Overactive IGF1/Insulin Receptors and NRASQ61R Mutation Drive Mechanisms of Resistance to Pazopanib and Define Rational Combination Strategies to Treat Synovial Sarcoma

Abstract

Pazopanib is approved for treatment of advanced soft tissue sarcomas, but primary and secondary drug resistance limits its clinical utility. We investigated the molecular mechanisms mediating pazopanib resistance in human synovial sarcoma (SS) models. We found reduced cell sensitivity to pazopanib associated with inefficient inhibition of the two critical signaling nodes, AKT and ERKs, despite strong inhibition of the main drug target, PDGFRα. In the CME-1 cell line, overactivation of IGF1 and Insulin receptors (IGF1R/InsR) sustained AKT activation and pazopanib resistance, which was overcome by a combination treatment with the double IGF1R/InsR inhibitor BMS754807. In the highly pazopanib resistant MoJo cell line, NRASQ61R mutation sustained constitutive ERK activation. Transfection of the NRAS mutant in the pazopanib sensitive SYO-1 cell line increased the drug IC50. MoJo cells treatment with pazopanib in combination with the MEK inhibitor trametinib restored ERK inhibition, synergistically inhibited cell growth, and induced apoptosis. The combination significantly enhanced the antitumor efficacy against MoJo orthotopic xenograft abrogating growth in 38% of mice. These findings identified two different mechanisms of intrinsic pazopanib resistance in SS cells, supporting molecular/immunohistochemical profiling of tumor specimens as a valuable approach to selecting patients who may benefit from rational drug combinations.

Sarcoma treatment in the era of molecular medicine

“…Despite a strong inhibition of the main target of pazopanib, PDGFRα/β, the activation of the AKT and ERK signaling pathways was only partially impaired, possibly due to the over activation of other tyrosine kinase receptors, including the insulin‐like growth factor receptor type 1 (IGF1R) and insulin receptor (IR). Similarly, in another SS cell line, the presence of an NRAS mutation sustained ERK activation and caused resistance to pazopanib treatment (Lanzi et al , 2019). Thus, a combination treatment with either an IGF1R/IR inhibitor or a MEK inhibitor has been suggested to restore the inhibition of the PDGFRα/β pathways and effectively promote apoptosis (Lanzi et al , 2019).…”

“…Similarly, in another SS cell line, the presence of an NRAS mutation sustained ERK activation and caused resistance to pazopanib treatment (Lanzi et al , 2019). Thus, a combination treatment with either an IGF1R/IR inhibitor or a MEK inhibitor has been suggested to restore the inhibition of the PDGFRα/β pathways and effectively promote apoptosis (Lanzi et al , 2019). Phosphoproteomic profiling of pazopanib‐resistant cells identified the inhibition of HSP90 as a therapeutic route to overcome resistance (Vyse et al , 2018).…”

“…In non‐GIST STSs, the currently approved targeted therapies are limited to the multi‐target TKI pazopanib, which targets VEGFR‐1, VEGFR‐2, and VEGFR‐3, PDGFRα and PDGFR‐β; and KIT (Lee et al , 2019). It has been demonstrated that anti‐angiogenic TKIs, including pazopanib, do not succeed in targeting sarcoma stem cells (Canter et al , 2014), whereas treatment with pazopanib in a human SS model promotes the development of resistance (Lanzi et al , 2019). Despite a strong inhibition of the main target of pazopanib, PDGFRα/β, the activation of the AKT and ERK signaling pathways was only partially impaired, possibly due to the over activation of other tyrosine kinase receptors, including the insulin‐like growth factor receptor type 1 (IGF1R) and insulin receptor (IR).…”

Cooperation between SS18-SSX1 and miR-214 in Synovial Sarcoma Development and Progression

“…These changes were not observed in eMCs expressing SYT-SSX1, suggesting that the expression modulation of the Atf2/Tle1/Egr1 axis might be achieved at a later stage. However, Igf1, which is downstream of the Cyclin D1 pathway, was upregulated both in tumors and eMCs expressing SS18-SSX1, which might be important for drug resistance in synovial sarcoma treatment [22]. Overall comparison of differentially expressed genes between SS18-SSX1-expressing eMCs and tumors showed that 3155 and 485 genes were upregulated in synovial sarcoma versus eMCs with an empty vector; fold change > 2.0, and in SS18-SSX1-expressing eMCs versus eMCs with an empty vector; fold change > 2.0, ( Figure 2E and Table S2).…”

“…Retroviral integration sites were identified by the inverse-PCR method previously described [22,23]. Genomic DNA sequences flanking the retrovirus were cloned into the pGEM T-easy plasmid (Promega, Madison, WI, USA), sequenced, and mapped on mouse chromosome.…”