Study On The Efficacy And Safety Of Apatinib In The Treatment Of Metastatic Colorectal Cancer And Synergistic Effect Of Combined PD-1 Inhibitor

PartⅠ The efficacy and safety of apatinib as a single agent in the posterior line treatment of metastatic colorectal cancer:a single-center,retrospective cohort studyBackground:The incidence of colorectal cancer ranks third among all malignant tumors,and the mortality rate ranks fifth in China.The 5-year survival rate for early colorectal cancer is about 90%,and the 5-year survival rate for advanced colorectal cancer is only 13.9%.In the past ten years,the survival of patients with advanced colorectal cancer has improved significantly.The emergence of new treatment options has extended the overall survival of advanced colorectal cancer to 3 years.Currently,chemotherapy is the standard treatment for metastatic colorectal cancer.Combination of anti-vascular endothelial growth factor with chemotherapy can prolong survival.Multi-target tyrosine kinase inhibitors such as regorafenib and fruquintinib are standard third-line treatments.However,many patients who fail the third-line treatment are still in good physical condition and have a strong desire to survive.There is an urgent need for safe and effective follow-up treatment options.Apatinib is a specific inhibitor of vascular endothelial growth factor 2 and is approved for the treatment of advanced metastatic gastric cancer and gastroesophageal junction adenocarcinoma refractory to chemotherapy.Some preclinical trials and clinical trials have shown that apatinib has a wide raryle of anti-tumor effects,including metastatic colorectal cancer,but the published literature is limited,and the relevant mechanism is still not fully understood.Objectives:This study is a single-center and retrospective cohort study.The purpose of the study is to provide more clinical evidence of apatinib in the treatment of metastatic colorectal cancer,and to find new safe treatment options for prolonging the overall survival of patients with metastatic colorectal cancer.Research Method:We collected 77 patients with metastatic colorectal cancer who failed the second or third-line treatments.Among them,35 patients failed the third-line treatment with regorafenib or fruquintinib.After the third-line,they received the chemotherapy regimen prescribed by the doctor.The other 42 patients received apatinib treatment.18 patients failed the second-line chemotherapy and 24 patients failed the third-line standard treatment.The primary endpoint is progression-free survival(PFS),and the secondary endpoints are disease control rate(DCR),objective response rate(ORR),overall survival(OS),and safety.The follow-up time was up to July 1,2019.Results:1.The median PFS of apatinib after the second-line application in metastatic colorectal cancer was 4.1 months,DCR was 57.1%,ORR was 13.9%,and median OS was 8.3 months,which were not statistically significant compared with the chemotherapy group.After inverse probability of treatment weighting(IPTW)adjustment,the mPFS was 5.0 months and mOS was 10.0 months of apatinib group,which were still not statistically significant compared with the chemotherapy group.2.In this study,there was no significant correlation between PFS and subgroup factors including the patient’s age,gender,ECOG PS,primary site,RAS status,liver metastasis,degree of differentiation and baseline serum CEA.3.The adverse reactions of patients in the apatinib group were mostly grade 1-2,and there were no grade 4 and 5 adverse reactions.the tolerability of Apatinib was acceptable.Hypertension,hand-foot syndrome,proteinuria,and hypothyroidism were only observed in the apatinib group.Peripheral neurotoxicity and cardiotoxicity were the only AEs observed in the chemotherapy group.In the common AEs of the two groups,the incidence of nausea,vomiting,granulocytopenia,anemia and fatigue in the chemotherapy group were higher than those in the apatinib group.Conclusion:Apatinib has the same efficacy as chemotherapy in the subsequent treatment of metastatic colorectal cancer,and the tolerability was acceptable.It provides an acceptable safety program for patients with metastatic colorectal cancer who are contraindicated