Cisplatin-induced Cell Death Research Articles

Adipose-derived stem cell conditioned medium attenuates cisplatin-triggered apoptosis in tongue squamous cell carcinoma.

Autologous fat grafting procedures have noted a markedly increased frequency, not only for cosmetic purposes, but also for deformities after head and neck cancer and breast cancer surgery. Carcinogenesis is always a major concern in cell therapy-related issues. However, there is no literature discussing this issue in head and neck squamous cell carcinoma patients. To evaluate the interaction of tongue cancer cells and adipose-derived stem cells, we performed a series of invitro experiments. Our results demonstrated that cisplatin significantly reduced the viabilities of SCC‑25 and CAL‑27 cells in a concentration-dependent manner, but it had low cytotoxicity in cisplatin-resistant CAL‑27 (CAR) cells. There was no significant difference in terms of viability among the SCC‑25, CAL‑27, and CAR cells in the adipose-derived stem cell conditioned medium and control groups. There was also no significant difference in terms of cell migration as determined by wound healing assay of SCC‑25, CAL‑27, and CAR cells between the adipose-derived stem cell conditioned medium treatment and control treatment. Importantly, the adipose-derived stem cell conditioned medium attenuated cisplatin-triggered cell death in the SCC‑25 and CAL‑27 cells. Moreover, adipose-derived stem cell conditioned medium markedly inhibited cisplatin-induced apoptotic cell death (sub‑G1 phase) in the CAL‑27 cells. Western blot analyses indicated that cisplatin-induced reductions in pro‑caspase‑3, pro‑caspase‑9, phospho-BAD, phospho-IGF-1R, phospho-AKT, and phospho-ERK in CAL‑27 cells were reversed by adipose-derived stem cell conditioned medium supplement. Taken together, we provide evidence that adipose-derived stem cell conditioned medium protects CAL‑27 cells from cisplatin-induced cell death, possibly through upregulation of the IGF-1R/AKT/ERK signaling pathway.

Midkine derived from cancer-associated fibroblasts promotes cisplatin-resistance via up-regulation of the expression of lncRNA ANRIL in tumour cells

Midkine (MK) is a heparin-binding growth factor that promotes carcinogenesis and chemoresistance. The tumour microenvironment (TME) can affect chemotherapy sensitivity. However, the role of stromal-derived MK, especially in cancer-associated fibroblasts (CAFs), is unclear. Here, we confirmed that MK decreased cisplatin-induced cell death in oral squamous cell carcinoma (OSCC) cells, ovarian cancer cells and lung cancer cells. We also isolated primary CAFs (n = 3) from OSCC patients and found that CAFs secreted increased levels of MK, which abrogated cisplatin-induced cell death. Moreover, MK increased the expression of lncRNA ANRIL in the tumour cells. Normal tissues, matched tumour-adjacent tissues and OSCC tissues were analysed (n = 60) and showed that lncRNA ANRIL was indeed overexpressed during carcinogenesis and correlated with both high TNM stage and lymph node metastasis (LNM). Furthermore, lncRNA ANRIL knockdown in tumour cells inhibited proliferation, induced apoptosis and increased cisplatin cytotoxicity of the tumour cells via impairment of the drug transporters MRP1 and ABCC2, which could be restored by treatment with human MK in a caspase-3/BCL-2-dependent manner. In conclusion, we firstly describe that CAFs in the TME contribute to the high level of MK in tumours and that CAF-derived MK can promote cisplatin resistance via the elevated expression of lncRNA ANRIL.

MiR-133b induces chemoresistance of osteosarcoma cells to cisplatin treatment by promoting cell death, migration and invasion.

