Cisplatin-resistant Lung Cancer Research Articles

Comprehensive analysis of circular RNA expression profiles in cisplatin-resistant non-small cell lung cancer cell lines.

Platinum-based drugs such as cisplatin are widely used in combination chemotherapy for non-small cell lung cancer (NSCLC) owing to their high clinical response rate; however, acquired resistance to cisplatin is eventually inevitable. Circular RNAs (circRNAs) are involved in the development of diverse types of cancers, but their connection to cisplatin-resistance in NSCLC has not been studied. In the present study, two cisplatin-resistant NSCLC cell lines (A549/DDP and PC9/DDP) were established by gradually increasing concentrations of cisplatin in the media. The resulting cell lines possessed high resistance to cisplatin and strong proliferation, migration, and colony formation abilities compared to the parental cells. Microarray analysis identified 19,161 circRNAs that were dysregulated in cisplatin-resistant cell lines (fold change abs>2), including 11,915 up-regulated and 7246 down-regulated circRNAs. The expression of the top five up-regulated and down-regulated circRNAs was validated using real-time quantitative polymerase chain reaction. A circRNA-micro RNA (miRNA) network of the top 20 dysregulated circRNAs and their predicted miRNAs was constructed using Cytoscape. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed that the host genes of the identified circRNAs were involved in the regulation of MAP kinase kinase kinase kinase activity, 6-phosphofructo-2-kinase activity, focal adhesion, ErbB signaling, and ECM-receptor interactions, which may contribute to cisplatin resistance in NSCLC. In summary, this is the first report on circRNA profiling in cisplatin-resistant NSCLC cells and it provides new potential targets for the reversal of cisplatin resistance in NSCLC.

Shenmai Injection Supresses Glycolysis and Enhances Cisplatin Cytotoxicity in Cisplatin-Resistant A549/DDP Cells via the AKT-mTOR-c-Myc Signaling Pathway.

Tumor cells, especially drug-resistant cells, predominately support growth by glycolysis even under the condition of adequate oxygen, which is known as the Warburg effect. Glucose metabolism reprogramming is one of the main factors causing tumor resistance. Previous studies on Shenmai injection (SMI), a Chinese herbal medicine, have shown enhanced efficacy in the treatment of tumors in combination with chemotherapy drugs, but the mechanism is not clear. In this study, we investigated the effect of SMI combined with cisplatin on cisplatin-resistant lung adenocarcinoma A549/DDP cells. Our results showed that cisplatin-resistant A549/DDP cells exhibited increased glucose consumption, lactate production, and expression levels of key glycolytic enzymes, including hexokinase 2 (HK2), pyruvate kinase M1/2 (PKM1/2), pyruvate kinase M2 (PKM2), glucose transporter 1 (GLUT1), and lactate dehydrogenase A (LDHA), compared with cisplatin-sensitive A549 cells. SMI combined with cisplatin in A549/DDP cells, led to significantly lower expression levels of key glycolytic enzymes, such as HK2, PKM1/2, GLUT1, and pyruvate dehydrogenase (PDH). In addition, we found that the combination of SMI and cisplatin could inhibit cell proliferation and promote apoptosis by reducing the expression levels of p-Akt, p-mTOR, and c-Myc, and then, it reduced the glycolysis level. These results suggest that SMI enhances the antitumor effect of cisplatin via glucose metabolism reprogramming. Therefore, the combination of SMI and cisplatin may be a potential therapeutic strategy to treat cisplatin-resistant nonsmall cell lung cancer.

Knockdown of ubiquitin‑specific protease 51 attenuates cisplatin resistance in lung cancer through ubiquitination of zinc‑finger E‑box binding homeobox 1.

