Hilla Solomon

Mutant p53 Prolongs NF-κB Activation and Promotes Chronic Inflammation and Inflammation-Associated Colorectal Cancer

The tumor suppressor p53 is frequently mutated in human cancer. Common mutant p53 (mutp53) isoforms can actively promote cancer through gain-of-function (GOF) mechanisms. We report that mutp53 prolongs TNF-α-induced NF-κB activation in cultured cells and intestinal organoid cultures. Remarkably, when exposed to dextran sulfate sodium, mice harboring a germline p53 mutation develop severe chronic inflammation and persistent tissue damage, and are highly prone to inflammation-associated colon cancer. This mutp53 GOF is manifested by rapid onset of flat dysplastic lesions that progress to invasive carcinoma with mutp53 accumulation and augmented NF-κB activation, faithfully recapitulating features frequently observed in human colitis-associated colorectal cancer (CAC). These findings might explain the early appearance of p53 mutations in human CAC.

p53‐repressed miRNAs are involved with E2F in a feed‐forward loop promoting proliferation

Normal cell growth is governed by a complicated biological system, featuring multiple levels of control, often deregulated in cancers. The role of microRNAs (miRNAs) in the control of gene expression is now increasingly appreciated, yet their involvement in controlling cell proliferation is still not well understood. Here we investigated the mammalian cell proliferation control network consisting of transcriptional regulators, E2F and p53, their targets and a family of 15 miRNAs. Indicative of their significance, expression of these miRNAs is downregulated in senescent cells and in breast cancers harboring wild‐type p53. These miRNAs are repressed by p53 in an E2F1‐mediated manner. Furthermore, we show that these miRNAs silence antiproliferative genes, which themselves are E2F1 targets. Thus, miRNAs and transcriptional regulators appear to cooperate in the framework of a multi‐gene transcriptional and post‐transcriptional feed‐forward loop. Finally, we show that, similarly to p53 inactivation, overexpression of representative miRNAs promotes proliferation and delays senescence, manifesting the detrimental phenotypic consequence of perturbations in this circuit. Taken together, these findings position miRNAs as novel key players in the mammalian cellular proliferation network.

Mutant p53 R175H upregulates Twist1 expression and promotes epithelial-mesenchymal transition in immortalized prostate cells

A mutation within one allele of the p53 tumor suppressor gene can inactivate the remaining wild-type allele in a dominant-negative manner and in some cases can exert an additional oncogenic activity, known as mutant p53 ‘gain of function’ (GOF). To study the role of p53 mutations in prostate cancer and to discriminate between the dominant-negative effect and the GOF activity of mutant p53, we measured, using microarrays, the expression profiles of three immortalized prostate epithelial cultures expressing wild-type, inactivated p53 or mutated p53. Analysis of these gene expression profiles showed that both inactivated p53 and p53R175H mutant expression resulted in the upregulation of cell cycle progression genes. A second group, which was upregulated exclusively by mutant p53R175H, was predominantly enriched in developmental genes. This group of genes included the Twist1, a regulator of metastasis and epithelial–mesenchymal transition (EMT). Twist1 levels were also elevated in metastatic prostate cancer-derived cell line DU145, in immortalized lung fibroblasts and in a subset of lung cancer samples, all in a mutant p53-dependent manner. p53R175H mutant bearing immortalized epithelial cells showed typical features of EMT, such as higher expression of mesenchymal markers, lower expression of epithelial markers and enhanced invasive properties in vitro. The mechanism by which p53R175H mutant induces Twist1 expression involves alleviation of the epigenetic repression. Our data suggest that Twist1 expression might be upregulated following p53 mutation in cancer cells.

