Novel derivatives of anaplastic lymphoma kinase inhibitors: Synthesis, radiolabeling, and preliminary biological studies of fluoroethyl analogues of crizotinib, alectinib, and ceritinib

Abstract

Anaplastic lymphoma kinase (ALK), an oncogenic receptor tyrosine kinase, is a therapeutic target in various cancers, including non-small cell lung cancer. Although several ALK inhibitors, including crizotinib, ceritinib, and alectinib, are approved for cancer treatment, their long-term benefit is often limited by the cancer's acquisition of resistance owing to secondary point mutations in ALK. Importantly, some ALK inhibitors cannot cross the blood-brain barrier (BBB) and thus have little or no efficacy against brain metastases. The introduction of a lipophilic moiety, such as a fluoroethyl group may improve the drug's BBB penetration. Herein, we report the synthesis of fluoroethyl analogues of crizotinib 1, alectinib 4, and ceritinib 9, and their radiolabeling with ¹⁸F for pharmacokinetic studies. The fluoroethyl derivatives and their radioactive analogues were obtained in good yields with high purity and good molar activity. A cytotoxicity screen in ALK-expressing H2228 lung cancer cells showed that the analogues had up to nanomolar potency and the addition of the fluorinated moiety had minimal impact overall on the potency of the original drugs. Positron emission tomography in healthy mice showed that the analogues had enhanced BBB penetration, suggesting that they have therapeutic potential against central nervous system metastases.

Keywords: Anaplastic lymphoma kinase; Brain metastasis; Lung cancer; PET; Targeted therapeutics.

Fig. 1.

Structures of several well-known ALK inhibitors.

Fig. 2.

Western blotting of ALK expression in lung cancer cells. Lane 1, molecular weight markers; lane 2, H2228 cell lysate; lane 3, H441 cell lysate. GAPDH was probed as a loading control.

Fig. 3.

Cytotoxicity of compounds 1 (A), 4 (B), and 9 (C) and their parent compounds in ALK-expressing H2228 lung cancer cells, and cytotoxicity of compound 4 and its parent compound in non-ALK-expressing H441 lung cancer cells (D). Data are means ± standard deviations.

Fig. 4.

Uptake of [¹⁸F]1 (A), [¹⁸F]4 (B), and [¹⁸F]9 (C) by ALK-positive H2228 cells and ALK-negative H441 cells. Data are means ± standard deviations.

Fig. 5.

Mean time-activity curves and distributions of [¹⁸F]1 (A), [¹⁸F]4 (B), and [¹⁸F]9 (C) in major organs, including brain, heart, liver, kidney, and muscle, in normal nude mice. Data are means ± standard deviations.

Fig. 6.

Maximum intensity projection PET images show the biodistribution of [¹⁸F]1 in a representative animal at 5, 10, 30, and 60 min after injection. B=brain, K=kidney, H=heart, and L=liver.

Fig. 7.

Maximum intensity projection PET images show the biodistribution of [¹⁸F]4 of in a representative animal at 5, 10, 30, and 60 min after injection. B=brain, K=kidney, H=heart, and L=liver.

Fig. 8.

Maximum intensity projection PET images show the biodistribution of [¹⁸F]9 in a representative animal at 5, 10, 30 and 60 min after injection. B=brain, K=kidney, H=heart, and L=liver.

Scheme 1.

Synthesis of fluoroethyl crizotinib 1 by Methods 1 and 2.

Scheme 2.

Radiosynthesis of [¹⁸F]fluoroethyl crizotinib ([¹⁸F]1) by Methods 1 and 2.

Scheme 3.

Synthesis of fluoroethyl alectinib 4 by Methods 1 and 2.

Scheme 4:

Radiosynthesis of [¹⁸F]fluoroethyl alectinib ([¹⁸F]4) by Method 2.