Combination of Metformin and Valproic acid for prostate cancer treatment

Author: Linh N.K Tran

Tran, Linh N.K, 2018 Combination of Metformin and Valproic acid for prostate cancer treatment, Flinders University, College of Medicine and Public Health

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Prostate cancer (PCa) is the most common cancer in developed countries. Despite aggressive local treatments such as radical prostatectomy and high-dose radiotherapy, 20-30% of men will relapse after 5-10 years. Androgen deprivation therapy and chemotherapy are used for the treatment of metastatic disease, but castration-resistant prostate cancer (CRPC) invariably emerges resulting in fatal outcomes. There is no durably effective targeted therapy for this stage of the disease. At present, FDA-approved treatments for CRPC consist of abiraterone acetate, enzalutamide, docetaxel, cabazitaxel, radium 223 and Sipuleucel-T, each of which only extend survival by a mean of 3-6 months. So, new treatment options are urgently required.

Metformin (MET; N’, N-dimethylbiguanide) is widely used in diabetic patients as first-line therapy for control of blood glucose. Recent studies show pleiotropic beneficial effects of MET including in cancer prevention. The antiepileptic drug, valproic acid (VPA) is a broad-range histone deacetylase inhibitor (HDACi) targeting HDAC class I (Ia, Ib) and class II (IIa) enzymes, and has been shown to have potential as an anti-cancer therapeutic. MET and VPA, when used alone, have shown varying anti-cancer effects in PCa. However, high doses are required which results in unacceptable toxicity. The mechanisms of both drugs have been widely studied.

Repurposing of commonly used drugs is an attractive approach for rapid development of new systemic cancer therapy options. Both MET and VPA have limitations when used alone as anti-cancer agents, however combinatorial therapy of MET and VPA could potentially be more effective than monotherapy as: (1) MET and VPA induce cell cycle arrest, and antiproliferative and proapoptotic effects via different biological pathways; (2) Lower doses of VPA and MET in combination could reduce the toxicity observed with higher doses of the single drugs; (3) Translation into the clinic would be rapid as MET and VPA are currently widely used drugs for non-cancer purposes.

Here the potential of combining MET and VPA (MET+VPA) was investigated in order to determine if the combination could be used to enhance anti-tumour responses in PCa. In this thesis, the findings demonstrated that MET+VPA induced a synergistic anti-proliferative effect in vitro (human PCa cell lines LNCaP and PC-3), inhibited proliferation in vivo (nude mouse xenografts bearing LNCaP and PC-3 cells) compared to either drug alone, and reduced proliferation ex vivo (human prostate tumour explants) compared to vehicle treatment. MET+VPA induced synergistic apoptosis in LNCaP cells but not PC-3 cells. Increased apoptosis was also observed in human prostate tumour explants. MET+VPA induced an increase in cytochrome c release from the mitochondria to the cytoplasm in LNCaP, but not PC-3 cells, suggesting the involvement of intrinsic apoptosis in the response to the drug combination in LNCaP. The drugs did not have the same effect on normal PrEC cells. Additionally, MET+VPA, at the doses used here, did not cause weight change or tissue toxicity (liver and kidney) in nude mice.

Knock-down of p53 in LNCaP (p53+) reduced the MET+VPA-induced synergistic apoptotic response, and p53 plasmid transfection into PC3 (p53-) increased apoptosis, indicating a role for p53 in the response to MET+VPA. The p53-dependent response to MET+VPA may be explained by dual action of MET-induced p53 phosphorylation (Ser15) and VPA-induced acetylation (Lys373-382), which activates p53 and induces intrinsic apoptosis. Androgen receptor (AR) inhibition in LNCaP, an androgen-sensitive cell line, reduced the synergistic apoptosis response to MET+VPA, while having no effect in PC-3, an androgen-resistant cell line. The highest induced synergistic apoptotic effect observed was in the presence of both functional p53 and AR suggesting that MET+VPA is acting through these pathways and could be particularly effective in tumours with functional p53 and AR signalling which have critical roles in prostate carcinogenesis. However, in terms of anti-proliferative effects, MET+VPA consistently reduced proliferation in both LNCaP and PC-3, in cell culture and as nude mouse xenografts, indicating that MET+VPA can provide an anti-cancer effect regardless of p53 and AR status.

The demonstration of efficacy of MET+VPA in vitro, in vivo, and ex vivo supports translation of the drug combination into clinical trials as a novel systemic therapy for PCa. The results of this thesis have formed the basis for a Phase I clinical trial (ACTRN 12616001021460) investigating MET+VPA as a neoadjuvant therapy which is currently underway.

Keywords: metformin, valproic acid, prostate cancer, chemotherapy, mTOR, AMPK, androgen signalling pathway, p53 protein, PTEN, ETS gene fusion, ERG overexpression, histone deacetylase inhibitor.

Subject: Medicine thesis

Thesis type: Doctor of Philosophy
Completed: 2018
School: College of Medicine and Public Health
Supervisor: Pamela Sykes