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Branched-chain amino acid transaminase 1 (BCAT1) is upregulated selectively in human isocitrate dehydrogenase (IDH) wildtype (WT) but not mutant glioblastoma multiforme (GBM) and promotes IDHWT GBM growth. Through a metabolic synthetic lethal screen, we report here that α-ketoglutarate (AKG) kills IDHWT GBM cells when BCAT1 protein is lost, which is reversed by reexpression of BCAT1 or supplementation with branched-chain α-ketoacids (BCKA), downstream metabolic products of BCAT1. In patient-derived IDHWT GBM tumors in vitro and in vivo, cotreatment of BCAT1 inhibitor gabapentin and AKG resulted in synthetic lethality. However, AKG failed to evoke a synthetic lethal effect with loss of BCAT2, BCKDHA, or GPT2 in IDHWT GBM cells. Mechanistically, loss of BCAT1 increased the NAD+/NADH ratio but impaired oxidative phosphorylation, mTORC1 activity, and nucleotide biosynthesis. These metabolic alterations were synergistically augmented by AKG treatment, thereby causing mitochondrial dysfunction and depletion of cellular building blocks, including ATP, nucleotides, and proteins. Partial restoration of ATP, nucleotides, proteins, and mTORC1 activity by BCKA supplementation prevented IDHWT GBM cell death conferred by the combination of BCAT1 loss and AKG. These findings define a targetable metabolic vulnerability in the most common subset of GBM that is currently incurable., Significance: Metabolic synthetic lethal screening in IDHWT glioblastoma defines a vulnerability to ΑΚG following BCAT1 loss, uncovering a therapeutic strategy to improve glioblastoma treatment. See related commentary by Meurs and Nagrath, p. 2354., (©2022 American Association for Cancer Research.)

Hypoxia induces thousands of mRNAs and miRNAs to mediate tumor malignancy. However, hypoxia-induced long noncoding RNA (lncRNA) transcriptome and their role in triple-negative breast cancer (TNBC) have not been defined. Here we identified hypoxia-induced lncRNA transcriptome in two human TNBC cell lines by whole transcriptome sequencing. AC093818.1 was one of 26 validated lncRNAs and abundantly expressed in TNBC in vitro and in vivo . 5'- and 3'-rapid amplification of cDNA ends assays revealed that the isoform 2 was a dominant AC093818.1 transcript in TNBC cells and thus referred to as lncIHAT ( lnc RNA i nduced by h ypoxia and a bundant in T NBC). Hypoxia-inducible factor 1 (HIF1) but not HIF2 bound to the hypoxia response element at the promoter of lncIHAT to activate its transcription in hypoxic TNBC cells. LncIHAT promoted TNBC cell survival in vitro and tumor growth and lung metastasis in mice. Mechanistically, lncIHAT was required for the expression of its proximal neighboring oncogenic genes PDK1 and ITGA6 in TNBC cells and tumors. Reexpression of PDK1 and ITGA6 rescued survival and growth of lncIHAT knockdown TNBC cells in vitro . Collectively, these findings uncovered lncIHAT as a new hypoxia-induced oncogenic cis -acting lncRNA in TNBC. IMPLICATIONS: This study systematically identified hypoxia-induced lncRNA transcriptome in TNBC and sheds light on multiple layers of regulatory mechanisms of gene expression under hypoxia., (©2020 American Association for Cancer Research.)

