MTORC1 is aberrantly activated in renal cell carcinoma (RCC) and is targeted by rapalogs. As for other targeted therapies, rapalogs clinical utility is limited by the development of resistance. Resistance often results from target mutation, but mTOR mutations are rarely found in RCC. As in humans, prolonged rapalog treatment of RCC tumorgrafts (TGs) led to resistance. Unexpectedly, explants from resistant tumors became sensitive both in culture and in subsequent transplants in mice. Notably, resistance developed despite persistent mTORC1 inhibition in tumor cells. In contrast, mTORC1 became reactivated in the tumor microenvironment (TME). To test the role of the TME, we engineered immunocompromised recipient mice with a resistance mTOR mutation (S2035T). Interestingly, TGs became resistant to rapalogs in mTOR ^S2035T mice. Resistance occurred despite mTORC1 inhibition in tumor cells and could be induced by coculturing tumor cells with mutant fibroblasts. Thus, enforced mTORC1 activation in the TME is sufficient to confer resistance to rapalogs. These studies highlight the importance of mTORC1 inhibition in nontumor cells for rapalog antitumor activity and provide an explanation for the lack of mTOR resistance mutations in RCC patients.

Patients with renal cell carcinoma (RCC) and hypercalcemia (HC) have worse outcomes. HC often involves parathyroid hormone–related protein (PTHrP), and the role of hypoxia-inducible factor 2 (HIF-2) is incompletely understood. Leveraging RCC tumorgraft (TG) models of HC, which were characterized by tumor cell–autonomous inflammatory/immune signatures, we show that HIF-2 inhibition with PT2399 frequently normalized calcium, downregulated circulating PTHrP, and reduced HIF-2 binding to the PTHLH (PTHrP) promoter. Likely contributing to the selective induction of PTHrP in a subset of HIF-2–dependent tumors, the PTHLH locus was generally more accessible in TG(HC). However, PTHLH chromatin accessibility was grossly unaffected by PT2399, unlike elsewhere (including the EPO locus in a TG with paraneoplastic polycythemia). As in TGs, paraneoplastic HC in patients was associated with clear-cell (cc)RCC (and sarcomatoid/rhabdoid differentiation) and was rapidly corrected by PT2977/belzutifan, which unlike bisphosphonates downregulated PTHrP. Our data support evaluating HIF-2 antagonists for ccRCC patients with paraneoplastic HC, which may serve as a predictive biomarker. Significance: This study uncovers a direct role for HIF-2 in driving humoral HC of malignancy in ccRCC through transcriptional activation of PTHLH (encoding PTHrP), identifies HC (and PTHrP) as potentially predictive biomarkers of HIF-2 engagement, and sets a foundation for the evaluation of HIF-2 antagonists for HC management in patients.

ATAC profile of hypercalcemic TGs

Clinicopathological features of metastatic RCC patients with high circulating PTHrP levels