Dipyridamole potentiates the in vitro activity of MTA (LY231514) by inhibition of thymidine transport

The novel pyrrolopyrimidine-based antifolate LY231514 (MTA), inhibits multiple folate-requiring enzymes including thymidylate synthase, glycinamide ribonucleotide formyltransferase and dihydrofolate reductase. Both thymidine and hypoxanthine are required to reverse MTA growth inhibition in leukaemia and colon cancer cells. Prevention of MTA growth inhibition by thymidine and/or hypoxanthine was investigated in two human lung (A549, COR L23) and two breast (MCF7, T47D) tumour cell lines, and the effect of the nucleoside/base transport inhibitor dipyridamole (DP) on thymidine and hypoxanthine rescue defined. MTA IC50 values (continuous exposure three population doublings) were: A549-640 nM, COR L23-28 nM, MCF7-52 nM and T47D-46 nM. Thymidine (1 microM) completely prevented growth inhibition at the MTA IC50 in all cell lines. At 10 x IC50, growth inhibition was only partially reversed by thymidine (< or = 10 microM); both thymidine and hypoxanthine (30 microM) being required for complete reversal, reflecting the multi-targeted nature of MTA. Growth inhibition by MTA was not affected by hypoxanthine alone. A non-toxic concentration (1 microM) of DP prevented thymidine/hypoxanthine rescue of MTA indicating that DP may potentiate MTA activity by preventing nucleoside and/or base salvage. Thymidine transport was inhibited by > or = 89% by 1 microM DP in all cell lines, whereas hypoxanthine transport was inhibited only in A549 and MCF7 cells. Therefore, prevention of end-product reversal of MTA-induced growth inhibition by DP can be explained by inhibition of thymidine transport alone.     

Pemetrexed disodium in recurrent locally advanced or metastatic squamous cell carcinoma of the head and neck

This phase II study determined response rate of patients with locally advanced or metastatic head and neck cancer treated with pemetrexed disodium, a new multitargeted antifolate that inhibits thymidylate synthase, dihydrofolate reductase and glycinamide ribonucleotide formyl transferase. 35 patients with local or metastatic relapse of squamous cell carcinoma of the head and neck (31 male, 4 female; median age 53 years) were treated with pemetrexed 500 mg m(2)administered as a 10-minute infusion on day 1 of a 21-day cycle. Patients received 1 to 8 cycles of therapy. 9 patients (26.5%) had an objective response, with a median response duration of 5.6 months (range 2.9-20 months). 15 (44.1%) had stable disease, and 8 (23.5%) had progressive disease. 2 patients were not assessable for response. Median overall survival was 6.4 months (range 0.7-28.1 months; 95% CI: 3.9-7.7 months). 24 patients (68.6%) experienced grade 3/4 neutropenia, with febrile neutropenia in 4 (11.4%). Grade 3/4 anaemia and thrombocytopenia occurred in 11 (34.3%) and 6 (17.1%) patients, respectively. The most frequent non-haematological toxicity was grade 3/4 mucositis (17.1%; 6 patients). In conclusion, pemetrexed is active in squamous cell carcinoma of the head and neck. Although substantial haematological toxicities were experienced by patients, subsequent studies have shown that these toxicities can be proactively managed by folic acid and vitamin B(12)supplementation.     

A long noncoding RNA AB073614 promotes tumorigenesis and predicts poor prognosis in ovarian cancer

