Efficacy and Tolerability of Pyrazolo[1,5- a]pyrimidine RET Kinase Inhibitors for the Treatment of Lung Adenocarcinoma

Abstract

RET (REarranged during Transfection) kinase gain-of-function aberrancies have been identified as potential oncogenic drivers in lung adenocarcinoma, along with several other cancer types, prompting the discovery and assessment of selective inhibitors. Internal mining and analysis of relevant kinase data informed the decision to investigate a pyrazolo[1,5-a]pyrimidine scaffold, where subsequent optimization led to the identification of compound WF-47-JS03 (1), a potent RET kinase inhibitor with >500-fold selectivity against KDR (Kinase insert Domain Receptor) in cellular assays. In subsequent mouse in vivo studies, compound 1 demonstrated effective brain penetration and was found to induce strong regression of RET-driven tumor xenografts at a well-tolerated dose (10 mg/kg, po, qd). Higher doses of 1, however, were poorly tolerated in mice, similar to other pyrazolo[1,5-a]pyrimidine compounds at or near the efficacious dose, and indicative of the narrow therapeutic windows seen with this scaffold.

Figure 1

X-ray cocrystal structure of 7 and RET reveals significant water-mediated interactions of R₂ substituent NH with key kinase residues E775, K758, D892, and S891.

Figure 2

RET inhibitor compound 1 (WF-47-JS03) significantly inhibits tumor growth in RIE KIF5B-RET xenograft mice and is well tolerated at 1, 3, and 10 mg/kg in the 10 day study.

Figure 3

Western Blot confirmation of intratumoral RET target engagement in RIE KIF5B-RET xenograft mice at tumor stasis dose (3 mg/kg, po, MC/Tween80, suspension) at the 5 h time point after 10 days of compound 1 administration.

Figure 4

Treatment of LC-2/ad tumor-bearing mice with 1 (WF-47-JS03) for 21 days is well-tolerated and leads to significant regression at 10 mg/kg.

Figure 5

Treatment with RET inhibitor compound 9 (YA-14-WJ39) demonstrates moderate tumor regression in RIE KIF5B-RET xenograft mice and is well tolerated at 30 mg/kg in the 8 day study.

Figure 6

Significant intratumoral RET target engagement is demonstrated in RIE KIF5B-RET xenograft mice at the tumor regression dose (30 mg/kg, po, MC/Tween80, suspension) at the 5 h time point after 8 days of compound 9 administration. Tumor volume was extremely small in one mouse from the 5 h cohort, leading to likely profiling of stroma tissue.