Efficacy and safety of EGFR-TKIs in combination with angiogenesis inhibitors as first-line therapy for advanced EGFR-mutant non-small-cell lung cancer: a systematic review and meta-analysis.

BACKGROUND: For patients with advanced non-small-cell lung cancer (NSCLC) with EGFR mutations, the suggested course of action is epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs). Even with a high disease control rate, a majority of patients develop acquired EGFR-TKIs resistance and eventually advance. To increase the benefits of treatment, clinical trials are increasingly exploring the value of EGFR-TKIs combined with angiogenesis inhibitors as a first-line treatment in advanced NSCLC carrying EGFR mutations. METHOD: Using PubMed, EMBASE and Cochrane Library, to locate published full-text articles in print or online, a thorough literature search was done from the database's inception to February 2021. Additionally, oral presentation RCTs from ESMO and ASCO were obtained. We sifted out RCTs that used EGFR-TKIs along with angiogenesis inhibitors as first-line therapy for advanced EGFR-mutant NSCLC. ORR, AEs, OS, and PFS were the endpoints. Review Manager version 5.4.1 was used for data analysis. RESULTS: One thousand eight hundred twenty-one patients were involved in 9 RCTs. According to the results, combining EGFR-TKIs with angiogenesis inhibitors therapy prolonged PFS of advanced EGFR-mutation NSCLC patients on the whole [HR:0.65 (95%CI: 0.59~0.73, P<0.00001)]. No significant statistical difference was identified between the combination group and single drug group in OS(P=0.20) and ORR (P=0.11). There are more adverse effects when EGFR-TKIs are used in combination with angiogenesis inhibitors than when used alone. CONCLUSION: The combination of EGFR-TKIs and angiogenesis inhibitors prolonged PFS in patients with EGFR-mutant advanced NSCLC, but the OS and ORR benefit was not significant, and the risk of adverse events was higher, more pronounced with hypertension and proteinuria; PFS in subgroups suggested that the combination was associated with better PFS in the smoking, liver metastasis, and no brain metastasis groups, and the included studies suggested that the smoking group , liver metastasis group, and brain metastasis group may have a potential OS benefit.

Programmed death ligand 1 expression and CD8+ tumor-infiltrating lymphocyte density differences between paired primary and brain metastatic lesions in non-small cell lung cancer.

Immunotherapy targeting the programmed cell death-1/programmed death ligand 1(PD-L1) pathway has shown promising antitumor activity in brain metastases (BMs) of non-small cell lung cancer (NSCLC) patients with an acceptable safety profile; however, the response rates often differ between primary lesions and intracranial lesions. Studies are necessary to identify detailed characterizations of the response biomarkers. In this study, we aimed to compare the differences of PD-L1 expression and CD8+ tumor-infiltrating lymphocyte (TIL) density, two major response biomarkers of PD-1/PD-L1 blockade, between paired primary and brain metastatic lesions in advanced NSCLC. We observed that among primary lesions or BMs, only a small number of patients harbored common PD-L1 expression on both tumor cells and tumor-infiltrating immune cells. Additionally, we found that the numbers of CD8+ TILs were significantly fewer in BMs than in primary lung cancers. Low stromal CD8+ TIL numbers in BMs were associated with significantly shorter overall survival compared to high stromal CD8+ TIL counts. Notably, we demonstrated a discrepancy in PD-L1 expression and CD8+ TIL density between primary lung cancers and their corresponding BMs. Such heterogeneities are significantly associated with the time at which BMs occurred. Our study emphasizes the spatial and temporal heterogeneity of biomarkers for anti-PD-1/PD-L1 therapy, which should be concerned in clinical practice.

Echinacoside, a phenylethanoid glycoside from Cistanche deserticola, extends lifespan of Caenorhabditis elegans and protects from Aß-induced toxicity.

