T790M mutation sensitizes non-small cell lung cancer cells to radiation via suppressing SPOCK1.

Background: Approximately 50% of patients harbor the T790M mutation after developing first-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) resistance. Evidence has showed the major treatment failure is local relapses and limited metastases. Several studies have demonstrated the value of radiotherapy in metastatic non-small cell lung cancer (NSCLC) with the EGFR T790M mutation after the development of TKI resistance. The aim of this study was to explore the role of radiation in T790M-mutant NSCLC and the value of early radiotherapy for NSCLC with T790M-mediated EGFR-TKI resistance. Methods: Gefitinib-resistant NSCLC cell lines were established via stepwise exposure to increasing concentrations of gefitinib (PC-9-GR). Droplet digital PCR was used to determine the relative T790M subclone abundance. In vitro and in vivo models were established using different mixtures of PC-9-GR and PC-9 cells. Differentially expressed genes were identified using RNA sequencing. Two research models were constructed (salvage and prophylactic radiotherapy) to determine the effects of early radiotherapy on gefitinib-resistant cells. Results: PC-9-GR cells exhibited higher radiosensitivity than PC-9 cells (sensitivity enhancement ratio = 1.5). Salvage radiation reduced the number of T790M-mutant subclones, and the relative T790M abundance was significantly lower than that without radiation at 90 days (10.94% vs. 21.54%). Prophylactic radiation prevented the development of T790M subclones. These results were also confirmed in vivo. qRT-PCR revealed threefold elevation of miR-1243 in PC-9-GR cells, and the increased radiosensitivity of PC-9-GR cells was inhibited when miR-1243 was knocked down. RNA sequencing revealed that SPOCK1 was downregulated in PC-9-GR cells. Interestingly, bioinformatic analysis showed that SPOCK1 was a target gene of miR-1243. SPOCK1 knockdown markedly increased the radiosensitivity of PC-9 cells. Conclusion: Gefitinib-resistant NSCLC with the T790M mutation had higher radiosensitivity than that without the mutation, possibly mediated by SPOCK1. Early radiotherapy can eliminate T790M subclones, providing evidence for the benefit of early local treatment in patients with TKI-resistant NSCLC.

Corrigendum to "Identifies microtubule-binding protein CSPP1 as a novel cancer biomarker associated with ferroptosis and tumor microenvironment" [Comput Struct Biotechnol J 20 (2022) 3322-3335].

[This corrects the article DOI: 10.1016/j.csbj.2022.06.046.].

Risk factors for thrombocytopenia in patients receiving linezolid therapy: a systematic review and meta-analysis.

PURPOSE: The incidence of linezolid-induced thrombocytopenia (LIT) has been reported to vary widely across studies. We performed a meta-analysis to identify the risk factors for thrombocytopenia among patients who received linezolid treatment. METHODS: The PubMed, Embase and Cochrane Library databases were searched from inception to November 2022 to identify eligible studies. Data on the potential predictors of incidence in LIT were pooled using a random effects model. Sensitivity analyses were performed to determine the robustness of the results when significant heterogeneity was observed. RESULTS: Forty observational studies involving 6454 patients treated with linezolid were included in the analysis. LIT was estimated to occur in 37% of patients. The following important factors were associated with the incidence of LIT: advanced age, body mass index, concurrent renal impairment or liver disease, abnormal laboratory parameters (including white blood cell count, serum creatinine, baseline platelet count, albumin, creatinine clearance rate, and estimated glomerular filtration rate), treatment duration and renal replacement therapy. CONCLUSIONS: A variety of risk factors related to the occurrence of LIT were revealed in our analysis. Early identification of these factors could help patients improve clinical outcomes.

Dynamic Regulation Genes at Microtubule Plus Ends: A Novel Class of Glioma Biomarkers.

