Abstract:
Increased proliferation and reduced apoptosis of pulmonary artery smooth muscle cells (PASMCs) is recognised as a universal hallmark of pulmonary arterial hypertension (PAH), in part related to the association with reduced pyruvate dehydrogenase (PDH) activity, resulting in decreased oxidative phosphorylation of glucose and increased aerobic glycolysis (Warburg effect). Perhexiline is a well-recognised carnitine palmitoyltransferase-1 (CPT1) inhibitor used in cardiac diseases, which reciprocally increases PDH activity, but is associated with variable pharmacokinetics related to polymorphic variation of the cytochrome P450-2D6 (CYP2D6) enzyme, resulting in the risk of neuro and hepatotoxicity in \'slow metabolisers\' unless blood levels are monitored and dose adjusted. We have previously reported that a novel perhexiline fluorinated derivative (FPER-1) has the same therapeutic profile as perhexiline but is not metabolised by CYP2D6, resulting in more predictable pharmacokinetics than the parent drug. We sought to investigate the effects of perhexiline and FPER-1 on PDH flux in PASMCs from patients with PAH. We first confirmed that PAH PASMCs exhibited increased cell proliferation, enhanced phosphorylation of AKTSer473, ERK 1/2Thr202/Tyr204 and PDH-E1αSer293, indicating a Warburg effect when compared to healthy PASMCs. Pre-treatment with perhexiline or FPER-1 significantly attenuated PAH PASMC proliferation in a concentration-dependent manner and suppressed the activation of the AKTSer473 but had no effect on the ERK pathway. Perhexiline and FPER-1 markedly activated PDH (seen as dephosphorylation of PDH-E1αSer293), reduced glycolysis, and upregulated mitochondrial respiration in these PAH PASMCs as detected by Seahorse analysis. However, both perhexiline and FPER-1 did not induce apoptosis as measured by caspase 3/7 activity. We show for the first time that both perhexiline and FPER-1 may represent therapeutic agents for reducing cell proliferation in human PAH PASMCs, by reversing Warburg physiology.
摘要:
肺动脉平滑肌细胞(PASMCs)的增殖增加和凋亡减少被认为是肺动脉高压(PAH)的普遍标志,部分与丙酮酸脱氢酶(PDH)活性降低有关,导致葡萄糖的氧化磷酸化减少和有氧糖酵解增加(Warburg效应)。Perhexiline是公认的用于心脏疾病的肉碱棕榈酰转移酶-1(CPT1)抑制剂,反过来增加PDH活性,但与与细胞色素P4502D6(CYP2D6)酶的多态性变异相关的可变药代动力学相关,除非监测血液水平并调整剂量,否则会导致慢代谢者的神经和肝毒性风险。我们先前已经报道了一种新型的全氟辛宁衍生物(FPER-1)具有与全氟辛宁相同的治疗方案,但不被CYP2D6代谢,导致比母体药物更可预测的药代动力学。我们试图研究帕西汀和FPER-1对PAH患者PASMC中PDH通量的影响。我们首先证实PAHPASMCs表现出增加的细胞增殖,AKTSer473,ERK1/2和PDH-E1αSer293的磷酸化增强,表明与健康的PASMC相比具有Warburg效应。使用通己素或FPER-1预处理以浓度依赖性方式显着减弱PAHPASMC的增殖,并抑制AKTSer473的激活,但对ERK途径没有影响。Perhexiline和FPER-1显著激活PDH(被视为PDH-E1αSer293的去磷酸化),糖酵解减少,并通过海马分析检测到这些PAHPASMC中的线粒体呼吸上调。然而,通过caspase3/7活性测定,perhexiline和FPER-1均未诱导细胞凋亡.我们首次表明,perhexiline和FPER-1可以代表减少人PAHPASMC细胞增殖的治疗剂,通过逆转Warburg生理学。