C-reactive protein to albumin ratio predict responses to programmed cell death-1 inhibitors in hepatocellular carcinoma patients

BACKGROUND Over the years,programmed cell death-1(PD-1)inhibitors have been routinely used for hepatocellular carcinoma(HCC)treatment and yielded improved survival outcomes.Nonetheless,significant heterogeneity surrounds the outcomes of most studies.Therefore,it is critical to search for biomarkers that predict the efficacy of PD-1 inhibitors in patients with HCC.AIM To investigate the role of the C-reactive protein to albumin ratio(CAR)in evaluating the efficacy of PD-1 inhibitors for HCC.METHODS The clinical data of 160 patients with HCC treated with PD-1 inhibitors from January 2018 to November 2022 at the First Affiliated Hospital of Guangxi Medical University were retrospectively analyzed.RESULTS The optimal cut-off value for CAR based on progression-free survival(PFS)was determined to be 1.20 using x-tile software.Cox proportional risk model was used to determine the factors affecting prognosis.Eastern Cooperative Oncology Group performance status[hazard ratio(HR)=1.754,95%confidence interval(95%CI)=1.045-2.944,P=0.033],CAR(HR=2.118,95%CI=1.057-4.243,P=0.034)and tumor number(HR=2.932,95%CI=1.246-6.897,P=0.014)were independent prognostic factors for overall survival.CAR(HR=2.730,95%CI=1.502-4.961,P=0.001),tumor number(HR=1.584,95%CI=1.003-2.500,P=0.048)and neutrophil to lymphocyte ratio(HR=1.120,95%CI=1.022-1.228,P=0.015)were independent prognostic factors for PFS.Two nomograms were constructed based on independent prognostic factors.The C-index index and calibration plots confirmed that the nomogram is a reliable risk prediction tool.The ROC curve and decision curve analysis confirmed that the nomogram has a good predictive effect as well as a net clinical benefit.CONCLUSION Overall,we reveal that the CAR is a potential predictor of short-and long-term prognosis in patients with HCC treated with PD-1 inhibitors.If further verified,CAR-based nomogram may increase the number of markers that predict individualized prognosis.

Clinical significance of upregulated Rho GTPase activating protein 12 causing resistance to tyrosine kinase inhibitors in hepatocellular carcinoma

BACKGROUND Hepatocellular carcinoma(HCC)is a major health challenge with high incidence and poor survival rates in China.Systemic therapies,particularly tyrosine kinase inhibitors(TKIs),are the first-line treatment for advanced HCC,but resistance is common.The Rho GTPase family member Rho GTPase activating protein 12(ARHGAP12),which regulates cell adhesion and invasion,is a potential therapeutic target for overcoming TKI resistance in HCC.However,no studies on the expression of ARHGAP12 in HCC and its role in resistance to TKIs have been reported.AIM To unveil the expression of ARHGAP12 in HCC,its role in TKI resistance and its potential associated pathways.METHODS This study used single-cell RNA sequencing(scRNA-seq)to evaluate ARHGAP12 mRNA levels and explored its mechanisms through enrichment analysis.CellChat was used to investigate focal adhesion(FA)pathway regulation.We integrated bulk RNA data(RNA-seq and microarray),immunohistochemistry and proteomics to analyze ARHGAP12 mRNA and protein levels,correlating with clinical outcomes.We assessed ARHGAP12 expression in TKI-resistant HCC,integrated conventional HCC to explore its mechanism,identified intersecting FA pathway genes with scRNA-seq data and evaluated its response to TKI and immunotherapy.RESULTS ARHGAP12 mRNA was found to be highly expressed in malignant hepatocytes and to regulate FA.In malignant hepatocytes in high-score FA groups,MDK-[integrin alpha 6(ITGA6)+integrinβ-1(ITGB1)]showed specificity in ligand-receptor interactions.ARHGAP12 mRNA and protein were upregulated in bulk RNA,immunohistochemistry and proteomics,and higher expression was associated with a worse prognosis.ARHGAP12 was also found to be a TKI resistance gene that regulated the FA pathway.ITGB1 was identified as a crossover gene in the FA pathway in both scRNA-seq and bulk RNA.High expression of ARHGAP12 was associated with adverse reactions to sorafenib,cabozantinib and regorafenib,but not to immunotherapy.CONCLUSION ARHGAP12 expression is elevated in HCC and TKI-resistant HCC,and its regulatory role in FA may underlie the TKI-resistant phenotype.

