Arkaitz Carracedo

Inhibition of mTORC1 leads to MAPK pathway activation through a PI3K-dependent feedback loop in human cancer

Numerous studies have established a causal link between aberrant mammalian target of rapamycin (mTOR) activation and tumorigenesis, indicating that mTOR inhibition may have therapeutic potential. In this study, we show that rapamycin and its analogs activate the MAPK pathway in human cancer, in what represents a novel mTORC1-MAPK feedback loop. We found that tumor samples from patients with biopsy-accessible solid tumors of advanced disease treated with RAD001, a rapamycin derivative, showed an administration schedule-dependent increase in activation of the MAPK pathway. RAD001 treatment also led to MAPK activation in a mouse model of prostate cancer. We further show that rapamycin-induced MAPK activation occurs in both normal cells and cancer cells lines and that this feedback loop depends on an S6K-PI3K-Ras pathway. Significantly, pharmacological inhibition of the MAPK pathway enhanced the antitumoral effect of mTORC1 inhibition by rapamycin in cancer cells in vitro and in a xenograft mouse model. Taken together, our findings identify MAPK activation as a consequence of mTORC1 inhibition and underscore the potential of a combined therapeutic approach with mTORC1 and MAPK inhibitors, currently employed as single agents in the clinic, for the treatment of human cancers.

Tenets of PTEN Tumor Suppression

Since its discovery as the elusive tumor suppressor gene at the frequently mutated 10q23 locus, PTEN has been identified as lost or mutated in several sporadic and heritable tumor types. A decade of work has established that PTEN is a nonredundant phosphatase that is essential for regulating the highly oncogenic prosurvival PI3K/AKT signaling pathway. This review discusses emerging modes of PTEN function and regulation, and speculates about how manipulation of PTEN function could be used for cancer therapy.

The PTEN–PI3K pathway: of feedbacks and cross-talks

The tumor suppressor PTEN was originally identified as a negative regulator of the phosphoinositide 3-kinase (PI3K) signaling, a main regulator of cell growth, metabolism and survival. Yet this function of PTEN is extremely relevant for its tumor-suppressive ability, albeit the recent characterization of many PI3K-independent tumor-suppressive activities. PI3K-mediated PIP3 production leads to the activation of the canonical AKT-mTORC1 pathway. The implications of this signaling cascade in health and disease have been underscored by the high number of regulators within the pathway whose alterations give rise to different malignancies, including familiar syndromes, metabolic dysfunctions and cancer. Moreover, PI3K is tightly buffered at multiple levels by downstream components, which have turned this signaling pathway literally upside down. PI3K and its downstream components in turn cross-talk with a number of other pathways, thereby leading to a complex network of signals that may have dramatic consequences when perturbed. Here, we review the current status of the PTEN–PI3K signaling pathway with special emphasis on the most recent data on targets and regulation of the PTEN–PI3K axis. This provides novel provocative therapeutic implications based on the targeted modulation of PI3K-cross-talking signals.

Aberrant ERG expression cooperates with loss of PTEN to promote cancer progression in the prostate

Chromosomal translocations involving the ERG locus are frequent events in human prostate cancer pathogenesis; however, the biological role of aberrant ERG expression is controversial. Here we show that aberrant expression of ERG is a progression event in prostate tumorigenesis. We find that prostate cancer specimens containing the TMPRSS2-ERG rearrangement are significantly enriched for loss of the tumor suppressor PTEN. In concordance with these findings, transgenic overexpression of ERG in mouse prostate tissue promotes marked acceleration and progression of high-grade prostatic intraepithelial neoplasia (HGPIN) to prostatic adenocarcinoma in a Pten heterozygous background. In vitro overexpression of ERG promotes cell migration, a property necessary for tumorigenesis, without affecting proliferation. ADAMTS1 and CXCR4, two candidate genes strongly associated with cell migration, were upregulated in the presence of ERG overexpression. Thus, ERG has a distinct role in prostate cancer progression and cooperates with PTEN haploinsufficiency to promote progression of HGPIN to invasive adenocarcinoma.

Cannabinoid action induces autophagy-mediated cell death through stimulation of ER stress in human glioma cells

Autophagy can promote cell survival or cell death, but the molecular basis underlying its dual role in cancer remains obscure. Here we demonstrate that delta(9)-tetrahydrocannabinol (THC), the main active component of marijuana, induces human glioma cell death through stimulation of autophagy. Our data indicate that THC induced ceramide accumulation and eukaryotic translation initiation factor 2alpha (eIF2alpha) phosphorylation and thereby activated an ER stress response that promoted autophagy via tribbles homolog 3-dependent (TRB3-dependent) inhibition of the Akt/mammalian target of rapamycin complex 1 (mTORC1) axis. We also showed that autophagy is upstream of apoptosis in cannabinoid-induced human and mouse cancer cell death and that activation of this pathway was necessary for the antitumor action of cannabinoids in vivo. These findings describe a mechanism by which THC can promote the autophagic death of human and mouse cancer cells and provide evidence that cannabinoid administration may be an effective therapeutic strategy for targeting human cancers.

SIRT3 opposes reprogramming of cancer cell metabolism through HIF1α destabilization.

