Abstract 4667: pCR induced by triplet therapy with low-dose afatinib, cetuximab and bevcizumab in lung cancer cells harboring EGFR T790M
Authors: Ohashi Kadoaki , Kudo Kenichiro , Ichihara Eiki , Kayatani Hiroe , Higo Hisao , Kato Yuka , Kurata Yasuko , Minami Daisuke , Ninomiya Takashi , Kubo Toshio , Hotta Katsuyuki , Tanimoto Mitsune , Kiura Katsuyuki 
Published: July 2016

Abstract
Background. EGFR T790M plays a major role on the resistance for 1st or 2nd-generation EGFR-TKIs (e.g. erlotinib, afatinib) in lung tumors with activating EGFR mutations. 3rd generation EGFR-TKIs (e.g. AZD9291) show an excellent response in lung tumors with EGFR T790M, while the acquired resistance is inevitable (NEJM 2015). The combination of afatinib and cetuximab also induced significant response in the lung cancers in clinical trial (Cancer Discov. 2014). However, there is clearly still room for improvement in the combination therapy. Bevacizumab, a key drug in lung cancer treatment, is thought to have a wide variety of effects, including anti-vascularization and improved drug delivery. Thus, we hypothesized that the dosage of afatinib and cetuximab could be reduced by combining with bevacizumab, and that the triple therapy might produce better tumor inhibition with tolerable toxicity.
Methods. Two lung cancer cell lines, H1975, with EGFR L858R and T790M, and RPC-9, with EGFR exon 19 deletion and T790M, were injected subcutaneously into nude mice as xenograft models. Mice were divided into four groups 10 days after tumor cell inoculation (vehicle, afatinib, afatinib + cetuximab, afatinib + cetuximab + bevcacizumab). Compared with previous reports (afatinib 25 mg/kg, 5 times/week, cetuximab 1 mg/mouse, every 3 days; J. Clin. Invest. 2009), the low dose of afatinib (10 mg/kg, 5 times/week; i.e., 60% less drug) or cetuximab (0.1 mg/mouse, once/week; i.e., 90% less drug) were administered to the mice for 1 month. Bevacizumab was injected at 2 mg/kg twice/week. Following the therapies, the mice were observed for 1 month without treatment.
Results. Surprisingly, the triple therapy induced a pathological complete response (pCR) in H1975 and RPC-9 cell xenograft tumors; in contrast, the tumors treated with single or double therapy were inhibited only partially. AZD9291 monotherapy did not reproduce pCR. Furthermore pCR was maintained during the observation period in the triple therapy group. There was no body weight loss in any group. Frozen tumors were obtained from RPC-9 xenograft models after 1 week treatment with each regimen. Expression levels of pEGFR, pAKT, and pERK were decreased in the triple therapy group. CD31-positive blood vessels and Ki-67-positive cells in tumors with triple therapy were reduced significantly versus the tumors in other groups. Cleaved caspase-3 expression in the tumors with triple therapy was positive in a higher number of tumor cells than in the other groups. There were no difference of afatinib concentration among the tumors.
Conclusions. We demonstrated that a low-dose of afatinib and cetuximab combined with bevacizumab induced pCR in EGFR T790M mutated cell xenograft tumors with no obvious adverse events. We suggest that the suppression of neovascularization and induction of apoptosis may play important roles in the triple therapy.
This project is supported by KAKEN(No26893155)
Citation Format: Kadoaki Ohashi, Kenichiro Kudo, Eiki Ichihara, Hiroe Kayatani, Hisao Higo, Yuka Kato, Yasuko Kurata, Daisuke Minami, Takashi Ninomiya, Toshio Kubo, Katsuyuki Hotta, Mitsune Tanimoto, Katsuyuki Kiura. pCR induced by triplet therapy with low-dose afatinib, cetuximab and bevcizumab in lung cancer cells harboring EGFR T790M. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4667.