Roles of the tumor microenvironment in the resistance to programmed cell death protein 1 inhibitors in patients with gastric cancer

Despite the continuous developments and advancements in the treatment of gastric cancer(GC),which is one of the most prevalent types of cancer in China,the overall survival is still poor for most patients with advanced GC.In recent years,with the progress in tumor immunology research,attention has shifted toward immunotherapy as a therapeutic approach for GC.Programmed cell death protein 1(PD-1)inhibitors,as novel immunosuppressive medications,have been widely utilized in the treatment of GC.However,many patients are still resistant to PD-1 inhibitors and experience recurrence in the advanced stages of PD-1 immunotherapy.To reduce the occurrence of drug resistance and recurrence in GC patients receiving PD-1 immunotherapy,to maximize the clinical activity of immunosuppressive drugs,and to elicit a lasting immune response,it is essential to research the tumor microenvironment mechanisms leading to PD-1 inhibitor resistance in GC patients.This article reviews the progress in studying the factors influencing the resistance to PD-1 inhibitors in the GC tumor microenvironment,aiming to provide insights and a basis for reducing resistance to PD-1 inhibitors for GC patients in the future.

Molecular Docking Studies of Botanical Beverage Mix Berries (LIFEGREENTM) against Breast Cancer Cells from Targeted Protein 1QQG, 7B5Q & 7B5O & Uterine Fibroid from Targeted Protein 2AYR, 6T41 & 3GRF

Fibroids, also called leiomyomas or myomas, are communal tumors of the muscle or uterine wall that affect about 20% of females who are of reproductive age. They can look as if singly or in clusters, and they often cease to grow after menopause. Fibroids can be classified as intramural, sub serosal, pedunculated, or submucosal based on where they are positioned in the uterus. Although fibroids are benign, they can grow quickly and cause a range of symptoms, such as pelvic pressure, heavy menstrual flow, and infertility. As a result, fibroids are a main reason behind hysterectomy surgeries. The majority of cases of breast cancer are ductal and lobular cancers, making it the second utmost common cancer in women international. Gene mutations like those in BRCA1 or BRCA2 knowingly raise the risk of breast and other cancers, typically with an earlier cancer onset. Cancer risk is influenced by a complex interplay of genetic abnormalities, environmental factors, and lifestyle selections. Further research into these relations is domineering. Although they are common in uterine leiomyomas, especially multiple leiomyomas, MED12 mutations do not significantly correlate with tumor size. These mutations have also been noticed in smooth muscle tumors and leiomyosarcomas, two other types of uterine cancer. The identification of MED12 mutations as the sole genetic abnormality originates in leiomyomas raises the opportunity of a role in the genesis of cancer. 10% - 15% of women who are of reproductive age have endometriosis, which grants serious difficulties because of its chronic nature and range of clinical symptoms. Even after effective surgeries, issues reoccur often, adding to the enormous financial burden. The effects of MED12 mutations have been experiential in recent studies examining the molecular causes of endometriosis-associated infertility, which have shown anomalies in cellular connections and signaling cascades. Computational techniques were used in this study to investigate LifeGreenTM’s potential to prevent uterine fibroids and breast cancer. The efficacy of LifeGreenTM as a preventive measure or a treatment for common gynecological matters was examined and modeled. We investigated the mechanisms underlying LifeGreenTM’s benefits in the treatment of uterine fibroids and breast cancer using computational techniques. Our research contributes to our understanding of its potential therapeutic benefits for women’s health.

TMED2 Induces Cisplatin Resistance in Breast Cancer via Targeting the KEAP1-Nrf2 Pathway

Objective Cisplatin is the first-line treatment for breast cancer,but it faces challenges of drug resistance.This study investigated new molecular mechanisms underlying cisplatin resistance in breast cancer.Methods We analyzed sequencing data from the TCGA database to identify potential associations between transmembrane emp24 protein transport domain containing 2(TMED2)and breast cancer.Western blotting,real-time PCR,CCK-8,and TUNEL assays were used to measure the effects and molecular mechanism of TMED2 on cisplatin resistance in MCF-7 and MDA-MB-231 cell lines.Results TMED2 was overexpressed in breast cancer and associated with poor prognosis.TMED2 increased cisplatin resistance in breast cancer cells in vitro via promoting ubiquitination of Kelch-like ECH-associated protein 1(KEAP1),relieving inhibition of KEAP1 on nuclear factor erythroid 2-related factor 2(Nrf2),and increasing expression of downstream drug resistance related genes,such as heme oxygenase 1(HO-1)and NAD(P)H quinone oxidoreductase 1(NQO1).Conclusion We identified a new molecular mechanism by which TMED2 affects cisplatin resistance in breast cancer.Our results provide theoretical guidance for future clinical applications.

