Synergistic effects of crizotinib and radiotherapy in experimental EML4-ALK fusion positive lung cancer.

Non-small cell lung cancer (NSCLC) patients with chromosomal rearrangements of the anaplastic lymphoma kinase gene (ALK) are sensitive to the tyrosine kinase inhibitor crizotinib. We aimed to investigate the effects of combined radiotherapy and crizotinib in ALK-positive vs. wild type NSCLC models. Clonogenic survival, proliferation and apoptosis of cells exposed to crizotinib and radiotherapy (photon and carbon ions) were evaluated in ALK mutation positive (ALK+; H3122) and negative (ALK-; A549 and LLC) NSCLC lines. The syngeneic mouse (LLC) and human (H3122) xenograft tumor models were further studied in vivo. Tumor growth kinetics, microvascular density (MVD), perfusion and proliferation were assessed. Crizotinib exerted potent and selective anti-proliferative and pro-apoptotic effects in ALK+ H3122 cells which were augmented by radiotherapy. The synergistic effect of this combination in ALK+ NSCLC was confirmed by isobologram analysis. Crizotinib also sensitized H3122 cells to particle therapy with carbon ions. In H3122 xenografts, dual combination was most effective in reducing tumor proliferation, MVD and perfusion. In contrast, in the LLC model, crizotinib led only to a transient tumor growth inhibition and combined treatment was inferior to radiotherapy alone. Crizotinib elicits beneficial effects in combination with radiotherapy only in ALK-positive NSCLC.

Dai Y ，Wei Q ，Schwager C ，Moustafa M ，Zhou C ，Lipson KE ，Weichert W ，Debus J ，Abdollahi A ... -  《-》

Oncogene addiction and radiation oncology: effect of radiotherapy with photons and carbon ions in ALK-EML4 translocated NSCLC.

Dai Y ，Wei Q ，Schwager C ，Hanne J ，Zhou C ，Herfarth K ，Rieken S ，Lipson KE ，Debus J ，Abdollahi A ... -  《Radiation Oncology》

ALK inhibitor PF02341066 (crizotinib) increases sensitivity to radiation in non-small cell lung cancer expressing EML4-ALK.

Crizotinib (PF02341066) is a tyrosine kinase inhibitor of anaplastic lymphoma kinase (ALK) that has been shown to selectively inhibit growth of cancer cells that harbor the EML4-ALK fusion found in a subset of patients with non-small cell lung cancer (NSCLC). While in clinical trials, PF02341066 has shown a significant therapeutic benefit as a single agent; the effectiveness of combining it with other therapeutic modalities including ionizing radiation remains unknown. To further elucidate the role of PF02341066 in tumor inhibition, we examined its effects alone and in combination with radiation on downstream signaling, apoptosis, and radiosensitivity in two NSCLC cell lines in vitro: H3122, which harbors the EML4-ALK fusion, and H460, which does not. We also examined the in vivo effects of PF02341066 in H3122 mouse xenografts. In the H3122 cell line, PF02341066 inhibited phosphorylation of ALK and its downstream effectors: AKT, ERK, and STAT3. H3122 cells treated with a combination of PF02341066 and radiation showed an increase in cellular apoptosis and were sensitized to radiation therapy (dose enhancement ratio, 1.43; P < 0.0001). Moreover, in an H3122 xenograft model, the combined treatment resulted in greater tumor growth inhibition than either treatment alone (P < 0.05). None of these effects was observed in the EML4-ALK-negative H460 cells. Our findings indicate that PF02341066 acts as a radiation sensitizer in cells harboring the EML4-ALK fusion, providing a rationale for a clinical trial combining ALK inhibitor with radiation in the NSCLCs expressing ALK.

Sun Y ，Nowak KA ，Zaorsky NG ，Winchester CL ，Dalal K ，Giacalone NJ ，Liu N ，Werner-Wasik M ，Wasik MA ，Dicker AP ，Lu B ... -  《-》

Targeting stemness is an effective strategy to control EML4-ALK+ non-small cell lung cancer cells.

Oh SJ ，Noh KH ，Lee YH ，Hong SO ，Song KH ，Lee HJ ，Kim S ，Kim TM ，Jeon JH ，Seo JH ，Kim DW ，Kim TW ... -  《Oncotarget》

Development of potent ALK inhibitor and its molecular inhibitory mechanism against NSCLC harboring EML4-ALK proteins.

Here, we show the newly synthesized and potent ALK inhibitor having similar scaffold to KRCA-0008, which was reported previously, and its molecular mechanism against cancer cells harboring EML4-ALK fusion protein. Through ALK wild type enzyme assay, we selected two compounds, KRCA-0080 and KRCA-0087, which have trifluoromethyl instead of chloride in R2 position. We characterized these newly synthesized compounds by in vitro and in vivo assays. Enzyme assay shows that KRCA-0080 is more potent against various ALK mutants, including L1196M, G1202R, T1151_L1152insT, and C1156Y, which are seen in crizotinib-resistant patients, than KRCA-0008 is. Cell based assays demonstrate our compounds downregulate the cellular signaling, such as Akt and Erk, by suppressing ALK activity to inhibit the proliferation of the cells harboring EML4-ALK. Interestingly, our compounds induced strong G1/S arrest in H3122 cells leading to the apoptosis, which is proved by PARP-1 cleavage. In vivo H3122 xenograft assay, we found that KRCA-0080 shows significant reduction in tumor size compared to crizotinib and KRCA-0008 by 15-20%. Conclusively, we report a potent ALK inhibitor which shows significant in vivo efficacy as well as excellent inhibitory activity against various ALK mutants.

Kang CH ，Yun JI ，Lee K ，Lee CO ，Lee HK ，Yun CS ，Hwang JY ，Cho SY ，Jung H ，Kim P ，Ha JD ，Jeon JH ，Choi SU ，Jeong HG ，Kim HR ，Park CH ... -  《-》