Glioma is the most prevalent and aggressive primary nervous system tumor with an unfavorable prognosis. Microtubule plus-end-related genes (MPERGs) play critical biological roles in the cell cycle, cell movement, ciliogenesis, and neuronal development by coordinating microtubule assembly and dynamics. This research seeks to systematically explore the oncological characteristics of these genes in microtubule-enriched glioma, focusing on developing a novel MPERG-based prognostic signature to improve the prognosis and provide more treatment options for glioma patients. First, we thoroughly analyzed and identified 45 differentially expressed MPERGs in glioma. Based on these genes, glioma patients were well distinguished into two subgroups with survival and tumor microenvironment infiltration differences. Next, we further screened the independent prognostic genes (CTTNBP2, KIF18A, NAV1, SLAIN2, SRCIN1, TRIO, and TTBK2) using 36 prognostic-related differentially expressed MPERGs to construct a signature with risk stratification and prognostic prediction ability. An increased risk score was related to the malignant progression of glioma. Therefore, we also designed a nomogram model containing clinical factors to facilitate the clinical use of the risk signature. The prediction accuracy of the signature and nomogram model was verified using The Cancer Genome Atlas and Chinese Glioma Genome Atlas datasets. Finally, we examined the connection between the signature and tumor microenvironment. The signature positively correlated with tumor microenvironment infiltration, especially immunoinhibitors and the tumor mutation load, and negatively correlated with microsatellite instability and cancer stemness. More importantly, immune checkpoint blockade treatment and drug sensitivity analyses confirmed that this prognostic signature was helpful in anticipating the effect of immunotherapy and chemotherapy. In conclusion, this research is the first study to define and validate an MPERG-based signature closely associated with the tumor microenvironment as a reliable and independent prognostic biomarker to guide personalized choices of immunotherapy and chemotherapy for glioma patients.

Non-coding ribonucleic acid-mediated CAMSAP1 upregulation leads to poor prognosis with suppressed immune infiltration in liver hepatocellular carcinoma.

Liver hepatocellular carcinoma (LIHC) is well-known for its unfavorable prognosis due to the lack of reliable diagnostic and prognostic biomarkers. Calmodulin-regulated spectrin-associated protein 1 (CAMSAP1) is a non-centrosomal microtubule minus-end binding protein that regulates microtubule dynamics. This study aims to investigate the specific role and mechanisms of CAMSAP1 in LIHC. We performed systematical analyses of CAMSAP1 and demonstrated that differential expression of CAMSAP1 is associated with genetic alteration and DNA methylation, and serves as a potential diagnostic and prognostic biomarker in some cancers, especially LIHC. Further evidence suggested that CAMSAP1 overexpression leads to adverse clinical outcomes in advanced LIHC. Moreover, the AC145207.5/LINC01748-miR-101-3p axis is specifically responsible for CAMSAP1 overexpression in LIHC. In addition to the previously reported functions in the cell cycle and regulation of actin cytoskeleton, CAMSAP1-related genes are enriched in cancer- and immune-associated pathways. As expected, CAMSAP1-associated LIHC is infiltrated in the suppressed immune microenvironment. Specifically, except for immune cell infiltration, it is significantly positively correlated with immune checkpoint genes, especially CD274 (PD-L1), and cancer-associated fibroblasts. Prediction of immune checkpoint blockade therapy suggests that these patients may benefit from therapy. Our study is the first to demonstrate that besides genetic alteration and DNA methylation, AC145207.5/LINC01748-miR-101-3p-mediated CAMSAP1 upregulation in advanced LIHC leads to poor prognosis with suppressed immune infiltration, representing a potential diagnostic and prognostic biomarker as well as a promising immunotherapy target for LIHC.

Identifies microtubule-binding protein CSPP1 as a novel cancer biomarker associated with ferroptosis and tumor microenvironment.

Centrosome and spindle pole-associated protein (CSPP1) is a centrosome and microtubule-binding protein that plays a role in cell cycle-dependent cytoskeleton organization and cilia formation. Previous studies have suggested that CSPP1 plays a role in tumorigenesis; however, no pan-cancer analysis has been performed. This study systematically investigates the expression of CSPP1 and its potential clinical outcomes associated with diagnosis, prognosis, and therapy. CSPP1 is widely present in tissues and cells and its aberrant expression serves as a diagnostic biomarker for cancer. CSPP1 dysregulation is driven by multi-dimensional mechanisms involving genetic alterations, DNA methylation, and miRNAs. Phosphorylation of CSPP1 at specific sites may play a role in tumorigenesis. In addition, CSPP1 correlates with clinical features and outcomes in multiple cancers. Take brain low-grade gliomas (LGG) with a poor prognosis as an example, functional enrichment analysis implies that CSPP1 may play a role in ferroptosis and tumor microenvironment (TME), including regulating epithelial-mesenchymal transition, stromal response, and immune response. Further analysis confirms that CSPP1 dysregulates ferroptosis in LGG and other cancers, making it possible for ferroptosis-based drugs to be used in the treatment of these cancers. Importantly, CSPP1-associated tumors are infiltrated in different TMEs, rendering immune checkpoint blockade therapy beneficial for these cancer patients. Our study is the first to demonstrate that CSPP1 is a potential diagnostic and prognostic biomarker associated with ferroptosis and TME, providing a new target for drug therapy and immunotherapy in specific cancers.