Use of programmed cell death protein ligand 1 assay to predict the outcomes of non-small cell lung cancer patients treated with immune checkpoint inhibitors

The recent discovery of immune checkpoints inhibitors, especially anti-programmed cell death protein 1(PD-1)and anti-programmed cell death protein ligand 1(PD-L1) monoclonal antibodies, has opened new scenarios in the management of non-small cell lung cancer(NSCLC) and this new class of drugs has achieved a rapid development in the treatment of this disease. However, considering the costs of these drugs and the fact that only a subset of patients experience long-term disease control, the identification of predictive biomarkers for the selection of candidates suitable for treatment has become a priority. The research focused mainly on the expression of the PD-L1 receptor on both tumor cells and/or immune infiltrates determined by immunohistochemistry(IHC). However, different checkpoint inhibitors were tested, different IHC assays were used, different targets were considered(tumor cells, immune infiltrates or both) and different expression thresholds were employed in clinical trials. In some trials the assay was used prospectively to select the patients, while in other trials it was evaluated retrospectively. Some confusion emerges, which makes it difficult to easily compare the literature data and to translate them in practice management. This mini-review shows the possibilities and pitfalls of the PD-L1 expression to predict the activity and efficacy of anti PD1/PD-L1 monoclonal antibodies in the treatment of NSCLC.

Protein kinase small molecule inhibitors for rheumatoid arthritis: Medicinal chemistry/clinical perspectives

Medicinal chemistry strategies have contributed to the development, experimental study of and clinical trials assessment of the first type of protein kinase small molecule inhibitor to target the Janus kinase/Signal Transducers and Activators of Transcription(JAK/STAT) signaling pathway. The orally administered small molecule inhibitor, tofacitinib, is the first drug to target the JAK/STAT pathway for entry into the armamentarium of the medical therapy of rheumatoid arthritis. The introduction of tofacitinib into general rheumatologic practice coupled with increasing understanding that additional cellular signal transduction pathways including the mitogen-activated protein kinase and phosphatidylinositide-3-kinase/Akt/mammalian target of rapa-mycin pathways as well as spleen tyrosine kinase also contribute to immune-mediated inflammatory in rheumatoid arthritis makes it likely that further development of orally administered protein kinase small molecule inhibitors for rheumatoid arthritis will occur in the near future.

Relationship between inter-α-trypsin inhibitor heavy chain 4 and ovarian cancer

Objective:The inter-α-trypsin inhibitor heavy chain 4(ITIH4)protein is involved in the development of tumors.However,the relationship between ITIH4 and ovarian cancer(OC)has not been extensively examined.This study aimed to explore the effect of ITIH4 on OC and to identify its underlying mechanism.Methods:Expressions of ITIH4 in OC tissues and cells were determined using quantitative reverse transcription polymerase chain reaction(RT-qPCR)and western blots.The function of ITIH4 in the OC cell line HO8910 pm was tested via ITIH4 knockdown.The cell growth rate was measured using MTT and colony formation assays.Flow cytometry was performed to evaluate cell cycle progression.Cell migration and invasion abilities were observed using the transwell migration assay.Results:ITIH4 was downregulated in OC tissues and cells.ITIH4 knockdown promoted cell growth and cell cycle progression.Consistent with these results,inhibition of ITIH4 in OC cells significantly increased cell migration and invasion abilities.Cox regression analysis suggests that ITIH4 expression alone is not a good predictor of the prognosis of malignant ovarian tumors in patients.Conclusions:ITIH4 inhibits the progression of OC,suggesting that ITIH4 may be a useful biomarker for OC.This study may provide a potential novel target for the treatment of OC.

