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Molecular Medicine Reports Aug 2020Vascular complications are the primary reason for disability and mortality associated with diabetes mellitus (DM), and numerous microRNAs (miRNAs/miRs) are involved in...
Vascular complications are the primary reason for disability and mortality associated with diabetes mellitus (DM), and numerous microRNAs (miRNAs/miRs) are involved in the process, such as miR‑122, miR‑24 and miR‑423. It has been reported that miR‑328 regulates DM and cardiovascular disease; however, the role and mechanism of action underlying miR‑328 in HUVECs is not completely understood. The present study aimed to investigate the role and mechanism of action underlying the effects of miR‑328 on the functions of HUVECs. To simulate hyperglycemia combined with ischemia‑induced tissue starvation, HUVECs were cultured in endothelial cell medium with 25 mmol/l D‑glucose and 2% FBS for 24 h [high glucose (HG) + 2% FBS group]. HUVEC miR‑328 expression levels were detected by reverse transcription‑quantitative PCR. Cell migration, cytotoxicity and tube‑like structure formation were analyzed using wound healing, Cell Counting Kit‑8 and tube formation assays, respectively. Following transfection with miR‑328 inhibitor, miR‑328 expression was downregulated in HUVECs. Protein expression levels were determined by western blotting. Compared with the control group, the migration and tube‑like structure formation of HUVECs were decreased, and cell cytotoxicity was increased in the HG + 2% FBS group. The protein expression levels of vascular endothelial growth factor were also decreased, and the expression levels of miRNA‑328 in the HG + 2% FBS group were increased compared with the control group. However, miRNA‑328 downregulation reversed the aforementioned effects. Further experiments indicated that the AKT signaling pathway was inhibited in the HG + 2% FBS group; however, miR‑328 downregulation activated the AKT/mTOR signaling pathway, which was blocked by the AKT signaling pathway inhibitor, perifosine. Gene prediction and western blotting demonstrated that miR‑328 displayed a regulatory role via Pim‑1 proto‑oncogene, serine/threonine kinase (PIM1). In conclusion, miR‑328 expression was upregulated and angiogenesis was inhibited when HUVECs were subjected to high glucose and low serum conditions. miR‑328 downregulation enhanced angiogenesis by increasing PIM1 expression and activating the AKT/mTOR signaling pathway in HUVECs under high glucose and low serum conditions.
Topics: Binding Sites; Cell Movement; Cell Survival; Cells, Cultured; Culture Media; Databases, Genetic; Down-Regulation; Glucose; Human Umbilical Vein Endothelial Cells; Humans; MicroRNAs; Neovascularization, Physiologic; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-pim-1; Serum; Signal Transduction; TOR Serine-Threonine Kinases; Vascular Endothelial Growth Factor A
PubMed: 32626978
DOI: 10.3892/mmr.2020.11141 -
Cell Stress & Chaperones Nov 20205-Hydroxytryptamine receptor 2A (HTR2A) is a central regulator of fetal brain development and cognitive function in adults. However, the roles of HTR2A in the...
5-Hydroxytryptamine receptor 2A (HTR2A) is a central regulator of fetal brain development and cognitive function in adults. However, the roles of HTR2A in the cardiovascular system are not fully understood. Here in this study, we explored the function of HTR2A in cardiac hypertrophy. Significantly, the expression levels of HTR2A mRNA and protein levels were upregulated in hypertrophic hearts of human patients. Besides, the expression of HTR2A was also upregulated in isoproterenol (ISO)-induced cardiac hypertrophy in the mouse. Next, the expression of HTR2A was knocked down with shRNA or overexpressed with adenovirus in neonatal rat cardiomyocytes, and ISO was used to induce cardiomyocyte hypertrophy. We showed that HTR2A knockdown repressed ISO-induced cardiomyocyte hypertrophy, which was demonstrated by decreased cardiomyocyte size and repressed expression of hypertrophic fetal genes (e.g., myosin heavy chain beta (β-Mhc), atrial natriuretic peptide (Anp), and brain natriuretic peptide (Bnp)). By contrast, HTR2A overexpression promoted cardiomyocyte hypertrophy. Of note, we observed that HTR2A promoted the activation (phosphorylation) of AKT-mTOR (mammalian target of rapamycin) signaling in cardiomyocytes, and repression of AKT-mTOR with perifosine or rapamycin blocked the effects of HTR2A on cardiomyocyte hypertrophy. Finally, we showed that HTR2A regulated AKT-mTOR signaling through activating the PI3K-PDK1 pathway, and inhibition of either PI3K or PDK1 blocked the roles of HTR2A in regulating AKT-mTOR signaling and cardiomyocyte hypertrophy. Altogether, these findings demonstrated that HTR2A activated PI3K-PDK1-AKT-mTOR signaling and promoted cardiac hypertrophy.