SERPINE2/PN-1 regulates the DNA damage response and radioresistance by activating ATM in lung cancer.

Although the DNA damage response (DDR) is associated with the radioresistance characteristics of lung cancer cells, the specific regulators and underlying mechanisms of the DDR are unclear. Here, we identified the serine proteinase inhibitor clade E member 2 (SERPINE2) as a modulator of radiosensitivity and the DDR in lung cancer. Cells exhibiting radioresistance after ionizing radiation show upregulation of SERPINE2, and SERPINE2 knockdown improves tumor radiosensitivity in vitro and in vivo. Functionally, SERPINE2 deficiency causes a reduction in homologous recombination repair, rapid recovery of cell cycle checkpoints, and suppression of migration and invasion. Mechanistically, SERPINE2 knockdown inhibits the accumulation of p-ATM and the downstream repair protein RAD51 during DNA repair, and RAD51 can restore DNA damage and radioresistance phenotypes in lung cancer cells. Furthermore, SERPINE2 can directly interact with MRE11 and ATM to facilitate its phosphorylation in HR-mediated DSB repair. In addition, high SERPINE2 expression correlates with dismal prognosis in lung adenocarcinoma patients, and a high serum SERPINE2 concentration predicts a poor response to radiotherapy in non-small cell lung cancer patients. In summary, these findings indicate a novel regulatory mechanism by which SERPINE2 modulates the DDR and radioresistance in lung cancer.

Capilliposide C from Lysimachia capillipes Restores Radiosensitivity in Ionizing Radiation-Resistant Lung Cancer Cells Through Regulation of ERRFI1/EGFR/STAT3 Signaling Pathway.

AIMS: Radiation therapy is used as the primary treatment for lung cancer. Unfortunately, radiation resistance remains to be the major clinic problem for lung cancer patients. Lysimachia capillipes capilliposide C (LC-C), an extract from LC Hemsl, has demonstrated multiple anti-cancer effects in several types of cancer. Here, we investigated the potential therapeutic impacts of LC-C on radiosensitivity in lung cancer cells and their underlying mechanisms. METHODS: Non-small cell lung cancer cell lines were initially irradiated to generate ionizing radiation (IR)-resistant lung cancer cell lines. RNA-seq analysis was used to examine the whole-transcriptome alteration in IR-resistant lung cancer cells treated with or without LC-C, and the differentially expressed genes with most significance were verified by RT-qPCR. Colony formation assays were performed to determine the effect of LC-C and the target gene ErbB receptor feedback inhibitor 1 (ERRFI1) on radiosensitivity of IR-resistant lung cancer cells. In addition, effects of ERRFI1 on cell cycle distribution, DNA damage repair activity were assessed by flow cytometry and γ-H2AX immunofluorescence staining respectively. Western blotting was performed to identify the activation of related signaling pathways. Tumor xenograft experiments were conducted to observe the effect of LC-C and ERRFI1 on radiosensitivity of IR-resistant lung cancer cells in vivo. RESULTS: Compared with parental cells, IR-resistant lung cancer cells were more resistant to radiation. LC-C significantly enhanced the effect of radiation in IR-resistant lung cancer cells both in vitro and in vivo and validated ERRFI1 as a candidate downstream gene by RNA-seq. Forced expression of ERRFI1 alone could significantly increase the radiosensitivity of IR-resistant lung cancer cells, while silencing of ERRFI1 attenuated the radiosensitizing function of LC-C. Accordingly, LC-C and ERRFI1 effectively inhibited IR-induced DNA damage repair, and ERRFI1 significantly induced G2/M checkpoint arrest. Additional investigations revealed that down-regulation of EGFR/STAT3 pathway played an important role in radiosensitization between ERRFI1 and LC-C. Furthermore, the high expression level of ERRFI1 was associated with high overall survival rates in lung cancer patients. CONCLUSIONS: Treatment of LC-C may serve as a promising therapeutic strategy to overcome the radiation resistance and ERRFI1 may be a potential therapeutic target in NSCLC.