Interleukin-1 beta up-regulates tissue inhibitor of matrix metalloproteinase-1 mRNA and phosphorylation of c-jun N-terminal kinase and p38 in hepatic stellate cells

AIM： To study the relationship between interleukin-lbeta （IL-1β） up-regulating tissue inhibitor of matrix metalloproteinase-1 （TIMMP-1） mRNA expression and phosphorylation of both c-jun N-terminal kinase （INK） and p38 in rat heffatic stellate cells （HSC）. METHODS： RT-PCR was performed to measure the expression of TIMMP-1 mRNA in rat HSC. Western blot was performed to measure IL-1β-induced JNK and p38 activities in rat HSC. RESULTS： TIMMP-1 mRNA expression （1.191± 0.079） was much higher after treatment with IL-1β （10 ng/mL） for 24 h than in control group （0.545±0.091） （P〈0.01）. IL-1β activated INK and p38 in a time-dependent manner. After stimulation with IL-1β for 0, 5, 15, 30, 60 and 120 min, the INK activity was 0.982±0.299, 1.501±0.720, 2.133±0.882, 3.360±0.452, 2.181±0.789, and 1.385 ± 0.368, respectively. There was a significant difference in JNK activity at 15 min （P〈 0.01）, 30 min （P〈 0.01） and 60 min （P〈0.01） in comparison to that at 0 min. The p38 activity was 1.061±0.310, 2.050±0.863, 2.380±0.573, 2.973±0.953, 2.421±0.793, and 1.755 ± 0.433 at the 6 time points （0, 5, 15, 30, 60 and 120 min） respectively. There was a significant difference in p38 activity at 5 min （P〈0.05）, 15 min （P〈0.01）, 30 min （P〈0.01） and 60 min （P〈0.01） compared to that at 0 min. TIMMP-1 mRNA expression trended to decrease in 3 groups pretreated with different concentrations of SP600125 （10 μmol/L, 1.022±0.113; 20 μmol/L, 0.869±0.070; 40 μmol/L, 0.666±0.123）. Their decreases were all significant （P〈0.05, P〈0.01, P〈0.01） in comparison to control group （without SP600125 treatment, 1.163±0.107）. In the other 3 groups pretreated with different concentrations of SB203580 （10 μmol/L, 1.507±0.099; 20 μmol/L, 1.698±0.107; 40 μmol/L, 1.857±0.054）, the expression of TIMMP-1 mRNA increased. Their levels were higher than those in the control group （without SB203580 treatment, 1.027 ± 0.061） with a significant statistical significance （P〈 0.01）. CONCLUSION： IL-1β has a direct action on hepatic fibrosis by up-regulating TIMMP-1 mRNA expression in ratessionin in rate HSC.JNK and p38 mitogen-activated protein kinases （MAPKs） are involved in IL-1β-induced TIMMP-1 gene expression, and play a distinct role in this process, indicating that p38 and .INK pathways cooperatively mediate TIMP-1 mRNA expression in rat HSC.

Plasma matrix metalloproteinase-1 and tissue inhibitor of metalloproteinase-1 as biomarkers of ulcerative colitis activity

AIM:Overexpression of mucosal metalloproteinases(MMP) has been demonstrated recently in inflammatory bowel disease.Their activity can be counterbalanced by the tissue inhibitor of metalloproteinases(TIMP).The aim of this study was to evaluate the effect of ulcerative colitis(UC)on MMP- 1 and TIMP-1 plasma concentrations,as two possible biomarkers of the disease activity. METHODS:MMP-1 and TIMP-1 plasma concentrations were measured with an enzyme immunoassay in 16 patients with endoscopically confirmed active UC. RESULTS:Plasma concentrations of both MMP-1(13.7±0.2 ng/ml)and TIMP-1(799±140 ng/ml)were significantly elevated in UC patients in comparison to healthy controls (11.9±0.9 ng/ml and 220±7 ng/ml respectively).There was no correlation between TIMP-1 and MMP-1 concentrations (r=0.02).TIMP-1 levels revealed significant positive correlations with scored endoscopic degree of mucosal injury, disease activity index and clinical activity index values as well as C-reactive protein concentration.There was no correlation between MMP-1 and laboratory,clinical or endoscopic indices of the disease activity.CONCLUSION: These results confirm the role of both MMP- 1 and TIMP-1 in the pathogenesis of ulcerative colitis. However only TIMP-1 can be useful as a biomarker of the disease activity, demonstrating association with clinical and endoscopic pictures.

