The NAMPT Inhibitor FK866 Increases Metformin Sensitivity in Pancreatic Cancer Cells.

Pancreatic cancer (pancreatic ductal adenocarcinoma: PDAC) is one of the most aggressive neoplastic diseases. Metformin use has been associated with reduced pancreatic cancer incidence and better survival in diabetics. Metformin has been shown to inhibit PDAC cells growth and survival, both in vitro and in vivo. However, clinical trials using metformin have failed to reduce pancreatic cancer progression in patients, raising important questions about molecular mechanisms that protect tumor cells from the antineoplastic activities of metformin. We confirmed that metformin acts through inhibition of mitochondrial complex I, decreasing the NAD+/NADH ratio, and that NAD+/NADH homeostasis determines metformin sensitivity in several cancer cell lines. Metabolites that can restore the NAD+/NADH ratio caused PDAC cells to be resistant to metformin. In addition, metformin treatment of PDAC cell lines induced a compensatory NAMPT expression, increasing the pool of cellular NAD+. The NAMPT inhibitor FK866 sensitized PDAC cells to the antiproliferative effects of metformin in vitro and decreased the cellular NAD+ pool. Intriguingly, FK866 combined with metformin increased survival in mice bearing KP4 cell line xenografts, but not in mice with PANC-1 cell line xenografts. Transcriptome analysis revealed that the drug combination reactivated genes in the p53 pathway and oxidative stress, providing new insights about the mechanisms leading to cancer cell death.

Hypoxia-mediated stabilization of HIF1A in prostatic intraepithelial neoplasia promotes cell plasticity and malignant progression.

Prostate cancer (PCa) is a leading cause of cancer-related deaths. The slow evolution of precancerous lesions to malignant tumors provides a broad time frame for preventing PCa. To characterize prostatic intraepithelial neoplasia (PIN) progression, we conducted longitudinal studies on Pten(i)pe-/- mice that recapitulate prostate carcinogenesis in humans. We found that early PINs are hypoxic and that hypoxia-inducible factor 1 alpha (HIF1A) signaling is activated in luminal cells, thus enhancing malignant progression. Luminal HIF1A dampens immune surveillance and drives luminal plasticity, leading to the emergence of cells that overexpress Transglutaminase 2 (TGM2) and have impaired androgen signaling. Elevated TGM2 levels in patients with PCa are associated with shortened progression-free survival after prostatectomy. Last, we show that pharmacologically inhibiting HIF1A impairs cell proliferation and induces apoptosis in PINs. Therefore, our study demonstrates that HIF1A is a target for PCa prevention and that TGM2 is a promising prognostic biomarker of early relapse after prostatectomy.

Phenylethynylbenzyl-modified biguanides inhibit pancreatic cancer tumor growth.

We present the design and synthesis of a small library of substituted biguanidium salts and their capacity to inhibit the growth of pancreatic cancer cells. We first present their in vitro and membrane activity, before we address their mechanism of action in living cells and in vivo activity. We show that phenylethynyl biguanidium salts possess higher ability to cross hydrophobic barriers, improve mitochondrial accumulation and anticancer activity. Mechanistically, the most active compound, 1b, like metformin, activated AMPK, decreased the NAD+/NADH ratio and mitochondrial respiration, but at 800-fold lower concentration. In vivo studies show that compound 1b significantly inhibits the growth of pancreatic cancer xenografts in mice, while biguanides currently in clinical trials had little activity.

Senescence controls prostatic neoplasia driven by Pten loss.

We report that Pten (phosphatase and tensin homologue) ablation in prostatic epithelial cells of adult mice promotes cell proliferation to generate prostatic intraepithelial neoplasia. Moreover, our results demonstrate that proliferating Pten-deficient cells undergo replication stress and exhibit a DNA damage response, leading to cell senescence, as seen in oncogene-induced senescence.

Circumventing senescence is associated with stem cell properties and metformin sensitivity.

Most cancers arise in old individuals, which also accumulate senescent cells. Cellular senescence can be experimentally induced by expression of oncogenes or telomere shortening during serial passage in culture. In vivo, precursor lesions of several cancer types accumulate senescent cells, which are thought to represent a barrier to malignant progression and a response to the aberrant activation of growth signaling pathways by oncogenes (oncogene toxicity). Here, we sought to define gene expression changes associated with cells that bypass senescence induced by oncogenic RAS. In the context of pancreatic ductal adenocarcinoma (PDAC), oncogenic KRAS induces benign pancreatic intraepithelial neoplasias (PanINs), which exhibit features of oncogene-induced senescence. We found that the bypass of senescence in PanINs leads to malignant PDAC cells characterized by gene signatures of epithelial-mesenchymal transition, stem cells, and mitochondria. Stem cell properties were similarly acquired in PanIN cells treated with LPS, and in primary fibroblasts and mammary epithelial cells that bypassed Ras-induced senescence after reduction of ERK signaling. Intriguingly, maintenance of cells that circumvented senescence and acquired stem cell properties was blocked by metformin, an inhibitor of complex I of the electron transport chain or depletion of STAT3, a protein required for mitochondrial functions and stemness. Thus, our studies link bypass of senescence in premalignant lesions to loss of differentiation, acquisition of stemness features, and increased reliance on mitochondrial functions.

