reversed by the ferroptosis inhibitor liproxstatin-1. In vivo, co-administration of PT2385 and atorvastatin significantly reduced tumor growth and increased ferroptotic cell death in xenografts, confirming the mechanistic link. Collectively, these findings uncover a metabolic vulnerability of CRC to dual HIF-2α and cholesterol biosynthesis inhibition supporting a clinically actionable strategy that leverages safe, FDA-approved statins to potentiate HIF-2α-targeted therapy.

Pancreatic ductal adenocarcinoma (PDAC), the most common subtype of pancreatic cancer, is a deadly disease with a complex tumor microenvironment (TME). How chemotherapy alters the TME, and whether these changes drive chemoresistance, is poorly understood. We examined matched pre- and post-treatment tissue specimens and found near-universal enrichment of axonal guidance genes in cancer-associated fibroblasts (CAFs) after treatment. These CAFs were enriched near sites of perineural invasion, coinciding with regions of increased tumor cell proliferation, and were enriched in tumor areas distant from nerves after chemotherapy. Metastatic recurrence lesions had the highest prevalence of these CAFs versus primary tumors and untreated metastasis. These CAFs showed elevated non-canonical Wnt mediators and axonal guidance genes, which complemented matching cognate binding partners in tumor epithelial cells, suggesting a role in tumor-stroma crosstalk. Our findings implicate fibroblast-derived axonal-guidance genes in promoting PDAC invasion and point to a promising target for this disease. Statement of Significance:Therapeutic resistance remains a major challenge in pancreatic cancer. Our findings shed light on the changes in the complex tumor microenvironment in response to chemotherapy and identify fibroblasts high in axonal-guidance genes that may drive tumor progression and chemoresistance, thus uncovering a potential avenue for targeting pancreatic cancer.

