Phase 1 Trial of MLN0128 (Sapanisertib) and CB-839 HCl (Telaglenastat) in Patients With Advanced NSCLC (NCI 10327): Rationale and Study Design

IntroductionThere are currently no approved targeted therapies for lung squamous-cell carcinoma (LSCC) and KRAS-mutant lung adenocarcinoma (LUAD). About 30% of LSCC and 25% of KRAS-mutant LUAD exhibit hyperactive NRF2 pathway activation through mutations in NFE2L2 (the gene encoding NRF2) or its negative regulator, KEAP1. Preclinical data demonstrate that these tumors are uniquely sensitive to dual inhibition of glycolysis and glutaminolysis via mammalian target of rapamycin (mTOR) and glutaminase inhibitors. This phase 1 study was designed to assess safety and preliminary activity of the mTOR inhibitor MLN0128 (sapanisertib) in combination with the glutaminase inhibitor CB-839 HCl.MethodsPhase 1 dose finding will use the queue-based variation of the 3 + 3 dose escalation scheme with the primary endpoint of identifying the recommended expansion dose. To confirm the acceptable tolerability of the recommended expansion dose, patients will subsequently enroll onto 1 of 4 expansion cohorts (n = 14 per cohort): (1) LSCC harboring NFE2L2 or (2) KEAP1 mutations, or (3) LUAD harboring KRAS/(KEAP1 or NFE2L2) coalterations, or (4) LSCC wild type for NFE2L2 and KEAP1. The primary endpoint of the dose expansion is to determine the preliminary efficacy of MLN0128/CB-839 combination therapy.ConclusionThis phase 1 study will determine the recommended expansion dose and preliminary efficacy of MLN0128 and CB-839 in advanced non–small-cell lung cancer with a focus on subsets of LSCC and KRAS-mutant LUAD harboring NFE2L2 or KEAP1 mutations.     

糖酵解与肿瘤

肿瘤细胞主要通过糖酵解获取能量促进其生长,这与线粒体DNA变异和抑癌基因突变有关。抑制糖酵解具有抑制增殖和杀伤肿瘤细胞的作用。糖酵解限速酶和低氧诱导因子有望成为治疗肿瘤的新靶点。     

柠檬酸盐对人胃癌细胞株SGC7901糖酵解的抑制作用及其机制研究

背景：糖酵解是肿瘤细胞的主要能量来源。柠檬酸作为三羧酸循环的中间产物,对糖酵解具有抑制作用。目的：研究柠檬酸盐对人胃癌细胞糖酵解的抑制作用及其机制。方法：柠檬酸三钠以不同浓度、不同作用时间处理人胃癌细胞株SGC7901,以生化法检测细胞培养液乳酸含量,CCK-8法检测细胞增殖率,Hoechst凋亡染色观察细胞凋亡情况,ELISA法检测细胞内磷酸果糖激酶（PFK）含量,蛋白质印迹法检测caspase-3、-9、cleavedcaspase-3、Bc1-2、细胞色素（Cyt）、缺氧诱导因子-1d（HIF-1仪）、葡萄糖转运蛋白-1（GLUT-1）蛋白表达,定量RT—PCR法检测PFK亚型mRNA表达。结果：在糖培养条件下,经不同浓度、不同作用时间柠檬酸三钠处理的SGC7901细胞,细胞培养液乳酸含量、细胞增殖率、细胞内PFK含量、caspase-3、-9、Bc1-2、HIF-10【、GLUT-1蛋白表达、肌肉型PFK（PFK—M）比例较空白对照组显著降低,cleavedcaspase-3、Cyt蛋白表达较空白对照组显著增高,细胞内可见典型凋亡征象,上述作用多呈时间和浓度依赖性。结论：柠檬酸盐对人胃癌细胞糖酵解的抑制作用与抑制PFK、HIF-1仪、GLUT-1表达相关。柠檬酸盐可通过启动内源性凋亡途径诱导人胃癌细胞凋亡。     

miR-143 inhibits glycolysis and depletes stemness of glioblastoma stem-like cells

Glioblastomas rely mainly on aerobic glycolysis to sustain proliferation and growth; however, little is known about the regulatory mechanisms of metabolism in glioblastoma stem cells. We show that miR-143 is significantly down-regulated in glioma tissues and glioblastoma stem-like cells (GSLCs), while miR-143 over-expression inhibits glycolysis by directly targeting hexokinase 2, and promotes differentiation of GSLCs. Moreover, miR-143 inhibits proliferation of GSLCs under hypoxic conditions and decreases tumor formation capacity of GSLCs in vivo. We also show that a combination of miR-143 and 2-DG, a widely used glycolysis inhibitor, has synergistic effects against GSLCs. miR-143 is a potential therapeutic target for glioblastoma treatment.     