Discovery of a small-molecule bromodomain-containing protein 4 inhibitor that induces AMP-activated protein kinase-modulated autophagy-associated cell death in breast cancer

OBJECTIVE To discover a small-molecule bromodomain-containing protein 4(BRD4)inhibitor that induces AMP-activated protein kinase-modulated autophagy-associated cell death in breast cancer and exploreits potential mechanisms.METHODS BRD4 interactors were analyzed by PPI network prediction and The Cancer Genome Atlas(TCGA)analysis.The interaction between BRD4 and AMPK was confirmed by co-immunoprecipitation assay.Novel BRD4 inhibitors were designed and synthesized based upon pharmacophore analysis of BRD4(1),then screened by antiproliferative activity and Alpha Screen of BRD4(1).The selectivity of the best candidate compound 8f was validated by co-crystallization,FRET assay and co-immuno precipitation assay.The mechanisms of 8f were investigated by fluorescence microscopy,electron microscopy,Western blotting,immunocytochemistry,si RNA and GFP-m RFP-LC3 plasmid transfections,as well as immunohistochemistry and immunofluorescence.Potential mechanisms were discovered by i TRAQ-based proteomics analysis and the therapeutic effect of 8f was assessed by xenograft breast cancer mouse and zebrafish models.RESULTS We identified that BRD4 interacted with AMPK,which was remarkably downregulated in breast cancer.We next designed and synthesized 49 candidate compounds,and eventually discovered a selective small-molecule inhibitor of BRD4(8f).Subsequently,8f was discovered to induce autophagyassociated cell death(ACD)by BRD4-AMPK interaction,and thus activating AMPK-m TOR-ULK1-modulated autophagic pathway in breast cancer cells.Interestingly,the i TRAQ-based proteomics analyses revealed that 8f induced ACD pathways,involved in HMGB1,VDAC1/2 and e EF2.Moreover,8f displayed a therapeutic potential on both xenograft breast cancer mouse and zebrafish models.CONCLUSION We discovered a novel small-molecule inhibitor of BRD4 that induces BRD4-AMPK-modulated ACD in breast cancer,which may provide a candidate drug for future cancer therapy.

Human antigen R mediated post-transcriptional regulation of inhibitors of apoptosis proteins in pancreatic cancer

AIM To determine the association of human antigen R(HuR) and inhibitors of apoptosis proteins(IAP1, IAP2) and prognosis in pancreatic cancer.METHODS Protein and mRNA expression levels of IAP1, IAP2 and HuR in pancreatic ductal adenocarcinoma(PDAC) were compared with normal pancreatic tissue. The correlations among IAP1/IAP2 and HuR as well as their respective correlations with clinicopathological parameters were analyzed. The Kaplan-Meier method and log-rank tests were used for survival analysis. Immunoprecipitation assay was performed to demonstrate HuR binding to IAP1, IAP2 mRNA. PANC1 cells were transfected with either anti-HuR siRNA or control siRNA for 72 h and quantitative reverse transcription polymerase chain reaction(RT-PCR), western blot analysis was carried out.RESULTS RT-PCR analysis revealed that HuR, IAP1, IAP2 mRNA expression were accordingly 3.3-fold, 5.5-fold and 8.4 higher in the PDAC when compared to normal pancreas(P < 0.05). Expression of IAP1 was positively strongly correlated with HuR expression(P < 0.05, r = 0.783). Western blot analysis confirmed RTPCR results. High IAP1 expression, tumor resection status, T stage, lymph-node metastases, tumor differentiation grade, perineural and lymphatic invasion were identified as significant factors for shorter survival in PDAC patients(P < 0.05).Immunohistological analysis showed that HuR was mainly expressed in the ductal cancer cell's nucleus and less so in cytoplasm. RNA immunoprecipitation analysis confirmed IAP1 and IAP2 post-transcriptional regulation by HuR protein. Following siHuR transfection, IAP1 mRNA and protein levels were decreased, however IAP2 expression levels were increased.CONCLUSION HuR mediated overexpression of IAP1 significantly correlates with poor outcomes and early progression of pancreatic cancer. Further studies are needed to assess the underlying mechanisms.

