Abstract
BackgroundRenal cell carcinomas (RCC) harboring a TFE3 gene fusion (TfRCC) represent an aggressive subset of kidney tumors. Key signaling pathways of TfRCC are unknown and preclinical in vivo data are lacking. We investigated Akt/mTOR pathway activation and the preclinical efficacy of dual mTORC1/2 versus selective mTORC1 inhibition in TfRCC.MethodsLevels of phosphorylated Akt/mTOR pathway proteins were compared by immunoblot in TfRCC and clear cell RCC (ccRCC) cell lines. Effects of the mTORC1 inhibitor, sirolimus, and the dual mTORC1/2 inhibitor, AZD8055, on Akt/mTOR activation, cell cycle progression, cell viability and cytotoxicity were compared in TfRCC cells. TfRCC xenograft tumor growth in mice was evaluated after 3-week treatment with oral AZD8055, intraperitoneal sirolimus and respective vehicle controls.ResultsThe Akt/mTOR pathway was activated to a similar or greater degree in TfRCC than ccRCC cell lines and persisted partly during growth factor starvation, suggesting constitutive activation. Dual mTORC1/2 inhibition with AZD8055 potently inhibited TfRCC viability (IC50 = 20-50 nM) due at least in part to cell cycle arrest, while benign renal epithelial cells were relatively resistant (IC50 = 400 nM). Maximal viability reduction was greater with AZD8055 than sirolimus (80–90% versus 30–50%), as was the extent of Akt/mTOR pathway inhibition, based on significantly greater suppression of P-Akt (Ser473), P-4EBP1, P-mTOR and HIF1α. In mouse xenograft models, AZD8055 achieved significantly better tumor growth inhibition and prolonged mouse survival compared to sirolimus or vehicle controls.ConclusionsAkt/mTOR activation is common in TfRCC and a promising therapeutic target. Dual mTORC1/2 inhibition suppresses Akt/mTOR signaling more effectively than selective mTORC1 inhibition and demonstrates in vivo preclinical efficacy against TFE3-fusion renal cell carcinoma.
Highlights
Renal cell carcinomas (RCC) harboring a TFE3 gene fusion (TfRCC) represent an aggressive subset of kidney tumors
Immunoblotting Phosphorylated and total levels of Akt/mTOR pathway proteins were measured by immunoblot in TFE3–fusion renal cell carcinoma (TfRCC) and clear cell RCC (ccRCC) cell lines. ccRCC cell lines were used for comparison since we have previously shown that this RCC subtype has frequent constitutive activation of the Akt/mTOR pathway [20]
Akt/mTOR pathway activation in TfRCC cells Akt/mTOR pathway activation was observed in all serumsupplemented TfRCC cell lines (Fig. 1a)
Summary
Renal cell carcinomas (RCC) harboring a TFE3 gene fusion (TfRCC) represent an aggressive subset of kidney tumors. Renal cell carcinoma (RCC) consists of distinct subtypes with characteristic histologic features, genetic mutations and clinical behaviors [1]. The RCC subtype harboring an Xp11.2 chromosomal rearrangement (Xp11 Translocation RCC, TFE3-fusion RCC, TfRCC) comprises 1–5% of all RCC cases [2,3,4,5]. Rearrangements include an inversion or translocation of the TFE3 gene (Xp11.2), which is a member of the Microphthalmia-associated transcription factor (MiT) family that regulates growth and differentiation [6]. The diagnosis is suggested by young age, tumor histology and nuclear immunoreactivity for the TFE3 C-terminus; confirmation of diagnosis requires cytogenetic or molecular evidence of an Xp11 rearrangement or fusion transcript [8, 10, 11]
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