Abstract Pancreatic ductal adenocarcinoma (PDAC) is among the most aggressive human malignancies, with a five-year survival rate below 15%. In contrast to several other solid tumors where chemotherapy, radiotherapy, and immune checkpoint blockade have significantly improved outcomes, PDAC remains largely refractory to current therapeutic strategies. One hallmark of PDAC is its profoundly hypoxic tumor microenvironment. Extensive desmoplasia and limited vascularization restrict oxygen delivery to tumor tissues, creating regions of severe oxygen deprivation. KRAS, the primary oncogenic driver of PDAC, can activate transcriptional programs that rewire cellular metabolism and enable tumor cell survival under oxygen limitation. However, the precise molecular mechanisms connecting oncogenic KRAS signaling to hypoxia-adaptive transcriptional programs remain poorly defined. To identify the top transcriptional regulators associated with KRAS, we integrated clinical and pathological information with gene expression data derived from laser captured, microdissected human PDAC epithelium. Across 242 transcriptional profiles, including 197 PDAC tumors and 45 benign precursor lesions, regulatory network analysis identified BMAL2, a member of the PAS superfamily typically associated with the regulation of circadian rhythms, as the top KRAS-associated transcriptional regulator and one of the strongest drivers of tumor initiation, progression, and post-resection survival. We found that BMAL2 activity was consistently elevated in PDAC relative to precursor lesions and independently associated with worse clinical outcomes. Meta-analysis across multiple independent PDAC cohorts confirmed that BMAL2 activity is reproducibly increased in aggressive tumor subtypes and correlated with poor prognosis. Phylogenetic analysis showed that BMAL1 and BMAL2 are most closely related to ARNT (HIF1β) and ARNT2 (HIF2β), the obligate binding partners of HIF transcription factors. Consistent with this evolutionary relationship, BMAL2 activity correlated strongly with hypoxia transcriptional signatures compared with other PAS family members. Functional studies demonstrated that BMAL2 is a critical regulator of PDAC cell fitness. CRISPR/Cas9-mediated BMAL2 knockout across multiple PDAC cell lines significantly reduced cell viability, clonogenic capacity, and migratory behavior. In wound-healing assays, BMAL2-deficient cells displayed markedly impaired migration compared with control cells. Conversely, BMAL2 overexpression enhanced tumor cell growth and accelerated wound closure, supporting a model in which BMAL2 promotes proliferative and migratory phenotypes in PDAC cells. Seahorse metabolic profiling revealed that BMAL2 expression increases cellular respiratory capacity and overall metabolic activity, whereas BMAL2 loss reduced oxygen consumption rates and impaired metabolic responses associated with hypoxic adaptation. These findings are consistent with transcriptional and metabolomic data indicating that BMAL2 promotes glycolytic metabolism, including increased lactate production and expression of key glycolytic enzymes. Together, these observations position BMAL2 as an important mediator of metabolic plasticity in pancreatic cancer cells. Beyond metabolic regulation, BMAL2 also influenced epithelial plasticity. In PATU8902 cells, BMAL2 overexpression induced morphological changes characterized by spindle-like architecture and reduced epithelial cohesion. These changes were accompanied by increased expression of Vimentin, a well-known marker of epithelial-to-mesenchymal transition (EMT). Collectively, these findings suggest that BMAL2 promotes EMT-like phenotypic reprogramming that may enhance tumor cell adaptability and invasiveness within the hypoxic tumor microenvironment. Mechanistically, we found that BMAL2 regulated the stability and balance of hypoxia-inducible factor paralogs. Under hypoxic conditions, BMAL2 is required for stabilization of HIF1A, a central regulator of glycolytic metabolism. Loss of BMAL2 prevented HIF1A stabilization while simultaneously increasing HIF2A accumulation, suggesting that BMAL2 functions as a molecular switch between HIF1A- and HIF2A-dependent hypoxia programs. This regulatory mechanism provides a direct link between oncogenic KRAS signaling and the transcriptional machinery that enables tumor cells to survive and proliferate in oxygen-limited environments. Importantly, we found that BMAL2 activity was controlled by KRAS signaling through the RAF-MEK-ERK pathway. Pharmacologic inhibition of RAS using the pan-RAS(ON) inhibitor RMC-7977 significantly reduced BMAL2 activity both in vitro and in vivo without altering BMAL2 expression levels, indicating a post-translational regulatory mechanism. Complementarily, ERK inhibition also displayed reduction in BMAL2 activity in a panel of PDAC cell lines. This finding places BMAL2 downstream of oncogenic KRAS and identifies it as a transcriptional effector of RAS-MAPK signaling. Finally, the functional consequences of BMAL2 activity extended into living animals. In orthotopic xenograft models of PDAC, BMAL2 knockout markedly impaired tumor growth and reduced tumor engraftment. Tumors derived from BMAL2-deficient cells grew significantly more slowly than controls, and in some models tumor formation was severely compromised. Consistent with reduced tumor growth, animals bearing BMAL2-deficient tumors exhibited prolonged survival compared with those implanted with control cells. These findings demonstrate that BMAL2 is not only necessary for PDAC cell proliferation in vitro but is also essential for efficient tumor initiation and progression in vivo. Collectively, these findings identify BMAL2 as a previously unrecognized transcriptional effector downstream of oncogenic KRAS that functions as a central transcriptional node linking KRAS signaling to hypoxia adaptation, metabolic reprogramming, and phenotypic plasticity in pancreatic cancer. By coordinating transcriptional and metabolic programs required for survival under extreme hypoxic conditions, BMAL2 enables PDAC cells to adapt to and thrive within their uniquely hostile tumor microenvironment. These results provide mechanistic insight into how KRAS signaling drives pancreatic cancer biology and highlight BMAL2 as a key regulator of tumor fitness within the hypoxic tumor ecosystem. Citation Format: Alvaro Curiel Garcia, Sam R. Holmstrom, Melina Chen, Diana V. Morales, Kathryn Buscher, Carmine F. Palermo, Steven A. Sastra, Anthony Andren, Lorenzo Tomassoni, Li Zhang, Maya Stella Dixon, Tessa Y.S. Le Large, Irina Sagalovskiy, Winston Wong, Kaitlin Shaw, Jeanine Genkinger, Hanina Hibshoosh, Gulam A. Manji, Alina C. Iuga, Roland M. Schmid, Michael A. Badgley, Pasquale Laise, Iok In Christine Chio, Costas A. Lyssiotis, Yatrik M. Shah, Andrea Califano, H. Carlo Maurer, Kenneth P. Olive. BMAL2 is a KRAS-driven master regulator of hypoxic adaptation and EMT in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(8_Suppl):Abstract nr NG04.