Long noncoding RNA (lncRNA) profiles in ovarian cancer (OC) remain largely unknown. In the present study, we screened {"type":"entrez-nucleotide","attrs":{"text":"AB073614","term_id":"51555790","term_text":"AB073614"}}AB073614 as a new candidate lncRNA which promotes development of OC, in two independent datasets ({"type":"entrez-geo","attrs":{"text":"GSE18521","term_id":"18521"}}GSE18521 and {"type":"entrez-geo","attrs":{"text":"GSE38666","term_id":"38666"}}GSE38666) from the Gene Expression Omnibus (GEO). The level of {"type":"entrez-nucleotide","attrs":{"text":"AB073614","term_id":"51555790","term_text":"AB073614"}}AB073614 was then detected in 75 paired OC tissues and adjacent normal tissues by qRT-PCR. Results showed that {"type":"entrez-nucleotide","attrs":{"text":"AB073614","term_id":"51555790","term_text":"AB073614"}}AB073614 expression was significantly up-regulated in 85.3% (64/75) cancerous tissues compared with normal counterparts (P < 0.01). Further, the 5-year overall survival (OS) in OC patients with high expression of {"type":"entrez-nucleotide","attrs":{"text":"AB073614","term_id":"51555790","term_text":"AB073614"}}AB073614 was inferior to that with low expression (17.2 months vs 30.0 months, P = 0.0025). To investigate the functional role of {"type":"entrez-nucleotide","attrs":{"text":"AB073614","term_id":"51555790","term_text":"AB073614"}}AB073614, {"type":"entrez-nucleotide","attrs":{"text":"AB073614","term_id":"51555790","term_text":"AB073614"}}AB073614 siRNA was transfected into OC cell lines. Knockdown of {"type":"entrez-nucleotide","attrs":{"text":"AB073614","term_id":"51555790","term_text":"AB073614"}}AB073614 expression significantly inhibited cell proliferation and invasion, resulted in cell arrest in G₁ phase of cell cycle and a dramatic increase of apoptosis, both in HO-8910 and OVCAR3 cells. In vivo experiment also revealed that knockdown {"type":"entrez-nucleotide","attrs":{"text":"AB073614","term_id":"51555790","term_text":"AB073614"}}AB073614 inhibited OVCAR3 cells proliferation. Finally, western blot assays indicated that lncRNA {"type":"entrez-nucleotide","attrs":{"text":"AB073614","term_id":"51555790","term_text":"AB073614"}}AB073614 may exert its function by targeting ERK1/2 and AKT-mediated signaling pathway. In conclusion, our study suggests that lncRNA {"type":"entrez-nucleotide","attrs":{"text":"AB073614","term_id":"51555790","term_text":"AB073614"}}AB073614 acts as a functional oncogene in OC development.     

A phase I study of LY317615 (enzastaurin) and temozolomide in patients with gliomas (EORTC trial 26054)

We report a phase 1 study to examine the safety and recommended dose of the oral protein kinase C-beta inhibitor (anti-angiogenic) enzastaurin in combination with single-agent temozolomide. The study was conducted in patients with recurrent glioblastoma or newly diagnosed disease that was not treatable with standard (chemo)radiotherapy. Patients were treated with standard dose temozolomide (200 mg/m(2) for 5 days every 4 weeks) together with daily oral enzastaurin. Three dose levels of enzastaurin were investigated: 250 mg daily (OD), 500 mg OD, and 250 mg twice daily (BID). Dose-limiting toxicity was determined in the first 2 cycles, but treatment continued until limiting toxicity or disease progression was identified. Twenty-eight patients were enrolled. No dose-limiting toxicity was noted at 250 mg OD or 500 mg OD. However, at 250 mg BID, 2 dose-limiting episodes of thrombocytopenia were noted. The recommended dose for enzastaurin in combination with standard 4-weekly temozolomide is therefore 500 mg OD. The pharmacokinetics of enzastaurin in combination with temozolomide was evaluated. Temozolomide did not appear to effect enzastaurin exposures at the 250 mg or 500 mg OD dose levels.     

Downregulation of LINC01508 contributes to cisplatin resistance in ovarian cancer via the regulation of the Hippo-YAP pathway

BackgroundSome long non-coding RNAs (lncRNAs) have been found to contribute to cisplatin resistance. Here, we identified a novel lncRNA that was downregulated in cisplatin-resistant to ovarian cancer (OC) cells and aimed to examine the contribution of LINC01508 to cisplatin resistance in OC cells.MethodsDifferences in the lncRNA expression profile between OV2008 and C13K cells were assessed by lncRNA expression microarray. The expression of LINC01508 in ovarian epithelial cells, four OC cells, and OC, benign ovary tumor and normal ovary, cisplatin-resistant and non-resistant OC specimens were evaluated by quantitative real-time polymerase chain reaction (qPCR). The role of LINC01508 in OC cisplatin-resistant was evaluated by cell counting kit-8 (CCK-8), flow cytometry, colony formation, wound healing, Transwell, and tumor growth inhibition study in vivo. The clinical associations of LINC01508 in OC were evaluated using correlation analysis. The effects of verteporfin (VP) on cisplatin were explored to reveal the function of the hippo-YAP pathway on the cisplatin tolerance of C13K.ResultsLINC01508 was downregulated in cisplatin-resistant OC cells and platinum-resistant OC tissue (p<0.01). LINC01508 downregulation was correlated with tumor size, residual tumor, and platinum resistance. The overexpression of LINC01508 improves in vitro and in vivo sensitivity to cisplatin while predicts the poor overall survival which need further follow-up research. The increased level of LINC01508 could suppress the cisplatin resistance of OC cells through the inhibition of the hippo-YAP pathway.ConclusionsThe study proposes that dysregulation of LINC01508 expression results in resistance of OC to cisplatin through the inhibition of the hippo-YAP pathway.     