Suberoylanilide hydroxamic acid enhances the radiosensitivity of lung cancer cells through acetylated wild-type and mutant p53-dependent modulation of mitochondrial apoptosis.

OBJECTIVE: Suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor, has shown potential as a candidate radiosensitizer for many types of cancers. This study aimed to explore the radiosensitization mechanism of SAHA in lung cancer cells. METHODS: Mutations in p53 were generated by site-directed mutagenesis using polymerase chain reaction. Transfection was performed to generate H1299 cells carrying wild-type or mutant p53. The radiosensitizing enhancement ratio was determined by clonogenic assays. Mitochondrial apoptosis was detected using JC-1 staining and flow cytometry analysis. RESULTS: Our results showed that SAHA induced radiosensitization in H1299 cells expressing wild-type p53, p53R175H or p53P223L, but this enhanced clonogenic cell death was not observed in parental H1299 (p53-null) cells or H1299 cells expressing p53 with K120R, A161T and V274R mutations. In SAHA-sensitized cells, mitochondrial apoptosis was induced following exposure to irradiation. Additionally, we observed that a secondary mutation at K120 (K120R) could eliminate p53-mediated radiosensitization and mitochondrial apoptosis. CONCLUSIONS: The results of this study suggest that wild-type and specific mutant forms of p53 mediate SAHA-induced radiosensitization by regulating mitochondrial apoptosis, and the stabilization of K120 acetylation by SAHA is the molecular basis contributing to radiosensitization in lung cancer cells.

Identification of somatic copy number variations in plasma cell free DNA correlating with intrinsic resistances to EGFR targeted therapy in T790M negative non-small cell lung cancer.

BACKGROUND: About 20-30% EGFR-mutant non-small lung cancer show intrinsic resistance to EGFR targeted therapies. Compared to T790M positive in acquired resistance patients, little is known about EGFR-TKI intrinsic resistance for T790M negative patients. METHODS: Thirty-one patients with advanced stage lung cancer, including 18 patients with intrinsic resistance (PFS <6 months) and 13 patients with acquired resistance (PFS >36 months) but are negative for plasma T790M were recruited in the study. Plasma cell free DNA was profiled by low coverage whole genome sequencing with median genome coverage of 1.86X by Illumina X10. Sequencing coverage across chromosomes was summarized by samtools, and normalized by segmentation analysis as provided by R package 'DNACopy'. RESULTS: The most frequent chromosomal changes were found on chr7, chr1 and chr8. Among them, chr7p gains were found in 12 (66.7%) intrinsic resistance and 4 (30.7%) acquired resistance patients. The gene EGFR was found located on the focal amplification peak of chr7p. The performance of 7p gain to predict intrinsic resistance reaches AUC =0.902. Similarly, focal amplifications were also found on chromosome 5, 16 and 22, where tumor related gene PCDHA@, ADAMTS18 and CRKL were located. Focal deletions were also found in chr1, 8, 10 and 16, where genes SFTPA1/2, DLC1, PTEN and CDH1 are located. CONCLUSIONS: The results suggest cell free DNA copy number might be a useful peripheral blood tumor biomarker for predicting intrinsic resistance of EGFR targeted therapy and prognosis.

Genome-wide profiling reveals alternative polyadenylation of mRNA in human non-small cell lung cancer.