As an important chemotherapeutic agent for the treatment of osteosarcoma, the effectiveness of cisplatin is considered to be due to its unique properties, which allow it to penetrate the cell membrane and form various DNA-platinum adducts, resulting in genetic alterations or DNA damage. However, chemoresistance to cisplatin remains a major challenge for its use and chemotherapeutic effects. In the present study, an isogenic model of a cisplatin resistant osteosarcoma cell line, MG63-DDP, was generated from the original MG63 cell line. The expression level of microRNA (miR)-133b in the MG63-DDP cisplatin-resistant osteosarcoma cell line was analyzed by reverse transcription-quantitative polymerase chain reaction (PCR). Cisplatin-DNA adduct formation, cell death (carboxyfluorescein succinimidyl ester/propidium iodide staining) and clonogenic survival assays (crystal violet staining) were performed, comparing various cell types. The effect of miR-133b on migration (scratch wound assay) and invasion (Transwell assay) was also evaluated. Characterization studies have previously revealed an increased level of miR-133b in MG63-DDP cells compared with normal MG63 cells. Upregulation of miR-133b was associated with the accumulation of cisplatin-DNA adducts and an increase in cisplatin-induced cell death. Furthermore, increased miR-133b expression levels enhanced the migration and invasion of MG63 cells under cisplatin stress. Concordantly, in MG63-DDP cells the neutralization of miR-133b demonstrated opposite effects, as compared with the upregulation of miR-133b. To the best of our knowledge, the present study demonstrated for the first time that cisplatin-resistant MG63 cells exhibit an increased level of miR-133b expression. The endogenous expression level of miR-133b is sufficient for inducing cisplatin resistance, which suggests that miR-133b may be a biomarker for cisplatin resistance in osteosarcoma.

MLKL-PITPα signaling-mediated necroptosis contributes to cisplatin-triggered cell death in lung cancer A549 cells

Necroptosis has been reported to be involved in cisplatin-induced cell death, but the mechanisms underlying the occurrence of necroptosis are not fully elucidated. In this study, we show that apart from apoptosis, cisplatin induces necroptosis in A549 cells. The alleviation of cell death by two necroptosis inhibitors—necrostatin-1 (Nec-1) and necrosulfonamide (NSA), and the phosphorylation of mixed lineage kinase domain-like protein (MLKL) at serine 358, suggest the involvement of receptor-interacting protein kinase 1 (RIPK1)-RIPK3-MLKL signaling in cisplatin-treated A549 cells. Additionally, the initiation of cisplatin-induced necroptosis relies on autocrine tumor necrosis factor alpha (TNF-α). Furthermore, we present the first evidence that phosphatidylinositol transfer protein alpha (PITPα) is involved in MLKL-mediated necroptosis by interacting with the N terminal MLKL on its sixth helix and the preceding loop, which facilitates MLKL oligomerization and plasma membrane translocation in necroptosis. Silencing of PITPα expression interferes with MLKL function and reduces cell death. Our data elucidate that cisplatin-treated lung cancer cells undergo a new type of programmed cell death called necroptosis and shed new light on how MLKL translocates to the plasma membrane.

Isoliquiritigenin pretreatment attenuates cisplatin induced proximal tubular cells (LLC-PK1) death and enhances the toxicity induced by this drug in bladder cancer T24 cell line

Cisplatin is an effective antineoplastic agent widely used in the treatment of solid tumors, however, it induces nephrotoxicity. Cisplatin-induced nephrotoxicity is associated with increased reactive oxygen species (ROS) production and decreased antioxidant system defense in kidney. Isoliquiritigenin (IsoLQ) is a chalcone, which is characterized by its antioxidant and antiinflammatory properties. Herein, we investigated the protective effect of IsoLQ on LLC-PK1 proximal tubular cells against cisplatin-induced death and its effect on the antineoplastic activity of cisplatin on bladder cancer T24 cell line. It was found that pretreatment with IsoLQ attenuates cisplatin-induced cell death, ROS production, and activation of caspase-3. IsoLQ also induced heme oxygenase-1 (HO-1) expression that may be associated with nuclear factor (erythroid-derived 2)-like 2 (Nrf2) nuclear translocation. The protective effect of IsoLQ was abrogated by tin mesoporphyrin (SnMP), an HO inhibitor. Further, bilirubin and carbon monoxide releasing molecule-2 also showed a protective effect against cisplatin-induced cell death. In addition, IsoLQ induced in a dose-dependent way, death of T24 cells and exacerbated cisplatin-induced cell death. These results suggest that IsoLQ has a protective effect on cisplatin-induced toxicity in LLC-PK1 cells, in part through induction of HO-1, without interfering with the antineoplastic activity of this agent in T24 cells.