SMAC exhibits anti-tumor effects in ECA109 cells by regulating expression of inhibitor of apoptosis protein family

BACKGROUND The poor prognosis and rising incidence of esophageal cancer highlight the need for improved therapeutics that are essential prior to treatment.LCL161 is an SMAC(second mitochondrial activator of caspases)mimic and inhibitor of apoptosis protein(IAP)antagonist which exhibits anti-tumor effects and improves the chemical sensitivity of many cancers.AIM To ascertain the effects and mechanisms of the SMAC analog LCL161 on esophageal cancer cells.METHODS MTT assay and TUNEL assay were used to detect cell proliferation and apoptosis,respectively.Western blot analysis was used to study the molecular mechanisms of LCL161-induced death of ECA109 cells.RESULTS LCL161 decreased ECA109 cell proliferation in dose-and time-dependent manner and induced apoptosis of ECA109 cells in a dose-dependent manner.Also,LCL161 induced a significant decrease in the expression of the XIAP and significant increase in the expression of Caspase-3.In addition,Bax increased significantly with increasing concentrations of LCL161,and the relative expression of Bax was significantly different between groups.CONCLUSION These findings support the hypothesis that LCL161 can inhibit proliferation and induce apoptosis in esophageal cancer cells by regulating the expression of IAP family members,suggesting that it has potential to be an effective treatment for esophageal squamous cell carcinoma.

Structural Insight into the Design on Oleanolic Acid Derivatives as Potent Protein Tyrosine Phosphatase 1B Inhibitors

Oleanolic acid derivatives act as newer protein tyrosine phosphatase 1B （PTP-1B） inhibitors for type 2 diabetes mellitus （T2DM）. In order to understand the structural requirement of PTP-1B inhibitors, 52 oleanolic acid derivatives were divided into a training set （34 compounds） and a test set （18 compounds）. The highly reliable and predictive 3D-QSAR models were constructed by CoMFA, CoMSIA and topomer CoMFA methods, respectively. The results showed that the cross validated coefficient （q2） and non-cross-validated coefficient （R2） were 0.554 and 0.999 in the CoMFA model, 0.675 and 0.971 in the CoMSIA model, and 0.628 and 0.939 in the topomer CoMFA model, which suggests that three models are robust and have good exterior predictive capabilities. Furthermore, ten novel inhibitors with much higher inhibitory potency were designed. Our design strategy was that （i） the electronegative substituents （Cl, -CH2OH, OH and -CH2Cl） were introduced into the double bond of ring C, （ii） the hydrogen bond acceptor groups （C≡N and N atom）, electronegative groups （C≡N, N atom, -COOH and -COOCH3） and bulky substituents （C6H5N） were connected to the C-3 position, which would result in generating potent and selective PTP-1B inhibitors. We expect that the results in this paper have the potential to facilitate the process of design and to develop new potent PTP-1B inhibitors.

Expression of second mitochondria-derived activator of caspases, X-linked inhibitor of apoptosis protein, and caspase-3 in pituitary adenomas

