ABSTRACT Inactivation of pVHL tumor suppressor in clear cell Renal Cell Carcinoma (ccRCC) increases the abundance of Histone H3 lysine 27 acetylation (H3K27ac). We hypothesized that H3K27ac, a marker of transcriptional activation, drives the expression of critical oncogenes in ccRCC. Using H3K27ac ChIP-Seq; RNA-Seq; an in vivo positive selection screen; cell-based functional studies; and clinical validations; here, we report the identification of the SLC1A1/EAAT3 aspartate (Asp) and glutamate (Glu) transporter as a ccRCC oncogene. pVHL loss promotes SLC1A1 expression in a HIF-independent manner. Importantly, SLC1A1 inactivation depletes Asp/Glu-derived metabolites, impedes ccRCC growth both in vitro and in vivo , and sensitizes ccRCCs to metabolic therapeutics (e.g., glutaminase blockers). Finally, in human ccRCC biospecimens, higher SLC1A1 expression is associated with metastatic disease and clusters with elevated expression of other solute carriers, but not HIF/Hypoxia pathways. Altogether, our studies identify a HIF-independent metabolic hub in ccRCC and credential SLC1A1 as an actionable ccRCC oncogene. STATEMENT OF SIGNIFICANCE Targeting chronic HIF activation underlies many therapeutic strategies in ccRCC; but, unfortunately, is not curative. SLC1A1, instead, represents a HIF-independent ccRCC dependency, which is targetable alone and together with other antimetabolites, such as glutaminase inhibitors. These observations identify an actionable metabolic program that functions independent of HIF in ccRCC.
<div>Abstract<p>Glioblastoma (GBM) is the most common and lethal primary brain tumor and remains incurable. This is in part due to the cellular heterogeneity within these tumors, which includes a subpopulation of treatment-resistant cells called cancer stem–like cells (CSC). We previously identified that the anaphase-promoting complex/cylosome (APC/C), a key cell-cycle regulator and tumor suppressor, had attenuated ligase activity in CSCs. Here, we assessed the mechanism of reduced activity, as well as the efficacy of pharmacologically targeting the APC/C in CSCs. We identified hyperphosphorylation of CDH1, but not pseudosubstrate inhibition by early mitotic inhibitor 1 (EMI1), as a major mechanism driving attenuated APC/C<sup>CDH1</sup> activity in the G<sub>1</sub>-phase of the cell cycle in CSCs. Small-molecule inhibition of the APC/C reduced viability of both CSCs and nonstem tumor cells (NSTCs), with the combination of proTAME and apcin having the biggest impact. Combinatorial drug treatment also led to the greatest mitotic arrest and chromosomal abnormalities.</p>Implications:<p>Our findings demonstrate how the activity of the APC/C<sup>CDH1</sup> tumor suppressor is reduced in CSCs and also validates small-molecule inhibition of the APC/C as a promising therapeutic target for the treatment of GBM.</p></div>
<div>AbstractPurpose:<p>Advanced/metastatic forms of clear-cell renal cell carcinomas (ccRCC) have limited therapeutic options. Genome-wide genetic screens have identified cellular dependencies in many cancers. Using the Broad Institute/Novartis combined short hairpin RNA (shRNA) dataset, and cross-validation with the CRISPR/Cas9 DepMap (21Q3) dataset, we sought therapeutically actionable dependencies in kidney lineage cancers.</p>Experimental Design:<p>We identified preferential genetic dependencies in kidney cancer cells versus other lineages. <i>BCL2L1</i>, which encodes the BCL-X<sub>L</sub> antiapoptotic protein, scored as the top actionable dependency. We validated this finding using genetic and pharmacologic tools in a panel of ccRCC cell lines. Select BCL-X<sub>L</sub>–dependent (versus independent) cell lines were then transcriptionally profiled to identify biomarkers and mechanistic drivers of BCL-X<sub>L</sub> dependence. Cell-based studies (<i>in vitro</i> and <i>in vivo</i>) and clinical validations were used to address physiologic relevance.</p>Results:<p>Inactivation of BCL-X<sub>L</sub>, but not BCL-2, led to fitness defects in renal cancer cells, and sensitized them to chemotherapeutics. Transcriptomic profiling identified a “BCL-X<sub>L</sub> dependency” signature, including an elevated mesenchymal gene signature. A mesenchymal state was both necessary and sufficient to confer increased BCL-X<sub>L</sub> dependence. The “BCL-X<sub>L</sub> dependency” signature was observed in approximately 30% of human ccRCCs, which were also associated with worse clinical outcomes. Finally, an orally bioavailable BCL-X<sub>L</sub> inhibitor, A-1331852, showed antitumor efficacy <i>in vivo</i>.</p>Conclusions:<p>Our studies uncovered an unexpected link between cell state and BCL-X<sub>L</sub> dependence in ccRCC. Therapeutic agents that specifically target BCL-X<sub>L</sub> are available. Our work justifies testing the utility of BCL-X<sub>L</sub> blockade to target, likely, a clinically aggressive subset of human kidney cancers.</p><p><i><a href="https://aacrjournals.org/clincancerres/article/doi/10.1158/1078-0432.CCR-22-2104" target="_blank">See related commentary by Wang et al., p. 4600</a></i></p></div>
Abstract Glioblastoma (GBM) is a malignant brain tumor with nearly universal recurrence. GBM cancer stem cells (CSCs), a subpopulation of radio- and chemo-resistant cancer cells capable of self-renewal, contribute to the high rate of recurrence. The anti-cancer agent, CBL0137, inhibits the FACT (facilitates chromatin transcription) complex leading to cancer cell specific cytotoxicity. Here, we show that CBL0137 sensitized GBM CSCs to radiotherapy using both in vitro and in vivo models. Treatment of CBL0137 combined with radiotherapy led to increased DNA damage in GBM patient specimens and failure to resolve the damage led to decreased cell viability. Using clonogenic assays, we confirmed that CBL0137 radiosensitized the CSCs. To validate that combination therapy impacted CSCs, we used an in vivo subcutaneous model and showed a decrease in the frequency of cancer stem cells present in tumors as well as decreased tumor volume. Using an orthotopic model of GBM, we confirmed that treatment with CBL0137 followed by radiotherapy led to significantly increased survival compared to either treatment alone. Radiotherapy remains a critical component of patient care for GBM, even though there exists a resistant subpopulation. Radio-sensitizing agents, including CBL0137, pose an exciting treatment paradigm to increase the efficacy of irradiation, especially by inclusively targeting CSCs.