Heat Shock Protein 27, a Novel Regulator of Transforming Growth Factor β Induced Resistance to Cisplatin in A549 Cell

Lung cancer is one of the major causes of cancer morbidity and mortality around the world, and the resistance to cisplatin is a critical issue to chemotherapy in lung cancer patients. Transforming growth factor β (TGF-β) signal pathway abnormality is widely observed in drug resistance during lung cancer chemotherapy. Here, we investigated the effects of heat-shock protein 27 (HSP27) in the TGF-β-induced cisplatin resistance in lung cancer cell. In this study, our results indicated that the mRNA and protein expression of HSP27 were significantly increased in human lung cancer tissues. TGF-β induced the mRNA and protein expression of HSP27 in human lung cancer cell (A549). Treatment of TGF-β-induced cisplatin resistance in A549 cell through blocking the cisplatin-induced apoptosis and cell death, which characterized as the increasing of cell viability and decreasing of PARP and caspase3 cleavage in the cisplatin-treated cell. Knockdown of SMAD3 attenuated the TGF-β-induced HSP27 expression and restored the TGF-β-induced cisplatin resistance in A549 cell. Additionally, the knockdown of HSP27 blocked TGF-β-induced cisplatin resistance via decreasing cell viability and increasing cell apoptosis in A549 cell. These data therefore suggested that HSP27 is critical to lung cancer progression and TGF-β-induced cisplatin resistance in human lung cancer cell, and may provide an effective clinical strategy in lung cancer patients with resistance to chemotherapy.

The TLR3 Agonist Inhibit Drug Efflux and Sequentially Consolidates Low-Dose Cisplatin-Based Chemoimmunotherapy while Reducing Side Effects.

The traditional maximum dose density chemotherapy renders the tumor patients not only the tumor remission but the chemotherapy resistance and more adverse side effects. According to the widely positive expression of Toll-like receptor (TLR)-3 in oral squamous cell carcinoma (OSCC) patients (n = 166), we here provided an alternative strategy involved the orderly treatment of TLR3 agonist polyinosine-polycytidylic acid (PIC) and low-dose cisplatin. The optimal dose of cisplatin, the novel role of PIC and the side effects of the combined chemotherapy were determined in vitro and in distinct human tumor models in vivo The results in vitro indicated that preculture with PIC downregulated drug transporters (e.g., P-gp and MRP-1) and increased the cytoplasmic residence of cisplatin, and dramatically strengthened the low-dose cisplatin-induced cell death in TLR3- and caspase-3-dependent manner. Meanwhile, the spleen immunocytes were activated but the immunosuppressive cancer-associated fibroblasts (CAF) were dampened. These findings were confirmed in human tumor models in vivo Pretreatment with PIC promoted the low-dose cisplatin residence for tumor regression with decreased myeloid-suppressive cells (MDSC), tumor-associated macrophages (TAM) and CAFs, and alleviated adverse side effects in the OSCC model, which was further enhanced by the Cetuximab safely. This strategy also repressed the progression of melanoma and lymphoma. Moreover, TLR3 negatively manipulated the inflammation-related long noncoding RNA lnc-IL7R, which was upregulated during this chemotherapy. Knockdown of lnc-IL7R improved the chemotherapy sensitivity. Overall, this study provided preclinically new instructions for the PIC/cisplatin utilization to target tumor microenvironment and strengthen the low-dose cisplatin-based chemotherapy with reduced side effects. Mol Cancer Ther; 16(6); 1068-79. ©2017 AACR.

NOTCH1 Signaling Regulates Self-Renewal and Platinum Chemoresistance of Cancer Stem-like Cells in Human Non-Small Cell Lung Cancer.

Cancer stem-like cells (CSC) are thought to drive tumor initiation, metastasis, relapse, and therapeutic resistance, but their specific pathogenic characters in many cancers, including non-small cell lung cancer (NSCLC), have yet to be well defined. Here, we develop findings that the growth factor HGF promotes CSC sphere formation in NSCLC cell populations. In patient-derived sphere-forming assays (PD-SFA) with HGF, CD49f and CD104 were defined as novel markers of lung CSC (LCSC). In particular, we isolated a subpopulation of CD166+CD49fhiCD104-Lin- LCSC present in all human specimens of NSCLC examined, regardless of their histologic subtypes or genetic driver mutations. This specific cell population was tumorigenic and capable of self-renewal, giving rise to tumor spheres in vitro and orthotopic lung tumors in immune-compromised mice. Mechanistic investigations established that NOTCH1 was preferentially expressed in this cell subpopulation and required for self-renewal via the transcription factor HES1. Through a distinct HES1-independent pathway, NOTCH1 also protected LCSCs from cisplatin-induced cell death. Notably, treatment with a γ-secretase inhibitor that blunts NOTCH1 function ablated self-renewing LCSC activity and restored platinum sensitivity in vitro and in vivo Overall, our results define the pathogenic characters of a cancer stem-like subpopulation in lung cancer, the targeting of which may relieve platinum resistance in this disease. Cancer Res; 77(11); 3082-91. ©2017 AACR.