Inter-organellar communication is critical for cellular metabolism. One of the most abundant inter-organellar interactions occurs at the endoplasmic reticulum and mitochondria contact sites (ERMCSs). However, an understanding of the mechanisms governing ERMCS regulation and their roles in cellular metabolism is limited by a lack of tools that permit temporal induction and reversal. Through screening approaches, we identified fedratinib, an FDA-approved drug that dramatically increases ERMCS abundance by inhibiting the epigenetic modifier BRD4. Fedratinib rapidly and reversibly modulates mitochondrial and ER morphology, induces a distinct ER-mitochondria envelopment structure, and alters metabolic homeostasis. Moreover, ERMCS modulation depends on mitochondrial electron transport chain complex III function. Comparison of fedratinib activity to other reported inducers of ERMCSs revealed common mechanisms of induction and function, providing clarity to a growing body of experimental observations. In total, our results uncovered a novel epigenetic signaling pathway and an endogenous metabolic regulator that connects ERMCSs and cellular metabolism.

Gliomas account for ∼80% of primary malignant brain tumors. Many CNS WHO grade 2-3 and some grade 4 gliomas harbor mutant isocitrate dehydrogenase 1 (mIDH1), which causes a gain-of-function mutation (IDH1 R132H) leading to the production of 2-hydroxyglutarate (2HG). Mutant IDH1-induced 2HG, through epigenetic reprogramming elicits an immune-permissive tumor microenvironment (TME). An immunosuppressive mechanism in the glioma TME involves adenosine production via the ectoenzyme CD73. This study investigates mIDH1's influence on CD73 expression and adenosine levels. We demonstrate that mIDH1 glioma cells exhibit reduced CD73 expression, driven by DNA hypermethylation, leading to reduced adenosine levels. Since wtIDH1 gliomas have high CD73 expression, we evaluated CD73 blockade as an immunotherapy target. We show that CD73 inhibition used as monotherapy did not improve survival in wtIDH1 glioma-bearing mice. However, when combined with immune-stimulatory Ad-TK (adenoviral vectors encoding herpes simplex virus thymidine kinase) and Ad-Flt3L (adenoviral vectors encoding FMS-like tyrosine kinase 3 ligand) gene therapy, CD73 blockade significantly enhanced therapeutic efficacy and increased anti-glioma effector T cell activity. These findings reveal that CD73 inhibition used in combination with immune-stimulatory Ad-TK/Ad-Flt3L gene therapy may be an effective treatment for wtIDH1 gliomas, which could be readily translated to the clinical arena.

Colorectal cancer (CRC) cells are addicted to iron, which fuels nucleotide synthesis, mitochondrial respiration, and proliferation. Yet paradoxically, high intracellular iron is cytotoxic to most cells, raising the question of how CRC cells tolerate and exploit iron-rich environments. Ferroptosis, an iron-dependent form of cell death, is thought to mediate iron toxicity. However, whether most ferroptosis regulators, identified through synthetic chemical screens or small molecule activators, play a role in modulating iron toxicity, particularly in vivo, remains unclear. Here, using multi-omics profiling, CRISPR screening, and in vivo models, we uncover a heme-succinate dehydrogenase (SDH)-coenzyme Q (CoQ) axis that enables CRC cells to buffer iron-induced oxidative stress. Heme-dependent SDH reduces CoQ, which redistributes to mitochondrial and plasma membranes to detoxify lipid reactive oxygen species (ROS) as a radical-trapping antioxidant. These findings reveal that CRCs co-opt metabolic cofactors both for growth and for survival under physiologically toxic iron levels, uncovering new vulnerabilities for therapy.