Prostate stromal cells produce CXCL-1, CXCL-2, CXCL-3 and IL-8 in response to epithelia-secreted IL-1

It is well accepted that tumor microenvironment is essential for tumor cells survival, cancer progression and metastasis. However, the mechanisms by which tumor cells interact with their surrounding at early stages of cancer development are largely unidentified. The aim of this study was to identify specific molecules involved in stromal-epithelial interactions that might contribute to early stages of prostate tumor formation. Here, we show that conditioned medium (CM) from immortalized non-transformed prostate epithelial cells stimulated immortalized prostate stromal cells to express cancer-related molecules. CM obtained from epithelial cells triggered stromal cells to express and secrete CXCL-1, CXCL-2, CXCL-3 and interleukin (IL)-8 chemokines. This effect was predominantly mediated by the cytokines of the IL-1 family secreted by the epithelial cells. Thus, prostate epithelial cells induced the secretion of proinflammatory and cancer-promoting chemokines by prostate stromal cells. Such interactions might contribute to prostatic inflammation and progression at early stages of prostate cancer formation.

Modulated expression of WFDC1 during carcinogenesis and cellular senescence

Fibroblasts located adjacent to the tumor [cancer-associated fibroblasts (CAFs)] that constitute a large proportion of the cancer-associated stroma facilitate the transformation process. In this study, we compared the biological behavior of CAFs that were isolated from a prostate tumor to their normal-associated fibroblast (NAF) counterparts. CAFs formed more colonies when seeded at low cell density, exhibited a higher proliferation rate and were less prone to contact inhibition. In contrast to the general notion that high levels of α-smooth muscle actin serve as a marker for CAFs, we found that prostate CAFs express it at a lower level compared with prostate NAFs. Microarray analysis revealed a set of 161 genes that were altered in CAFs compared with NAFs. We focused on whey acidic protein four-disulfide core domain 1 (WFDC1), a known secreted protease inhibitor, and found it to be downregulated in the CAFs. WFDC1 expression was also dramatically downregulated in highly prolific mesenchymal cells and in various cancers including fibrosarcomas and in tumors of the lung, bladder and brain. Overexpression of WFDC1 inhibited the growth rate of the fibrosarcoma HT1080 cell line. Furthermore, WFDC1 level was upregulated in senescent fibroblasts. Taken together, our data suggest an important role for WFDC1 in inhibiting proliferation of both tumors and senescent cells. Finally, we suggest that the downregulation of WFDC1 might serve as a biomarker for cellular transformation.

p53 Regulates the Ras Circuit to Inhibit the Expression of a Cancer-Related Gene Signature by Various Molecular Pathways

In this study, we focus on the analysis of a previously identified cancer-related gene signature (CGS) that underlies the cross talk between the p53 tumor suppressor and Ras oncogene. CGS consists of a large number of known Ras downstream target genes that were synergistically upregulated by wild-type p53 loss and oncogenic H-Ras(G12V) expression. Here we show that CGS expression strongly correlates with malignancy. In an attempt to elucidate the molecular mechanisms underling the cooperation between p53 loss and oncogenic H-Ras(G12V), we identified distinguished pathways that may account for the regulation of the expression of the CGS. By knocking-down p53 or by expressing mutant p53, we revealed that p53 exerts its negative effect by at least two mechanisms mediated by its targets B-cell translocation gene 2 (BTG2) and activating transcription factor 3 (ATF3). Whereas BTG2 binds H-Ras(G12V) and represses its activity by reducing its GTP loading state, which in turn causes a reduction in CGS expression, ATF3 binds directly to the CGS promoters following p53 stabilization and represses their expression. This study further elucidates the molecular loop between p53 and Ras in the transformation process.

Mutant p53 gain of function is interwoven into the hallmarks of cancer.