Cistanche deserticola has been found to exert protection against aging and age-related diseases, but mechanisms underlying its longevity effects remain largely unclear. Here, the multicellular model organism Caenorhabditis elegans was employed to identify lifespan extending and protective effects against ß-amyloid (Aß) induced toxicity by echinacoside (ECH), a phenylethanoid glycoside isolated from C. deserticola. Our results showed that ECH extends the mean lifespan of worms and increases their survival under oxidative stress. Levels of intracellular reactive oxygen species and fat accumulation were also significantly suppressed by ECH. Moreover, ECH-mediated lifespan extension was found to be dependent on mev-1, eat-2, daf-2, and daf-16, but not sir-2.1 or hsf-1 genes. Furthermore, ECH triggered DAF-16 nuclear localization and upregulated two of its downstream targets, sod-3 and hsp-16.2. In addition, ECH significantly improved the survival of CL4176 worms in response to proteotoxic stress induced by Aß protein aggregation. Collectively, these findings suggested that reactive oxygen species scavenging, dietary restriction, and insulin/insulin-like growth factor signaling pathways could be partly involved in ECH-mediated lifespan extension. Thus, ECH may target multiple longevity mechanisms to extend lifespan and have a potency to prevent Alzheimer's disease progression.

miR-491 Inhibits Osteosarcoma Lung Metastasis and Chemoresistance by Targeting αB-crystallin.

Dysregulated microRNAs (miRNAs) play an important role in osteosarcoma (OS) progression. In the present study, we investigate the clinical significance of serum miR-491 level and the potential role of miR-491 in OS lung metastasis and chemoresistance. Clinical data show that the level of miR-491 was decreased in serum from OS patients compared with healthy control subjects, and that a decreased serum miR-491 level is correlated with increased metastasis, poor chemoresponse, and lower survival rate in OS patients. In vitro and in vivo experiments show that overexpression of miR-491 suppresses OS cell lung metastasis, whereas it enhances cisplatin (CDDP)-induced tumor growth inhibition and apoptosis. In contrast, inhibition of miR-491 stimulates OS cell lung metastasis and suppresses CDDP-induced tumor growth inhibition and apoptosis. Furthermore, we demonstrate that miR-491 exerts its role by directly targeting αB-crystallin (CRYAB) in OS. Our findings suggest that serum level of miR-491 has potential as a biomarker for predicting OS progression and prognosis of OS patients. Additionally, restoration of miR-491 may be a novel strategy for inhibiting OS lung metastasis and overcoming OS cell resistance to chemotherapy.

Central nervous system progression in advanced non-small cell lung cancer patients with EGFR mutations in response to first-line treatment with two EGFR-TKIs, gefitinib and erlotinib: a comparative study.

BACKGROUND: Central nervous system (CNS) brain metastasis of advanced non-small cell lung cancer (NSCLC) patients confers a worse quality of life and prognosis. The efficacy comparison of two first-generation epidermal growth factor receptor (EGFR) inhibitors erlotinib or gefitinib as first-line treatment for CNS metastasis NSCLC patients with EGFR-sensitizing mutations is yet to be elucidated. METHODS: A retrospective analysis was done on cerebral metastasis rate after erlotinib or gefitinib as first-line treatment for advanced NSCLC patients with EGFR-sensitizing mutations. Time to neurological progression (nTTP) and median progression-free survival (mPFS) were calculated. RESULTS: The study involved 279 patients (erlotinib group: 108, gefitinib group: 171). After a median follow-up of 22 months, 27 patients (25%) in the erlotinib group and 60 patients (35.1%) in the gefitinib group showed CNS progression. The HR of CNS progression for erlotinib versus gefitinib was 0.695 [95% confidence interval (CI), 0.406-1.190], suggesting a risk reduction of 30.5% although not achieving statistical significance. The 6-, 12- and 18-month cumulative CNS progression rates were 0.9, 3.7 and 12% for erlotinib compared with corresponding rates of 5.8, 9.4 and 17% for gefitinib (P = 0.181). However, for those patients with preexisting brain metastases prior to EGFR-TKI treatment, erlotinib as first line treatment significantly extended the median nTTP in comparison to gefitinib (30 months vs 15.8 months, p = 0.024). CONCLUSIONS: Our data show that nTTP can be effectively extended in preexisting brain metastases patients with EGFR-sensitizing mutations initially treated with erlotinib compared with gefitinib. If confirmed, our results indicate that erlotinib may play an important role in controlling CNS progression from EGFR mutation-positive NSCLC.