Abstract NTOC-091: INVOLVEMENT OF TIE–1 TYROSINE KINASE RECEPTOR IN CHEMO–RESISTANCE: POTENTIAL OF TIE1 AS A NOVEL THERAPEUTIC TARGET

Abstract OBJECTIVES: Platinum-resistance is one of the most challenging difficulties in the treatment of ovarian cancer patients. To overcome this problem, we have explored a target molecule which can conquer platinum-resistance of ovarian cancer cells utilizing a functional genomics approach. MATERIALS AND METHODS: High-throughput functional siRNA screening was designed to target 6550 genes in cisplatin-resistant A2780 CP ovarian cancer cells. Cell viability was assessed by luminescent cell viability assay. After identifying a candidate molecule, cisplatin uptake was determined by atomic absorption spectrometry. DNA damages were determined by the western blotting and immunofluorescent staining using γH2AX antibodies. RESULTS: Through a functional screening, receptor tyrosine kinase TIE 1 was identified as a top candidate gene, of which inhibition give rise to enhancement of cisplatin sensitivity in ovarian cancer cells. Conversely, over-expression of TIE 1 gene significantly decreased susceptibility to cisplatin-induced cell death without affecting cisplatin uptake. DNA damages induced by cisplatin was significantly suppressed in TIE 1 over-expressed cells, raising novel potential mechanisms of TIE 1 in nucleotide excision repair system that removes chemicals adduct to DNA. In addition, over-expression of TIE 1 increased the expression of XPC, which is responsible for nucleotide excision repair. CONCLUSION: We have identified TIE 1 as a molecular target to overcome platinum-resistance in ovarian cancer cells. TIE 1 contribute platinum- resistance in ovarian cancer cells by promoting XPC-dependent DNA repairing system. Citation Format: Masumi Ishibashi, Masafumi Toyoshima, Mahy Egiz, Xuewei Zhang, Junko Minato, Shogo Shigeta, Kazuyuki Kitatani, Nobuo Yaegashi. INVOLVEMENT OF TIE–1 TYROSINE KINASE RECEPTOR IN CHEMO–RESISTANCE: POTENTIAL OF TIE1 AS A NOVEL THERAPEUTIC TARGET [abstract]. In: Proceedings of the 11th Biennial Ovarian Cancer Research Symposium; Sep 12-13, 2016; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(11 Suppl):Abstract nr NTOC-091.

Anti-cancer effect of Scutellaria baicalensis in combination with cisplatin in human ovarian cancer cell

BackgroundOvarian cancer is one of the major causes of death among females in worldwide. Cisplatin is a primary anti-cancer drug against ovarian cancer, but the recurrent tumors after treatment frequently show acquired chemoresistance. Extract of Scutellaria baicalensis (SbE) has been reported to have functional compounds including baicalin, which has anti-cancer effects. However, the anti-cancer effects of SbE in ovarian cancer and its underlying mechanisms are elusive.MethodsWe investigated that the effects of SbE and/or cisplatin on cell death in the cisplatin sensitive ovarian cancer cell line A2780 (CSC) and the counterpart cell line that has cisplatin resistance (CRC). Molecular mechanisms of the effects, focusing on apoptosis and autophagy, were examined.ResultsTreatment of cisplatin or SbE reduced cell viability significantly in CSC and too much lesser extent in CRC. Cisplatin-induced cell death in CSC was mediated by p53-induced apoptosis acompanied by expresson of damage-regulated autophagy modulator (DRAM). In CRC, decreased DRAM expression (p < 0.01) hindered p21-mediated cell death and contributed to cisplatin resistance. Treatment of SbE also induced cell death in CSC by p53-dependent apoptosis, not in CRC. Autophagy was not induced by neither cisplatin nor SbE. Intriguingly, the combinational treatment of SbE and cisplatin significantly decreased cell viability in CRC. The cell death was mediated by autophagy with increased expression of Atg5 and Atg12 (p < 0.05), rather than p53-dependent pathway with repressed expression of p21 (p < 0.001) through HDAC1 activation.ConclusionsThe combined treatment of SbE with cisplatin was effective in CRC, leading to cell death via Beclin1-independent autophagy, suggesting that SbE treatment in combination with cisplatin has a potential as a chemotherapeutic agent in cisplatin-resistant ovarian cancer.