in chemotherapy or who are unwilling to receive chemotherapy.PartⅡ Study on the mechanism of apatinib mesylate combined with PD-X inhibitor to control colon cancer growthObjectives: In the first part,we concluded that the efficacy of apatinib in the treatment of metastatic colorectal cancer is only not inferior to chemotherapy.The development of new treatment methods is crucial to enable patients to obtain more survival benefits.Immunotherapy is a new and promising therapeutic method that uses immunomodulatory antibodies as the target to enhance the anti-tumor response of T cells.Immunotherapy has shown impressive clinical effects in colorectal cancer with highly unstable microsatellites.However,for colorectal cancer patients with low microsatellite instability or stable microsatellites,the therapeutic effect needs to be further improved,and the literature reports that only about 5% of colorectal cancer patients are highly unstable with microsatellites.Generally,combination therapy has a higher response rate,better progression-free survival and overall survival compared with monotherapy.The combination of anti-angiogenesis and immune checkpoint inhibitors has seen efficacy in other tumors,but there is no relevant research on apatinib combined with immune checkpoint inhibitors in colorectal cancer.This study used apatinib combined with PD-1 inhibitor to study the effectiveness of colon cancer in preclinical models,to analyze the effects of apatinib on the tumor immunomicroenvironment and to explore the preliminary mechanism of apatinib and immune checkpoint inhibitors in the treatment of colorectal cancer.Research Method: Cultivation and expansion of mouse colon cancer cell line CT26.WT cells,inoculation of CT26.WT cells into 40 female BALB/c mice on the right inguinal area to induce a mouse colon cancer heterotopic xenograft model.When the mice were induced to form tumors,they were randomly divided into 4 groups: the first group(IgG control group),the second group(PD-1 inhibitor group),the third group(apatinib group),the fourth group(PD-1 inhibitor combined with apatinib group)and given different treatments.After the treatment,the mice were sacrificed,the subcutaneous tumors of each group of mice were collected to calculate the tumor volume,the tumors and spleen tissues of each group of mice were collected,and the proportion of CD4+,CD8+ T cells and DC cells were analyzed by flow cytometry;the immune group was used Chemically observe the changes of CD4+ and CD8+ T cells in tumor tissues;separate fresh tumor tissues to detect mRNA differential expression.Survival analysis was performed on the remaining six mice in each group.Results:1.Apatinib and PD-1 inhibitors combination treatment demonstrated a dramatic improvement in tumor regression and animal survival in CT26 tumor models.2.PD-1 inhibitors combined with apatinib up-regulates the ratio of tumor microenvironment CD8+ T cells and DC cells.3.Tumor mRNA expression analysis confirmed: compared with apatinib,the combination therapy has angiogenesis-related genes()/Z3,g/t,Jaglt Dili),antigen presentation and adhesion molecule-related genes(Clecla,Cd209e),cytokine and chemokine related genes(Ccr6,IllO,Ccr5,1l16,1l18,Ccrl,Tnf,Cc1l7, Ccr7,Ccl22,Ifng,Ccl5,Cx3crl,Ccr2,Cxcll6), immune checkpoint molecular related genes(Cd80,Pvrt,Tnfrsf9,Cd276,Cd274,Cd200),TLR pathway related genes(Tlr3,Tlr7,Tlr9,Tlrll,Tlrl2)expression changes.GSEA analysis showed that the enrichment of autophagy and mitophagy pathways in apatinib group was up-regulated,while the enrichment of Notch pathway in apatinib combined with PD-1 inhibitor was down-regulated.Conclusion:1.Our data indicate that using apatinib in combination with anti-PD-1 therapy can lead to dramatic improvement in antitumor immune response over either agent used as monotherapy.2.The combination of PD-1 inhibitors and apatinib can increase the proportion of DC and CD8+ T cells in the tumor,enhance the ability of antigen presentation and the ability to kill tumor cells,and improve the host’s anti-tumor immune response.3.The combination of apatinib and PD-1 inhibitors may down-regulate the expression of angiogenesis pathway-related genes,up-regulate the expression