Study On The Enhancement Of Anti-tumor Effect Of High-LET Carbon Ions Combined With Autophagy Inhibition Via Promoting Er Stress-related Apoptosis

Cancer is a commonly occurring disease that presents a serious threat to human life and health.More than 100 types of cancer have thus far been identified.The 8 hallmarks of cancer are as follows: sustaining proliferative signaling,evading growth suppressors,enabling replicative immortality,resisting cell death,activating invasion and metastasis,inducing angiogenesis,evading immune destruction,and reprogramming energy metabolism.Autophagy is an evolutionarily conserved catabolic process in which components of the cytoplasm are sequestered within double-or multi-membraned vesicles referred to as autophagosomes.These components are then delivered to lysosomes for bulk degradation by vacuolar or lysosomal hydrolases.Directly related to human health and various diseases,autophagy helps cells under stress to cope with severe metabolic demand by degradation of basic cellular components.Diverse physiological or pathological changes may elicit an imbalance between endoplasmic reticulum(ER)protein folding load and capability,leading to the aggregation of unfolded or misfolded proteins in the ER lumen,a situation referred to as “ER stress.” In response to ER stress,Bi P disassociates from sensor proteins PERK,ATF6,and IRE1 and then activates a protective cellular stress response known as the unfolded protein response(UPR).Once the UPR is activated,signaling downstream reduces proteins transported into the ER and enhances the degradation of abnormal proteins by ER-associated degradation(ERAD)to restore ER function.The UPR essentially aims at reestablishing proper ER homeostasis and eventually induces autophagy and apoptosis during acute or persistent ER stress.At the same time,autophagy can effectively remove the unfolded proteins that cannot be degraded by the ERAD system,and maintain physiological and biochemical homeostasis.Charged carbon ions(CI)deposit their maximum energy at the end of the range,an effect known as the Bragg peak.High-dose radiation may be delivered to a tumor while minimizing the dose delivered to the tissues and organs at risk.Charged CI has higher relative biological effectiveness(RBE),decreased oxygen enhancement ratio(OER),reduced cell cycle and oxygen dependency radiosensitivity,and provides great advantages over conventional radiations in cancer therapy,such as X-or γ-rays.In this dissertation,by using the S180 mouse sarcoma cell line,combined in vitro and in vivo experiments after high linear energy transfer(LET)CI or X-ray irradiation,we examined how inhibition of autophagy induced by ER stress enhances apoptosis and increases the antitumor effect of ionizing radiation.We obtained the following results:1.The combination of ionizing radiation and chloroquine(CQ)could inhibit the proliferation of S180 cells,promote cell apoptosis,reduce the volume and weight of transplanted tumor,and inhibit the growth of transplanted tumor.For the S180 solid tumor,treatment with CQ 50 mg/kg a day alone could promote tumor growth.2.ER stress induced by ionizing radiation induces autophagy via the IRE1/JNK/p-Bcl-2/Beclin-1 pathway,which can alleviate ER stress and maintain the proliferation of tumor cells and tumor growth.After the inhibition of autophagy by CQ,intracellular unfolded protein or misfolded proteins cannot be cleared quickly and effectively,thus accumulating in the cells.This process leads to apoptosis of tumor cells and enhances the radiosensitivity of tumor cells and xenografts.3.CI irradiation can induce UPR signaling,and IRE1 upregulates the expression of proapoptotic protein CHOP,demonstrating the involvement of IRE1 in the apoptotic signaling pathway.Promotion of CHOP leads to the downregulation of the antiapoptotic mitochondrial protein Bcl-2 and increases the expression of apoptotic protein Bax.Meanwhile,X-ray irradiation exerted no effect on CHOP expression,which may be one of the reasons CI provides a greater advantage in tumor suppression,compared with X-ray.4.The killing effect of single CI irradiation on tumor is an acute reaction.After 15 days,the killing effect gradually weakened and even disappeared.Apoptosis in the group treated with irradiation and CQ was significantly higher than that in the the control group.Apoptotic cells in the group treated with CI irradiation and continuous CQ were significantly higher in number than that in the group treated with single-dose irradiation and CQ.This finding indicated that CI irradiation combined with continuous administration of CQ can more effectively suppress tumor.In summary,this dissertation showed that high-LET CI combined with CQ,which was used to inhibit autophagy,could enhance the antitumor effect of CI radiation via the aggravation of ER stress-related apoptosis,increasing the radiosensitivity of tumor cells.Thus,our findings could shed new light on approaches to promoting the efficacy of heavy ion radiotherapy.