Table S3. Single cell RNA sequencing datasets

Amino acid (AA) uptake is essential for T cell metabolism and function, but how tissue sites and inflammation affect CD4+ T cell subset requirements for specific AAs remains uncertain. Here, we tested CD4+ T cell AA demands with in vitro and in vivo CRISPR screens and identified subset- and tissue-specific dependencies on the AA transporter SLC38A1 (SNAT1). While dispensable for T cell persistence and expansion in vivo in lung inflammation, SLC38A1 was critical for Th1, but not Th17, cell-driven experimental autoimmune encephalomyelitis (EAE) and contributed to Th1 cell-driven inflammatory bowel disease. SLC38A1 deficiency reduced mTORC1 signaling and glycolytic activity in Th1 cells, in part by reducing glutamine uptake and disrupting hexosamine biosynthesis and redox regulation. Pharmacological inhibition of SLC38 transporters also delayed Th1-mediated EAE but did not affect lung inflammation. CD4+ T cells thus have subset- and tissue-specific nutrient transporter dependencies that may guide new metabolic approaches for selective immunotherapies.

Abstract Pancreatic ductal adenocarcinoma (PDAC) is characterized by profound metabolic rewiring and a strongly immunosuppressive tumor microenvironment, both of which contribute to poor therapeutic responses. Immunogenic cell death (ICD) represents a potential strategy to overcome immune suppression by coupling tumor cell death to anti-tumor immune activation. Here, we investigated whether targeting amino acid metabolism in PDAC can induce ICD and promote tumor immunity. Through a focused metabolic screen in a panel of syngeneic mouse cancer cell lines, we identified cysteine restriction as a robust inducer of multiple damage-associated molecular patterns (DAMPs) in vitro, hallmark features of ICD. In addition to driving DAMPs, cystine-deprived tumor cells also promoted dendritic cell phagocytosis, maturation, and proinflammatory cytokine production in vitro. Because cysteine deprivation is a known trigger of ferroptosis, we further demonstrated that pharmacologic inhibition of glutathione peroxidase 4 (GPX4) similarly elicited ICD-associated features, which were reversible by the ferroptosis inhibitor Ferrostatin-1 (Fer-1). To define additional immune-modulatory signals associated with ferroptosis, we performed metabolomic and lipidomic profiling of cells undergoing, but not yet committed to, ferroptotic death. These analyses revealed selective release of immunosuppressive metabolites and oxidized phospholipids. Consistent with this, conditioned media from ferroptotic cells impaired CD8⁺ T cell proliferation and cytotoxicity in vitro. Thus, together our results indicated that the induction of ferroptotic immunogenic cell death led to the release of both pro- and anti-inflammatory signals. Subsequent analysis in vivo revealed that ferroptotic tumor cells predominantly contributed to a tumor-protective environment. In particular, tumors inoculated with ferroptotic cells were enriched with immunosuppressive myeloid cells and exhibited reduced populations of tumor-infiltrating CD8+ T cells. Further investigation using immune compromised mice suggested that ferroptotic cells may suppress both adaptive and innate immune responses. Collectively, these results underscore the complex and highly context-dependent effects of ferroptosis on tumor immunity, highlighting the critical importance of in vivo models to determine true immunogenic potential within the tumor microenvironment.