Lung cancer is a devastating cancer with high morbidity and mortality. Ubiquitin-specific protease (USP) is a type of deubiquitinating enzyme (DUB) that has been implicated in numerous cancers, including colorectal, myeloma and breast. In the present study, the expression of USP51 was determined in the lung cancer cell line A549 and cisplatin (also known as DDP)-resistant lung cancer strain A549/DDP. The expression of zinc-finger E-box binding homeobox 1 (ZEB1), a transcriptional repressor, was also examined. The effects of USP51 knockdown or overexpression on proliferation and apoptosis, as well as the impact of ZEB1 overexpression and USP51 interference on apoptosis and ubiquitination were then assessed. Notably, increased expression of USP51 and ZEB1 in A549/DDP cells was observed, and treatment with DDP significantly inhibited proliferation in A549/DDP cells. In addition, knockdown of USP51 in A549/DDP cells significantly induced apoptosis, decreased ZEB1 expression and increased cleaved poly ADP-ribose polymerase 1 (PARP1) and cleaved caspase-3 levels. Consistently, USP51 overexpression in A549 cells displayed the opposite effects and potently attenuated DDP-induced apoptosis. Notably, overexpression of ZEB1 in A549/DDP cells potently attenuated the effects of USP51 knockdown on apoptosis, and co-IP experiments further demonstrated interaction between USP51 and ZEB. Lastly, knockdown of USP51 promoted ZEB1 ubiquitination, leading to ZEB1 degradation. Collectively, the present findings demonstrated that USP51 inhibition attenuated DDP resistance in A549/DDP cells via ubiquitin-mediated degradation of ZEB1. Hence, targeting USP51 may serve as a novel therapeutic target for DDP resistance in lung cancer.

Evaluating the immunoproteasome as a potential therapeutic target in cisplatin-resistant small cell and non-small cell lung cancer.

We evaluated the expression of proteasome subunits to assess whether the proteasome could be a therapeutic target in cisplatin-resistant lung cancer cells. Cisplatin-resistant (CR) variants were established from three non-small cell lung cancer (NSCLC) cell lines (A549, H1299, and H1975) and two small cell lung cancer (SCLC) cell lines (SBC3 and SBC5). The expression of proteasome subunits, the sensitivity to immunoproteasome inhibitors, and 20S proteasomal proteolytic activity were examined in the CR variants of the lung cancer cell lines. All five CR cell lines highly expressed one or both of the immunoproteasome subunit genes, PSMB8 and PSMB9, while no clear trend was observed in the expression of constitutive proteasome subunits. The CR cells expressed significantly higher levels of PSMB8 and PSMB9 proteins, as well. The CR variants of the H1299 and SBC3 cell lines were more sensitive to immunoproteasome inhibitors, and had significantly more proteasomal proteolytic activity than their parental counterparts. The immunoproteasome may be an effective therapeutic target in a subset of CR lung cancers. Proteasomal proteolytic activity may be a predictive marker for the efficacy of immunoproteasome inhibitors in cisplatin-resistant SCLC and NSCLC.

Enhanced histone H3 acetylation of the PD-L1 promoter via the COP1/c-Jun/HDAC3 axis is required for PD-L1 expression in drug-resistant cancer cells

BackgroundDrug resistance is a major obstacle to treating cancers because it desensitizes cancer cells to chemotherapy. Recently, attention has been focused on changes in the tumor immune landscape after the acquisition of drug resistance. Programmed death-ligand-1 (PD-L1) is an immune suppressor that inhibits T cell-based immunity. Evidence has shown that acquired chemoresistance is associated with increased PD-L1 expression in cancer cells. However, the underlying mechanism is still largely unknown.MethodsPD-L1 expression in three drug-resistant A549/CDDP, MCF7/ADR and HepG2/ADR cell lines was detected by qRT-PCR, western blotting and flow cytometry, and a T cell proliferation assay was performed to test its functional significance. Then, the potential roles of JNK/c-Jun, histone H3 acetylation, histone deacetylase 3 (HDAC3) and the E3 ligase COP1 in the PD-L1 increase were explored through ChIP assays and gain- and loss-of-function gene studies. Furthermore, murine xenograft tumor models were used to verify the role of JNK/c-Jun and HDAC3 in PD-L1 expression in A549/CDDP cells in vivo. Finally, the correlations of PD-L1, c-Jun and HDAC3 expression in clinical cisplatin-sensitive and cisplatin-resistant non-small cell lung cancer (NSCLC) tissues were analyzed by immunohistochemistry and Pearson’s correlation coefficient.ResultsPD-L1 expression was significantly increased in A549/CDDP, MCF7/ADR and HepG2/ADR cells and was attributed mainly to enhanced JNK/c-Jun signaling activation. Mechanistically, decreased COP1 increased c-Jun accumulation, which subsequently inhibited HDAC3 expression and thereby enhanced histone H3 acetylation of the PD-L1 promoter. Furthermore, PD-L1 expression could be inhibited by JNK/c-Jun inhibition or HDAC3 overexpression in vivo, which could largely reverse inhibited CD3+ T cell proliferation in vitro. PD-L1 expression was significantly increased in the cisplatin-resistant clinical NSCLC samples and positively correlated with c-Jun expression but negatively correlated with HDAC3 expression.ConclusionsEnhanced histone H3 acetylation of the PD-L1 promoter via the COP1/c-Jun/HDAC3 axis was crucial for the PD-L1 increase in drug-resistant cancer cells. Our study reveals a novel regulatory network for the PD-L1 increase in drug-resistant cancer cells and that combined PD-L1-targeting strategies could improve T cell-based immunity in drug-resistant cancers.