Topics: 3-Phosphoinositide-Dependent Protein Kinases; Animals; Animals, Newborn; Cardiomegaly; Humans; Isoproterenol; Male; Mice, Inbred C57BL; Models, Biological; Myocytes, Cardiac; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Rats, Sprague-Dawley; Receptor, Serotonin, 5-HT2A; Signal Transduction; TOR Serine-Threonine Kinases
PubMed: 32519137
DOI: 10.1007/s12192-020-01124-x -
EXCLI Journal 2020Endometrial cancer is the most common cancer of the female reproductive system. Combination treatment with specific agents has been widely used as a targeted therapy for...
Endometrial cancer is the most common cancer of the female reproductive system. Combination treatment with specific agents has been widely used as a targeted therapy for cancer. In this study, we aimed to investigate the anti-proliferative and apoptotic effects of varying concentrations of perifosine and vitamin D on the human endometrial cancer cell line (HEC-1A). HEC-1A cells were exposed to perifosine (10 μM, 30 μM), vitamin D (50 nM, 200 nM) and combinations of both for 48 h and 72 h. Monitoring of cell proliferation in a time-dependent manner was performed with the xCELLigence RTCA DP system. The levels of BCL2, BAX and P53 mRNA expression were examined using RT-qPCR. Apoptosis was determined using Annexin V, which were followed by flow cytometry analysis. Ultra-structural morphology of cells was analyzed by transmission electron microscopy (TEM) for 72 h. The anti-proliferative and apoptotic effects of the perifosine+vitamin D combination (30 μM + 200 nM at 48 h and 10 μM + 200 nM at 72 h) on HEC-1A cells were higher than in perifosine and vitamin D alone. It was observed that perifosine has increased the expression of BAX mRNA in HEC-1A cells in a dose-dependent manner. While perifosine+vitamin D combinations increased P53 mRNA expression in HEC-1A cells we did not find any significant change in BCL2, BAX mRNA expression levels. In TEM examinations of HEC-1A cells, perifosine appeared to lead autophagic cell death, whereas vitamin D caused paraptosis-like cell death and combination of perifosine+vitamin D caused apoptotic and non-apoptotic (paraptotic, autophagic and necrotic) cell death. Therefore, it is considered that the combination of both drugs in the treatment of endometrial cancer might be an alternative and effective treatment option through activating the apoptotic and non-apoptotic cell death mechanisms in cancer cells.
PubMed: 32483402
DOI: 10.17179/excli2019-1834 -
Pharmaceutical Research May 2020Hemolysis is a serious side effect of antitumor alkylphospholipids (APLs) that limits dose levels and is a constraint in their use in therapeutic regimen. Nine prodrugs...
PURPOSE
Hemolysis is a serious side effect of antitumor alkylphospholipids (APLs) that limits dose levels and is a constraint in their use in therapeutic regimen. Nine prodrugs of promising APLs (miltefosine, perifosine, and erufosine) were synthesized so as to decrease their membrane activity and improve their toxicity profile while preserving their antineoplastic potency.
METHODS
The synthesis of the pro-APLs was straightforwardly achieved in one step starting from the parent APLs. The critical aggregation concentration of the prodrugs, their hydrolytic stability under various pH conditions, their blood compatibility and cytotoxicity in three different cell lines were determined and compared to those of the parent antitumor lipids.
RESULTS
The APL prodrugs display antitumor activity which is similar to that of the parent alkylphospholipids but without associated hemolytic toxicity.