High expression of autophagy-related gene EIF4EBP1 could promote tamoxifen resistance and predict poor prognosis in breast cancer

BACKGROUND Breast cancer(BC) remains a public health problem. Tamoxifen(TAM) resistance has caused great difficulties for treatment of BC patients. Eukaryotic translation initiation factor 4E binding protein 1(EIF4EBP1) plays critical roles in the tumorigenesis and progression of BC. However, the expression and mechanism of EIF4EBP1 in determining the efficacy of TAM therapy in BC patients are still unclear.AIM To investigate the expression and functions of EIF4EBP1 in determining the efficacy of TAM therapy in BC patients.METHODS High-throughput sequencing data of breast tumors were downloaded from the Gene Expression Omnibus database. Differential gene expression analysis identified EIF4EBP1 to be significantly upregulated in cancer tissues. Its prognostic value was analyzed. The biological function and related pathways of EIF4EBP1 was analyzed. Subsequently, the expression of EIF4EBP1 was determined by real-time reverse transcription polymerase chain reaction and western blotting. Cell Counting Kit-8 assays, colony formation assay and wound healing assay were used to understand the phenotypes of function of EIF4EBP1.RESULTS EIF4EBP1 was upregulated in the TAM-resistant cells, and EIF4EBP1 was related to the prognosis of BC patients. Gene Set Enrichment Analysis showed that EIF4EBP1 might be involved in Hedgehog signaling pathways. Decreasing the expression of EIF4EBP1 could reverse TAM resistance, whereas overexpression of EIF4EBP1 promoted TAM resistance.CONCLUSION This study indicated that EIF4EBP1 was overexpressed in the BC and TAM-resistant cell line, which increased cell proliferation, invasion, migration and TAM resistance in BC cells.

Differential expression of breast cancer-resistance protein,lung resistance protein,and multidrug resistance protein 1 in retinas of streptozotocin-induced diabetic mice

AIM：To investigate the altering expression profiles of efflux transporters such as breast cancer-resistance protein（BCRP）,lung resistance protein（LRP）,and multidrug resistance protein 1（MDR1） at the inner blood-retinal barrier（BRB） during the development of early diabetic retinopathy（DR） and/or aging in mice.METHODS：Relative m RNA and protein expression profiles of these three efflux transporters in the retina during the development of early DR and/or aging in mice were examined.The differing expression profiles of Zonula occludens 1（ ZO-1） and vascular endothelial growth factor-A（ VEGFA） in the retina as well as the perfusion characterization of fluorescein isothiocyanate（FITC）-dextran and Evans blue were examined to evaluate the integrity of the inner BRB.RESULTS：There were significant alterations in these three efflux transporters＇ expression profiles in the m RNA and protein levels of the retina during the development of diabetes mellitus and/or aging.The development of early DR was confirmed by the expression profiles of ZO-1 and VEGFA in the retina as well as the compromised integrity of the inner BRB.CONCLUSION：The expression profiles of some efflux transporters such as BCRP,LRP,and MDR1 in mice retina during diabetic and/or aging conditions are tested,and the attenuated expression of BCRP,LRP,and MDR1 along with the breakdown of the inner BRB is found,which may be linked to the pathogenesis of early DR.

Breast Cancer Resistance Protein Expression and 5-Fluorouracil Resistance

Objective To filtrate breast cancer resistance protein （BCRP）-mediated resistant agents and to investigate clinical relationship between BCRP expression and drug resistance. Methods MTT assay was performed to filtrate BCRP-mediated resistant agents with BCRP expression cell model and to detect chemosensitivity of breast cancer tissue specimens to these agents. A high performance liquid chromatography （HPLC） assay was established, and was used to measure the relative dose of intracellular retention resistant agents. RT-PCR and immunohistochemistry （IHC） were employed to investigate the BCRP expression in breast cancer tissue specimens. Results MTT assay showed that the expression of BCRP increased with the increasing resistance of 5-fluorouracil （5-Fu） （P〈0.05, n=3） in the cell model, while HPLC assay indicated that the intracellular retention dose of 5-Fu was significantly correlated with the expression of BCRP （t=-0.897, P〈0.05, n=3）. A total of 140 breast cancer tissue specimens were collected. BCRP-positive expression was detected in forty-seven specimens by both RT-PCR and IHC. As shown by MTT assay subsequently, the resistance index （RI） of 47 BCRP-positive breast cancer tissue specimens to 5-Fu was 7-12 times as high as that of adjacent normal tissue samples. BCRP expression was related to 5-Fu resistance （R2=0.8124, P〈0.01）. Conclusion Resistance to 5-Fu can be mediated by BCRR Clinical chemotherapy for breast cancer patients can be optimized based on BCRP-positive expression.