Tumor cells exhibit aberrant metabolism characterized by high glycolysis even in the presence of oxygen. This metabolic reprogramming, known as the Warburg effect, provides tumor cells with the substrates required for biomass generation. Here, we show that the mitochondrial NAD-dependent deacetylase SIRT3 is a crucial regulator of the Warburg effect. Mechanistically, SIRT3 mediates metabolic reprogramming by destabilizing hypoxia-inducible factor-1α (HIF1α), a transcription factor that controls glycolytic gene expression. SIRT3 loss increases reactive oxygen species production, leading to HIF1α stabilization. SIRT3 expression is reduced in human breast cancers, and its loss correlates with the upregulation of HIF1α target genes. Finally, we find that SIRT3 overexpression represses glycolysis and proliferation in breast cancer cells, providing a metabolic mechanism for tumor suppression.

Subtle variations in Pten dose determine cancer susceptibility

Cancer susceptibility has been attributed to at least one heterozygous genetic alteration in a tumor suppressor gene (TSG). It has been hypothesized that subtle variations in TSG expression can promote cancer development. However, this hypothesis has not yet been definitively supported in vivo. Pten is a TSG frequently lost in human cancer and mutated in inherited cancer-predisposition syndromes. Here we analyze Pten hypermorphic mice (Ptenhy/+), expressing 80% normal levels of Pten. Ptenhy/+ mice develop a spectrum of tumors, with breast tumors occurring at the highest penetrance. All breast tumors analyzed here retained two intact copies of Pten and maintained Pten levels above heterozygosity. Notably, subtle downregulation of Pten altered the steady-state biology of the mammary tissues and the expression profiles of genes involved in cancer cell proliferation. We present an alterative working model for cancer development in which subtle reductions in the dose of TSGs predispose to tumorigenesis in a tissue-specific manner.

High frequency of PTEN, PI3K, and AKT abnormalities in T-cell acute lymphoblastic leukemia.

To more comprehensively assess the pathogenic contribution of the PTEN-PI3K-AKT pathway to T-cell acute lymphoblastic leukemia (T-ALL), we examined diagnostic DNA samples from children with T-ALL using array comparative genomic hybridization and sequence analysis. Alterations of PTEN, PI3K, or AKT were identified in 47.7% of 44 cases. There was a striking clustering of PTEN mutations in exon 7 in 12 cases, all of which were predicted to truncate the C2 domain without disrupting the phosphatase domain of PTEN. Induction chemotherapy failed to induce remission in 3 of the 4 patients whose lymphoblasts harbored PTEN deletions at the time of diagnosis, compared with none of the 12 patients with mutations of PTEN exon 7 (P = .007), suggesting that PTEN deletion has more adverse therapeutic consequences than mutational disruptions that preserve the phosphatase domain. These findings add significant support to the rationale for the development of therapies targeting the PTEN-PI3K-AKT pathway in T-ALL.

The deubiquitinylation and localization of PTEN are regulated by a HAUSP–PML network

Nuclear exclusion of the PTEN (phosphatase and tensin homologue deleted in chromosome 10) tumour suppressor has been associated with cancer progression. However, the mechanisms leading to this aberrant PTEN localization in human cancers are currently unknown. We have previously reported that ubiquitinylation of PTEN at specific lysine residues regulates its nuclear–cytoplasmic partitioning. Here we show that functional promyelocytic leukaemia protein (PML) nuclear bodies co-ordinate PTEN localization by opposing the action of a previously unknown PTEN-deubiquitinylating enzyme, herpesvirus-associated ubiquitin-specific protease (HAUSP, also known as USP7), and that the integrity of this molecular framework is required for PTEN to be able to enter the nucleus. We find that PTEN is aberrantly localized in acute promyelocytic leukaemia, in which PML function is disrupted by the PML–RARα fusion oncoprotein. Remarkably, treatment with drugs that trigger PML–RARα degradation, such as all-trans retinoic acid or arsenic trioxide, restore nuclear PTEN. We demonstrate that PML opposes the activity of HAUSP towards PTEN through a mechanism involving the adaptor protein DAXX (death domain-associated protein). In support of this paradigm, we show that HAUSP is overexpressed in human prostate cancer and is associated with PTEN nuclear exclusion. Thus, our results delineate a previously unknown PML–DAXX–HAUSP molecular network controlling PTEN deubiquitinylation and trafficking, which is perturbed by oncogenic cues in human cancer, in turn defining a new deubiquitinylation-dependent model for PTEN subcellular compartmentalization.

A PML-PPAR-δ pathway for fatty acid oxidation regulates hematopoietic stem cell maintenance

Stem-cell function is an exquisitely regulated process. Thus far, the contribution of metabolic cues to stem-cell function has not been well understood. Here we identify a previously unknown promyelocytic leukemia (PML)–peroxisome proliferator-activated receptor δ (PPAR-δ)–fatty-acid oxidation (FAO) pathway for the maintenance of hematopoietic stem cells (HSCs). We have found that loss of PPAR-δ or inhibition of mitochondrial FAO induces loss of HSC maintenance, whereas treatment with PPAR-δ agonists improved HSC maintenance. PML exerts its essential role in HSC maintenance through regulation of PPAR signaling and FAO. Mechanistically, the PML–PPAR-δ–FAO pathway controls the asymmetric division of HSCs. Deletion of Ppard or Pml as well as inhibition of FAO results in the symmetric commitment of HSC daughter cells, whereas PPAR-δ activation increased asymmetric cell division. Thus, our findings identify a metabolic switch for the control of HSC cell fate with potential therapeutic implications.