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Reversal of the glycolytic phenotype of primary effusion lymphoma cells by combined targeting of cellular metabolism and PI3K/Akt/ mTOR signaling.

ABSTRACT: PEL is a B-cell non-Hodgkin lymphoma, occurring predominantly as a lymphomatous effusion in body cavities, characterized by aggressive clinical course, with no standard therapy. Based on previous reports that PEL cells display a Warburg phenotype, we hypothesized that the highly hypoxic environment in which they grow in vivo makes them more reliant on glycolysis, and more vulnerable to drugs targeting this pathway. We established here that indeed PEL cells in hypoxia are more sensitive to glycolysis inhibition. Furthermore, since PI3K/Akt/mTOR has been proposed as a drug target in PEL, we ascertained that pathway-specific inhibitors, namely the dual PI3K and mTOR inhibitor, PF-04691502, and the Akt inhibitor, Akti 1/2, display improved cytotoxicity to PEL cells in hypoxic conditions. Unexpectedly, we found that these drugs reduce lactate production/extracellular acidification rate, and, in combination with the glycolysis inhibitor 2-deoxyglucose (2-DG), they shift PEL cells metabolism from aerobic glycolysis towards oxidative respiration. Moreover, the associations possess strong synergistic cytotoxicity towards PEL cells, and thus may reduce adverse reaction in vivo, while displaying very low toxicity to normal lymphocytes. Finally, we showed that the association of 2-DG and PF-04691502 maintains its cytotoxic and proapoptotic effect also in PEL cells co-cultured with human primary mesothelial cells, a condition known to mimic the in vivo environment and to exert a protective and pro-survival action. All together, these results provide a compelling rationale for the clinical development of new therapies for the treatment of PEL, based on combined targeting of glycolytic metabolism and constitutively activated signaling pathways.

SUBMITTER: Mediani L

PROVIDER: S-EPMC4868703 | biostudies-other | 2016 Feb

REPOSITORIES: biostudies-other

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Reversal of the glycolytic phenotype of primary effusion lymphoma cells by combined targeting of cellular metabolism and PI3K/Akt/ mTOR signaling.

Mediani Laura L Gibellini Federica F Bertacchini Jessika J Frasson Chiara C Bosco Raffaella R Accordi Benedetta B Basso Giuseppe G Bonora Massimo M Calabrò Maria Luisa ML Mattiolo Adriana A Sgarbi Gianluca G Baracca Alessandra A Pinton Paolo P Riva Giovanni G Rampazzo Enrico E Petrizza Luca L Prodi Luca L Milani Daniela D Luppi Mario M Potenza Leonardo L De Pol Anto A Cocco Lucio L Capitani Silvano S Marmiroli Sandra S

Oncotarget 20160201 5

PEL is a B-cell non-Hodgkin lymphoma, occurring predominantly as a lymphomatous effusion in body cavities, characterized by aggressive clinical course, with no standard therapy. Based on previous reports that PEL cells display a Warburg phenotype, we hypothesized that the highly hypoxic environment in which they grow in vivo makes them more reliant on glycolysis, and more vulnerable to drugs targeting this pathway. We established here that indeed PEL cells in hypoxia are more sensitive to glycol ...[more]