Antagonism of adenosine A2A receptor expressed by lung adenocarcinoma tumor cells and cancer associated fibroblasts inhibits their growth

Recently it has become clear that the cost associated with the Warburg effect, which is inefficient production of ATP, is offset by selective advantages that are produced by resultant intracellular metabolic alterations. In fact tumors may be addicted to the Warburg effect. In addition these alterations result in changes in the extracellular tumor microenvironment that can also produce selective advantages for tumor cell growth and survival. One such extracellular alteration is increased adenosine concentrations that have been shown to impair T cell mediated rejection and support angiogenesis. The expression of the A2A receptor in non-small cell cancer (NSCLC) tissues, cell lines and cancer associated fibroblasts (CAF) was determined by performing immunohistrochemistry and immunoblot analysis. The efficacy of the A2A receptor antagonists in vivo was evaluated in a PC9 xenograft model. To determine the mode of cell death induced by A2A receptor antagonists flow cytometry, immunoblot, and cytotoxic analysis were performed. We found that a significant number of lung adenocarcinomas express adenosine A2A receptors. Antagonism of these receptors impaired CAF and tumor cell growth in vitro and inhibited human tumor xenograft growth in mice. These observations add to the rationale for testing adenosine A2A receptor antagonists as anticancer therapeutics. Not only could there be prevention of negative signaling in T cells within the tumor microenvironment and inhibition of angiogenesis, but also an inhibitory effect on tumor-promoting, immunosuppressive CAFs and a direct inhibitory effect on the tumor cells themselves.     

Inhibition of CXCR4–CXCL12 chemotaxis in melanoma by AMD11070

Background:

Despite intensive research and novel adjuvant therapies, there is currently no cure for metastatic melanoma. The chemokine receptor CXCR4 controls metastasis to sites such as the liver; however, the therapeutic blockade with the existing agents has proven difficult.

Methods:

{"type":"entrez-protein","attrs":{"text":"AMD11070","term_id":"985559755"}}AMD11070, a novel orally bioavailable inhibitor of CXCR4, was tested for its ability to inhibit the migration of melanoma cells compared with the commonly described antagonist AMD3100.

Results:

{"type":"entrez-protein","attrs":{"text":"AMD11070","term_id":"985559755"}}AMD11070 abrogated melanoma cell migration and was significantly more effective than AMD3100. Importantly for the clinical context, the expression of B-RAF-V600E did not the affect the sensitivity of {"type":"entrez-protein","attrs":{"text":"AMD11070","term_id":"985559755"}}AMD11070.

Conclusion:

Liver-resident myofibroblasts excrete CXCL12, which is able to promote the migration of CXCR4-expressing tumour cells from the blood into the liver. Blockade of this axis by {"type":"entrez-protein","attrs":{"text":"AMD11070","term_id":"985559755"}}AMD11070 thus represents a novel therapeutic strategy for both B-RAF wild-type and mutated melanomas.     