Rosmarinic acid reverses non-small cell lung cancer cisplatin resistance by activating the MAPK signaling pathway.

Cisplatin (DDP) is one of the first‐line chemotherapeutic agents for non‐small cell lung cancer (NSCLC). However, repeated use of cisplatin in clinical practice often induces chemoresistance. The aims of this study were to investigate whether rosmarinic acid (RA) could reverse multidrug resistance (MDR) in NSCLC and to explore the underlying mechanisms. Our data demonstrated that RA significantly inhibited NSCLC cell proliferation and cell colony formation in a dose‐dependent manner, induced G1 phase cell cycle arrest and apoptosis, and increased the sensitivity of cell lines resistant to DDP. Mechanistically, RA inhibited NSCLC cell growth, arrested cell cycle, and induced apoptosis by activating MAPK and inhibiting the expression of P‐gp and MDR1, which correspondingly enhanced p21 and p53 expression. We observed that the growth of xenograft tumors derived from NSCLC cell lines in nude mice was significantly inhibited by combination therapy. We demonstrate that RA is a potentially effective MDR reversal agent for NSCLC, based on downregulation of MDR1 mRNA expression and P‐gp. Together, these results emphasize the putative role of RA as a resistance reversal agent in NSCLC.

Targeting the Kynurenine Pathway for the Treatment of Cisplatin-Resistant Lung Cancer.

Cisplatin resistance is a major barrier in the effective treatment of lung cancer. Cisplatin-resistant (CR) lung cancer cells do not primarily use glucose but rather consume amino acids such as glutamine and tryptophan (Trp) for survival. CR cells activate the kynurenine (KYN) pathway (KP) to cope with excessive reactive oxygen species (ROS) and maintain homeostasis for growth and proliferation. Consequently, indoleamine 2,3-dioxygenase-1 (IDO1) becomes an essential enzyme for CR cells' survival because it initiates and regulates the first step in the KP. Increased IDO1 activities and ROS levels are found in CR cells versus cisplatin-sensitive lung cancer. Importantly, significantly greater KYN/Trp ratio (P = 0.005) is detected in serum of patients who fail cisplatin when compared with naïve treatment. Knocking down IDO1 using shRNA or IDO1 inhibitors heightens ROS levels and results in a significant growth inhibitory effect only on CR cells and not on cisplatin-sensitive cells. Exposing CR cells to antioxidant (TIRON) results in suppression of IDO1 activity and confers resistance to IDO1 inhibition, indicating an interrelationship between ROS and IDO1. Because KYN plays a critical role in reprogramming naïve T cells to the immune-suppressive regulatory T-cell (T-reg) phenotype, we observed higher expression of TGFβ, FoxP3, and CD4+CD25+ in mice bearing CR tumors compared with tumors from cisplatin-sensitive counterparts. IMPLICATIONS: Findings suggest that the enzyme-inhibitory activity and antitumor efficacy of IDO1 inhibitors rely in part on ROS levels, arguing that IDO1 expression alone may be insufficient to determine the clinical benefits for this class of experimental cancer drugs. Importantly, IDO1 inhibitors may be more suitable to treat patients with lung cancer who failed cisplatin therapy than naïve treatment patients.

Heterobimetallic propargyl gold complexes with π-bound copper or silver with enhanced anticancer activity.