Studies concerning correlations between pituitary adenomas and cell apoptosis have mainly focused on upstream apoptosis signaling, but seldom on downstream mediators. In the present study, second mitochondria-derived activator of caspases （Smac）, X-linked inhibitor of apoptosis protein （XIAP）, and caspase-3 protein were qualitatively analyzed using immunohistochemistry, and quantified by western blot. Smac, XIAP, and caspase-3 mRNA expressions were detected by reverse transcription-PCR. Results showed that XIAP protein and mRNA expressions were greater in the invasive pituitary adenoma group compared with the noninvasive pituitary adenoma group. However, Smac and caspase-3 protein and mRNA expressions were lower in the invasive pituitary adenoma group compared with the noninvasive pituitary adenoma group. In the invasive pituitary adenomas, Smac expression was positively correlated with caspase-3 protein and mRNA expression （Protein： r = 0.55, P 0.01; mRNA： r = 0.50, P 0.01）. Smac and caspase-3 expressions were negatively correlated with XIAP protein and mRNA expression （Protein： r = -0.56, -0.64, P 0.01; mRNA： r = -0.69, -0.67, P 0.01）. However, no significant differences in correlation among Smac, XIAP, and caspase-3 were detectable in noninvasive pituitary adenomas. These data indicated that high expression of XIAP and low expression of Smac and caspase-3 suppressed cell apoptosis and led to enhanced invasiveness of pituitary adenomas. Thus, Smac, XIAP, and caspase-3 may be useful markers in determining the invasive behavior of pituitary adenomas.

The expression and localization of a novel protein phosphatase inhibitor 2810408A11Rik in mouse testis and sperm

This study investigated the expression and distribution of 2810408A11Rik in mouse testis and sperm, and explored its role in sperrnatogenesis and sperm function. The expression levels of 2810408A11Rik mRNA in multiple tissue samples were analyzed using bioinformatic resources and RT-PCR technique. A specific rabbit polyclonal antibody was prepared by prokaryotic expression of 2810408A11Rik recombinant protein and utilized for animal immunization. Western blotting, immunohistochemistry and immunofluorescence were used to detect the expression and distribution of 2810408A11Rik. The results of the bioinformatic analysi and RT-PCR showed that 2810408A11Rik mRNA was specifically expressed in mouse testis, and 2810408AllRik protein included a protein phosphatase inhibitor domain. Western blotting assays, immunohistochemistry and immunofluorescence confirmed the expression of 2810408A11Rik protein in mouse testis, especially in post-meiosis round and long spermatids, and that it is localized in the acrosome and the post-nucleus area of sperm. Our findings suggest that 2810408A11Rik may play an important role in spermatogenesis, sperm capacitation and fertilization.

Storage Proteins and Trypsin Inhibitors of an Underutilized Legume, <i>Mucuna</i>: Variability and Their Stability during Germination

The proteins and trypsin inhibitors were isolated from the seeds of different varieties/accessions of an underutilized legume, Mucuna. The crude protein content of all the germplasms of Mucuna is varied from 15% - 26%, showed little variation and contain higher crude protein when compared with other Mucuna species reported earlier and the pulse crops commonly consumed in India. The seeds of all the varieties of Mucuna exhibited trypsin inhibitor activity. The trypsin inhibitor activity varied from 11 - 14 TIA/mg of protein. Not much variation was observed in trypsin inhibitory activities in soaked seeds compared to dry seeds. Germination of Mucuna pruriens has been carried out and the change in the protein content and trypsin inhibitors were monitored. The protein content of the endosperm increased up to 72 hrs of germination and then decreased. The trypsin inhibitory activity decreased with increase in germination time. The trypsin inhibitor activity was decreased from 14.81 TIA/mg to 2.62 TIA/mg (82% reduction in the trypsin inhibitor activity) after 144 hrs germination.

Protein kinase inhibitors affect spermatogenic functions and blood testis barrier remodelling:A scoping review

Objective:To identify the role of protein kinase in male reproduction in animal models and human spermatogenic function.Methods:This study assessed the protein kinase of male reproduction in animal models and human using different reviewed paper indexed in PubMed,Science Direct,EBSCO,Scopus,Cochrane Library,Sage Journals,and Google Scholar.Data were charted based on author,year of publication published between 1893 and 2023,country,purpose,data collection,key findings,and research focus/domain.Results:The MAPK pathway contributed to the growth,maturation,and functionality of male germ cells.We also found out that certain influencing factors categorized into hormonal/non hormonal factors and chemotoxicant,as well as heat stress expressed an inhibitory mechanism on protein kinase,thus affecting spermatogenic functions and maintenance/remodeling of the blood testis barrier,as well as the physiology of the Sertoli cells necessary for nutritional support of spermatogenesis.However,activating protein kinases pathway like the mTOR pathway as well as increased expression of peroxiredoxin-4 and L-carnitine mediated protein kinases may be useful for treating or managing male reproductive dysfunction.Conclusions:Protein kinase plays an important role in spermatogenic functions and blood testis remodeling in animal and human.Its assessment provides essential information that can guide treatment strategies aimed at improving male reproductive potential.Taken together,these recent advances highlight a future therapeutic intervention in assessing male reproductive potential.It might also be possible to look at potential targets for male contraceptives in the MAPK pathway.

