Kimal Rajapakshe

Background: Sporadic aortic aneurysm and dissection (AAD), caused by progressive aortic smooth muscle cell (SMC) loss and extracellular matrix degradation, is a highly lethal condition. Identifying mechanisms that drive aortic degeneration is a crucial step in developing an effective pharmacologic treatment to prevent disease progression. Recent evidence has indicated that cytosolic DNA and abnormal activation of the cytosolic DNA sensing adaptor STING (stimulator of interferon genes) play a critical role in vascular inflammation and destruction. Here, we examined the involvement of this mechanism in aortic degeneration and sporadic AAD formation. Methods: The presence of cytosolic DNA in aortic cells and activation of the STING pathway were examined in aortic tissues from patients with sporadic ascending thoracic AAD. The role of STING in AAD development was evaluated in Sting -deficient ( Sting gt/gt ) mice in a sporadic AAD model induced by challenging mice with a combination of a high-fat diet and angiotensin II. We also examined the direct effects of STING on SMC death and macrophage activation in vitro. Results: In human sporadic AAD tissues, we observed the presence of cytosolic DNA in SMCs and macrophages and significant activation of the STING pathway. In the sporadic AAD model, Sting gt/gt mice showed significant reductions in challenge-induced aortic enlargement, dissection, and rupture in both the thoracic and abdominal aortic regions. Single-cell transcriptome analysis revealed that aortic challenge in wild-type mice induced the DNA damage response, the inflammatory response, dedifferentiation and cell death in SMCs, and matrix metalloproteinase expression in macrophages. These changes were attenuated in challenged Sting gt/gt mice. Mechanistically, nuclear and mitochondrial DNA damage in SMCs and the subsequent leak of DNA to the cytosol activated STING signaling, which induced cell death through apoptosis and necroptosis. In addition, DNA from damaged SMCs was engulfed by macrophages in which it activated STING and its target interferon regulatory factor 3, which directly induced matrix metalloproteinase-9 expression. We also found that pharmacologically inhibiting STING activation partially prevented AAD development. Conclusions: Our findings indicate that the presence of cytosolic DNA and subsequent activation of cytosolic DNA sensing adaptor STING signaling represent a key mechanism in aortic degeneration and that targeting STING may prevent sporadic AAD development.

KRAS inhibitors demonstrate clinical efficacy in pancreatic ductal adenocarcinoma (PDAC); however, resistance is common. Among patients with KRASG12C-mutant PDAC treated with adagrasib or sotorasib, mutations in PIK3CA and KRAS, and amplifications of KRASG12C, MYC, MET, EGFR, and CDK6 emerged at acquired resistance. In PDAC cell lines and organoid models treated with the KRASG12D inhibitor MRTX1133, epithelial-to-mesenchymal transition and PI3K-AKT-mTOR signaling associate with resistance to therapy. MRTX1133 treatment of the KrasLSL-G12D/+; Trp53LSL-R172H/+; p48-Cre (KPC) mouse model yielded deep tumor regressions, but drug resistance ultimately emerged, accompanied by amplifications of Kras, Yap1, Myc, Cdk6, and Abcb1a/b, and co-evolution of drug-resistant transcriptional programs. Moreover, in KPC and PDX models, mesenchymal and basal-like cell states displayed increased response to KRAS inhibition compared to the classical state. Combination treatment with KRASG12D inhibition and chemotherapy significantly improved tumor control in PDAC mouse models. Collectively, these data elucidate co-evolving resistance mechanisms to KRAS inhibition and support multiple combination therapy strategies. Significance: Acquired resistance may limit the impact of KRAS inhibition in patients with PDAC. Using clinical samples and multiple preclinical models, we define heterogeneous genetic and non-genetic mechanisms of resistance to KRAS inhibition that may guide combination therapy approaches to improve the efficacy and durability of these promising therapies for patients. See related commentary by Marasco and Misale, p. 2018.

Somatic missense mutations in the substrate-binding pocket of the E3 ubiquitin ligase adaptor SPOP are present in up to 15% of human prostate adenocarcinomas, but are rare in other malignancies, suggesting a prostate-specific mechanism of action. SPOP promotes ubiquitination and degradation of several protein substrates, including the androgen receptor (AR) coactivator SRC-3. However, the relative contributions that SPOP substrates may make to the pathophysiology of SPOP-mutant (mt) prostate adenocarcinomas are unknown. Using an unbiased bioinformatics approach, we determined that the gene expression profile of prostate adenocarcinoma cells engineered to express mt-SPOP overlaps greatly with the gene signature of both SRC-3 and AR transcriptional output, with a stronger similarity to AR than SRC-3. This finding suggests that in addition to its SRC-3-mediated effects, SPOP also exerts SRC-3-independent effects that are AR-mediated. Indeed, we found that wild-type (wt) but not prostate adenocarcinoma-associated mutants of SPOP promoted AR ubiquitination and degradation, acting directly through a SPOP-binding motif in the hinge region of AR. In support of these results, tumor xenografts composed of prostate adenocarcinoma cells expressing mt-SPOP exhibited higher AR protein levels and grew faster than tumors composed of prostate adenocarcinoma cells expressing wt-SPOP. Furthermore, genetic ablation of SPOP was sufficient to increase AR protein levels in mouse prostate. Examination of public human prostate adenocarcinoma datasets confirmed a strong link between transcriptomic profiles of mt-SPOP and AR. Overall, our studies highlight the AR axis as the key transcriptional output of SPOP in prostate adenocarcinoma and provide an explanation for the prostate-specific tumor suppressor role of wt-SPOP.