Several propargyl functionalised substrates with different heteroatoms (N, O or S) have been used for the preparation of propargyl gold(i) phosphine complexes. The complexes were prepared in high yields either by reaction of the substrate with [Au(acac)PPh3] or by reaction of [AuCl(PPh3)] with potassium hydroxide and the substrate in methanol. Several of the complexes have been characterised by X-ray diffraction showing the presence of secondary bonds such as π-stacking and aurophilic interactions. The reaction of the propargyl gold(i) phosphine complexes with [Cu(NO3)(PPh3)2] or [Ag(OTf)(PPh3)2] afforded heterobimetallic complexes with π-coordination of {Cu(PPh3)2} or {Ag(PPh3)2} to the alkyne bond. When the substituent of the propargyl unit contained more strongly coordinating pyridine moieties, [(PyCH2)2NCH2C[triple bond, length as m-dash]CAuPPh3], coordination of the heterometal to the pyridine units occurred, displacing the phosphine groups and giving rise to a dimeric structure. The antiproliferative activity of the complexes against cisplatin resistant lung cancer cell line A549 was determined by MTT assay. The mononuclear gold complexes showed excellent activities with IC50 values < 14 μM. Coordination of copper of silver to the alkynyl fragment resulted in a significant increase in activity suggesting a synergistic effect between the two metal centres.

Cytoplasm-localized SIRT1 downregulation attenuates apoptosis and cell cycle arrest in cisplatin-resistant lung cancer A549 cells.

Objective: We propose that sirtuin (SIRT) may induce a pro-apoptotic effect by deacetylating transcription factors in A549 cells: depletion of sirtuin-1 (SIRT1) induced cell cycle arrest in cisplatin-resistant A549 (A549/CADD) cells.Methods: Protein and mRNA levels of SIRT1 were investigated using western blot and RT-PCR. In A549 and A549/CADD cells, the cytotoxicity of cisplatin administration was evaluated by MTT assay, proliferation was measured by ECIS, and the cell cycle distribution was analyzed using FACS. Cells were transfected with pcDNA3.1-Myc-SIRT1 or pcDNA3.1-Myc-Control vectors to analyze the impact of SIRT-1 on cisplatin induced drug resistance. SIRT1 localization was studied using immunofluorescence analysis. In addition, immunoprecipitation and 20S proteasome activity assay were performed to examine the relationship of SIRT1 with the proteasome complex.Results: A549/CADD cells exhibited a mesenchymal-like cell characteristic. SIRT1 expression was markedly decreased in A549/CADD cells. We observed that cisplatin regulates p53 stability through the depletion of ubiquitination following SIRT1 downregulation. Furthermore, cisplatin treatment increased proteasomal activity and significantly decreased cytoplasmic SIRT1 protein levels in A549/CADD cells.Conclusion: In this study, we found SIRT1 to be depleted in A549/CADD cells and also determined the underlying resistance mechanism which may act as novel therapeutic targets in overcoming drug resistance.

Differential Expression And Effects Of Peroxiredoxin-6 On Drug Resistance And Cancer Stem Cell-Like Properties In Non-Small Cell Lung Cancer.

ObjectiveCancer stem-like cells (CSC) are thought to be involved in the cisplatin resistance of tumors. This study was designed to investigate the effect of PRDX6 on CSCs present in cisplatin-resistant non-small cell lung cancer (NSCLC) tumors.Materials and methodsCD133+/ABCG2+ H1299 CSCs and A549 CSCs were isolated. The IC50 values for cisplatin in treatment of CSCs were detected using the CCK8 assay. Then the isolated cells were identified using CD133. Wnt/β-catenin expression was evaluated by Western blot assays. Specimens of tumor and adjacent para-carcinoma tissue were collected from 30 NSCLC patients and examined by immunohistochemistry (IHC), qRT-PCR, and Western blotting to determine and compare their levels of PRDX6 and CD133 expression. Finally, siRNA-mediated silencing of PRDX6 was employed with both types of CSCs to determine the impact of PRDX6 on CD133 enrichment by flow cytometry, cell viability, and sphere formation ability.ResultsHigh levels of PRDX6 and CD133 expression were detected in samples of tumor tissue from NSCLC patients, and expression of PRDX6 and CD13 presented a positive relationship. Increasing levels of cisplatin resistance and upregulated levels of PRDX6, ABCG2, Wnt, and β-catenin expression were detected in CD133+/ABCG2+ H1299 and A549 CSCs. Transfection with siRNA targeting PRDX6 changed these cellular characteristics by decreasing the levels of PRDX6, ABCG2, Wnt, and β-catenin expression. We further demonstrated that exogenous silencing of PRDX6 effectively inhibited the sphere formation ability of CSCs and re-sensitized them to cisplatin.ConclusionOur results strongly suggest that PRDX6 promotes cisplatin resistance in human lung cancer cells by promoting the stem-like properties of cancer cells. Our findings also suggest PRDX6 as a target for treating cisplatin resistant NSCLC.