Emerging studies indicate that DNA damage in cancer cells triggers antitumor immunity, but its intrinsic regulatory mechanism in breast cancer cells remains poorly understood. Here, we show that ZMYND8 is upregulated and inhibits micronucleus formation and DNA damage in breast cancer cells. Loss of ZMYND8 triggered activation of the DNA sensor cyclic guanosine monophosphate-adenosine monophosphate synthase in micronuclei, leading to further activation of the downstream signaling effectors stimulator of IFN genes and NF-κB, but not TANK-binding kinase 1 and IFN regulatory factor 3, thereby inducing the expression of IFNβ and IFN-stimulated genes (ISG) in breast cancer cells in vitro and tumors in vivo . ZMYND8 knockout (KO) in breast cancer cells promoted infiltration of CD4 + and CD8 + T cells, leading to tumor inhibition in syngeneic mouse models, which was significantly attenuated by treatment of anti-CD4/CD8-depleting antibodies or anti-IFNAR1 antibody and in immunodeficient Rag1 KO mice. In human breast tumors, ZMYND8 was negatively correlated with ISGs, CD4, CD8A, CD8B, and the tumor-lymphocyte infiltration phenotype. Collectively, these findings demonstrate that maintenance of genome stability by ZMYND8 causes breast cancer cells to evade cytotoxic T-lymphocyte surveillance, which leads to tumor growth. SIGNIFICANCE: These findings show that ZMYND8 is a new negative and intrinsic regulator of the innate immune response in breast tumor cells, and ZMYND8 may be a possible target for antitumor immunotherapy., (©2020 American Association for Cancer Research.)

Recruitment of RNA polymerase II to hypoxia-inducible factor (HIF) target genes under normoxia is a prerequisite for HIF-mediated transactivation. However, the underlying mechanism of this recruitment remains unknown. Here we report that chromodomain helicase DNA-binding protein 4 (CHD4) physically interacts with α and β subunits of HIF1 and HIF2 and enhances HIF-driven transcriptional programs to promote breast cancer progression. Loss of HIF1/2α abolished CHD4-mediated breast tumor growth in mice. In breast cancer cells under normoxia, CHD4 enrichment at HIF target gene promoters increased RNA polymerase II loading through p300. Hypoxia further promoted CHD4 binding to the chromatin via HIF1/2α, where CHD4 in turn enhanced recruitment of HIF1α, leading to HIF target gene transcription. CHD4 was upregulated and correlated with HIF target gene expression in human breast tumors; upregulation of CHD4 and other known HIF coactivators in human breast tumors was mutually exclusive. Furthermore, CHD4 was associated with poor overall survival of patients with breast cancer. Collectively, these findings reveal a new fundamental mechanism of HIF regulation in breast cancer, which has clinical relevance. SIGNIFICANCE: This study identifies CHD4 as a HIF coactivator and elucidates the fundamental mechanism underlying CHD4-mediated HIF transactivation in breast tumors., (©2020 American Association for Cancer Research.)

Hypoxia induces a vast array of long noncoding RNAs (lncRNA) in breast cancer cells, but their biological functions remain largely unknown. Here, we identified a hitherto uncharacterized hypoxia-induced lncRNA RAB11B-AS1 in breast cancer cells. RAB11B-AS1 is a natural lncRNA upregulated in human breast cancer and its expression is induced by hypoxia-inducible factor 2 (HIF2), but not HIF1, in response to hypoxia. RAB11B-AS1 enhanced the expression of angiogenic factors including VEGFA and ANGPTL4 in hypoxic breast cancer cells by increasing recruitment of RNA polymerase II. In line with increased angiogenic factors, conditioned media from RAB11B-AS1-overexpressing breast cancer cells promoted tube formation of human umbilical vein endothelial cells in vitro . Gain- and loss-of-function studies revealed that RAB11B-AS1 increased breast cancer cell migration and invasion in vitro and promoted tumor angiogenesis and breast cancer distant metastasis without affecting primary tumor growth in mice. Taken together, these findings uncover a fundamental mechanism of hypoxia-induced tumor angiogenesis and breast cancer metastasis. SIGNIFICANCE: This study reveals the molecular mechanism by which the lncRNA RAB11B-AS1 regulates hypoxia-induced angiogenesis and breast cancer metastasis, and provides new insights into the functional interaction between a lncRNA and tumor microenvironment. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/5/964/F1.large.jpg., (©2020 American Association for Cancer Research.)