FokI polymorphism in vitamin D receptor gene and risk of breast cancer among Caucasian women.

Vitamin D plays a central role in cellular proliferation, apoptosis induction, and tumor growth suppression. The vitamin D receptor (VDR) is a crucial mediator for the cellular effects of vitamin D. A series of epidemiological studies have examined the association between the VDR FokI polymorphism and breast cancer risk, but the findings remain inconclusive. Fifteen eligible case-control studies involving 15,681 cancer cases and 20,632 control subjects were identified through searching PubMed, Embase, and Web of Science. Odds ratios (ORs) and 95% confidence intervals (CIs) were used to assess the association. Heterogeneity across studies was examined with the chi-square-based Q test and the I (2) index. Begg's and Egger's test were also performed to determine publication bias. All statistical data were analyzed by STATA software. The combined estimates did not show significant risks correlated with the FokI polymorphism. However, we found an increased risk in the subgroup analysis by source of control (OR = 1.11, 95% CI = 1.01-1.22; heterogeneity test: P = 0.116, I(2) = 0.0% for ff vs FF; OR = 1.10, 95% CI = 1.01-1.21; heterogeneity test: P = 0.832, I(2) = 0.0% for ff vs Ff + FF). This meta-analysis suggests that the presence of FokI polymorphism may contribute to the risk of breast cancer in Caucasians.

Human apurinic/apyrimidinic endonuclease 1 translocalizes to mitochondria after photodynamic therapy and protects cells from apoptosis.

Photodynamic therapy (PDT) is an effective therapeutic regime for lung cancer. Mitochondrial functional failure is considered to be one of the most important factors causing cell death after PDT. However, the detailed mechanisms that are involved are still unclear. We previously reported that apurinic/apyrimidinic endonuclease (APE1) plays a critical role in regulating sensitivity to PDT in the lung cancer A549 cell line. An important mitochondrial regulatory role for APE1 has recently been reported, so therefore we explored the role of APE1 in cell survival after PDT-induced oxidative stress through regulation of mitochondrial function. We first observed that photoirradiation induced the mitochondrial translocation of APE1. The ability of APE1 to regulate mitochondrial membrane potential and reactive oxygen species (ROS) production after photoirradiation was tested in APE1 knockdown A549 cells. APE1-deficient A549 cells were characterized as having a lower mitochondrial membrane potential and higher ROS production, which led to increased apoptosis through the mitochondrial pathway after PDT. Additionally, unexpected activity of APE1 was observed in mitochondria: the control of mitochondrial transcriptional activity by redox regulation of mitochondrial transcription factor A (TFAM). Furthermore, two dominant-negative mutants of APE1 were overexpressed to enhance their individual activities in mitochondria. The results suggest that both these APE1 activities play a role in the regulation of mitochondrial function but through different mechanisms. The present study not only provides possible mechanisms for APE1 in regulating survival after photoirradiation but also uncovers a new activity of APE1 in mitochondria.

[nm23-H1-siRNA enhances the chemosensitivity to liposome-encapsulated paclitaxel in lung adenocarcinoma cells in vitro].