Cisplatin-resistant A549 non-small cell lung cancer cells can be identified by increased mitochondrial mass and are sensitive to pemetrexed treatment

BackgroundCisplatin plus pemetrexed combination therapy is considered the standard treatment for patients with advanced, non-squamous, non-small-cell lung cancer (NSCLC). However, advanced NSCLC has a 5-year survival rate of below 10%, which is mainly due to therapy resistance. We previously showed that the NSCLC cell line A549 harbors different subpopulations including a mesenchymal-like subpopulation characterized by increased chemo- and radiotherapy resistance. Recently, therapy resistance in hematological and solid tumors has been associated with increased mitochondrial activity. Thus, the aim of this study was to investigate the role of the mitochondrial activity in NSCLC chemotherapy resistance.MethodsBased on MitoTracker staining, subpopulations characterized by the highest 10% (Mito-High) or lowest 10% (Mito-Low) mitochondrial mass content were sorted by FACS (Fluorescence-Activated Cell Sorting) from paraclonal cultures of the NSCLC A549 cell line . Mitochondrial DNA copy numbers were quantified by real-time PCR whereas basal cellular respiration was measured by high-resolution respirometry. Cisplatin and pemetrexed response were quantified by proliferation and colony formation assay.ResultsPemetrexed treatment of parental A549 cells increased mitochondrial mass over time. FACS-sorted paraclonal Mito-High cells featured increased mitochondrial mass and mitochondrial DNA copy number compared to the Mito-Low cells. Paraclonal Mito-High cells featured an increased proliferation rate and were significantly more resistant to cisplatin treatment than Mito-Low cells. Interestingly, cisplatin-resistant, paraclonal Mito-High cells were significantly more sensitive to pemetrexed treatment than Mito-Low cells. We provide a working model explaining the molecular mechanism underlying the increased cisplatin- and decreased pemetrexed resistance of a distinct subpopulation characterized by high mitochondrial mass.ConclusionsThis study revealed that cisplatin resistant A549 lung cancer cells can be identified by their increased levels of mitochondrial mass. However, Mito-High cells feature an increased sensitivity to pemetrexed treatment. Thus, pemetrexed and cisplatin target reciprocal lung cancer subpopulations, which could explain the increased efficacy of the combination therapy in the clinical setting.

1-Hydroxy-8-methoxy-anthraquinon Reverses Cisplatin Resistance by Inhibiting 6PGD in Cancer Cells.

Targeting 6-phosphogluconate dehydrogenase (6PGD) can inhibit cancer cell proliferation and tumor growth. However, the relationship between 6PGD and cisplatin resistance still needs further study. Cisplatin-sensitive and cisplatin-resistant ovarian cancer OV2008 and C13* lines and lung cancer A549 and A549DDP lines were treated with different concentrations of cisplatin and cell viability was evaluated. We also compared the growth rates and the cell cycle distributions between cisplatin-sensitive and cisplatin-resistant cells. The expression level of 6PGD was detected by immunoblotting. The Chou-Talalay method was used to evaluate the effect of a combination treatment using cisplatin and the small molecule inhibitor 1-Hydroxy-8-methoxy-anthraquinon (S3) that targets 6PGD. The cisplatin-resistant ovarian and lung cancer cell lines grew faster than the cisplatin- sensitive cell lines, with more cells in S and G2 phases in cisplatin-resistant cell lines. The expression level of 6PGD in cisplatin-resistant cell lines was significantly increased compared with cisplatin-sensitive cell lines. Furthermore, treatment of cells with the S3 small molecule inhibitor of 6PGD together with cisplatin could overcome cisplatin resistance. The expression level of 6PGD in cisplatin-resistant cells lines was significantly upregulated, and the resistance to cisplatin of drug-resistant cells lines could be overcome when treated with the small molecule inhibitor S3 that specifically targets 6PGD.

Erastin/sorafenib induces cisplatin-resistant non-small cell lung cancer cell ferroptosis through inhibition of the Nrf2/xCT pathway.