of antigen presentation and adhesion molecule-related genes,cytokine and chemokine-related genes,and down-regulate immune checkpoint molecules.The expression of genes reshapes the tumor microenvironment,regulates tumor immunity,and then synergistically inhibits tumor growth.PartⅢ Case report of apatinib combined with PD-1 inhibitor in the posterior line treatment of metastatic colorectal cancerObjectives: By constructing a mouse subcutaneous tumor model,we preliminarily confirmed that apatinib and PD-1 inhibitors have a stronger anti-tumor effect.It is also necessary to further verify the efficacy and safty of the combination therapy from the clinical perspective.Research Method and Results: In the third part of this study,we collected post-line metastatic colorectal cancer patients who were hospitalized at Qianfoshan Hospital in Shandong Province from January 1,2020 to July 1,2020,of which 2 Case received apatinib combined with carrelizumab treatment.We evaluate the efficacy and safety.One of them was an elderly male who was confirmed by histology to have liver metastasis from rectal cancer.He had received XELOX,bevacizumab combined with FOLFIRI,regorafenib,bevacizumab combined with raltitrexed regimen in the prior treatment.Received carrelizumab combined with apatinib combined treatment,and evaluated the efficacy twice as SD,and the adverse reactions were evaluated as grade2 hypertension and grade 1 hand-foot syndrome.So far,the PFS is 10 months,and the safety is good.The other patient was a middle-aged woman who was diagnosed with colon cancer with abdominal metastasis.She had received mFOLFOX6,XELOX,bevacizumab+FOLFIRI,bevacizumab+SOX,and fruquintinib in the past,and received apatinib combined with carrelizumab after progression.The PFS for 11 months and is well tolerated.Follow-up will continue to be closely monitored.Conclusion:Both patients have obtained good efficacy and good safety.Due to the small number of cases,more clinical cases are urgently needed for apatinib combined with immune checkpoint inhibitor,in order to provide more clinical evidence.PartⅣ Clinical observation of apatinib-related hypothyroidism in patients with advanced malignanciesObjectives: Thyroid dysfunction has been previously reported during treatment with certain small-molecule multi-tyrosine kinase inhibitors,including sunitinib and sorafenib.Apatinib,which has a similar mechanism of action to these inhibitors,has reportedly induced hypothyroidism during treatment.Fully elucidating drug-associated adverse events could aid in patient monitoring and recommendations of suitable manage-ment strategies.The current 2-year observational study monitored patients with solid tumors who were prescribed apatinib.A total of 149 patients treated with apatinib from February 2015 to January 2016 were included.Their thyroid function and thyroid ultrastructure were evaluated for at least 24 months or until death.The primary objective of the current study was evaluating accepted thyroid replacement treatment.Secondary objective was ultrastructural changes in the thyroid gland.Research Method: We collected 149 patients who received apatinib from February2015 to January 2016 and 42 patients who received apatinib in the retrospective study of Part I.We observed and evaluated thyroid function and thyroid structure for at least24 months or until death.The primary study endpoint was to evaluate the number of patients receiving thyroid replacement therapy after apatinib treatment and the start time.The secondary study endpoint is the changes in the ultrasound structure of the thyroid.Results:1.A total of 64(39.27%)patients developed hypothyroidism,which varied from subclinical(21 cases;10.99%)to clinical(43 cases;22.51%).Thyroid nodules were noted in 19 cases(9.95%).Furthermore,5 cases(2.62%)had thyroid imaging reporting and data system scores of 4a/4b/4c and 14 cases(7,33%)had scores of 1，2 and 3.2.A total of 42 patients(21.99%)experienced 1-2 grade fatigue and 6 patients(3.14%)reported 3-4 grade fatigue.There was no reported association between disease control rate and hypothyroidism.Conclusion:Apatinib increased the risk of clinically relevant hypothyroidism and altered thyroid gland structure.