Cancer is viewed as being governed by several aberrant biological events defined by Weinberg and Hanahan as ‘hallmarks’. In most human cancers the tumour suppressor p53 is mutated, leading to its malfunction and to the acquirement of oncogenic activities, termed ‘gain of function’. This commentary links mutant p53 activities to the hallmarks of cancer, describing its involvement in resistance to apoptosis, genomic instability, aberrant cell cycle, invasion and metastasis, tumour microenvironment, and inflammation. Recent work published in The Journal of Pathology by Acin and colleagues, summarized here, reveals an interesting mechanism by which mutant p53 accelerates mitosis entry. Collectively, the growing body of evidence relating mutant p53 and the hallmarks of cancer reinforces the notion that targeting mutant p53 pathways might be beneficial for anti‐cancer therapy. Copyright

Mutant p53R273H attenuates the expression of phase 2 detoxifying enzymes and promotes the survival of cells with high levels of reactive oxygen species

Summary Uncontrolled accumulation of reactive oxygen species (ROS) causes oxidative stress and induces harmful effects. Both high ROS levels and p53 mutations are frequent in human cancer. Mutant p53 forms are known to actively promote malignant growth. However, no mechanistic details are known about the contribution of mutant p53 to excessive ROS accumulation in cancer cells. Herein, we examine the effect of p53R273H, a commonly occurring mutated p53 form, on the expression of phase 2 ROS-detoxifying enzymes and on the ability of cells to readopt a reducing environment after exposure to oxidative stress. Our data suggest that p53R273H mutant interferes with the normal response of human cells to oxidative stress. We show here that, upon oxidative stress, mutant p53R273H attenuates the activation and function of NF-E2-related factor 2 (NRF2), a transcription factor that induces the antioxidant response. This effect of mutant p53 is manifested by decreased expression of phase 2 detoxifying enzymes NQO1 and HO-1 and high ROS levels. These findings were observed in several human cancer cell lines, highlighting the general nature of this phenomenon. The failure of p53R273H mutant-expressing cells to restore a reducing oxidative environment was accompanied by increased survival, a known consequence of mutant p53 expression. These activities are attributable to mutant p53R273H gain of function and might underlie its well-documented oncogenic nature in human cancer.

Various p53 mutant proteins differently regulate the Ras circuit to induce a cancer-related gene signature

Summary Concomitant expression of mutant p53 and oncogenic Ras, leading to cellular transformation, is well documented. However, the mechanisms by which the various mutant p53 categories cooperate with Ras remain largely obscure. From this study we suggest that different mutant p53 categories cooperate with H-Ras in different ways to induce a unique expression pattern of a cancer-related gene signature (CGS). The DNA-contact p53 mutants (p53R248Q and p53R273H) exhibited the highest level of CGS expression by cooperating with NF&kgr;B. Furthermore, the Zn+2 region conformational p53 mutants (p53R175H and p53H179R) induced the CGS by elevating H-Ras activity. This elevation in H-Ras activity stemmed from a perturbed function of the p53 transcription target gene, BTG2. By contrast, the L3 loop region conformational mutant (p53G245S) did not affect CGS expression. Our findings were further corroborated in human tumor-derived cell lines expressing Ras and the aforementioned mutated p53 proteins. These data might assist in future tailor-made therapy targeting the mutant p53–Ras axis in cancer.

The paradigm of mutant p53-expressing cancer stem cells and drug resistance.

It is well accepted that expression of mutant p53 involves the gain of oncogenic-specific activities accentuating the malignant phenotype. Depending on the specific cancer type, mutant p53 can contribute to either the early or the late events of the multiphase process underlying the transformation of a normal cell into a cancerous one. This multifactorial system is evident in ~50% of human cancers. Mutant p53 was shown to interfere with a variety of cellular functions that lead to augmented cell survival, cellular plasticity, aberration of DNA repair machinery and other effects. All these effects culminate in the acquisition of drug resistance often seen in cancer cells. Interestingly, drug resistance has also been suggested to be associated with cancer stem cells (CSCs), which reside within growing tumors. The notion that p53 plays a regulatory role in the life of stem cells, coupled with the observations that p53 mutations may contribute to the evolvement of CSCs makes it challenging to speculate that drug resistance and cancer recurrence are mediated by CSCs expressing mutant p53.