OBJECTIVE: To study the chemosensitivity of lung adenocarcinoma cell line A549 cells to liposome-encapsulated paclitaxel after treatment by nm23-H1-small interference RNA (nm23-H1-siRNA) in vitro. METHODS: The A549 cells were divided into two groups: non-transfected group and nm23-H1-siRNA-transfected group. Western blot analysis was used to detect the expression of nm23-H1. MTT and flow cytometry were used to determine the cell mortality rate, apoptosis rate and cell cycle after liposome-encapsulated paclitaxel treatment in both groups. RESULTS: The expression of nm23-H1 in A549 cells was significantly decreased after transfection with nm23-H1-siRNA. After treatment for 48 hours with liposome-encapsulated paclitaxel, the cell mortality rate was increased with the increasing concentration of liposome-encapsulated paclitaxel in both groups, but increased higher in the nm23-H1-siRNA-transfected group. When the concentration of liposome-encapsulated paclitaxel was above 5 µg/ml, the cell mortality rate was significantly higher than that in the non-transfected group (P < 0.05). The proportion of apoptotic cells also increased in the nm23-H1-siRNA-transfected group, compared with that of the non-transfected group (t = 3.812, P < 0.05), while the proportion of cells at S and G(2)/M phase decreased after transfection with nm23-H1-siRNA (S phase:t = 8.356, P < 0.05; G(2)/M phase:t = 7.256, P < 0.05). CONCLUSIONS: Nm23-H1 is related with the chemoresistance to liposome-encapsulated paclitaxel in lung adenocarcinoma cell line A549 cells. Inhibition of the expression of nm23-H1 by nm23-H1-siRNA can improve the in vitro chemosensitivity of A549 cells to liposome-encapsulated paclitaxel.

Gastric metastasis and transformation of primary lung adenocarcinoma to small cell cancer after acquired resistance to epidermal growth factor receptor tyrosine kinase inhibitors: A case report.

RATIONALE: Transformation to small cell lung cancer (SCLC) is one of the mechanisms of resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs). However, no standard treatment is available after the transformation. In addition, gastric metastasis of primary lung cancer is rarely observed; thus, little is known about its metastatic characteristics. PATIENT CONCERNS: A 58-year-old male patient was treated with gefitinib (0.25 g /day) as the 1st line treatment due of recurrence after surgical resection for EGFR exon 19 mutation pulmonary adenocarcinoma. However, he experienced recurrence with positive T790 M, and osimertinib (80 mg/day) was administered as the 2nd line therapy. DIAGNOSIS: One year and 6 months after osimertinib initiation, he complained of stomachache, and a diagnostic gastroscopy biopsy confirmed small cell lung cancer in the gastric body, indicating osimertinib-induced phenotypic transformation. INTERVENTIONS AND OUTCOMES: The patient was treated with etoposide and platinum chemotherapy and maintenance therapy with osimertinib. Finally, the patient achieved a partial response after 4 cycles. LESSONS: Timely second biopsies should be considered in the diagnosis of phenotypic transformation. After transformation, chemotherapeutic treatment with etoposide and platinum and maintenance therapy with osimertinib inhibited the progression of the disease.

Knock down of the dual functional protein apurinic /apyrimidinic endonuclease 1 enhances the killing effect of hematoporphrphyrin derivative-mediated photodynamic therapy on non-small cell lung cancer cells in vitro and in a xenograft model.

Photodynamic therapy (PDT) is considered to be effective treatment for many cancers including lung cancer, head and neck cancers, and prostate cancer. It uses the combination of nontoxic photosensitizers and harmless visible light to generate reactive oxygen species and kill cells. However, DNA repair and reactive oxygen species-induced signaling pathway activation play crucial roles in cellular response to PDT and may also result in therapeutic limitation of PDT. To improve the cancer therapeutic efficacy of PDT, we targeted apurinic/apyrimidinic endonuclease (APE1), which is essential for both DNA repair and redox regulation of gene transcription, as a potential candidate for PDT combined gene therapy. In our study, an adenovirus-mediated APE1 silencing strategy was introduced to test its therapeutic enhancement for the non-small cell lung cancer cell line A549 both in vitro and in vivo after hematoporphrphyrin derivative (HpD)-mediated PDT. The adenovirus vector Ad5/F35-shAPE1 was validated to significantly suppress the protein expression of APE1 in cultured A549 cell and in its xenograft of nude mice. Ad5/F35-shAPE1 effectively inhibited APE1 protein upregulation induced by PDT and resulted in an increase in A549 cell killing by photoirradiation compared with the hematoporphrphyrin derivative-PDT alone group. Ad5/F35-shAPE1 suppressed the DNA repair capacity for single-strand breaks and abolished the activation of some stress-related transcription factors such as hypoxia-induced factor (HIF)-1 that consequently lead to increased cell apoptosis after PDT. Additionally, knock down of APE1 enhanced the tumor suppression efficacy of PDT on the A549 xenograft. Our study indicated that APE1-targeted gene therapy combined with PDT is a promising strategy for enhancement of the efficacy of PDT in treatment of non-small cell lung cancer.