Effect of Ursolic Acid on Breast Cancer Resistance Protein-mediated Transport of Rosuvastatin In Vivo and Vitro

Objective To evaluate whether ursolic acid can inhibit breast cancer resistance protein(BCRP)-mediated transport of rosuvastatin in vivo and in vitro. Methods Firstly, we explored the pharmacokinetics of 5-fluorouracil(5-FU, a substrate of BCRP) in rats in the presence or absence of ursolic acid. Secondly, we studied the pharmacokinetics of rosuvastatin in rats in the presence or absence of ursolic acid or Ko143(inhibitor of BCRP). Finially, the concentration-dependent transport of rosuvastatin and the inhibitory effects of ursolic acid and Ko143 were examined in Madin-Darby Canine Kidney(MDCK) Ⅱ-BCRP421CC(wild type) cells and MDCKⅡ-BCRP421AA(mutant type) cells. Results As a result, significant changes in pharmacokinetics parameters of 5-FU were observed in rats following pretreatment with ursolic acid. Both ursolic acid and Ko143 could significantly affect the pharmacokinetics of rosuvastatin. The rosuvastatin transport in the BCRP overexpressing system was increased in a concentration-dependent manner. However, there was no statistical difference in BCRP-mediated transport of rosuvastatin betweent the wild type cells and mutant cells. The same as Ko143, ursolic acid inhibited BCRP-mediated transport of rosuvastatin in vitro. Conclusion Ursolic acid appears to be a potent modulator of BCRP that affects the pharmacokinetic of rosuvastatin in vivo and inhibits the transport of rosuvastatin in vitro.

Effects of long non-coding RNA Opa-interacting protein 5 antisense RNA 1 on colon cancer cell resistance to oxaliplatin and its regulation of micro RNA-137

BACKGROUND The incidence of colon cancer(CC)is currently high,and is mainly treated with chemotherapy.Oxaliplatin(L-OHP)is a commonly used drug in chemotherapy;however,long-term use can induce drug resistance and seriously affect the prognosis of patients.Therefore,this study investigated the mechanism of Opainteracting protein 5 antisense RNA 1(OIP5-AS1)on L-OHP resistance by determining the expression of OIP5-AS1 and micro RNA-137(miR-137)in CC cells and the effects on L-OHP resistance,with the goal of identifying new targets for the treatment of CC.AIM To study the effects of long non-coding RNA OIP5-AS1 on L-OHP resistance in CC cell lines and its regulation of miR-137.METHODS A total of 114 CC patients admitted to China-Japan Union Hospital of Jilin University were enrolled,and the expression of miR-137 and OIP5-AS1 in tumor tissues and corresponding normal tumor-adjacent tissues was determined.The influence of OIP5-AS1 and miR-137 on the biological behavior of CC cells was evaluated.Resistance to L-OHP was induced in CC cells,and their activity was determined and evaluated using cell counting kit-8.Flow cytometry was used to analyze the apoptosis rate,Western blot to determine the levels of apoptosisrelated proteins,and dual luciferase reporter assay combined with RNA-binding protein immunoprecipitation to analyze the relationship between OIP5-AS1 and miR-137.RESULTS OIP5-AS1 was up-regulated in CC tissues and cells,while miR-137 was downregulated in CC tissues and cells.OIP5-AS1 was inversely correlated with miR-137(P<0.001).Silencing OIP5-AS1 expression significantly hindered the proliferation,invasion and migration abilities of CC cells and markedly increased the apoptosis rate.Up-regulation of miR-137 expression also suppressed these abilities in CC cells and increased the apoptosis rate.Moreover,silencing OIP5-AS1 and up-regulating miR-137 expression significantly intensified growth inhibition of drug-resistant CC cells and improved the sensitivity of CC cells to LOHP.OIP5-AS1 targetedly inhibited miR-137 expression,and silencing OIP5-AS1 reversed the resistance of CC cells to L-OHP by promoting the expression of miR-137.CONCLUSION Highly expressed in CC,OIP5-AS1 can affect the biological behavior of CC cells,and can also regulate the resistance of CC cells to L-OHP by mediating miR-137 expression.