Abstract Background Pancreatic Ductal Adenocarcinoma remains one of the deadliest cancers, with an overall 5 year survival rate of 13 percent. The majority of PDAC cases are driven by mutations in KRAS, making it a critical therapeutic target. Although many KRAS inhibitors have been developed and show promise in preclinical and clinical settings as single agents, the emergence of resistance and adaptive mechanisms has highlighted the need for combinatorial therapies. We recently demonstrated that the concurrent targeting of KRAS-MAPK and the lipid kinase PIKfyve disrupts lipid homeostasis in PDAC and has potent antitumor effects, curing the majority of mice in a syngeneic orthotopic mouse model. Interestingly, PIKfyve and KRAS-MAPK inhibition independently increase MHC Class 1 surface expression and enhance effectiveness of anti-PD-1 therapy. Thus, building on these findings, we hypothesized that dual inhibition of PIKfyve and KRAS-MAPK signaling modulates the immune landscape of PDAC. Methods & Results Using a syngeneic orthotopic PDAC model, mice underwent 19 days of treatment with PIKfyve inhibitor ESK981 in combination with RAS inhibitor RMC-6236. Those that achieved a complete cure at the end-point were left untreated for 2 months, and it was observed that tumors grew back for all mice, highlighting the need for additional therapy. Notably, PIKfve or KRAS-MAPK inhibition alone increased expression of immune signaling markers MHC Class I and PD-L1, suggesting that the addition of anti-PD-1 therapy could further enhance efficacy. To investigate the effect of PIKfyve and RAS inhibition on the PDAC tumor microenvironment, we performed single-cell RNA sequencing on tumors harvested after 5 days of treatment with ESK981 and RMC-6236. The combination therapy resulted in an increased proportion of immune cells, particularly T and B cells, along with a decreased proportion of epithelial cells and fibroblasts, which are associated with immune evasion. These suggest that PIKfyve and RAS inhibition alter immune function and prime the microenvironment for immune therapy. Together, our preliminary findings suggest that the combined inhibition of PIKfyve and KRAS-MAPK exerts both direct antitumor effects and enhances the immune response in PDAC. Our goal is to build upon this strategy by leveraging its immune-enhancing effects, particularly with the addition of immune checkpoint blockade. Citation Format: Jasmine P. Wisniewski, Caleb Cheng, Rüya Pakkan, Sydney Peters, Gabriel Cruz, Yuanyuan Qiao, Costas A. Lyssiotis, Arul M. Chinnaiyan, . Immune implications of dual PIKfyve and KRAS inhibition in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 1575.

Table S1. Antibodies used for immunostaining, flow cytometry and CyTOF

Neuroendocrine prostate cancer (NEPC) persists in a profoundly hypoxic microenvironment, yet the mechanisms enabling tumor adaptation to this metabolically challenging niche remain undefined. Here, we identify the lipid kinase PIKfyve as overexpressed in NEPC, functioning as a central node in a stress-adaptive lipid kinase axis that supports adaptation to persistent endoplasmic reticulum (ER) stress. Mechanistically, NEPC requires PIKfyve-mediated lysosomal degradation and lipid recycling to maintain metabolic homeostasis under hypoxia. PIKfyve inhibition disrupts lysosomal function, exacerbates ER stress, and activates a compensatory sterol regulatory element-binding protein (SREBP)-dependent de novo lipogenesis program essential for NEPC survival. This stress-lipid axis creates a synthetic vulnerability between PIKfyve and fatty acid synthase (FASN), where dual inhibition synergistically amplifies ER stress, triggers the terminal unfolded protein response, and induces tumor cell death. These findings reveal a metabolic adaptation in NEPC and provide preclinical evidence that co-targeting PIKfyve and FASN can overcome hypoxia-associated stress adaptation.

Nitrogen metabolism is more complex and less mapped than carbon pathways owing to the diversity of nitrogen sources and disjointed routing. Savani et al. develop a multiplexed nitrogen labelling platform to trace system-wide nitrogen flows, revealing how undifferentiated and mature cells select nitrogen sources for pyrimidine nucleotide synthesis.