The Mechanism Of FGFR1 Regulating The Cancer Stem Cell Like-phenotype In Lung Squamous Cell Cancer

Objective:Lung squamous cell cancer(LSCC)is one of the major subtype of non-small cell lung cancer(NSCLC),accounts for 30% of all lung cancer cases.Compared with the lung adenocarcinoma(LADC),the mainstream treatment regimens for lung SCC(LSCC)are still platinum-based chemotherapeutics,and the advance of targeted therapy in LSCC is rare.Thus,it is essential to further explore the molecular mechanism underlying the occurrence and development of lung cancer,especially in LSCC,and find out the druggable driver genes for the future intervention strategies.FGFR1 amplification is a prominent gene alteration in NSCLC,especially in LSCC.However its biological function in the occurance and development of LSCC is still an unknown field waiting to be explored.In our studies,we focus on the role of FGFR1 in maintaining the stem cell-like phenotype in lung squamous cancer cells,and the molecular mechanism underlying this course.Based on these results,we will discuss the cause of primary resistance against FGFR1 TKI in LSCC preliminarily.Methods and Results:(1)We isolated and enriched the oncosphere from FGFR1 amplified cell lines in serum-free culture medium at low adherence.And through RT-PCR,flowcytometry clonal expansion expreiments of individual cells and subcutaneous transplanted model,we found that these FGFR1 amplified cells can form oncosphere in the serum-free culture medium and exhibit the stem cell-like phenotype compared with these parenatal cells.Further more,we established the role of FGFR1 as a prominent oncogene in Chinese LSCC patients,rather than other members of FGFR1 family.Then,we found that treatment with FGFR1 inhibitor AZD4547 suppressed the growth of tumor spheres and reduced ALDH positive proportion in FGFR1-amplified lung cancer cells in vitro,as well as inhibited the growth of oncospheres and parental cells in xenograft models.To explore the role of FGFR1 directly,we established the stable trasfected FGFR1 knockdown cell lines,and we found knockdown of FGFR1 recaptured the similar effect as AZD4547 in vitro.(2)To explore the role of GLI2 in maintaining the stem cell-like phenotype of FGFR1 amplified cells,we established the stable trasfected FGFR1 knockdown cell lines.We found that knockdown of GLI2 dramatically inhibited the growth of these oncospheres,reduced ALDH positive proportion,and inhibited the stem cell markers expression in FGFR1-amplified lung cancer cells in vitro.Later,we verified that FGFR1 regulates the expression of GLI2 through ERK pathway.And then,we found that overexpression of GLI2 sufficiently rescued the phenotype caused by FGFR1 knockdown.Notably we also identified a correlation between FGFR1 and GLI2 expressions from clinical data,as well as an inverse relationship with progression free survival(PFS).(3)Based on further studies,we foud there were as correlationship between FGFR1,SOX2 and GLI2 in LSCC tissues.And then we found that FGFR1 can regulated the expression of SOX2,and SOX2 can regulated the expression of GLI2 further.And overexpression of SOX2 can weaken the sensitivity of FGFR1 TKI against these FGFR1 amplified lung cancer cells.Conclusions:We found that FGFR1 can regulate the stem cell-like phenotype and the self-renewal of FGFR1-amplified lung cancer cells through GLI2 pathways.we also identified a correlation between FGFR1 and GLI2 expressions from clinical data,as well as an inverse relationship with progression free survival(PFS).Further more,we found the existance of coexpression between FGFR1,SOX2 and GLI2.FGFR1 regulated the expression of GLI2 may through SOX2.We also found that SOX2 overexpression may lead to FGFR1 TKI resistance in LSCC.