Non-small cell lung cancer (NSCLC) has long been one of the most lethal types of cancer due to its lack of typical clinical symptoms at early stages and high risk of tumour recurrence, even following complete surgical resection. Multicourse chemotherapy based on cisplatin (CDDP) is the standard adjuvant treatment for NSCLC; however, its benefits for the overall survival of patients are limited. In this study, NSCLC cells possessing CDDP-resistant characteristics (N5CP cells), obtained from surgical resection of clinical specimens of patients with NSCLC, were cultured and screened to generate research models. This study aimed to identify the mechanism underlying tumour cell resistance to CDDP and to identify a novel treatment for NSCLC following CDDP failure. CDDP-mediated NF-E2 related factor 2 (Nrf2)/light chain of System xc− (xCT) pathway activation was associated with the resistance of cells to CDDP. Therefore, erastin/sorafenib regulation of Nrf2 or xCT expression may alter the sensitivity of tumour cells to CDDP. The small molecules erastin and sorafenib effectively induced N5CP cell ferroptosis, which was mediated by the accumulation of intracellular lipid reactive oxygen species. Additionally, low doses of erastin or sorafenib could be used in association with CDDP to effectively trigger N5CP cell ferroptosis. Furthermore, it was indicated that erastin and sorafenib, alone or in combination with a low dose of CDDP, effectively inhibited the growth of N5CP cells in vivo. Therefore, ferroptosis inducers, including erastin and sorafenib, may be considered a novel treatment regimen for patients with NSCLC, particularly patients with CDDP failure.

Mutated p53 Promotes the Symmetric Self-Renewal of Cisplatin-Resistant Lung Cancer Stem-Like Cells and Inhibits the Recruitment of Macrophages.

It has been proposed that mutant p53 is correlated with the recurrence of lung cancer. Recently, a small population of cells with asymmetric or symmetric self-renewal potential has been identified in lung cancer, which was termed as cancer stem-like cells (CSCs) and was speculated to be the reason for cancer recurrence after chemotherapy. In this study, we used lung cancer cell lines with different TP53 backgrounds to elucidate the potential role of mutant p53 in regulating lung CSC self-renewal and on lung cancer recurrence. Cisplatin-resistant lung cancer cells with different TP53 backgrounds were generated in vitro by exposing A549, H460, and H661 lung cancer cell lines repeatedly to cisplatin. CD44+/CD90+ stem-like cells were identified in above cisplatin-resistant lung cancers (termed as cisplatin-resistant lung cancer stem-like cells, (Cr-LCSCs)) and stained with PKH26 dye which was used to define the self-renewal pattern. The proportion of symmetric divisions was significantly higher in Cr-LCSCs with mutant (mt) p53 compared with Cr-LCSCs with wild-type (wt) p53, and forced expression of mt p53 promoted the symmetric division of Cr-LCSCs. Furthermore, fewer macrophages accumulated in subcutaneously implanted xenografts consisting of mt p53 Cr-LCSCs compared with wt p53 Cr-LCSCs. These results indicated that mt p53 might accelerate the recurrence of lung cancer by regulating the self-renewal kinetics of Cr-LCSCs as well as the recruitment of macrophages.

SHCBP1 promotes cisplatin induced apoptosis resistance, migration and invasion through activating Wnt pathway

Lung cancer is the leading cause for cancer death due to refractory nature to current treatment strategies, understanding the regulatory mechanism of therapy resistance of lung cancer is important for lung cancer therapy. Here, we aimed to study the role of SHCBP1 in lung cancer cisplatin resistance, we found SHCBP1 was upregulated in lung cancer tissues and cells, patients with high SHCBP1 had poor prognosis. SHC binding and spindle associated 1 (SHCBP1) overexpression promoted cisplatin induced apoptosis resistance, migration and invasion determined by apoptosis assay and transwell assay with or without Matrigel, while SHCBP1 knockdown inhibited cisplatin induced apoptosis resistance, migration and invasion. Wnt pathway promoted lung cancer progression, we found SHCBP1 activated Wnt pathway, characterized by promoting β-catenin nuclear translocation. Inhibition of Wnt pathway in SHCBP1 overexpression cells reversed the effect of SHCBP1 overexpression, confirming SHCBP1 promoted lung cancer progression through activating Wnt pathway. We also found SHCBP1 expression was positively corrected with Wnt pathway activity in lung cancer samples. In summary, we found SHCBP1 promoted cisplatin induced apoptosis resistance, migration and invasion through activating Wnt pathway, providing a potential target for lung cancer therapy.