Microarray analysis of DNA damage repair gene expression profiles in cervical cancer cells radioresistant to 252Cf neutron and X-rays.

BACKGROUND: The aim of the study was to obtain stable radioresistant sub-lines from the human cervical cancer cell line HeLa by prolonged exposure to 252Cf neutron and X-rays. Radioresistance mechanisms were investigated in the resulting cells using microarray analysis of DNA damage repair genes. METHODS: HeLa cells were treated with fractionated 252Cf neutron and X-rays, with a cumulative dose of 75 Gy each, over 8 months, yielding the sub-lines HeLaNR and HeLaXR. Radioresistant characteristics were detected by clone formation assay, ultrastructural observations, cell doubling time, cell cycle distribution, and apoptosis assay. Gene expression patterns of the radioresistant sub-lines were studied through microarray analysis and verified by Western blotting and real-time PCR. RESULTS: The radioresistant sub-lines HeLaNR and HeLaXR were more radioresisitant to 252Cf neutron and X-rays than parental HeLa cells by detecting their radioresistant characteristics, respectively. Compared to HeLa cells, the expression of 24 genes was significantly altered by at least 2-fold in HeLaNR cells. Of these, 19 genes were up-regulated and 5 down-regulated. In HeLaXR cells, 41 genes were significantly altered by at least 2-fold; 38 genes were up-regulated and 3 down-regulated. CONCLUSIONS: Chronic exposure of cells to ionizing radiation induces adaptive responses that enhance tolerance of ionizing radiation and allow investigations of cellular radioresistance mechanisms. The insights gained into the molecular mechanisms activated by these "radioresistance" genes will lead to new therapeutic targets for cervical cancer.

Role of PARP1-mediated autophagy in EGFR-TKI resistance in non-small cell lung cancer.

Resistance to epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) has become the main clinical challenge of advanced lung cancer. This research aimed to explore the role of PARP1-mediated autophagy in the progression of TKI therapy. PARP1-mediated autophagy was evaluated in vitro by CCK-8 assay, clonogenic assay, immunofluorescence, and western blot in the HCC-827, H1975, and H1299 cells treated with icotinib (Ico), rapamycin, and AZD2281 (olaparib) alone or in combination. Our results and GEO dataset analysis confirmed that PARP1 is expressed at lower levels in TKI-sensitive cells than in TKI-resistant cells. Low PARP1 expression and high p62 expression were associated with good outcomes among patients with NSCLC after TKI therapy. AZD2281 and a lysosomal inhibitor reversed resistance to Ico by decreasing PARP1 and LC3 in cells, but an mTOR inhibitor did not decrease Ico resistance. The combination of AZD2281 and Ico exerted a markedly enhanced antitumor effect by reducing PARP1 expression and autophagy in vivo. Knockdown of PARP1 expression reversed the resistance to TKI by the mTOR/Akt/autophagy pathway in HCC-827IR, H1975, and H1299 cells. PARP1-mediated autophagy is a key pathway for TKI resistance in NSCLC cells that participates in the resistance to TKIs. Olaparib may serve as a novel method to overcome the resistance to TKIs.

Targeting truncated APE1 in mitochondria enhances cell survival after oxidative stress.