Hsa-miR-485-5p reverses epithelial to mesenchymal transition and promotes cisplatin-induced cell death by targeting PAK1 in oral tongue squamous cell carcinoma.

Oral squamous cell carcinoma (OSCC) is currently a highly prevalent disease worldwide. Cisplatin (CDDP) is widely used for the chemotherapy of OSCC. Yet, the molecular mechanisms responsible for cisplatin resistance have not been fully elucidated. In this study, we showed that overexpression of p21 (RAC1) activated kinase 1 (PAK1) induced epithelial to mesenchymal transition (EMT) and significantly promoted the invasion and migration of oral squamous cell carcinoma SCC25 cells. Emerging evidence indicates a strong link between resistance to therapy and the induction of EMT in cancer. We showed that overexpression of PAK1 induced cisplatin resistance in SCC25 cells. ERCC1 and YAP can promote cisplatin resistance in human OSCC. We showed that ERCC1 and YAP protein were upregulated by PAK1 in SCC25 cells. We found that miR-485-5p inhibited PAK1 protein expression in the SCC25 cells. Contrary to PAK1, we demonstrated that overexpression of miR-485-5p reversed EMT and significantly inhibited invasion and migration. Moreover, its overexpression sensitized SCC25-CR cells (cisplatin-resistant cells) to cisplatin. Thus, we conclude that miR-485-5p reverses EMT and promotes cisplatin-induced cell death by targeting PAK1 in oral tongue squamous cell carcinoma. This study suggests that PAK1 plays an essential role in the progression of OSCC and it is a potential therapeutic target for OSCC.

Protective effect of lanostane triterpenoids from the sclerotia of Poria cocos Wolf against cisplatin-induced apoptosis in LLC-PK1 cells

Cisplatin-induced nephrotoxicity is a serious adverse effect that limits the use of cisplatin in cancer patients. In the present study, we investigated the protective effect of lanostane triterpenoids (1–10) isolated from the ethanolic extract of Poria cocos Wolf against cisplatin-induced cell death in LLC-PK1 kidney tubular epithelial cells. Treatment of cisplatin induced significant cell death, which was suppressed by treatment with dehydroeburicoic acid monoacetate (1) and 3β-acetoxylanosta-7,9(11),24-trien-21-oic acid (9). Compound 1 exhibited the highest efficacy among the tested compounds and was thus subjected to further mechanistic studies. The increase in the percentage of apoptotic cells induced by cisplatin reduced by 4.3% after co-treatment of cells with compound 1 (50 and 100μM). Furthermore, phosphorylation of the mitogen-activated protein kinases JNK, ERK, and p38, and caspase-3, which characterize oxidative stress-mediated apoptosis, increased significantly after treatment with cisplatin, and decreased after treatment with compound 1. These results indicate that the renoprotective effects of compound 1 may be mediated by its anti-apoptotic activity.

Methionine Sulfoxide Reductase A Deficiency Exacerbates Cisplatin-Induced Nephrotoxicity via Increased Mitochondrial Damage and Renal Cell Death.