Paclitaxel-induced stress granules increase LINE-1 mRNA stability to promote drug resistance in breast cancer cells

Abnormal expression of long interspersed element-1(LINE-1)has been implicated in drug resistance,while our previous study showed that chemotherapy drug paclitaxel(PTX)increased LINE-1 level with unknown mechanism.Bioinformatics analysis suggested the regulation of LINE-1 mRNA by drug-induced stress granules(SGs).This study aimed to explore whether and how SGs are involved in drug-induced LINE-1 increase and thereby promotes drug resistance of triple negative breast cancer(TNBC)cells.We demonstrated that SGs increased LINE-1 expression by recruiting and stabilizing LINE-1 mRNA under drug stress,thereby adapting TNBC cells to chemotherapy drugs.Moreover,LINE-1 inhibitor efavirenz(EFV)could inhibit drug-induced SG to destabilize LINE-1.Our study provides the first evidence of the regulation of LINE-1 by SGs that could be an important survival mechanism for cancer cells exposed to chemotherapy drugs.The findings provide a useful clue for developing new chemotherapeutic strategies against TNBCs.

Breast cancer resistance protein （Bcrp） and the testis--an unexpected turn of events

Breast cancer resistance protein （Bcrp） is an ATP-dependent efflux drug transporter. It has a diverse spectrum of hydrophilic and hydrophobic substrates ranging from anticancer, antiviral and antihypertensive drugs, to organic anions, antibiotics, phytoestrogens （e.g., genistein, daidzein, coumestrol）, xenoestrogens and steroids （e.g., dehydroepiandrosterone sulfate）. Bcrp is an integral membrane protein in cancer and normal cells within multiple organs （e.g., brain, placenta, intestine and testis） that maintains cellular homeostasis by extruding drugs and harmful substances from the inside of cells. In the brain, Bcrp is a major component of the blood- brain barrier located on endothelial cells near tight junctions （TJs）. However, Bcrp is absent at the Sertoli cell blood-testis barrier （BTB）; instead, it is localized almost exclusively to the endothelial TJ in microvessels in the interstitium and the peritubular myoid cells in the tunica propria. Recent studies have shown that Bcrp is also expressed stage specifically and spatiotemporally by Sertoli and germ cells in the seminiferous epithelium of rat testes, limited only to a testis-specific cell adhesion ultrastructure known as the apical ectoplasmic specialisation （ES） in stage VI-early VIII tubules. These findings suggest that Bcrp is equipped by late spermatids and Sertoli cells to protect late-stage spermatids completing spermiogenesis. Furthermore, Bcrp was found to be associated with F （filamentous）-actin and several actin regulatory proteins at the apical ES and might be involved in the organisation of actin filaments at the apical ES in stage VII-VIII tubules. These findings will be carefully evaluated in this brief review.

GP88 (Progranulin) Confers Fulvestrant (Faslodex, ICI 182,780) Resistance to Human Breast Cancer Cells

The 88 kDa glycoprotein known as GP88, Progranulin or PC cell derived growth factor is an autocrine growth factor with a unique cysteine rich motif that is over expressed in breast cancer whereas it is negative in normal mammary epithelial cells. It has been shown to play a major role in estrogen independence, tamoxifen resistance and tumorigenesis of breast cancer cells. In the present study, we investigated the effect of GP88 overexpression on the response of the human breast cancer MCF-7 cells to the pure estrogen receptor antagonist fulvestrant (ICI 182,780). While fulvestrant effectively inhibited cell proliferation of empty vector transfected cells, it had no inhibitory effect on the proliferation of GP88 overexpressing breast cancer cells. Mouse xenograft experiments in athymic ovariectomized nude mice showed that GP88 over expressing cells were fulvestrant resistant in vivo in contrast to low GP88 expressing cells. We show that the ability of fulvestrant to induce apoptosis determined by measuring cleavage of poly (ADP-ribose) polymerase was inhibited by GP88. Anti-apoptotic activity of GP88 was associated with sustained expression of bcl-2 and bcl-xL after fulvestrant treatment. In contrast, fulvestrant was still able to inhibit the ability of estrogen to stimulate ERE-luciferase reporter gene activity as well as vEGF expression in GP88 over expressing MCF-7 cells similarly to control MCF-7 cells. Collectively, our data suggest that GP88 prevents apoptosis induced by faslodex and contributes to antiestrogen resistance in human breast cancer.

