Heat shock proteins in ovarian cancer: a potential target for therapy.

Ovarian cancer is the leading cause of death from gynecologic cancer in Western countries. Most of the patients are chemosensitive, and they will present a complete response after initial treatment, but most of them will relapse within 2 years. The resistance to standard chemotherapy represents a significant clinical challenge. Therefore, a better understanding of the molecular mechanisms involved in the drug resistance should allow to improve the treatment in this disease. Interestingly, recent data showed essential roles of heat shock proteins (HSPs) in various processes of carcinogenesis and their association with resistance to anticancer drugs. Here, we report the main investigations on HSPs in ovarian cancer with specific emphasis on clinical application.

PTPL1/PTPN13 regulates breast cancer cell aggressiveness through direct inactivation of Src kinase.

The protein tyrosine phosphatase PTPL1/PTPN13, the activity of which is decreased through allelic loss, promoter methylation, or somatic mutations in some tumors, has been proposed as a tumor suppressor gene. Moreover, our recent clinical study identified PTPL1 expression level as an independent prognostic indicator of a favorable outcome for patients with breast cancer. However, how PTPL1 can affect tumor aggressiveness has not been characterized. Here, we first show that PTPL1 expression, assessed by immunohistochemistry, is decreased in breast cancer and metastasis specimens compared with nonmalignant tissues. Second, to evaluate whether PTPL1 plays a critical role in breast cancer progression, RNA interference experiments were performed in poorly tumorigenic MCF-7 breast cancer cells. PTPL1 inhibition drastically increased tumor growth in athymic mice and also enhanced several parameters associated with tumor progression, including cell proliferation on extracellular matrix components and cell invasion. Furthermore, the inhibition of Src kinase expression drastically blocked the effects of PTPL1 silencing on cell growth. In PTPL1 knockdown cells, the phosphorylation of Src on tyrosine 419 is increased, leading to the activation of its downstream substrates Fak and p130cas. Finally, substrate-trapping experiments revealed that Src tyrosine 419 is a direct target of the phosphatase. Thus, by identification of PTPL1 as the first phosphatase able to inhibit Src through direct dephosphorylation in intact cells, we presently describe a new mechanism by which PTPL1 inhibits breast tumor aggressiveness.

Downregulation of protein tyrosine phosphatase PTP-BL represses adipogenesis.

The insulin/insulin-like growth factor 1 (IGF-1) signaling pathway is a major regulator of adipose tissue growth and differentiation. We recently demonstrated that human protein tyrosine phosphatase (PTP) L1, a large cytoplasmic phosphatase also known as PTP-BAS/PTPN13/PTP-1E, is a negative regulator of IGF-1R/IRS-1/Akt pathway in breast cancer cells. This triggered us to investigate the potential role of PTPL1 in adipogenesis. To evaluate the implication of PTP-BL, the mouse orthologue of PTPL1, in adipose tissue biology, we analyzed PTP-BL mRNA expression in adipose tissue in vivo and during proliferation and differentiation of 3T3-L1 pre-adipocytes. To elucidate the role of PTP-BL and of its catalytic activity during adipogenesis we use siRNA techniques in 3T3-L1 pre-adipocytes, and mouse embryonic fibroblasts that lack wildtype PTP-BL and instead express a variant without the PTP domain (Delta P/Delta P MEFs). Here we show that PTP-BL is strongly expressed in white adipose tissue and that PTP-BL transcript and protein levels increase during proliferation and differentiation of 3T3-L1 pre-adipocytes. Strikingly, knockdown of PTP-BL expression in 3T3-L1 adipocytes caused a dramatic decrease in adipogenic gene expression levels (PPAR gamma, aP2) and lipid accumulation but did not interfere with the insulin/Akt pathway. Delta P/Delta P MEFs differentiate into the adipogenic lineage as efficiently as wildtype MEFs. However, when expression of either PTP-BL or PTP-BL Delta P was inhibited a dramatic reduction in the number of MEF-derived adipocytes was observed. These findings demonstrate a key role for PTP-BL in 3T3-L1 and MEF-derived adipocyte differentiation that is independent of its enzymatic activity.

The putative tumor suppressor gene PTPN13/PTPL1 induces apoptosis through insulin receptor substrate-1 dephosphorylation.

The protein tyrosine phosphatase (PTP) PTPL1/PTPN13 is a candidate tumor suppressor gene. Indeed, PTPL1 activity has been reported recently to be decreased through somatic mutations, allelic loss, or promoter methylation in some tumors. We showed previously that its expression was necessary for inhibition of Akt activation and induction of apoptosis by antiestrogens in breast cancer cells. Implications of the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway in cancer progression are now well established, and our study was therefore designed to define whether PTPL1 is sufficient to inhibit this pathway and, if so, to identify a direct substrate of this PTP, which may trigger a proapoptotic effect. We first show by complementary approaches that PTPL1 specifically dephosphorylates insulin receptor substrate-1 (IRS-1) in vitro and in cellulo. Next, our experiments using a dominant-negative mutant and RNA interference confirm the crucial role of PTPL1 in IRS-1 dephosphorylation. Finally, we report that PTPL1 expression is sufficient to block the IRS-1/PI3K/Akt signaling pathway, to inhibit the insulin-like growth factor-I effect on cell survival, and to induce apoptosis. Altogether, these data provide the first evidence for a direct positive role of the putative tumor suppressor gene PTPL1/PTPN13 on apoptosis and identify its target in the IRS-1/PI3K/Akt signaling pathway.

The diagnostic accuracy of reverse transcription-PCR quantification of cytokeratin mRNA in the detection of sentinel lymph node invasion in oral and oropharyngeal squamous cell carcinoma: a comparison with immunohistochemistry.

PURPOSE: The main goal of sentinel lymph node (SLN) detection in head and neck squamous cell carcinomas is to limit neck dissections to pN+ cases only. However, intraoperative + diagnosis cannot be routinely done using the current gold standard, serial step sectioning with immunohistochemistry. Real-time quantitative reverse transcription-PCR (RT-PCR) is potentially compatible with intraoperative use, proving highly sensitive in detecting molecular markers. This study postoperatively assessed the accuracy of quantitative RT-PCR in staging patients from their SLN. EXPERIMENTAL DESIGN: A combined analysis on the same SLN by serial step sectioning with immunohistochemistry and quantitative RT-PCR targeting cytokeratins 5, 14, and 17 was done in 18 consecutive patients with oral or oropharyngeal squamous cell carcinoma and 10 control subjects. RESULTS: From 71 lymph nodes examined, mRNA levels (KRT) were linked to metastasis size for the three cytokeratins studied (Pearson correlation coefficient, r = 0.89, 0.73, and 0.77 for KRT 5, 14, and 17 respectively; P < 0.05). Histopathology-positive SLNs (macro- and micrometastases) showed higher mRNA values than negative SLNs for KRT 17 (P < 10(-4)) and KRT 14 (P < 10(-2)). KRT 5 showed nonsignificant results. KRT 17 seemed to be the most accurate marker for the diagnosis of micrometastases of a size >450 mum. Smaller micrometastases and isolated tumor cells did not provide results above the background level. Receiver operating characteristic curve analysis for KRT 17 identified a cutoff value where patient staging reached 100% specificity and sensitivity for macro- and micrometastases. CONCLUSION: Quantitative RT-PCR for SLN staging in cN(0) patients with oral and oropharyngeal squamous cell carcinoma seems to be a promising approach.