Methionine sulfoxide reductase A (MsrA), which is abundantly localized in the mitochondria, reduces methionine-S-sulfoxide, scavenging reactive oxygen species (ROS). Cisplatin, an anticancer drug, accumulates at high levels in the mitochondria of renal cells, causing mitochondrial impairment that ultimately leads to nephrotoxicity. Here, we investigated the role of MsrA in cisplatin-induced mitochondrial damage and kidney cell death using MsrA gene-deleted (MsrA-/-) mice. Cisplatin injection resulted in increases of ROS production, methionine oxidation, and oxidative damage in the kidneys. This oxidative stress was greater in MsrA-/- mouse kidneys than in wild-type (MsrA+/+) mouse kidneys. MsrA gene deletion exacerbated cisplatin-induced reductions in the expression and activity of MsrA and MsrBs, and the expression of thioredoxin 1, glutathione peroxidase 1 and 4, mitochondrial superoxide dismutase, cystathionine-β-synthase, and cystathionine-γ-lyase. Cisplatin induced swelling, cristae loss, and fragmentation of mitochondria with increased lipid peroxidation, more so in MsrA-/- than in MsrA+/+ kidneys. The ratio of mitochondrial fission regulator (Fis1) to fusion regulator (Opa1) was higher in MsrA-/- than MsrA+/+ mice. MsrA deletion exacerbated cisplatin-induced increases in Bax to Bcl-2 ratio, cleaved caspase-3 level, and apoptosis, whereas MsrA overexpression attenuated cisplatin-induced oxidative stress and apoptosis. MsrA gene deletion in mice exacerbates cisplatin-induced renal injury through increases of mitochondrial susceptibility, whereas MsrA overexpression protects cells against cisplatin. This study demonstrates that MsrA protects kidney cells against cisplatin-induced methionine oxidation, oxidative stress, mitochondrial damage, and apoptosis, suggesting that MsrA could be a useful target protein for the treatment of cisplatin-induced nephrotoxicity. Antioxid. Redox Signal. 27, 727-741.

P53 modulates the effect of ribosomal protein S6 kinase1 (S6K1) on cisplatin toxicity in chronic myeloid leukemia cells

Chronic myeloid leukemia (CML) is characterized by the expression of the oncoprotein, BCR-ABL. BCR-ABL inhibitors revolutionized CML chemotherapy while blast crisis (BC) CML patients are less responsive. Since suppression of ribosomal protein S6 kinase1 (S6K1) phosphorylation reverses the resistance to BCR-ABL inhibitor in CML cells and S6K1 inhibitors augment cisplatin toxicity in lung cancer cells, we speculated that combination of S6K1 inhibitor and cisplatin may be beneficial for eliminating BC CML cells. To our surprise, S6K1 inhibition decreased cisplatin-induced DNA damage and cell death only in p53-/− BC CML cells but not in p53+/+ BC CML cells. During the progression of CML, p53 expression either decreases or mutates. Moreover, the expression of p53 affects drug response of CML cells. Our results confirmed that S6K1 inhibition reversed cisplatin toxicity is dependent on p53 expression in CML cells. Moreover, p53 attenuated the phosphorylation and localization of S6K1 via attenuating 3-phosphoinositide dependent protein kinase-1 (PDK1) phosphorylation. Furthermore, S6K1 acts via DNA-PKcs to regulate H2AX phosphorylation and PARP cleavage, respectively. Taken together, our results suggest that p53/PDK1/S6K1 is a novel pathway regulating cisplatin toxicity in BC CML cells.

Abstract B25: Discovery of molecular probes inhibiting La:RNA interactions in cancer cells

Abstract The RNA-binding protein La is overexpressed in a number of tumor tissues and is proposed to support tumorigenesis via binding to mRNAs encoding tumor-promoting and anti-apoptotic factors to facilitate their translation. Our data suggest that those mRNAs might need La to be translated mainly in a cap-independent mechanism in cancer cells. Hence, small molecules able to block the binding of La to specific RNAs could represent a novel therapeutic strategy for cancer treatment. In an attempt toward this approach we developed a high-throughput fluorescence polarization assay to screen small compound libraries for molecules inhibiting the binding of La to an RNA element derived from cyclin D1 mRNA. Herein we describe a robust fluorescence polarization assay and the validation of primary hits by electrophoretic mobility shift assays. Using isothermal titration calorimetry we show that a compound binds to La and molecular modeling suggests an interaction between the compound and the RNA recognition motif of La. We have shown recently that La protects cells against cisplatin treatment by stimulating the protein synthesis of the anti-apoptotic factor Bcl2. Here we show by RNA immunoprecipitation experiments that one small compound specifically impairs the association of La with Bcl2 mRNA, reduces Bcl2 protein expression and sensitizes cells for cisplatin-induced cell death. Taken together, herein we report the development of a high-throughput fluorescence polarization assay to identify small compounds able to impair the binding of cancer-associated La to RNAs encoding tumor-promoting and anti-apoptotic factors in vitro and cancer cells. Citation Format: Gunhild Sommer, Alena Fedarovich, Venkatesh Kota, Yuri K. Peterson, Tilman Heise. Discovery of molecular probes inhibiting La:RNA interactions in cancer cells. [abstract]. In: Proceedings of the AACR Special Conference on Translational Control of Cancer: A New Frontier in Cancer Biology and Therapy; 2016 Oct 27-30; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2017;77(6 Suppl):Abstract nr B25.