Correlation of Kif2a and HPK1 expression in breast cancer with the oncogene and drug resistance gene expression

Objective: To investigate the correlation of Kif2a and HPK1 expression in breast cancer with the oncogene and drug resistance gene expression. Methods: A total of 91 patients with breast cancer and 85 patients with breast adenoma who accepted surgical treatment in our hospital between August 2016 and February 2018 were selected, and the breast cancer tissues and breast adenoma tissues were collected respectively as the research samples. Fluorescence quantitative PCR method was used to detect the expression of Kif2a and HPK1 genes as well as oncogenes and drug resistance genes in sample tissues, and Pearson test was used to evaluate the inner link of Kif2a and HPK1 gene expression in breast cancer tissue with oncogene and drug resistance gene expression. Results: Kif2a mRNA expression in breast cancer tissues was higher than that in breast adenoma tissues whereas HPK1 mRNA expression was lower than that in breast adenoma tissues;oncogenes DEK, iASPP-SV, Stat3, MDM2 and Fra-1 mRNA expression were higher than those in breast adenoma tissues;drug resistance genes ESR1, MDR1, P-gp, MRP1 and GST- mRNA expression were higher than those in breast adenoma tissues whereas BCRP mRNA expression was lower than that in breast adenoma tissues. Correlation analysis showed that the Kif2a and HPK1 gene expression in breast cancer tissues were directly correlated with the expression of oncogenes and drug resistance genes. Conclusion: Kif2a gene is abnormally highly expressed whereas HPK1 gene is abnormally lowly expressed in breast cancer tissues, and they are involved in the regulation of oncogene and drug resistance gene expression.

Serum Y-Box Binding Protein 1 (YBX-1) and Interleukin 6 (IL-6) Are Associated with Metastasis in Breast Cancer Patients

Objectives: The aim of this study was to assess the levels of Y-box binding protein 1 (YBX-1) and interleukin 6 (IL-6) in the sera of metastatic and non-metastatic breast cancer patients (BC), investigate their clinicopathological significance and to analyze their potential use as biomarkers of breast cancer metastasis. Methods: The study included ninety subjects sub-grouped equally into metastatic BC, non-metastatic BC and healthy volunteers. Serum YBX-1 and IL-6 were quantified using ELISA technique while CA 15-3 was quantified using IRMA kit. Clinical data were collected from patients’ records. Results: YBX-1 (p < 0.001), IL-6 (p < 0.001) and CA15-3 (p = 0.017, 0.001) were significantly elevated in metastatic and non-metastatic BC patients compared to healthy controls, however, only YBX-1 (p 0.001) showed a significant difference with cancer metastasis. Generally, YBX-1 and IL-6 were correlated with worse histological grade and late clinical stage in breast cancer patients and they were also associated with axillary lymph nodes involvement and positive vascular invasion in metastatic BC patients. Serum YBX-1 and IL-6 levels were positively correlated to each other (rs = 0.615, p < 0.001) and they showed high sensitivity and specificity compared to CA 15-3 (p < 0.001 and p = 0.004 for YBX-1 and IL-6 respectively) for predicting cancer metastasis. Conclusions: Serum YBX-1 and IL-6 are potential biomarkers of breast cancer patients with significant correlation with bad clinicopathological characteristics. Serum YBX-1 and IL-6 have superior sensitivity and specificity compared to CA15-3 and can serve as potential follow up and prognostic markers.

AR、SKP2、SOX10、PD-L1及TIL表达在三阴性乳腺癌中的意义

目的:探索雄激素受体(androgen receptor,AR)、S期激酶相关蛋白2(S-phase kinase-associated protein 2,SKP2)、性别决定区Y相关的HMG盒含因子10(sry-related HMG box-containing factor 10,SOX10)、程序性死亡配体1(programmed death-ligand 1,PD-L1)及肿瘤浸润性淋巴细胞(tumor infiltrating lymphocyte,TIL)在三阴性乳腺癌(triple negative breast cancer,TNBC)表达与临床病理特征和预后的关系。方法:根据苏木精-伊红染色(hematoxylineosin, HE)染色切片评判109例TNBC瘤巢内TIL的比例,采用Leica Bond-Max全自动免疫组化仪检测TNBC组织中AR、SKP2、SOX10、PD-L1的表达。分析以上各生物指标与临床病理特征间的关系,并采用kaplan-Meier、Log-rank进行生存分析。结果:95例患者获得随访,中位随访时间为48个月,中位无病生存时间(disease-free survival, DFS)为42个月,中位总生存时间(overall survival, OS)48个月。在TNBC中,AR阳性表达与淋巴结转移阴性(P=0.009)、肿瘤最大径<2 cm(P=0.008)相关,TIL高表达与低级别TNBC相关(P=0.007),SKP2阳性表达与神经/脉管侵犯阳性(P=0.011)、高级别TNBC相关(P=0.002),SOX10阳性表达与淋巴结转移阳性(P=0.022)、高级别TNBC(P=0.005)相关,PD-L1阳性表达与淋巴结转移阳性(P=0.020)、神经/脉管侵犯阳性(P=0.006)、高级别TNBC(P=0.042)相关。生存分析显示,SKP2、SOX10阳性表达与更差的DFS(P=0.007、P<0.001)和OS(P=0.013、P<0.001)相关,TIL高表达与更好的DFS(P=0.016)及OS(P=0.004)相关。在生物表志物的联合表达中,AR+/SKP2-、AR+/SOX10-与更好的DFS(P=0.004、P<0.001)及OS(P=0.007、P=0.001)相关,SOX10+/低TIL、PD-L1+/低TIL与更差的DFS(P<0.001、P=0.008)及OS(P=0.001、P=0.002)相关,AR-/低TIL者具有更差的OS(P=0.014)。SKP2(HR=4.143,95%CI为1.578~10.875)、SOX10(HR=7.578,95%CI为2.067~27.782)的阳性表达是影响TNBC患者DFS的独立预后因子,SKP2(HR=3.758,95%CI为1.400~10.084)、SOX10(HR=5.131,95%CI为1.316~20.000)及TIL(HR=0.375,95%CI为0.154~0.917)的阳性表达是TNBC患者OS的独立预后因子(P均<0.05)。结论:在TNBC中,AR阳性、TIL高表达与具有更好预后的临床病理特征相关,SKP2、SOX10和PD-L1与具侵袭性的临床病理特征相关。SKP2、SOX10及TIL表达与TNBC预后相关,提示这些生物指标可能成为TNBC新的预后因子,同时它们也有可能成为潜在的治疗靶点。