SNHG7 mediates cisplatin-resistance in non-small cell lung cancer by activating PI3K/AKT pathway.

The aim of this study was to clarify the function of long noncoding ribonucleic acids (lncRNAs) small nucleolar RNA host gene 7 (SNHG7) in cisplatin-resistant non-small cell lung cancer (NSCLC), and to explore the potential mechanism. SNHG7 expression in NSCLC and para-cancerous tissues was determined by quantitative real-time polymerase chain reaction (qRT-PCR). Meanwhile, the correlation between SNHG7 expression with clinical stage and cisplatin-resistance in NSCLC patients was analyzed. After transfection of si-SNHG7 or p-complementary deoxyribonucleic acid (pcDNA)-SNHG7, changes in cellular behaviors of A549/DDP cells were evaluated, including cell viability, apoptosis, migration, invasion and cell cycle. The regulatory effects of SNHG7 on the expressions of genes were determined by qRT-PCR as well. Furthermore, Western blot was conducted to determine the protein expressions of drug-resistance genes minimal residual disease1 (MRD1), P-glycoprotein (P-gp), BCRP and relative genes in phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway. Compared with adjacent normal tissues, SNHG7 was highly expressed in NSCLC tissues. Moreover, SNHG7 expression was significantly higher in advanced-stage NSCLC patients than those in early-stage. SNHG7 level remained significantly higher in DDP-resistant NSCLC tissues and cell lines as well. Knockdown of SNHG7 remarkably enhanced cisplatin-resistance in NSCLC cells, manifesting as decreased cell viability, migratory and invasive rates, DNA synthesis capacity, and promoted apoptosis. Meanwhile, SNHG7 knockdown down-regulated the mRNA levels of matrix metalloprotein2 (MMP2), MMP7 and MMP9 in vitro. After SNHG7 knockdown, the expressions of drug-resistant and relative genes in the PI3K/AKT pathway were notably down-regulated. SNHG7 induces the development of cisplatin-resistance in NSCLC through upregulating MRD1 and BCRP via PI3K/AKT pathway.

The Extract of Cordyceps Militaris Inhibited the Proliferation of Cisplatin-Resistant A549 Lung Cancer Cells by Downregulation of H-Ras.

We investigated the antitumor effect of Cordyceps militaris extract (CME) on A549 cisplatin-resistant (CR) lung cancer cells. The proliferation of A549/CR cells was suppressed by CME. Apoptosis of the cells was induced by CME. The cell cycle arrest was observed in the sub-G1 phase in the cells treated with CME. Proteomic profile analysis showed that H-Ras was downregulated in CME-treated cells and it was confirmed by western blot analysis. Collectively, these data demonstrated that CME is an alternative treatment for the anticancer effect.

A PolH Transcript with a Short 3'UTR Enhances PolH Expression and Mediates Cisplatin Resistance.

Platinum-based anticancer drugs are widely used as a first-line drug for cancers, such as non-small cell lung carcinoma (NSCLC) and bladder cancer. However, the efficacy is limited due to intrinsic or acquired resistance to these drugs. DNA polymerase eta (PolH, Polη) belongs to the Y-family of DNA polymerases and mediates DNA translesion synthesis, a major mechanism for DNA damage tolerance. Here, we showed that a high level of PolH is associated with cisplatin resistance in lung and bladder cancer. Consistent with this, loss of PolH markedly attenuates cisplatin resistance in both cisplatin-sensitive and cisplatin-resistant lung cancer cells. Interestingly, we found that due to the presence of multiple polyadenylation sites, alternative polyadenylation (APA) produces three major PolH transcripts with various lengths of 3'untranslated region (3'UTR; 427-/2516-/6245-nt). We showed that the short PolH transcript with 427-nt 3'UTR is responsible for high expression of PolH in various cisplatin-resistant lung and bladder cancer cell lines. Importantly, loss of the short PolH transcript significantly sensitizes cancer cells to cisplatin treatment. Moreover, we found that miR-619 selectively inhibits the ability of the long PolH transcript with 6245-nt 3'UTR to produce PolH protein and, subsequently, PolH-dependent cell growth. Together, our data suggest that PolH expression is controlled by APA and that the short PolH transcript produced by APA can escape miR-619-mediated repression and, subsequently, confers PolH-mediated cisplatin resistance. SIGNIFICANCE: A short PolH transcript produced by alternative polyadenylation escapes repression by miR-619 and confers resistance to cisplatin.