Poly ADP-ribose polymerase inhibition suppresses cisplatin toxicity in chronic myeloid leukemia cells.

Cancer cells may acquire drug resistance by activating DNA repair signaling. Poly ADP-ribose polymerase (PARP) plays an important role in DNA repair and it is overexpressed in many cancers including chronic myeloid leukemia (CML). PARP inhibitors have been used either alone or with other drugs to augment cancer cell death. However, whether PARP inhibitors may also augment cell death induced by chemotherapeutic agents in CML cells has not been studied. K562 cells with or without PARP-1 knockdown were treated with cisplatin alone or together with olaparib. The cell death was investigated by propidium iodide staining and apoptosis-related proteins were detected by western blotting. Olaparib suppressed cisplatin-induced cell death in K562 and MEG01 cells. Moreover, PARP-1 knockdown also attenuated cisplatin toxicity in CML cells. Inhibition of PARP decreased cisplatin toxicity by attenuating caspase-3 and caspase-9 activity. These results indicated that the toxicity of cisplatin in CML cells requires PARP activity. Therefore, PARP inhibitors may not be useful with DNA-damaging agents such as cisplatin in CML treatment.

Involvement of mitochondrial dynamics in the antineoplastic activity of cisplatin in murine leukemia L1210 cells.

Leukemia is a type of hematopoietic stem cell malignant cloned disease with high mortality. Cisplatin-based chemotherapy is one of the most common treatments for leukemia. Similar to other chemotherapeutic agents, cisplatin resistance has become a serious issue in cancer therapy. In the present study, we investigated the role of mitochondrial dynamics in the antineoplastic activity of cisplatin in murine leukemia L1210 cells. Firstly, the L1210 cell line resistant to cisplatin (L1210/DDP) was established. Compared to its parental cell line, the IC50 value of cisplatin in the L1210/DDP cells was increased 10-fold. Mitofusins (Mfn1 and Mfn2), mitochondrial outer membrane fusion proteins, were markedly upregulated in the L1210/DDP cells, whereas the expression of fission protein Drp1 and inner membrane fusion protein OPA1 were not significantly altered. In addition, mitofusins were also upregulated in the parental L1210 cells subjected to cisplatin stress. To investigate the role of mitochondrial dynamics in the antineoplastic activity of cisplatin, the effect of mitochondrial division inhibitor (Mdivi)-1 on cisplatin‑induced cell death, caspase-3 cleavage and ROS production was examined in L1210 cells. We found that 5µM of Mdivi-1 efficiently attenuated cisplatin-induced cell death, caspase activation and intracellular ROS increase in L1210 cells. Our data indicated that mitochondrial dynamics play an important role in the antineoplastic activity of cisplatin, and mitofusin-mediated mitochondrial fusion may be involved in the process of cisplatin resistance in leukemia cells. Therefore, the present study revealed that mitochondrial dynamics may be a potential target used to improve the antineoplastic activity of cisplatin in leukemia in the future.