The high steady-state level of mitochondrial DNA (mtDNA) oxidative lesions is assumed to be the result of high susceptibility to DNA damage attack and limited DNA repair capacity in mitochondria. As a key enzyme of base excision repair (BER), human apurinic/apyrimidinic endonuclease (APE1) is often scarce in mitochondria, and mitochondria-targeted APE1 with robust repair activity represents a promising therapeutic candidate. In this study, overexpression vectors of mitochondria-targeted truncated APE1 (mtAPE1) and that of full-length APE1 (flAPE1) were constructed and transfected to human umbilical vein endothelial cells to test their protective effects after hydrogen peroxide-induced oxidative stress. The overexpression of truncated APE1 was achieved at protein and enzyme activity levels in mitochondria of mtAPE1-transfected cells. In parallel, enhanced mtDNA repair capacity and increased cell survival were observed. MtAPE1 transfection also prevented apoptosis by blocking mitochondria-dependent pathways. In contrast, flAPE1 transfection rendered slight elevation of nuclear APE1 protein level and nuclear APE activity but no benefits for cell resistance to oxidative stress. The present results suggest that overexpression of the truncated APE1 in mitochondria appears to be a viable approach to protecting healthy cells from some deleterious effects of oxidative stress.

Lonicera japonica extends lifespan and healthspan in Caenorhabditis elegans.

Lonicera japonica (LJ) is widely used as the local medicine to improve body and prevent ills in China, but mechanisms of its healthy beneficial effects remain largely unclear. Here, we evaluated the anti-aging and healthspan promoting activities of 75% ethanol extract of LJ (LJ-E) in the animal model Caenorhabditis elegans. Our results showed that LJ-E (500 µg/mL) treatment enhanced the mean lifespan of worms by over 21.87% and significantly improved age-associated physiological functions in C. elegans. The 500 µg/mL concentration of LJ-E enhanced the survival rates under oxidative and thermal stresses, and decreased reactive oxygen species (ROS) levels and fat accumulation in the worms. Gene-specific mutant studies showed that LJ-E-mediated lifespan extension was dependent on mev-1, daf-2, daf-16, and hsf-1, but not eat-2 genes. LJ-E could upregulate stress-inducible genes, viz., hsp-16.2, sod-3 and mtl-1. Moreover, we found that the D1086.10 protein interacted with superoxide dismutase (SOD)-3 by functional protein association networks analysis according to RNA-sequencing results. It was confirmed that D1086.10 was needed to promote longevity, and positively regulated expression of sod-3 by using D1086.10 mutants. Furthermore, LJ-E significantly delayed amyloid ß-protein induced paralysis in CL4176 strain. Given the important role of autophagy in aging and protein homeostasis, we observed that LJ-E could remarkably increase the mRNA expression of autophagy gene bec-1 in CL4176 strain, and decrease expression of autophagy substrate p62 protein by more than 40.0% in BC12921 strain. Finally, we found that combination composed of three major compounds (54 µg/mL chlorogenic acid, 15 µg/mL 1,5-dicaffeoylquinic acid and 7.5 µg/mL 1,3-dicaffeoylquinic acid) of 500 µg/mL LJ-E could significantly delay paralysis in CL4176 worms caused by Aß toxicity, comparable to that of LJ-E. Overall, our study may have important implications in using Lonicera japonica to promote healthy aging and have a potency to design therapeutics for age-related diseases.

Genistein promotes ionizing radiation-induced cell death by reducing cytoplasmic Bcl-xL levels in non-small cell lung cancer.

Genistein (GEN) has been previously reported to enhance the radiosensitivity of cancer cells; however, the detailed mechanisms remain unclear. Here, we report that GEN treatment inhibits the cytoplasmic distribution of Bcl-xL and increases nuclear Bcl-xL in non-small cell lung cancer (NSCLC). Interestingly, our in vitro data show that ionizing radiation IR treatment significantly increases IR-induced DNA damage and apoptosis in a low cytoplasmic Bcl-xL NSCLC cell line compared to that of high cytoplasmic Bcl-xL cell lines. In addition, clinical data also show that the level of cytoplasmic Bcl-xL was negatively associated with radiosensitivity in NSCLC. Furthermore, we demonstrated that GEN treatment enhanced the radiosensitivity of NSCLC cells partially due to increases in Beclin-1-mediated autophagy by promoting the dissociation of Bcl-xL and Beclin-1. Taken together, these findings suggest that GEN can significantly enhance radiosensitivity by increasing apoptosis and autophagy due to inhibition of cytoplasmic Bcl-xL distribution and the interaction of Bcl-xL and Beclin-1 in NSCLC cells, respectively.