BACKGROUND: Lung cancer is the second most common cancer with an extremely poor overall survival rate. Post-transcriptional regulation of gene expression play many important roles in human cancer, and one of the potential mechanisms underlying this is alternative mRNA maturation at its 3' untranslated regions (3'-UTRs). METHODS: Cancer tissues and paired adjacent normal lung tissues from 26 patients diagnosed with non-small cell lung cancer (NSCLC) were analyzed by in vitro transcription-sequencing alternative polyadenylation sites (IVT-SAPAS). 41,773,101 reads in average were obtained from each paired sample. A potential regulation of Cleavage Stimulation Factor Subunit 2 (CSTF2) on 3'UTR length of genes was tested in H460 cells. RESULTS: 1439 (10.26%) genes showed up-regulated expression and 1364 (9.72%) genes showed down-regulated expression in lung cancer tissue versus normal lung tissue, and shorten 3'UTR in cancer tissue was detected in cancer tissues collected from 96.2% (25/26) patients, indicating lung cancer tend to have shortened 3'UTRs of these identified genes. KEGG analysis showed 1855 genes with shorten 3'UTR were enriched in mTOR signaling, ubiquitin mediated proteolysis and RNA degradation. Knocking down CSTF2 expression in H460 cells results in 3'UTR elongation of genes that was identified to be with shortened length in cancer tissues. CONCLUSION: Alternative polyadenylation (APA) site-switching of 3'UTRs is prevalent in NSCLC, and CSTF2 may serve as an oncogene regulates the 3'UTR length of cancer related genes in NSCLC.

Intraperitoneal perfusion chemotherapy and whole abdominal hyperthermia using external radiofrequency following radical D2 resection for treatment of advanced gastric cancer.

BACKGROUND: The peritoneum is the most frequent site of disease recurrence in gastric cancer, and the prognosis remains poor. This study assessed the role of adjuvant intraperitoneal (IP) chemotherapy with whole abdominal hyperthermia using external radiofrequency in gastric cancer patients after D2 dissection. METHODS: Patients with gastric cancer who underwent gastrectomy with D2 regional lymph node dissection were enrolled in the study. Patients received IP chemotherapy with whole abdominal hyperthermia. Preheated normal saline containing 75 mg/m2 of cisplatin was delivered into the abdominal cavity through a Tenckhoff catheter at McBurney's point. Regional hyperthermia was performed using two sets of orthogonal radiofrequency waves immediately after all saline was irrigated into the abdominal cavity. For each patient, recurrent or metastatic sites and adverse events were evaluated. RESULTS: A total of 22 patients were finally included. All patients tolerated hyperthermia well. Only two patients experienced grade 1 superficial thermal injury. The most frequent grade 3/4 adverse events were myelosuppression, nausea/vomiting, trichomadesis and liver dysfunction. We also found IP chemotherapy with whole abdominal hyperthermia could reduce the total recurrent/metastatic rate, especially peritoneal metastasis (4.5%). CONCLUSIONS: This hypothesis-generating study indicated that IP chemotherapy with whole abdominal hyperthermia might be feasible for gastric cancer patients after D2 resection.

S-allylcysteine suppresses ovarian cancer cell proliferation by DNA methylation through DNMT1.

BACKGROUND: The anti-tumor effects of S-allylcysteine (SAC), a water-soluble garlic derivative, on human ovarian cancer cells have been previous studied in vitro and in vivo models but the precise epigenetic molecular mechanisms are still unclear. This study aimed to investigate the epigenetic mechanism of SAC. METHODS: Human epithelial ovarian cancer cell line A2780 was selected. Cell proliferation and cell cycle was analyzed. DNA methylation, DNA methyltransferase (DNMT) activity, tumor suppressor gene expressions, as well as protein expression were analyzed. RESULTS: SAC could inhibit the proliferation of A2780 cells in dose- and time-dependent manners (the IC50 was 16.25 mmol/L and 5.25 mmol/L at 48 h and 72 h). Treatment of A2780 cells with SAC resulted in G1/S phase arrest. SAC treatment decreased global DNA methylation levels in A2780 cells in a dose-dependent manner. SAC decreased the levels of 5-methylcytosine, DNMT activity, messenger RNA (mRNA) and protein levels of DNMT1. Additionally, SAC treatment resulted in re-expression of the mRNA and proteins of silenced tumor suppressor gene CDKN1A accompany with reduced cell division control 2 expression. CONCLUSION: Our data indicated the potential therapeutic effects of SAC on the human ovarian carcinoma cell line A2780 in vitro. The epigenetic mechanism of action of SAC may have important implications for epigenetic therapy.

Therapeutical effect of intrapleural perfusion with hyperthermic chemotherapy on malignant pleural effusion under video-assisted thoracoscopic surgery.