Cisplatin-induced necroptosis in TNFα dependent and independent pathways

Cisplatin is a chemotherapeutic drug for treatment of many solid tumors. It has been shown to induce apoptosis and/or necrosis in different types of cancer cells. However, the underlying mechanisms remain elusive. In this study, we provide evidences that cisplatin induces necroptosis in receptor-interacting protein 3 (RIP3)-expressing cell lines, but not in cell lines lacking RIP3 protein expression. Deficiency of core components of necroptotic pathway, RIP1, RIP3, or mixed lineage kinase domain-like protein (MLKL) blocked cisplatin-induced cell death in L929 cells. This phenomenon is dependent on RIP1/RIP3/MLKL necrosome formation and translocation to mitochondria-associated membrane (MAM), but only partially via autocrine production of tumor necrosis factor α (TNFα). Moreover, we demonstrate that the mitochondrial permeability transition pore opening (mPTP) opening and reactive oxygen species (ROS) generation is a critical downstream event of the formation of necrosome in cisplatin-induced necroptosis, which is TNFα independent. Deficiency of cyclophilin-D (CypD) partially reduced cisplatin-induced cell death, indicating CypD mediated-mPTP opening plays an important role during cisplatin-induced necroptosis. Both deletion of CypD and TNFα completely blocked cisplatin-induced cell death, suggesting that cisplatin could induce necroptosis through TNFα dependent and independent pathway. These findings provide new insight into the molecular mechanisms underlying cisplatin-induced necroptosis.

Effect of Ecklonia Cava Polyphenol Extract in House Ear Institute-Organ of Corti 1 Cells Against Cisplatin Ototoxicity: A Preliminary Study.

Cisplatin is a widely used agent for the treatment of adult and childhood malignancies. Side effects such as nephrotoxicity, neurotoxicity, and ototoxicity lead to dose limitations. Ecklonia cava polyphenol extract (ECP) is a molecule obtained from algae that live in seawater in the Far East. ECP has recently been shown to have protective effects against oxidative stress. The aim of this study was to evaluate the possible protective effects of ECP on cisplatin ototoxicity. In this study, we investigated the protective effects of ECP against cisplatin-induced cell death in mouse-derived House Ear Institute Organ of Corti (HEI-OC1) cochlear cells. Cisplatin (100 μM) and 1, 10, and 25 μM doses of ECP were administered to the cells, and the protective effects of ECP at 24 and 72 hours were investigated. Cell viability was evaluated by the WST-1 (water soluble tetrazolium salt). Cisplatin (100 μM) reduced cell viability in both the 24th and 72nd hour evaluation. Although the 25 μM dose of ECP showed otoprotective effects in the 24th hour, in the 72nd hour this effect disappeared. Other doses of ECP showed no otoprotective effects in the 24th and 72nd hours. Although ECP showed some protective effects in the 24th hour against cisplatin ototoxicity, these effects disappeared by the 72nd hour. Further studies using recurrent and higher doses of ECP are required.

Ameliorative Effect of Daidzein on Cisplatin-Induced Nephrotoxicity in Mice via Modulation of Inflammation, Oxidative Stress, and Cell Death.

Oxidative stress and inflammation are part and parcel of cisplatin-induced nephrotoxicity. The purpose of this work is to study the role of soy isoflavone constituent, daidzein, in cisplatin-induced renal damage. Cisplatin-induced nephrotoxicity was evident by the histological damage in proximal tubular cells and by the increase in serum neutrophil gelatinase-associated lipocalin (NGAL), blood urea nitrogen (BUN), creatinine, and urinary kidney injury molecule-1 (KIM-1). Cisplatin-induced cell death was shown by TUNEL staining and caspase-3/7 activity. Daidzin treatment reduced all kidney injury markers (NGAL, BUN, creatinine, and KIM-1) and attenuated cell death (apoptotic markers). In cisplatin-induced kidney injury, renal oxidative/nitrative stress was manifested by the increase in lipid peroxidation and protein nitration. Cisplatin induced the reactive oxygen species-generating enzyme NOX-2 and impaired antioxidant defense enzyme activities such as glutathione peroxidase (GPX) and superoxide dismutase (SOD) activities. Cisplatin-induced oxidative/nitrative stress was attenuated by daidzein treatment. Cisplatin induced CD11b-positive macrophages in kidneys and daidzein attenuated CD11b-positive cells. Daidzein attenuated cisplatin-induced inflammatory cytokines tumor necrosis factor α (TNFα), interleukin 10 (IL-10), interleukin 18 (IL-18), and monocyte chemoattractant protein-1 (MCP-1). Daidzein attenuated cell death in vitro. Our data suggested that daidzein attenuated cisplatin-induced kidney injury through the downregulation of oxidative/nitrative stress, immune cells, inflammatory cytokines, and apoptotic cell death, thus improving kidney regeneration.