CONCLUSION
The pro-APL compounds may be considered as intravenously injectable derivatives of APLs. They could thus address one of the major issues met in cancer therapies involving antitumor lipids and restricting their utilization to oral and topical administration because of limited maximum tolerated dose.
Topics: Administration, Intravenous; Antineoplastic Agents; Cell Line, Tumor; Drug Screening Assays, Antitumor; Drug Stability; Hemolysis; Humans; Maximum Tolerated Dose; Neoplasms; Organophosphates; Phosphorylcholine; Prodrugs; Quaternary Ammonium Compounds
PubMed: 32462253
DOI: 10.1007/s11095-020-02830-y -
Journal of Pharmacological Sciences Jul 2020The optimum strategy for heart failure (HF) treatment has yet to be elucidated. This study intended to test the benefit of a combination of valsartan (VAL) and...
The optimum strategy for heart failure (HF) treatment has yet to be elucidated. This study intended to test the benefit of a combination of valsartan (VAL) and perifosine (PER), a specific AKT inhibitor, in protecting against pressure overload induced mouse HF. Mouse were subjected to aortic banding (AB) surgery to establish HF models and then were given vehicle (HF), VAL (50 mg/kg/d), PER (30 mg/kg/d) or combination of VAL and PER for 4 weeks. Mouse with sham surgery treated with VEH were used for control (VEH). VAL or PER treatment could significantly alleviate mouse heart weight, attenuate cardiac fibrosis and improve cardiac function. The combination treatment of VAL and PER presented much better benefit compared with VAL or PER group respectively. PER treatment significantly inhibited AKT/GSK3β/mTORC1 signaling. Besides the classic AT1 inhibition, VAL treatment significantly inhibited MAPK (ERK1/2) signaling. Furthermore, VAL and PER treatment could markedly prevent neonatal rat cardiomyocyte hypertrophy and the activation of neonatal rat cardiac fibroblast. Combination of VAL and PER also presented superior beneficial effects than single treatment of VAL or PER in vitro experiments respectively. This study presented that the combination of valsartan and PER may be a potential treatment for HF prevention.
Topics: Animals; Disease Models, Animal; Drug Therapy, Combination; Glycogen Synthase Kinase 3 beta; Heart Failure; MAP Kinase Signaling System; Male; Mice, Inbred C57BL; Organ Size; Phosphorylcholine; Pressure; Proto-Oncogene Proteins c-akt; Signal Transduction; Valsartan
PubMed: 32414690
DOI: 10.1016/j.jphs.2020.04.001 -
Frontiers in Pharmacology 2020Autophagy is a highly conserved multistep process and functions as passage for degrading and recycling protein aggregates and defective organelles in eukaryotic cells.... (Review)
Review
Autophagy is a highly conserved multistep process and functions as passage for degrading and recycling protein aggregates and defective organelles in eukaryotic cells. Based on the nature of these materials, their size and degradation rate, four types of autophagy have been described, chaperone mediated autophagy, microautophagy, macroautophagy, and selective autophagy. One of the major regulators of this process is mTOR, which inhibits the downstream pathway of autophagy following the activation of its complex 1 (mTORC1). Alkylphosphocholine (APC) derivatives represent a novel class of antineoplastic agents that inhibit the serine-threonine kinase Akt ( protein kinase B), which mediates cell survival and cause cell cycle arrest. They induce autophagy through inhibition of the Akt/mTOR cascade. They interfere with phospholipid turnover and thus modify signaling chains, which start from the cell membrane and modulate PI3K/Akt/mTOR, Ras-Raf-MAPK/ERK and SAPK/JNK pathways. APCs include miltefosine, perifosine, and erufosine, which represent the first-, second- and third generation of this class, respectively. In a high fraction of human cancers, constitutively active oncoprotein Akt1 suppresses autophagy and . mTOR is a down-stream target for Akt, the activation of which suppresses autophagy. However, treatment with APC derivatives will lead to dephosphorylation (hence deactivation) of mTOR and thus induces autophagy. Autophagy is a double-edged sword and may result in chemotherapeutic resistance as well as cancer cell death when apoptotic pathways are inactive. APCs display differential autophagy induction capabilities in different cancer cell types. Therefore, autophagy-dependent cellular responses need to be well understood in order to improve the chemotherapeutic outcome.