Experimental study enhancing the chemosensitivity of multiple myeloma to melphalan by using a tissue-specific APE1-silencing RNA expression vector.

PURPOSE: Because of a developing resistance to chemotherapy agents, multiple myeloma (MM) has been an incurable disease until now. As a means to overcome MM tumor cell resistance and/or sensitize tumor cells to chemotherapeutic treatments currently used, we examined the role of human apurinic/apyrimidinic endonuclease 1 (APE1) in resistance and prognosis in patients with MM. PATIENTS AND METHODS: Multiple myeloma cells were analyzed by using bone marrow specimens from 32 patients with MM and 10 normal volunteers. RESULTS: The positive rate of APE1 protein expression was 65.6% in the bone marrow specimens of patients with MM with known clinical outcome. Positive rate of APE1 expression beyond grade 2 in the relapsed/refractory group was significantly higher than that in the untreated group. No positive results of grade > 2 were detected in bone marrow specimens from patients with noncancerous disease. It was also confirmed that the amount of APE1 protein in KM3 cells was positively correlated with the dose and action time of melphalan. Because APE1 was overexpressed in refractory/relapsed MM cells, siRNA-targeted technology was used to decrease APE1 levels in KM3 cells, with protein levels deceasing to 80%-90% within 24 hours and continuing to decease for 72 hours. The best dose and time of inhibiting expression of APE1 protein were 3 mug and 2 days long. A decrease in APE1 levels in siRNA-treated KM3 cells led to enhanced cell sensitization to melphalan. CONCLUSION: The findings herein present prognostic and therapeutic implications for treating relapsed/refractory MM. The APE1-silencing RNA results demonstrate the feasibility of the therapeutic modulation of APE1 using a variety of molecules and approaches.

miR-135b Stimulates Osteosarcoma Recurrence and Lung Metastasis via Notch and Wnt/ß-Catenin Signaling.

Cancer stem cells (CSCs) play an important role in osteosarcoma (OS) metastasis and recurrence, and both Wnt/ß-catenin and Notch signaling are essential for the development of the biological traits of CSCs. However, the mechanism that underlies the simultaneous hyperactivation of both Wnt/ß-catenin and Notch signaling in OS remains unclear. Here, we report that expression of miR-135b correlates with the overall and recurrence-free survival of OS patients, and that miR-135b has an activating effect on both Wnt/ß-catenin and Notch signaling. The overexpression of miR-135b simultaneously targets multiple negative regulators of the Wnt/ß-catenin and Notch signaling pathways, including glycogen synthase kinase-3 beta (GSK3ß), casein kinase 1a (CK1α), and ten-eleven translocation 3 (TET3). Therefore, upregulated miR-135b promotes CSC traits, lung metastasis, and tumor recurrence in OS. Notably, antagonizing miR-135b potently inhibits OS lung metastasis, cancer cell stemness, CSC-induced tumor formation, and recurrence in xenograft animal models. These findings suggest that miR-135b mediates the constitutive activation of Wnt/ß-catenin and Notch signaling, and that the inhibition of miR-135b is a novel strategy to inhibit tumor metastasis and prevent CSC-induced recurrence in OS.

APE1 promotes antioxidant capacity by regulating Nrf-2 function through a redox-dependent mechanism.

APE1 is a multifunctional protein that has recently been implicated in protecting cells from oxidative stress. In the current study, we confirmed that APE1׳s effect on cellular antioxidant capacity is related to its redox activity through the use of an APE1 functional mutant, and we investigated the mechanism through which this multifunctional protein affects the function of the transcription factor Nrf-2 in regulating oxidative stress-induced genes. Using a pair of mutants for both the redox activity and the acetylation-regulated activity of APE1, in vitro assays showed that the redox activity of APE1 is crucial for its nuclear association with Nrf-2 and subsequent activation of Nrf-2׳s transcription of several downstream genes during oxidative challenge. Important oxidative stress genes are affected by APE1 redox activity, including Hmox1, Gstm1, and Txnrd1. In addition, utilizing human non-small-cell lung cancer sample tissue as well as a nude mouse xenograft model, we determined that APE1 expression levels are inversely correlated to oxidative stress in vivo. These findings indicated that interference with these crucial functions of APE1 shows promise in preventing resistance to certain radiotherapies and that further research is necessary to understand APE1׳s complex roles in regulating both the basal redox status and the oxidative stress state of the cellular environment.