BACKGROUND: Patients with malignant pleural effusions (MPEs) have limited life expectancy. This study aims to investigate the feasibility of intrapleural perfusion with hyperthermic chemotherapy (IPHC) under video-assisted thoracoscopic surgery on MPE patients. METHODS: MPE patients were enrolled in the study and treated with IPHC. The treatment response was classified as complete response (CR, no re-accumulation of pleural fluid for 4 weeks), partial response (PR, re-accumulation above the post-IPHC level but below the pre-IPHC level for four weeks), no response (NR; re-accumulation or above the pre-IPHC level). The change of Karnofsky performance score (KPS) and tumour markers were also recorded. Follow-up was done every two weeks during first month and monthly thereafter until death. RESULTS: Eighty patients included 46 males and 34 females were included in the study. The total response rate was 100%, with 71.3% of CR and 28.7% of PR. The KPS scores were significantly elevated and the level of tumour markers in pleural effusion were dramatically decreased after IPHC. The median survival was 16.8 months ranged from 2.1 to 67.4 months. One-year and two-year survival rates were 82.5% and 23.8%, respectively. There were no serious clinical compilations during IPHC treatment. CONCLUSIONS: IPHC is a safety, effective and promising approach for MPE patients. It provides well survival benefit and minor toxicities.

Capilliposide from Lysimachia capillipes inhibits AKT activation and restores gefitinib sensitivity in human non-small cell lung cancer cells with acquired gefitinib resistance.

Most gefitinib-treated patients with non-small cell lung cancer (NSCLC) would eventually develop resistance. Lysimachia capillipes (LC) capilliposide extracts from LC Hemsl. show both in vitro and in vivo anti-cancer effects. In this study we investigated whether LC capilliposide in combination with gefitinib could overcome the resistance of NSCLC cells to gefitinib and identified the signaling pathways involved. Treatment with LC capilliposide alone inhibited the growth of a panel of NSCLC cell lines (PC-9, H460, H1975, H1299 and PC-9-GR) sensitive or resistant to gefitinib with IC50 values in the range of µg/mL. In the gefitinib-resistant PC-9-GR cells (which have a T790M EGFR mutation), LC capilliposide (at the IC30, i.e.1.2 µg/mL) markedly enhanced the inhibitory effects of gefitinib with its IC50 value being decreased from 6.80±1.00 to 0.77±0.12 µmol/L. By using the median effect analysis we showed that combination treatment of LC capilliposide and gefitinib could restore gefitinib sensitivity in PC-9-GR cells. Furthermore, LC capilliposide (1.2 µg/mL) significantly increased the apoptotic responses to gefitinib (0.77 µmol/L) in PC-9-GR cells, but did not affect gefitinib-induced G0/G1 arrest. Moreover, LC capilliposide (1.2 µg/mL) in combination with gefitinib (0.77, 1.0 µmol/L) markedly decreased the phosphorylation of the EGFR downstream signaling molecule AKT, which neither LC capilliposide nor gefitinib alone affected. In PC-9-GR cells with siRNA knockdown of AKT, addition of LC capilliposide was unable to increase gefitinib sensitivity. In a PC-9-GR xenograft mouse model, combination treatment with LC capilliposide (15 mg·kg-1·d-1, ip) and gefitinib (50 mg·kg-1·d-1, ip) dramatically enhanced tumor growth suppression (with a TGI of 109.3%), compared with TGIs of 22.6% and 56.6%, respectively, in mice were treated with LC capilliposide or gefitinib alone. LC capilliposide can restore the cells' sensitivity to gefitinib through modulation of pAKT levels, suggesting that a combination of LC capilliposide and gefitinib may be a promising therapeutic strategy to overcome gefitinib resistance in NSCLCs with a T790M mutation.

Efficacy of afatinib, an irreversible ErbB family blocker, in the treatment of intracerebral metastases of non-small cell lung cancer in mice.