Intracellular acidification induced by membrane depolarization in rat hippocampal slices: roles of intracellular Ca and glycolysis

To elucidate the mechanism of pH_i changes induced by membrane depolarization, the variations in pH_i and [Ca²⁺]_i induced by a number of depolarizing agents, including high K⁺, veratridine, N-methyl-

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-aspartate (NMDA) and ouabain, were investigated in rat hippocampal slices by the fluorophotometrical technique using BCECF or fura-2. All of these depolarizing agents elicited a decrease in pH_i and an elevation of intracellular calcium ([Ca²⁺]_i) in the CA1 pyramidal cell layer. The increases in [Ca²⁺]_i caused by the depolarizing agents almost completely disappeared in the absence of Ca²⁺ (0 mM Ca²⁺ with 1 mM EGTA). In Ca²⁺ free media, pH_i acid shifts produced by high K⁺, veratridine or NMDA were attenuated by 10–25%, and those produced by ouabain decreased by 50%. Glucose-substitution with equimolar amounts of pyruvate suppressed by two-thirds the pH_i acid shifts induced by both high K⁺ and NMDA. Furthermore, lactate contents were significantly increased in hippocampal slices by exposure to high K⁺, veratridine or NMDA but not by ouabain. These results suggest that the intracellular acidification produced by these depolarizing agents, with the exception of ouabain, is mainly due to lactate accumulation which may occur as a result of accelerated glycolysis mediated by increased Na⁺–K⁺ ATPase activity. A Ca²⁺-dependent process may also contribute to the intracellular acidification induced by membrane depolarization. Since an increase in H⁺ concentration can attenuate neuronal activity, glycolytic acid production induced by membrane depolarization may contribute to the mechanism that prevents excessive neuronal excitation.     

Intracerebroventricular Injection of Fructose Stimulates Feeding in Rats

Abstract

2-Deoxy-d-glucose (2DG) inhibits glycolysis and stimulates food intake. Previous work suggests that fructose may attenuate the hyperglycemic and hypothermic effects of 2DG. The current study examined the effect of intracerebroventricular fructose on 2DG-induced feeding. We found that concentrated fructose injected into the cerebroventricles enhanced food intake both in the presence and absence of 2DG. On the other hand, similar concentrations of glucose suppressed 2DG-induced food intake. These data suggest differences in metabolism of glucose and fructose and may provide insight into the metabolic steps monitored by brain glucoreceptors to control food intake.     

Glycolysis revisited

Summary Glycolysis is usually considered as a paradigm metabolic pathway, due to the fact that it is present in most organisms, and also because it is the pathway by which an important nutrient, glucose, is consumed. Far from being completely understood, the regulation of this pathway witnessed several important progresses during the last few years. One of these is the discovery of fructose 2,6-bisphosphate, a potent stimulator of phosphofructokinase and inhibitor of fructose-1,6-bisphosphatase. Originally found in the liver during the course of a study on the mechanism by which glucagon acts on gluconeogenesis, this compound is now recognized as a major element in the control of glycolysis and/or gluconeogenesis in many cell types and in various organisms. The other finding is that of a regulatory protein that modulates the activity of glucokinase, the enzyme that phosphorylates glucose in the liver and in the beta cells of pancreatic islets.Presented as the Minkowski Lecture, EASD Meeting Prague, Czechoslovakia 1992     

Role of glycolysis in the energy production for the non-mechanical myocardial work in isolated pig hearts*

Summary

Background: The dissociation of mechanical from non-mechanical energy utilisation can be studied using BDM (2,3-butanedione monoxime), which inhibits the actin-myosin interaction without inhibiting Ca²⁺ transport. The objective of the present study was to establish if increasing the non-mechanical energy demand of perfused isolated pig hearts by dobutamine stimulation requires glycolysis with increased exogenous glucose uptake.

Methods: Five isolated pig hearts (CTRL) were perfused for 90?min at constant flow (1?ml?g^?1?min^?1) with non-recirculating blood containing 30?mM BDM and 26?MBq/l of fluorine-18 2-fluoro-2-deoxyglucose (¹⁸FDG). This was compared with five hearts (DOBU) subjected to the same protocol for the first 30?min and then to dobutamine (1.5?μM) for the following 30?min and dobutamine (4?μM) for the last 30?min. Five other isolated hearts were perfused as for the DOBU group but without BDM (CTRLDOBU). Using a clinical PET scanner, glucose uptake was assessed by estimating ¹⁸FDG uptake using linear regression. The slope variations were compared using a global test of coincidence.

Results: Heart rate was at 100?±?2 b.p.m. in the CTRL group and at 180?±?7 b.p.m. in the DOBU group. ¹⁸FDG uptake was homogeneous within the whole myocardium and we observed a linear and regular increase in both the CTRL and DOBU groups (p, NS). In the CTRLDOBU group, ¹⁸FDG uptake was also homogeneous within the whole myocardium, but slopes of ¹⁸FDG uptake during dobutamine perfusion were higher than without dobutamine.

Conclusion: In blood-perfused isolated pig hearts, exogenous glucose is not necessarily required when non-mechanical energy is increased by dobutamine stimulation. These findings suggest that ATP derived from glycolysis is not necessary to preserve myocardial Ca²⁺ transport during β-adrenergic stimulation.     

Carnosine inhibits KRAS-mediated HCT116 proliferation by affecting ATP and ROS production

Carnosine is a natural dipeptide that has generated particular interest for its antioxidant, anti-aging and especially for its antiproliferative properties. In this study, we demonstrate that carnosine inhibits the proliferation of human HCT116 colon cancer cells. In this cell line, the activating KRAS mutation induces mitochondrial ROS, the signaling molecules for cell proliferation. We observed that 50-100 mM carnosine decreases ATP and ROS concentration and induces cell cycle arrest in G1 phase. In HCT116 cells these effects are related to decreased ERK1/2 phosphorylation and increased p21^waf1 protein. Our findings support the concept that carnosine could inhibit HCT116 cell growth via its antioxidant activity and its ability to affect glycolysis.