LDA-SVM-based EGFR mutation model for NSCLC brain metastases: an observational study.

Epidermal growth factor receptor (EGFR) activating mutations are a predictor of tyrosine kinase inhibitor effectiveness in the treatment of non-small-cell lung cancer (NSCLC). The objective of this study is to build a model for predicting the EGFR mutation status of brain metastasis in patients with NSCLC. Observation and model set-up. This study was conducted between January 2003 and December 2011 in 6 medical centers in Southwest China. The study included 31 NSCLC patients with brain metastases. Eligibility requirements were histological proof of NSCLC, as well as sufficient quantity of paraffin-embedded lung and brain metastases specimens for EGFR mutation detection. The linear discriminant analysis (LDA) method was used for analyzing the dimensional reduction of clinical features, and a support vector machine (SVM) algorithm was employed to generate an EGFR mutation model for NSCLC brain metastases. Training-testing-validation (3 : 1 : 1) processes were applied to find the best fit in 12 patients (validation test set) with NSCLC and brain metastases treated with a tyrosine kinase inhibitor and whole-brain radiotherapy. Primary and secondary outcome measures: EGFR mutation analysis in patients with NSCLC and brain metastases and the development of a LDA-SVM-based EGFR mutation model for NSCLC brain metastases patients. EGFR mutation discordance between the primary lung tumor and brain metastases was found in 5 patients. Using LDA, 13 clinical features were transformed into 9 characteristics, and 3 were selected as primary vectors. The EGFR mutation model constructed with SVM algorithms had an accuracy, sensitivity, and specificity for determining the mutation status of brain metastases of 0.879, 0.886, and 0.875, respectively. Furthermore, the replicability of our model was confirmed by testing 100 random combinations of input values. The LDA-SVM-based model developed in this study could predict the EGFR status of brain metastases in this small cohort of patients with NSCLC. Further studies with larger cohorts should be carried out to validate our findings in the clinical setting.

Genistein alleviates radiation-induced pneumonitis by depressing Ape1/Ref-1 expression to down-regulate inflammatory cytokines.

The aim of the study was to investigate the role of genistein in alleviating radiation-induced pneumonitis(RIP) through down-regulating levels of the inflammatory cytokines by inhibiting the expression of apurinic/apyrimidinic endonuclease 1/redox factor-1 (Ape1/Ref-1). Fifty female C57BL/6J mice (8 weeks old) were randomly divided into a control group, a pure irradiation (IR) group and a genistein + IR group. At the four time points after IR, hematoxylin, and Masson's trichrome stainings were used to examine the pathological changes and collagen fiber deposition. Flow cytometry was used to detect reactive oxygen system (ROS) changes, EMSA was used to estimate the nuclear factor kappa B (NF-κB) transcriptional activities and an ELISA assay was used to measure the levels of TGF-ß1, IL-1ß, TNF-α, and IL-6 in the serum and bronchoalveolar lavage fluid (BALF) 2 weeks after IR.The pathological detection results showed acute inflammatory/fibrinoid exudation of the thoracic tissue after IR,which was significantly alleviated with genistein. The IR inducedan APE1 protein expression increase and NF-jB was effectively suppressed by genistein (P < 0.05). The induction of the inflammatory cytokines TGF-ß1, IL-1ß,TNF-α, and IL-6 by IR were in turn inhibited in the serum and BALF of the genistein-pretreated mice (P < 0.05). In addition, the ROS production was significantly boosted in the A549 cells after IR, which could be down-regulated by the pretreatment of genistein. The results demonstrate that genistein alleviates RIP by attenuating the inflammatory response in the initiation of RIP. A possible target of genistein is the Ape1/ref-1, which regulates key inflammatory cytokines by activating the NF-κB.