Epigenetic regulation of CD38/CD48 by KDM6A mediates NK cell response in multiple myeloma.

Anti-CD38 monoclonal antibodies like Daratumumab (Dara) are effective in multiple myeloma (MM); however, drug resistance ultimately occurs and the mechanisms behind this are poorly understood. Here, we identify, via two in vitro genome-wide CRISPR screens probing Daratumumab resistance, KDM6A as an important regulator of sensitivity to Daratumumab-mediated antibody-dependent cellular cytotoxicity (ADCC). Loss of KDM6A leads to increased levels of H3K27me3 on the promoter of CD38, resulting in a marked downregulation in CD38 expression, which may cause resistance to Daratumumab-mediated ADCC. Re-introducing CD38 does not reverse Daratumumab-mediated ADCC fully, which suggests that additional KDM6A targets, including CD48 which is also downregulated upon KDM6A loss, contribute to Daratumumab-mediated ADCC. Inhibition of H3K27me3 with an EZH2 inhibitor resulted in CD38 and CD48 upregulation and restored sensitivity to Daratumumab. These findings suggest KDM6A loss as a mechanism of Daratumumab resistance and lay down the proof of principle for the therapeutic application of EZH2 inhibitors, one of which is already FDA-approved, in improving MM responsiveness to Daratumumab.

Systematic profiling of conditional pathway activation identifies context-dependent synthetic lethalities.

The paradigm of cancer-targeted therapies has focused largely on inhibition of critical pathways in cancer. Conversely, conditional activation of signaling pathways as a new source of selective cancer vulnerabilities has not been deeply characterized. In this study, we sought to systematically identify context-specific gene-activation-induced lethalities in cancer. To this end, we developed a method for gain-of-function genetic perturbations simultaneously across ~500 barcoded cancer cell lines. Using this approach, we queried the pan-cancer vulnerability landscape upon activating ten key pathway nodes, revealing selective activation dependencies of MAPK and PI3K pathways associated with specific biomarkers. Notably, we discovered new pathway hyperactivation dependencies in subsets of APC-mutant colorectal cancers where further activation of the WNT pathway by APC knockdown or direct ß-catenin overexpression led to robust antitumor effects in xenograft and patient-derived organoid models. Together, this study reveals a new class of conditional gene-activation dependencies in cancer.

Lenalidomide bypasses CD28 co-stimulation to reinstate PD-1 immunotherapy by activating Notch signaling.

Programmed cell death protein 1 (PD-1) checkpoint blockade therapy requires the CD28 co-stimulatory receptor for CD8+ T cell expansion and cytotoxicity. However, CD28 expression is frequently lost in exhausted T cells and during immune senescence, limiting the clinical benefits of PD-1 immunotherapy in individuals with cancer. Here, using a cereblon knockin mouse model that regains in vivo T cell response to lenalidomide, an immunomodulatory imide drug, we show that lenalidomide reinstates the anti-tumor activity of CD28-deficient CD8+ T cells after PD-1 blockade. Lenalidomide redirects the CRL4Crbn ubiquitin ligase to degrade Ikzf1 and Ikzf3 in T cells and unleashes paracrine interleukin-2 (IL-2) and intracellular Notch signaling, which collectively bypass the CD28 requirement for activation of intratumoral CD8+ T cells and inhibition of tumor growth by PD-1 blockade. Our results suggest that PD-1 immunotherapy can benefit from a lenalidomide combination when treating solid tumors infiltrated with abundant CD28- T cells.

S-nitrosylation of Hsp90 promotes cardiac hypertrophy in mice through GSK3ß signaling.

