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Novel complementary antitumour effects of celastrol and metformin by targeting IκBκB, apoptosis and NLRP3 inflammasome activation in diethylnitrosamine-induced murine hepatocarcinogenesis

Cancer Chemotherapy and Pharmacology Aims and scope Submit manuscript

Abstract

One promising strategy for minimizing chemotherapeutic resistance in hepatocellular carcinoma (HCC) is the use of effective chemosensitizers. We studied the complementary multi-targeted molecular mechanisms of metformin and celastrol in mice with diethylnitrosamine-induced HCC to investigate whether metformin could augment the sensitivity of HCC tissue to the effect of celastrol. Simultaneous administration of celastrol (2 mg/kg) and metformin (200 mg/kg) improved liver function, enhanced the histological picture and prolonged survival. Additionally, combination therapy exerted anti-inflammatory activity, as indicated by the decreased levels of TNF-α and IL-6. This protective role could be attributed to inhibition of inflammasome activation. Herein, our data revealed downregulated NLRP3 gene expression, suppressed caspase-1 activity and reduced levels of the active forms of IL-1β and IL-18. Under this condition, pyroptotic activity was suppressed. In contrast, in the celastrol and celastrol + metformin groups, the apoptotic potential was amplified, as revealed by the increase in the caspase-9 and caspase-3 levels and Bax:BCL-2 ratio. In addition to their repressive effect on the gene expression of NFκBp65, TNFR and TLR4, metformin and celastrol inhibited phosphorylation-induced activation of IκBκB and NFκBp65 and decreased IκBα degradation. Combination therapy with metformin and celastrol repressed markers of angiogenesis, metastasis and tumour proliferation, as revealed by the decreased hepatic levels of VEGF, MMP-2/9 and cyclin D1 mRNA, respectively. In conclusion, by inhibiting NLRP3 inflammasome and its prerequisite NFκB signalling, simultaneous administration of metformin and celastrol appears to have additive benefits in the treatment of HCC compared to cela monotherapy. This effect warrants further clinical investigation.

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Abbreviations

AFP:

Alpha-fetoprotein

AKT:

Protein kinase B

Bax:

BCL-2 associated × protein

BCL-2:

B-cell lymphoma-2

cel:

Celastrol

ECM:

Extracellular matrix

DEN:

Diethylnitrosamine

GAPDH:

Glyceraldehyde 3-phosphate dehydrogenase

HCC:

Hepatocellular carcinoma

IκBα:

IκBκB: the inhibitor κB kinase complex

MAPK:

Mitogen-activated protein kinase

MDA:

Malondialdehyde

met:

Metformin

MMP:

Matrix metalloproteinase

NFκB:

Nuclear transcription factor kappa B

NLRP3:

Nod-like receptor protein 3

PI3K:

Phosphoinositide 3-kinase

TLR:

Toll like receptor

TNF-α:

Tumour necrosis factor-alpha

VEGF:

Vascular endothelial growth factor

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Acknowledgements

My deepest sense of gratitude and respectful regards to Dr. Adel Bakeer, professor of pathology, Cairo University, for his indispensable help during the histopathological examination of liver tissues. His long time experience in the field had a remarkable influence on histological interpretations.

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Conceptualization of this research idea, methodology development, experiments, data collection, data analysis, survival analysis, editing and interpretation were implemented by SS; writing—original draft preparation, interpretation, literature review and analysis were implemented by AMG and EMA; qRT-PCR, ELISA, editing and final revision were implemented by EEA.

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Correspondence to Sameh Saber.

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Saber, S., Ghanim, A.M.H., El-Ahwany, E. et al. Novel complementary antitumour effects of celastrol and metformin by targeting IκBκB, apoptosis and NLRP3 inflammasome activation in diethylnitrosamine-induced murine hepatocarcinogenesis. Cancer Chemother Pharmacol 85, 331–343 (2020). https://doi.org/10.1007/s00280-020-04033-z

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