AdAPT-001, an oncolytic adenovirus armed with a TGF-β trap,overcomes in vivo resistance to PD-L1-immunotherapy

Monoclonal antibodies targeting the programmed cell death protein-1/programmed cell death-ligand 1 (PD-1/PD-L1) and cytotoxic T lymphocyte-associated protein-4 (CTLA-4) axes have permanently changed the therapeutic landscape for multiple tumor types previously associated with a dismal prognosis such as melanoma, non-small cell lung cancer, renal cell carcinoma, bladder cancer, head and neck squamous cell carcinoma, MSI-high colorectal carcinoma, Merkel cell carcinoma, and Hodgkin lymphoma. However, only a subset of patients initially benefits from these inhibitors, and increasing clinical experience indicates that in a substantial proportion of initial responders, lethal secondary resistance ultimately develops months or years later. In this paper we evaluated combination therapy with a Phase 1 oncolytic adenovirus called AdAPT-001, armed with a TGF-β “trap” that binds to and neutralizes the immunosuppressive cytokine, TGF-β, and a checkpoint inhibitor, anti-PD-L1, in PD-L1 resistant tumors. The study, which was performed in an immunocompetent syngeneic ADS-12 mouse model, demonstrated that the combination of AdAPT-001 with PD-L1 blockade reversed PD-L1 resistance, potentially representing a future paradigm shift for patients that are primarily or secondarily resistant to checkpoint inhibitors.     

苍术素通过激活RIPK1/RIPK3/MLKL 信号通路诱导非小细胞肺癌A549细胞程序性坏死并抑制裸鼠移植瘤生长

目的：探讨苍术素（ATR）通过调节受体相互作用蛋白激酶（RIPK）1/RIPK3/混合谱系激酶结构域样（MLKL）信号通路对非小细胞肺癌（NSCLC）A549 细胞程序性死亡及裸鼠移植瘤生长的影响。方法：使用0~160 μmol/L 的ATR 处理A549 细胞, MTT 法检测细胞存活率以确定后续实验给药浓度。使用ATR 和/或RIPK1 抑制剂Nec-1（necrostatin-1）、caspase 抑制剂Z-VAD-FMK处理A549 细胞,验证ATR是否诱导A549 细胞发生程序性坏死。将A549 细胞分为对照组、ATR-L 组、ATR-M 组、ATR-H 组（分别用0、10、20、40 μmol/L ATR 处理）、ATR+Nec-1 组（40 μmol/L ATR+50 μmol/L Nec-1 处理）,处理24 h 后,采用PI 单染及Hoechst33342/PI 双染法检测细胞死亡情况、透射电镜观察细胞死亡形态、DCFH-DA 荧光探针法检测细胞内ROS水平、JC-1染色法检测线粒体膜电位、WB法检测细胞中RIPK1/RIPK3/MLKL 信号通路相关蛋白质的表达水平。构建A549 细胞裸鼠移植瘤模型,用10 mg/kg ATR（溶于玉米油中）对裸鼠灌胃给药5 周,观察ATR 对移植瘤生长的影响,WB 法检测移植瘤组织中RIPK1/RIPK3/MLKL信号通路相关蛋白质的表达水平。结果：10~160 μmol/L的ATR可显著抑制A549细胞增殖,选择10、20、40 μmol/L的ATR进行后续实验。ATR组A549 细胞存活率显著低于对照组（P<0.01）和ATR+Nec-1组（P<0.01）,而ATR+z-VAD组细胞存活率显著低于z-VAD组（P<0.01）,说明ATR可诱导A549 细胞发生程序性坏死而非凋亡。与对照组比较,ATR处理组A549 细胞发生肿胀,线粒体内脊消失呈空泡化,细胞内容物向外泄漏,细胞核聚集,表现为坏死特征,ATR-L 组、ATR-M组、ATR-H 组A549 细胞死亡率、ROS水平及p-RIPK1、p-RIPK3、p-MLKL表达水平均显著升高,线粒体膜电位显著降低（均P<0.01）,且呈药物浓度依赖性;与ATR-H 组比较,ATR+Nec-1 组细胞死亡率、ROS 及p-RIPK1、p-RIPK3、p-MLKL 表达水平降低,线粒体膜电位显著升高（均P<0.01）。裸鼠移植瘤实验结果显示,与对照组比较,ATR 组裸鼠移植瘤体积、移植瘤质量均降低（P<0.05,或P<0.01）,而与瘤组织中p-RIPK1、p-RIPK3、p-MLKL 蛋白表达水平均显著升高（均P<0.01）。结论：ATR可能通过激活RIPK1/RIPK3/MLKL信号通路诱导A549细胞发生程序性坏死,抑制A549细胞及其裸鼠移植瘤的生长。