Cardiac hypertrophy, as one of the major predisposing factors for chronic heart failure, lacks effective interventions. Exploring the pathogenesis of cardiac hypertrophy will reveal potential therapeutic targets. S-nitrosylation is a kind of posttranslational modification that occurs at active cysteines of proteins to mediate various cellular processes. We here identified heat shock protein 90 (Hsp90) as a highly S-nitrosylated target in the hearts of rodents with hypertrophy, and the role of Hsp90 in cardiac hypertrophy remains undefined. The S-nitrosylation of Hsp90 (SNO-Hsp90) levels were elevated in angiotensin II (Ang II)- or phenylephrine (PE)-treated neonatal rat cardiomyocytes (NRCMs) in vitro as well as in cardiomyocytes isolated from mice subjected to transverse aortic constriction (TAC) in vivo. We demonstrated that the elevated SNO-Hsp90 levels were mediated by decreased S-nitrosoglutathione reductase (GSNOR) expression during cardiac hypertrophy, and delivery of GSNOR adeno-associated virus expression vectors (AAV9-GSNOR) decreased the SNO-Hsp90 levels to attenuate cardiac hypertrophy. Mass spectrometry analysis revealed that cysteine 589 (Cys589) might be the S-nitrosylation site of Hsp90. Delivery of the mutated AAV9-Hsp90-C589A inhibited SNO-Hsp90 levels and attenuated cardiac hypertrophy. We further revealed that SNO-Hsp90 led to increased interaction of glycogen synthase kinase 3ß (GSK3ß) and Hsp90, leading to elevated GSK3ß phosphorylation and decreased eIF2Bε phosphorylation, thereby aggravating cardiac hypertrophy. Application of GSK3ß inhibitor TWS119 abolished the protective effect of Hsp90-C589A mutation in Ang II-treated NRCMs. In conclusion, this study demonstrates a critical role of SNO-Hsp90 in cardiac hypertrophy, which may be of a therapeutic target for cardiac hypertrophy treatment.

Tumor evolution selectively inactivates the core microRNA machinery for immune evasion.

Cancer cells acquire genetic heterogeneity to escape from immune surveillance during tumor evolution, but a systematic approach to distinguish driver from passenger mutations is lacking. Here we investigate the impact of different immune pressure on tumor clonal dynamics and immune evasion mechanism, by combining massive parallel sequencing of immune edited tumors and CRISPR library screens in syngeneic mouse tumor model and co-culture system. We find that the core microRNA (miRNA) biogenesis and targeting machinery maintains the sensitivity of cancer cells to PD-1-independent T cell-mediated cytotoxicity. Genetic inactivation of the machinery or re-introduction of ANKRD52 frequent patient mutations dampens the JAK-STAT-interferon-γ signaling and antigen presentation in cancer cells, largely by abolishing miR-155-targeted silencing of suppressor of cytokine signaling 1 (SOCS1). Expression of each miRNA machinery component strongly correlates with intratumoral T cell infiltration in nearly all human cancer types. Our data indicate that the evolutionarily conserved miRNA pathway can be exploited by cancer cells to escape from T cell-mediated elimination and immunotherapy.

A dynamic and multilocus metabolic regulation strategy using quorum-sensing-controlled bacterial small RNA.

Metabolic regulation strategies have been developed to redirect metabolic fluxes to production pathways. However, it is difficult to screen out target genes that, when repressed, improve yield without affecting cell growth. Here, we report a strategy using a quorum-sensing system to control small RNA transcription, allowing cell-density-dependent repression of target genes. This strategy is shown with convenient operation, dynamic repression, and availability for simultaneous regulation of multiple genes. The parameters Ai, Am, and RA (3-oxohexanoyl-homoserine lactone [AHL] concentrations at which half of the maximum repression and the maximum repression were reached and value of the maximum repression when AHL was added manually, respectively) are defined and introduced to characterize repression curves, and the variant LuxRI58N is identified as the most suitable tuning factor for shake flask culture. Moreover, it is shown that dynamic overexpression of the Hfq chaperone is the key to combinatorial repression without disruptions on cell growth. To show a broad applicability, the production titers of pinene, pentalenene, and psilocybin are improved by 365.3%, 79.5%, and 302.9%, respectively, by applying combinatorial dynamic repression.