1STC1在乳腺癌中的作用研究进展

乳腺癌是目前全球女性恶性肿瘤发病率最高的一种疾病,是一种高度异质性疾病,可根据其不同分子指标表达分为4种分子亚型,具有不同的组织学、临床行为和预后特征。锡钙素-1(STC1)是一种分泌型糖蛋白激素,目前已被证实可调节钙和磷酸盐的稳态,STC1在哺乳动物的各种组织中表达,并参与多种生理和病理生理过程。同时越来越多的证据表明STC1在多种不同类型的肿瘤中起致癌作用。为了进一步理清STC1与乳腺癌之间的关系,通过大量文献阅读及总结,总结分析了STC1与乳腺癌现有的研究数据,探索STC1与乳腺癌发生、发展、治疗预后等一系列的相关证据,并讨论其在乳腺癌各亚型中的潜在作用,为今后的研究找寻方向。

Targeted inhibition of Notch1 gene enhances the killing effects of Paclitaxel on triple-negative breast cancer cells

Objective:To study the influence of targeted inhibition of Notch1 gene on the killing effects of Paclitaxel on triple-negative breast cancer cells.Methods:The triple-negative [estrogen receptor(ER)/progesterone receptor(PR)/human epidermal growth factor receptor 2(Her2)] breast cancer cell line MDA-MB-231 and ER/PR/HER-2-positive breast cancer cell line MCF-7 were cultured,transfected with Notch1-si RNA-overexpression plasmid and blank plasmid,and treated with different concentrations of paclitaxel,and then the cell proliferation activity and apoptosis rate as well as the m RNA expression of Caspase-3,Caspase-9 and Bcl-2 were determined.Results:Paclitaxel could decrease the MDA-MB-231 and MCF-7 cell proliferation activity as well as Bcl-2 mRNA expression,and increase MDA-MB-231 and MCF-7 cell apoptosis rate as well as Caspase-3 and Caspase-9 mRNA expression in dosedependent manners;with the same dose of paclitaxel treatment,the inhibitory effects on MDAMB-231 cell proliferation activity and Bcl-2 m RNA expression as well as the promoting effects on MDA-MB-231 cell apoptosis and mR NA expression of Caspase-3 and Caspase-9 were weaker than those on MCF-7 cell;after 0.5 μM paclitaxel combined with Notch1-siRNA treatment,MDA-MB-231 cell proliferation activity and Bcl-2 mRNA expression were significantly lower than those after 0.5 μM paclitaxel combined with control plasmid treatment while cell apoptosis rate and mR NA expression of Caspase-3 and Caspase-9 were higher than those after 0.5 μM paclitaxel combined with control plasmid treatment.Conclusions:Targeted inhibition of Notch1 gene may enhance the killing effects of paclitaxel on triple-negative breast cancer cells by up-regulating the expression of Caspase-3 and Caspase-9 and inhibiting the expression of Bcl-2.

