Abstract 362: Prevention of centrosome clustering by targeting the motor protein X: A promising therapeutic strategy for small cell lung cancer

Abstract Cancer cells often have more than two centrosomes (centrosome amplification). This is a cancer hallmark. Cancer cells cluster supernumerary centrosomes during mitosis so that they can undergo bipolar mitosis, similar to normal cells. We previously reported that inhibition of centrosome clustering by CDK2 antagonism in non-small lung cell cancer caused multipolar cell division and apoptotic death, termed anaphase catastrophe. Anaphase catastrophe preferentially affects cancer cells, sparing normal cells with two centrosomes. This therapeutic window implicates triggering anaphase catastrophe as a promising antineoplastic pathway. Motor protein X is reported to affect centrosome clustering but is not essential in normal cells. Small cell lung cancer (SCLC) has p53 inactivation and a poor prognosis with limited therapeutic options. Loss of p53 function disrupts the cell cycle checkpoint and is associated with centrosome amplification. We hypothesized that SCLC is vulnerable to mitotic abnormalities, including anaphase catastrophe. We explore here protein X as a novel therapeutic target that triggers anaphase catastrophe and apoptosis in SCLC. In vitro studies of multiple SCLC cell lines were performed. Immunofluorescence staining of pericentrin revealed higher frequency of centrosome amplification in SCLC than in normal human bronchial epithelial (NHBE) cells. Immunoblotting indicated that motor protein X was over-expressed in SCLC cells compared to NHBE cells. Functional analyses of protein X was done using siRNAs, CRISPR-Cas9 system, and a specific inhibitor. Protein X inhibition reduced cell proliferation and triggered apoptosis in the examined SCLC cell lines. In NHBE cells with infrequent centrosome amplification, these effects were rarely observed. To elucidate the mechanism for these antineoplastic effects, the mitotic status of SCLC cells was studied by staining for α-tubulin, pericentrin, and DAPI. Cells with clustered supernumerary centrosomes were statistically significantly decreased, while the number of multipolar mitotic cells were statistically significantly increased upon protein X inhibition. This indicated that targeting the protein X produced anaphase catastrophe in SCLC cells. We confirmed and extended these in vitro findings to the in vivo setting. The protein X inhibitor statistically significantly suppressed the growth of NCI-H146 tumor xenografts in mice. Pericentrin staining of resected treated xenografts revealed that the protein X inhibitor antagonized clustering of supernumerary centrosomes and led to multipolar mitosis in vivo. Taken together, prevention of supernumerary centrosome clustering by targeting the motor protein X is an attractive therapeutic strategy for SCLC. Citation Format: Natsuki Nakagawa, Masakatsu Tokunaga, Mirei Ka, Takahiro Iida, Takahiro Ando, Akiko Kunita, Kousuke Watanabe, Yasunori Ohta, Xi Liu, Ethan Dmitrovsky, Hidenori Kage, Masanori Kawakami. Prevention of centrosome clustering by targeting the motor protein X: A promising therapeutic strategy for small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 362.