PMID: 26575168

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Project description:Primary effusion lymphoma (PEL) is caused by Kaposi's sarcoma-associated herpesvirus (KSHV). PEL has a highly active mTOR pathway, which makes mTOR a potential therapeutic target. MLN0128 is an ATP-competitive inhibitor of mTOR that has entered clinical trials for solid tumors. Our results demonstrated that MLN0128 has a greater effect on inhibiting proliferation than the allosteric mTOR inhibitor rapamycin. MLN0128 has ∼30 nM 50% inhibitory concentration (IC50) across several PEL cell lines, including PEL that is resistant to conventional chemotherapy. MLN0128 induced apoptosis in PEL, whereas rapamycin induced G1 arrest, consistent with a different mechanism of action. MLN0128 inhibited phosphorylation of mTOR complex 1 and 2 targets, while rapamycin only partially inhibited mTOR complex 1 targets. PEL xenograft mouse models treated with MLN0128 showed reduced effusion volumes in comparison to the vehicle-treated group. Rapamycin-resistant (RR) clones with an IC50 for rapamycin 10 times higher than the parental IC50 emerged consistently after rapamycin exposure as a result of transcriptional adaptation. MLN0128 was nevertheless capable of inducing apoptosis in these RR clones. Our results suggest that MLN0128 might offer a new approach to the treatment of chemotherapy-resistant PEL.IMPORTANCE Primary effusion lymphoma (PEL) is an aggressive and incurable malignancy, which is usually characterized by lymphomatous effusions in body cavities without tumor masses. PEL has no established treatment and a poor prognosis, with a median survival time shorter than 6 months. PEL usually develops in the context of immunosuppression, such as HIV infection or post-organ transplantation. The optimal treatment for PEL has not been established, as PEL is generally resistant to traditional chemotherapy. The molecular drivers for PEL are still unknown; however, PEL displays a constitutively active mammalian target of rapamycin (mTOR) pathway, which is critical for metabolic and cell survival mechanisms. Therefore, the evaluation of novel agents targeting the mTOR pathway could be clinically relevant for the treatment of PEL.

Project description:Resistance to radiotherapy (IR), with consequent disease recurrence, continues to limit the efficacy of contemporary anticancer treatment for patients with hepatocellular carcinoma (HCC), especially in late stage. Despite accruing evidence implicating the PI3K/AKT signaling pathway in cancer-promoting hypoxia, cancerous cell proliferation and radiotherapy-resistance, it remains unclear which molecular constituent of the pathway facilitates adaptation of aggressive HCC cells to tumoral stress signals and drives their evasion of repeated IR-toxicity. This present study investigated the role of PDK1 signaling in IR-resistance, enhanced DNA damage repair and post-IR relapse, characteristic of aggressive HCC cells, while exploring potential PDK1-targetability to improve radiosensitivity. The study employed bioinformatics analyses of gene expression profile and functional protein-protein interaction, generation of IR-resistant clones, flow cytometry-based ALDH activity and side-population (SP) characterization, siRNA-mediated loss-of-PDK1function, western-blotting, immunohistochemistry and functional assays including cell viability, migration, invasion, clonogenicity and tumorsphere formation assays. We showed that the aberrantly expressed PDK1 characterizes poorly differentiated HCC CVCL_7955, Mahlavu, SK-HEP1 and Hep3B cells, compared to the well-differentiated Huh7 or normal adult liver epithelial THLE-2 cells, and independently activates the PI3K/AKT/mTOR signaling. Molecular ablation of PDK1 function enhanced susceptibility of HCC cells to IR and was associated with deactivated PI3K/AKT/mTOR signaling. Additionally, PDK1-driven IR-resistance positively correlated with activated PI3K signaling, enhanced HCC cell motility and invasiveness, augmented EMT, upregulated stemness markers ALDH1A1, PROM1, SOX2, KLF4 and POU5F1, increased tumorsphere-formation efficiency and suppressed biomarkers of DNA damage-RAD50, MSH3, MLH3 and ERCC2. Furthermore, the acquired IR-resistant phenotype of Huh7 cells was strongly associated with significantly increased ALDH activity, SP-enrichment, and direct ALDH1-PDK1 interaction. Moreover, BX795-mediated pharmacological inhibition of PDK1 synergistically enhances the radiosensitivity of erstwhile resistant cells, increased Bax/Bcl-2 apoptotic ratio, while suppressing oncogenicity and clonogenicity. We provide preclinical evidence implicating PDK1 as an active driver of IR-resistance by activation of the PI3K/AKT/mTOR signaling, up-modulation of cancer stemness signaling and suppression of DNA damage, thus, projecting PDK1-targeting as a putative enhancer of radiosensitivity and a potential new therapeutic approach for patients with IR-resistant HCC.

Dataset Information

Reversal of the glycolytic phenotype of primary effusion lymphoma cells by combined targeting of cellular metabolism and PI3K/Akt/ mTOR signaling.

Publications

Reversal of the glycolytic phenotype of primary effusion lymphoma cells by combined targeting of cellular metabolism and PI3K/Akt/ mTOR signaling.

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