食管鳞癌组织中BARD1的表达及与淋巴结转移的关系

目的分析食管鳞癌(ESCC)中乳腺癌易感基因1相关环指结构域蛋白1(BARD1)的表达及与淋巴结转移的关系。方法纳入2018年1月至2021年12月本院收治且经组织病理学检查确诊的114例ESCC患者,免疫组化EnVision两步法检测癌组织与癌旁组织中BARD1的表达情况并比较BARD1阳性率,分析BARD1表达与ESCC临床病理特征的关系,建立Logistic回归模型分析BARD1表达对ESCC淋巴结转移的影响,绘制受试者工作特征(ROC)曲线分析BARD1表达对ES⁃CC淋巴结转移的预测价值。结果114例ESCC组织中BARD1阳性78例、阴性36例,对应癌旁组织中BARD1阳性23例、阴性91例,ESCC组织的BARD1阳性率为68.42%,高于癌旁组织的20.18%(χ^(2)=53.769,P<0.001)。BARD1表达与ESCC患者的T分期、分化程度和淋巴结转移有关(P<0.05),其中淋巴结转移组织的BARD1阳性率为80.36%(45/56),高于未转移组织的56.90%(33/58)。经Phi系数分析发现,BARD1表达与ESCC淋巴结转移呈正相关(r=0.276,P=0.003)。BARD1阳性是ESCC患者淋巴结转移的独立危险因素(OR=3.099,95%CI:1.339~7.175,P=0.008),且BARD1表达对ESCC患者淋巴结转移具有一定预测价值(AUC=0.636,P=0.020)。结论BARD1在ESCC中高表达,其与ESCC淋巴结转移密切相关,BARD1阳性会增加淋巴结转移风险,在评估ESCC淋巴结转移上有一定价值。

长链非编码RNA淋巴细胞白血病缺失基因2/微小RNA-30c-5p/丝裂原活化蛋白激酶1轴在食管癌细胞紫杉醇耐药中的作用机制实验研究

目的:探讨长链非编码RNA(lncRNA)淋巴细胞白血病缺失基因2(DLEU2)/微小RNA(miR)-30c-5p/丝裂原活化蛋白激酶1(MAPK1)轴在食管癌细胞紫杉醇(PTX)耐药中的作用机制。方法:构建EC109/PTX细胞,检测食管癌组织和癌旁正常组织,以及食管癌细胞EC109和KYSE150细胞、食管上皮细胞SHEE和EC109/PTX细胞DLEU2、miR-30c-5p、MAPK1 mRNA表达。取生长良好的EC109/PTX细胞,分为沉默(siRNA)DLEU2组、阴性对照(siRNA NC)组、siRNA DLEU2+miR-30c-5p抑制剂(inhibitor)组、siRNA DLEU2+inhibitor NC组、siRNA DLEU2+过表达(pcDNA)MAPK1组、siRNA DLEU2+pcDNA NC组和空白组。验证miR-30c-5p与DLEU2、MAPK1的靶向关系。检测各组EC109/PTX细胞DLEU2、miR-30c-5p、MAPK1 mRNA表达水平。检测EC109/PTX细胞活性、凋亡情况及对PTX的耐药性。检测谷胱甘肽S-转移酶π(GST-π)、MAPK1、P-蛋白(P-gp)蛋白表达水平。结果:食管癌组织和EC109/PTX细胞miR-30c-5p表达降低,DLEU2、MAPK1 mRNA表达增加(均P<0.05)。DLEU2靶向调控miR-30c-5p,miR-30c-5p靶向调控MAPK1。siRNA DLEU2组miR-30c-5p表达和细胞凋亡率较siRNA NC组、空白组增加,DLEU2和MAPK1 mRNA表达、细胞增殖率、药物半数抑制浓度(IC 50)以及GST-π、MAPK1、P-gp蛋白表达降低(均P<0.05)。siRNA DLEU2+miR-30c-5p inhibitor组miR-30c-5p表达和细胞凋亡率较siRNA DLEU2+inhibitor NC组降低,MAPK1 mRNA表达、细胞增殖率、IC 50以及GST-π、MAPK1、P-gp蛋白表达增加(均P<0.05)。siRNA DLEU2+pcDNA MAPK1组细胞凋亡率较siRNA DLEU2+pcDNA NC组降低,MAPK1 mRNA表达、细胞增殖率、IC 50以及GST-π、MAPK1、P-gp蛋白表达增加(均P<0.05)。结论:干扰lncRNA DLEU2可减弱EC109/PTX细胞对PTX的耐药性,可能与调控miR-30c-5p/MAPK1轴有关。