Translational and HIF-1α-Dependent Metabolic Reprogramming Underpin Metabolic Plasticity and Responses to Kinase Inhibitors and Biguanides.

There is increasing interest in therapeutically exploiting metabolic differences between normal and cancer cells. We show that kinase inhibitors (KIs) and biguanides synergistically and selectively target a variety of cancer cells. Synthesis of non-essential amino acids (NEAAs) aspartate, asparagine, and serine, as well as glutamine metabolism, are major determinants of the efficacy of KI/biguanide combinations. The mTORC1/4E-BP axis regulates aspartate, asparagine, and serine synthesis by modulating mRNA translation, while ablation of 4E-BP1/2 substantially decreases sensitivity of breast cancer and melanoma cells to KI/biguanide combinations. Efficacy of the KI/biguanide combinations is also determined by HIF-1α-dependent perturbations in glutamine metabolism, which were observed in VHL-deficient renal cancer cells. This suggests that cancer cells display metabolic plasticity by engaging non-redundant adaptive mechanisms, which allows them to survive therapeutic insults that target cancer metabolism.

PTEN deletion in luminal cells of mature prostate induces replication stress and senescence in vivo.

Genetic ablation of the tumor suppressor PTEN in prostatic epithelial cells (PECs) induces cell senescence. However, unlike oncogene-induced senescence, no hyperproliferation phase and no signs of DNA damage response (DDR) were observed in PTEN-deficient PECs; PTEN loss-induced senescence (PICS) was reported to be a novel type of cellular senescence. Our study reveals that PTEN ablation in prostatic luminal epithelial cells of adult mice stimulates PEC proliferation, followed by a progressive growth arrest with characteristics of cell senescence. Importantly, we also show that proliferating PTEN-deficient PECs undergo replication stress and mount a DDR leading to p53 stabilization, which is however delayed by Mdm2-mediated p53 down-regulation. Thus, even though PTEN-deficiency induces cellular senescence that restrains tumor progression, as it involves replication stress, strategies promoting PTEN loss-induced senescence are at risk for cancer prevention and therapy.

Genetically engineered mouse models of prostate cancer.

Despite major improvement in treatment of early stage localised prostate cancer, the distinction between indolent tumors and those that will become aggressive, as well as the lack of efficient therapies of advanced prostate cancer, remain major health problems. Genetically engineered mice (GEM) have been extensively used to investigate the molecular and cellular mechanisms underlying prostate tumor initiation and progression, and to evaluate new therapies. Moreover, the recent development of conditional somatic mutagenesis in the mouse prostate offers the possibility to generate new models that more faithfully reproduce the human disease, and thus should contribute to improve diagnosis and treatments. The strengths and weaknesses of various models will be discussed, as well as future opportunities.

Isomer-specific retinoic acid biosynthesis in HeLa cells expressing recombinant class I aldehyde dehydrogenases.

Retinal dehydrogenase type 1 (RALDH1) catalyzes the oxidation of all-trans and 9-cis retinal to the respective retinoic acids (RAs), whereas another member of the aldehyde dehydrogenase family, the phenobarbital-induced aldehyde dehydrogenase (PB-ALDH), is very poorly active. We have previously generated chimeras between these two enzymes that displayed selectivity for retinal isomers in crude bacterial extracts. To examine whether the selectivity of the recombinant enzymes is retained in intact cells, we first assessed whether retinoid-isomerizing activity is present in cultured eukaryotic cells. Our results demonstrate that the only RA isomers detected in RALDH1-expressing or non-expressing cells corresponded to the same steric conformation as the supplied retinoids, indicating a lack of measurable 9-cis/all-trans retinoid-isomerizing activity. Finally, HeLa cells transfected with RALDH1 derivatives that were retinal isomer-selective in vitro produced only the corresponding RA isomers, establishing these enzymes as useful tools to assess the respective roles of the two RA isomers in vivo.

[Retinoid metabolism and cancer]. / Métabolisme des rétinoïdes et cancer.

Retinoids play important roles in cell differentiation and apoptosis, notably in epithelial tissues. Their utility in cancer therapy has been demonstrated in specific cancer types. Use of retinoic acid (RA) in the treatment of acute promyelocytic leukemia was the first successful example of retinoid-based differentiation therapy. RA has since been evaluated for treatment of other cancers, revealing variable effectiveness. The observation that expression of enzymes involved in RA biosynthesis is suppressed during tumorigenesis suggests that intra-tumor depletion in RA levels may contribute to tumor development and argues for the use of retinoids in cancer treatment. However, the induction of RA-inactivating enzymes is one of the mechanisms that may limit the efficacy of retinoid therapy and contribute to acquired resistance to RA treatment, suggesting that retinoic acid metabolism blocking agents may be effective agents in differentiation therapy.