Few effective therapeutic options are currently available for the treatment of non-small cell lung cancer (NSCLC) with brain metastases (BM). Recent evidence shows that NSCLC patients with BMs respond well to afatinib, but little is known about the underlying mechanisms. In this study, we evaluated the efficacy of afatinib in treatment of BMs in mice and investigated whether afatinib could actively penetrate the brain-blood barrier and bind to its target. NSCLC BM model was established in nude mice by intracerebral injection of PC-9.luc cells. The tumors were measured weekly using in vivo quantitative bioluminescence. The mice are administrated afatinib (15, 30 mg·kg-1·d-1, ig) for 14 d. The antitumor efficacy of afatinib was determined by tumor growth inhibition (TGI), which was calculated as [1-(change of tumor volume in treatment group/control group)×100]. Pharmacokinetic characteristics were measure in mice receiving a single dose of afatinib (30 mg/kg, ig). Pharmacodynamics of afatinib was also assessed by detecting the expression of pEGFR (Tyr1068) in brain tumor foci using immunohistochemistry. Administration of afatinib (15, 30 mg·kg-1·d-1) dose-dependently inhibited PC-9 tumor growth in the brain with a TGI of 90.2% and 105%, respectively, on d 14. After administration of afatinib (30 mg/kg), the plasma concentration of afatinib was 91.4±31.2 nmol/L at 0.5 h, reached a peak (417.1±119.9 nmol/L) at 1 h, and was still detected after 24 h. The cerebrospinal fluid (CSF) concentrations followed a similar pattern. The T1/2 values of afatinib in plasma and CSF were 5.0 and 3.7 h, respectively. The AUC(0-24 h) values for plasma and CSF were 2375.5 and 29.1 nmol/h, respectively. The plasma and CSF concentrations were correlated (r=0.844, P<0.01). Pharmacodynamics study showed that the expression levels of pEGFR were reduced by 90% 1 h after afatinib administration. The Emax was 86.5%, and the EC50 was 0.26 nmol/L. A positive correlation between CSF concentrations and pEGFR modulation was revealed. Afatinib penetrates the BBB in NSCLC BM mice and contributes to the brain tumor response. The CSF exposure level is correlated with the plasma level, which in turn is correlated with the modulation of pEGFR in the tumor tissues. The results support for the potential application of afatinib in NSCLC patients with BMs.

Role of p38 MAPK in enhanced human cancer cells killing by the combination of aspirin and ABT-737.

Regular use of aspirin after diagnosis is associated with longer survival among patients with mutated-PIK3CA colorectal cancer, but not among patients with wild-type PIK3CA cancer. In this study, we showed that clinically achievable concentrations of aspirin and ABT-737 in combination could induce a synergistic growth arrest in several human PIK3CA wild-type cancer cells. In addition, our results also demonstrated that long-term combination treatment with aspirin and ABT-737 could synergistically induce apoptosis both in A549 and H1299 cells. In the meanwhile, short-term aspirin plus ABT-737 combination treatment induced a greater autophagic response than did either drug alone and the combination-induced autophagy switched from a cytoprotective signal to a death-promoting signal. Furthermore, we showed that p38 acted as a switch between two different types of cell death (autophagy and apoptosis) induced by aspirin plus ABT-737. Moreover, the increased anti-cancer efficacy of aspirin combined with ABT-737 was further validated in a human lung cancer A549 xenograft model. We hope that this synergy may contribute to failure of aspirin cancer therapy and ultimately lead to efficacious regimens for cancer therapy.

S-allylcysteine, a garlic derivative, suppresses proliferation and induces apoptosis in human ovarian cancer cells in vitro.

AIM: To investigate the effects of S-allylcysteine (SAC), a water-soluble garlic derivative, on human ovarian cancer cells in vitro. METHODS: Human epithelial ovarian cancer cell line A2780 was tested. Cell proliferation was examined with CCK-8 and colony formation assays. Cell cycle was analyzed with flow cytometry. Cell apoptosis was studied using Hoechst 33258 staining and Annexin V/PI staining with flow cytometry. The migration and invasion of A2780 cells were examined with transwell and wound healing assays. The expression of relevant proteins was detected with Western blot assays. RESULTS: SAC (1-100 mmol/L) inhibited the proliferation of A2780 cells in dose- and time-dependent manners (the IC50 value was approximately 25 mmol/L at 48 h, and less than 6.25 mmol/L at 96 h). Furthermore, SAC dose-dependently inhibited the colony formation of A2780 cells. Treatment of A2780 cells with SAC resulted in G1/S phase arrest and induced apoptosis, accompanied by decreased expression of pro-caspase-3, Parp-1 and Bcl-2, and increased expression of active caspase-3 and Bax. SAC treatment significantly reduced the migration of A2780 cells, and markedly decreased the protein expression of Wnt5a, p-AKT and c-Jun, which were the key proteins involved in proliferation and metastasis. CONCLUSION: SAC suppresses proliferation and induces apoptosis in A2780 ovarian cancer cells in vitro.