土贝母苷甲逆转人三阴性乳腺癌MDA-MB-231细胞顺铂耐药及其机制探讨

目的:探究土贝母苷甲(tubeimoside-1,TBMS-1)对三阴性乳腺癌顺铂(cisplatin,DDP)耐药的逆转作用及可能的机制。方法:构建对DDP耐药的三阴性乳腺癌细胞株MDA-MB-231/DDP,并利用CCK-8法检测细胞的耐药指数(resistance index,RI)变化,评估TBMS-1的抑制效果以及对DDP耐药株的逆转效果。实验设置了MDA-MB-231/DDP细胞的对照组、DDP组、TBMS-1组和TBMS-1+DDP组。通过平板克隆实验检测各组细胞的增殖能力;DCFH-DA活性氧荧光探针检测各组细胞ROS水平;TUNEL染色和流式细胞术检测各组细胞的凋亡情况;采用Western blot法检测β-catenin总蛋白与核蛋白,以及Wnt3a、Bcl-2、Bax的表达水平。取MDA-MB-231/DDP细胞,设置对照组、DDP组、Adavivint组(Wnt/β-catenin通路抑制剂)和DDP+Adavivint组,采用Western blot法检测β-catenin总蛋白与核蛋白,以及Wnt3a、Bcl-2、Bax的表达水平。结果:TBMS-1呈浓度依赖性抑制MDA-MB-231/DDP细胞的增殖,降低DDP对MDA-MB-231/DDP细胞的IC_(50)值,逆转其耐药性(P<0.05)。TBMS-1可以增强DDP诱导MDA-MB-231/DDP细胞凋亡的效果和细胞ROS积累,并抑制Wnt/β-catenin的异常激活。结论:TBMS-1对MDA-MB-231/DDP细胞的DDP耐药具有良好的逆转作用,可能通过抑制Wnt/β-catenin信号通路的激活来实现。

lncRNA AC092718.4对HER2阳性乳腺癌耐药性的影响及其可能机制

目的探讨长链非编码RNA(lncRNA)AC092718.4对人类表皮生长因子受体2(HER2)阳性乳腺癌耐药性的影响及其可能机制。方法(1)获取曲妥珠单抗非耐药及耐药HER2阳性乳腺癌患者的乳腺癌组织(设为非耐药组、耐药组),检测其lncRNA AC092718.4、miR-135a-5p、S100钙结合蛋白P(S100P)mRNA和蛋白的表达水平。(2)以对曲妥珠单抗不敏感的HER2阳性乳腺癌细胞系MDA-MB-361细胞作为原发耐药细胞模型,以乳腺癌细胞株BT-474细胞为亲本,构建对曲妥珠单抗继发耐药的细胞模型(BT-474/TRA细胞)。检测3种细胞中lncRNA AC092718.4、miR-135a-5p、S100P mRNA和蛋白的表达水平。经同一浓度曲妥珠单抗干预48 h后,检测3种细胞的活力。(3)取MDA-MB-361细胞分为sh-AC092718.4组、sh-NC组、对照组进行实验,其中sh-AC092718.4组细胞和sh-NC组分别转染sh-AC092718.4和sh-NC,对照组细胞未经任何处理。经同一浓度曲妥珠单抗干预48 h后,检测3组细胞的活力。(4)采用starBase和TargetScan分别预测lncRNA AC092718.4和miR-135a-5p的潜在靶标。通过双荧光素酶报告基因实验验证lncRNA AC092718.4与miR-135a-5p之间、miR-135a-5p与S100P之间的靶向结合情况。结果(1)与非耐药组相比,耐药组lncRNA AC092718.4、S100P mRNA、S100P蛋白表达水平升高,miR-135a-5p表达水平降低(P<0.05)。(2)与BT-474细胞相比,BT-474/TRA细胞及MDA-MB-361细胞的lncRNA AC092718.4、S100P mRNA、S100P蛋白表达水平升高,miR-135a-5p表达水平降低,曲妥珠单抗干预48 h后的细胞活力更大(P<0.05)。(3)与对照组和sh-NC组比较,sh-AC092718.4组MDA-MB-361细胞活力降低(P<0.05)。(4)starBase预测结果显示,lncRNA AC092718.4与miR-135a-5p有靶向结合位点;TargetScan预测结果显示,miR-135a-5p与S100P有靶向结合位点。荧光素酶报告基因实验结果提示,lncRNA AC092718.4可与miR-135a-5p直接结合,S100P是miR-135a-5p的靶基因。结论LncRNA AC092718.4促进乳腺癌细胞对曲妥珠单抗产生耐药性,下调lncRNA AC092718.4表达可减轻MDA-MB-361细胞对曲妥珠单抗的耐药性,其机制可能涉及lncRNA AC092718.4作为竞争性内源RNA竞争性结合miR-135a-5p,从而上调S100P的表达。