PubMed: 32410999
DOI: 10.3389/fphar.2020.00547 -
Annals of Clinical and Translational... Apr 2020Malignant glioma (MG) is the most deadly primary brain cancer. Signaling though the PI3K/AKT/mTOR axis is activated in most MGs and therefore a potential therapeutic...
PURPOSE
Malignant glioma (MG) is the most deadly primary brain cancer. Signaling though the PI3K/AKT/mTOR axis is activated in most MGs and therefore a potential therapeutic target. The mTOR inhibitor temsirolimus and the AKT inhibitor perifosine are each well-tolerated as single agents but with limited activity reclinical data demonstrate synergistic anti-tumor effects from combined treatment. Therefore, we initiated a phase I trial of combined therapy in recurrent MGs to determine safety and a recommended phase II dose.
METHODS
Adults with recurrent MG, Karnofsky Performance Status ≥ 60 were enrolled, with no limit on the number of prior therapies. Temsirolimus dose was escalated using standard 3 + 3 design from 15 mg to 170 mg administered once weekly. Perifosine was fixed as a 600 mg load on day 1 followed by 100 mg nightly (single agent MTD) until dose level 7 when the load increased to 900 mg.
RESULTS
We treated 35 patients with with glioblastoma (17) or other MGs (18; including nine anaplastic astrocytoma, nine anaplastic oligodendroglioma, one anaplastic oligoastrocytoma, and two low grade astrocytomas with radiographic transformation to MG). We observed five dose-limiting toxicities (DLTs): one at dose level 3 (50mg temsirolimus), then two at dose level 7 expansion (170 mg temsirolimus), and then two more at dose level 6 expansion (170 mg temsirolimus). DLTs included thrombocytopenia (n = 3), intracerebral hemorrhage (n = 1) and lung infection (n = 1).
CONCLUSION
Combining the mTOR inhibitor temsirolimus dosed at 115 mg weekly and the AKT inhibitor perifosine dosed at 100 mg daily (following 600 mg load) is tolerable in heavily pretreated adults with recurrent MGs.
Topics: Adult; Aged; Antineoplastic Agents; Brain Neoplasms; Drug Therapy, Combination; Female; Glioblastoma; Humans; Male; Middle Aged; Neoplasm Recurrence, Local; Phosphorylcholine; Prospective Studies; Proto-Oncogene Proteins c-akt; Sirolimus; TOR Serine-Threonine Kinases; Young Adult
PubMed: 32293798
DOI: 10.1002/acn3.51009 -
ACS Pharmacology & Translational Science Feb 2020-amplified neuroblastoma is one of the deadliest forms of childhood cancer and remains a significant clinical challenge. Direct pharmacological inhibition of MYCN is not...
-amplified neuroblastoma is one of the deadliest forms of childhood cancer and remains a significant clinical challenge. Direct pharmacological inhibition of MYCN is not currently achievable. One strategy could be to target the AKT/GSK3β pathway, which directly regulates the stability of the MYCN protein. Numerous potent and isoform-specific small-molecule AKT inhibitors have been developed. However, the selection of the right drug combinations in the relevant indication will have a significant impact on AKT inhibitor clinical success. To maximally exploit the potential of AKT inhibitors, a better understanding of AKT isoform functions in cancer is crucial. Here using RNAi to downregulate specific AKT isoforms, we demonstrated that loss of total AKT activity rather than isoform-specific expression was necessary to decrease MYCN expression and cause a significant decrease in neuroblastoma cell proliferation. Consistent with these observations, isoform-specific pharmacological inhibition of AKT was substantially less effective than pan-AKT inhibition in combination with cytotoxic drugs in -amplified neuroblastoma. The allosteric pan-AKT inhibitor perifosine had promising and activity in combination with conventional cytotoxic drugs in -amplified neuroblastoma cells. Our results demonstrated that perifosine drug combination was able to induce apoptosis and downregulate ABC transporter expression. Collectively, this study shows that selecting pan-AKT inhibitors rather than isoform-specific drugs to synergize with first-line chemotherapy treatment should be considered for clinical trials for aggressive neuroblastoma and, potentially, other MYCN -driven cancers.