Regulatory Effects of X-linked Inhibitor of Apoptosis Protein and Pro-apoptotic Protein Smac on Apoptosis Resistance to Chemotherapy in Pancreatic Cancer Cells~*

Objective： To investigate the relation of X-linked inhibitor of apoptosis （XIAP） and second mitochondria-derived activator of caspase （Smac） signaling pathway to chemoresistance in human pancreatic cancer Panc-1 and BXPC-3 cells. Methods： Apoptosis and the changes of XIAP expression in permeabilized cells induced by cisplatin and 5-fluorouracil （FU） were measured by flow cytometry. The cytosolic expression of XIAP, Smac and caspase-3 was detected by Western blot. A recombinant plasmid vector pEGFP-N1/Smac was constructed and transfected into of Pancol cells. The effect of cytosolic overexpression of Smac on apoptosis of Panc-1 cells was evaluated by flow cytometry. Results： Panc-1 was more resistant to cisplatin or 5-FU induced apoptosis than BXPC-3. Western blot revealed that chemoresistant Panc-1 highly expressed XIAP, and increased cytosolic expression of Smac might be responsible for the marked down-regulation of XIAP in chemo-sensitive BXPC-3 cells after exposure to cisplatin or 5-FU. Furthermore, cytosolic overexpression of Smac could significantly down-regulate the levels of XIAP and promote the activity of caspase-3, as well as sensitize Panc-1 cells to anticancer drug-induced apoptosis. Conclusion： Anticancer drug-induced apoptosis requires mitochondrial release of Smac and downregulation of XIAP, which may be an important determinant of chemo-sensitivity in pancreatic cancer cells. Up-regulation of cytosolic expression of Smac may act as an effective modifying signal to overcome apoptosis resistance to chemotherapy in pancreatic cancer cells.

Targeting X-linked inhibitor of apoptosis protein inhibits pancreatic cancer cell growth through p-Akt depletion

AIM: To determine whether lentivirus-mediated shRNA targeting the X-linked inhibitor of apoptosis protein (XIAP) gene could be exploited in the treatment of pancreatic cancer. METHODS: Human pancreatic cancer cells Panc-1, Mia-paca2, Bxpc-3 and SW1990, infected with lentivirus, were analyzed by real-time polymerase chain reaction (PCR). Western blotting was used to examine XIAP protein levels, survivin and p-Akt to confirm the result of real-time PCR and determine the possible mechanism. The 3-(4,5-cimethylthiazol-2-yl)-2,5- diphenyl tetrazolium bromide (MTT) assay was used to measure IC50 to determine chemosensitivity to the chemotherapeutic drugs 5-fluorouracil (5-FU) and gemcitabine. A colony assay, MTT assay and a tumorigenicity experiment were used to study cell proliferation in vitro and in vivo . Caspase-3/7 activity, 4',6-diamidino-2-phenylindole-staining and flow cytometric measurements were used to study apoptosis in SW1990 cells. RESULTS: XIAP proteins were found to be differen- tially expressed among pancreatic cancer cell lines Panc-1, Mia-paca2, Bxpc-3 and SW1990. Data of real-time PCR and Western blotting showed that XIAP was reduced persistently and markedly by lentivirus-mediated shRNA. Downregulation of XIAP by transfection with XIAP shRNA resulted in decreased p-Akt expression. XIAP shRNA also inhibited the growth of pancreatic cancer cells in vitro and in vivo , enhanced drug-induced apoptosis and increased chemosensitivity to 5-FU and gemcitabine. Results also suggest that inhibition of XIAP and subsequent p-Akt depletion may have an anti-tumor effect through attenuating the ability of cancer cells to survive. CONCLUSION: Lentivirus-mediated gene therapy is an attractive strategy in the treatment of pancreatic cancer and justifies the use of lentivirus in pancreatic cancer gene therapy studies.