Abstract 4361: Kynurenine - Aryl hydrocarbon receptor axis: A crucial modulator of immunometabolism in cisplatin resistant lung cancer

Abstract The effect of tumor metabolism on the tumor microenvironment is not well established. Our previous data have shown that cisplatin resistant (CR) lung cancer cells increased secretion of thioredoxin-1 (TRX1) and kynurenine (KYN). Interestingly, high TRX1 and KYN levels in tumor microenvironment can enhance immunosuppressive environment. We have demonstrated that CR cells possessed lower levels of hypoxia-inducible factor-1α (HIF1α) due to metabolic re-programming. ARNT or HIF1β is a known binding partner of both HIF1α and aryl hydrocarbon (AHR). Importantly, recent study indicates that KYN can serve as an endogenous ligand for AHR in cancer cells. Thus, we hypothesize that in the absence of HIF1α, ARNT is now available to bind and form a new partner with KYN/AHR and initiate the transcription of genes which favor survival/proliferation of CR cells. Four pairs of NSCLC cell lines and their CR variants (ALC, FC, H460R, A549R) were used. Using immunofluorescence technique, our result showed that AHR localized primary in the nucleus of CR cells when compared with parental cell counterparts. We further determined that KYN was specific to AHR activation in CR cells by inhibiting AHR translocation using 10µM of dimethoxyflavone (DMF; an AHR antagonist). AHR was less accumulated inside nuclease after treatment. Importantly, treatment of DMF also resulted in suppression of Indoleamine 2,3-Dioxygenase-1 (IDO1) activities. To further verify these findings, we assayed the expression of AHR-target gene (LAT1 and CYP1B1) with and without adding KYN. Both genes expressions were higher in CR cells and were further augmented upon an addition of KYN. In contrast, we did not find an increase in LAT1 after exposure to KYN in parental cells. Using flow cytometer, we found that CR cells possessed higher surface-LAT1 levels when compared to parental cell counterparts, and treatment of KYN resulted in significant tryptophan uptake in CR cells. Importantly, treatment with IDO1 inhibitor (5.5uM) significantly suppressed LAT1 and CYP1B1 expression in CR cells and reverse cisplatin resistance in CR from 2.5 to 0.75 μg/ml (3.3 fold). Thus, our data strongly indicate that KYN/AHR/ARNT axis plays a unique modulator role in CR cells metabolism. Overall these results will have potential impact on (i) how one can effectively exploit the KP pathway to treat CR tumors, and (ii) improving understanding on how CR cells evade immune surveillance. Supported by Department of Veterans Affairs (BLR&amp;D Merit review and CDA2) Citation Format: Medhi Wangpaichitr, Dan JM Nguyen, Ying-Ying Li, Chunjing Wu, Lynn G. Feun, Niramol Savaraj. Kynurenine - Aryl hydrocarbon receptor axis: A crucial modulator of immunometabolism in cisplatin resistant lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4361.

Novel tetranuclear ruthenium(II) arene complexes showing potent cytotoxic and antimetastatic activity as well as low toxicity in vivo

Ruthenium complexes have attracted a surge of interest as anticancer drug candidates because of their low toxicity, diversity in mode-of-actions and non-cross drug resistance with conventional platinum-based agents. Despite remarkable advances, only a limited number of ruthenium complexes have been demonstrated to kill cancer cells and suppress metastasis simultaneously. Here, two organometallic tetranuclear Ru(II) arene complexes (Ru-1 and Ru-2) have been synthesized and evaluated for their in vitro activity against a panel of human cancer cell lines, including a cisplatin-resistant human lung cancer A549 cell line. A superior cytotoxic activity of the ruthenium complexes compared to cisplatin across distinct cell lines was observed. Further examination of the mechanism indicated that anticancer activity was accomplished by inducing apoptosis in cancer cells. In addition, we found that such compounds exhibited promising antimetastatic activity and reduced the invasiveness of cancer cells. Importantly, choosing Ru-1 as a target compound, a significantly enhanced safety profile relative to cisplatin in animals was validated, suggesting that these complexes can be used as promising candidates for cancer therapy and deserve further investigation.