PubMed: 32259094
DOI: 10.1021/acsptsci.9b00085 -
EJHaem Jul 2020Perifosine, an investigational, oral, synthetic alkylphospholipid, inhibits signal transduction pathways of relevance in multiple myeloma (MM) including PI3K/Akt....
Randomized, placebo-controlled, phase 3 study of perifosine combined with bortezomib and dexamethasone in patients with relapsed, refractory multiple myeloma previously treated with bortezomib.
Perifosine, an investigational, oral, synthetic alkylphospholipid, inhibits signal transduction pathways of relevance in multiple myeloma (MM) including PI3K/Akt. Perifosine demonstrated anti-MM activity in preclinical studies and encouraging early-phase clinical activity in combination with bortezomib. A randomized, double-blind, placebo-controlled phase 3 study was conducted to evaluate addition of perifosine to bortezomib-dexamethasone in MM patients with one to four prior therapies who had relapsed following previous bortezomib-based therapy. The primary endpoint was progression-free survival (PFS). The study was discontinued at planned interim analysis, with 135 patients enrolled. Median PFS was 22.7 weeks (95% confidence interval 16·0-45·4) in the perifosine arm and 39.0 weeks (18.3-50.1) in the placebo arm (hazard ratio 1.269 [0.817-1.969]; = .287); overall response rates were 20% and 27%, respectively. Conversely, median overall survival (OS) was 141.9 weeks and 83.3 weeks (hazard ratio 0.734 [0.380-1.419]; = .356). Overall, 61% and 55% of patients in the perifosine and placebo arms reported grade 3/4 adverse events, including thrombocytopenia (26% vs 14%), anemia (7% vs 8%), hyponatremia (6% vs 8%), and pneumonia (9% vs 3%). These findings demonstrate no PFS benefit from the addition of perifosine to bortezomib-dexamethasone in this study of relapsed/refractory MM, but comparable safety and OS.
PubMed: 35847734
DOI: 10.1002/jha2.4 -
Chemistry and Physics of Lipids Jul 2020Anti-tumour lipids are synthetic analogues of lysophosphatidylcholine. These drugs are both cytotoxic and cytostatic, and more interestingly, exert these effects...
Anti-tumour lipids are synthetic analogues of lysophosphatidylcholine. These drugs are both cytotoxic and cytostatic, and more interestingly, exert these effects preferentially in tumour cells. While the exact mechanism of action isn't fully elucidated, these drugs appear to preferentially partition into rigid lipid domains in cell membranes. Upon insertion, the compounds alter membrane domain organization, disrupt normal signal transduction, and cause cell death. Recently, it has been reported that these drugs induce accumulation of diacylglycerol in yeast cells which in turn sensitizes cells to the drugs. Conversely, phosphatidic acid accumulation appears to protect cells against the drugs. In the current work, the aim was to compare the biophysical effects of the drugs edelfosine, miltefosine and perifosine on monolayers of dimyristoyl phosphatidic acid, dimyristoyl glycerol and an equimolar mixture, to understand how these lipids modulate the mode of action. Surface pressure - area isotherms, compression moduli and Brewster angle microscopy were used to compare drug effects on lipid packing, monolayer compressibility and lateral domain organization of these films. Results suggest that edelfosine and miltefosine have stabilizing effects on all of the monolayers, while perifosine destabilizes dimyristoyl glycerol and the equimolar mixture. Additionally, all three drugs change the morphology of the domains observed. Based on these results the stabilization of diacylgylcerol by edelfosine and miltefosine may contribute to the mode of action as diacylglycerol is a known disruptor of bilayers. Perifosine however does not stabilize diacylglycerol, and therefore cell death may occur through a more direct inhibition of specific signal transduction. These results suggest that perifosine may illicit cytotoxicity through a different mechanism compared to the other antitumor lipid drugs.
Topics: Antineoplastic Agents; Diglycerides; Membrane Lipids; Membrane Microdomains; Microscopy; Phosphatidic Acids
PubMed: 32184083
DOI: 10.1016/j.chemphyslip.2020.104896