Baculoviral inhibitor of apoptosis protein repeatcontaining protein 3 delays early Wallerian degeneration after sciatic nerve injury

Wallerian degeneration is a complex biological process that occurs after nerve injury,and involves nerve degeneration and regeneration.Schwann cells play a crucial role in the cellular and molecular events of Wallerian degeneration of the peripheral nervous system.However,Wallerian degeneration regulating nerve injury and repair remains largely unknown,especially the early response.We have previously reported some key regulators of Wallerian degeneration after sciatic nerve injury.Baculoviral inhibitor of apoptosis protein repeat-containing protein 3(BIRC3)is an important factor that regulates apoptosis-inhibiting protein.In this study,we established rat models of right sciatic nerve injury.In vitro Schwann cell models were also established and subjected to gene transfection to inhibit and overexpress BIRC3.The data indicated that BIRC3 expression was significantly up-regulated after sciatic nerve injury.Both BIRC3 upregulation and downregulation affected the migration,proliferation and apoptosis of Schwan cells and affected the expression of related factors through activating c-fos and ERK signal pathway.Inhibition of BIRC3 delayed early Wallerian degeneration through inhibiting the apoptosis of Schwann cells after sciatic nerve injury.These findings suggest that BIRC3 plays an important role in peripheral nerve injury repair and regeneration.The study was approved by the Institutional Animal Care and Use Committee of Nantong University,China(approval No.2019-nsfc004)on March 1,2019.

Purification of Protein Kinase C Inhibitor from Bovine Spermatozoa

A protein kinase C inhibitor is found in bovine sper-matozoa. This inhibitor was purified by Sephadex G-200and isoelectrofocusion electrophoresis. The molecularweight of this protein kinase C inhibitor was about 63 000,the isoelectric point of the inhibitor was pH 4.5.

Hepatic arterial infusion chemotherapy with anti-angiogenesis agents and immune checkpoint inhibitors for unresectable hepatocellular carcinoma and meta-analysis

BACKGROUND Hepatic arterial infusion chemotherapy(HAIC)has been proven to be an ideal choice for treating unresectable hepatocellular carcinoma(uHCC).HAIC-based treatment showed great potential for treating uHCC.However,large-scale studies on HAIC-based treatments and meta-analyses of first-line treatments for uHCC are lacking.AIM To investigate better first-line treatment options for uHCC and to assess the safety and efficacy of HAIC combined with angiogenesis inhibitors,programmed cell death of protein 1(PD-1)and its ligand(PD-L1)blockers(triple therapy)under real-world conditions.METHODS Several electronic databases were searched to identify eligible randomized controlled trials for this meta-analysis.Study-level pooled analyses of hazard ratios(HRs)and odds ratios(ORs)were performed.This was a retrospective single-center study involving 442 patients with uHCC who received triple therapy or angiogenesis inhibitors plus PD-1/PD-L1 blockades(AIPB)at Sun Yat-sen University Cancer Center from January 2018 to April 2023.Propensity score matching(PSM)was performed to balance the bias between the groups.The Kaplan-Meier method and cox regression were used to analyse the survival data,and the log-rank test was used to compare the suvival time between the groups.RESULTS A total of 13 randomized controlled trials were included.HAIC alone and in combination with sorafenib were found to be effective treatments(P values for ORs:HAIC,0.95;for HRs:HAIC+sorafenib,0.04).After PSM,176 HCC patients were included in the analysis.The triple therapy group(n=88)had a longer median overall survival than the AIPB group(n=88)(31.6 months vs 14.6 months,P<0.001)and a greater incidence of adverse events(94.3%vs 75.4%,P<0.001).CONCLUSION This meta-analysis suggests that HAIC-based treatments are likely to be the best choice for uHCC.Our findings confirm that triple therapy is more effective for uHCC patients than AIPB.