Carcinogenesis of PIK3CA

PIK3CA is the most frequently mutated oncogene in human cancers. PIK3CA is phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha. It controls cell growth, proliferation, motility, survival, differentiation and intracellular trafficking. In most of human cancer alteration occurred frequently in the alpha isoform of phosphatidylinositol 3 kinase. PIK3CA mutations were most frequent in endometrial, ovarian, colorectal, breast, cervical, squamous cell cancer of the head and neck, chondroma, thyroid carcinoma and in cancer family syndrome. Inhibition of PI3K signaling can diminish cell proliferation, and in some circumstances, promote cell death. Consequently, components of this pathway present attractive targets for cancer therapeutics. A number of PI3K pathway inhibitors have been developed and used. PI3K inhibitors (both pan-PI3K and isoform-specific PI3K inhibitors), dual PI3K-mTOR inhibitors that are catalytic site inhibitors of the p110 isoforms and mTOR (the kinase component of both mTORC1 and mTORC2), mTOR catalytic site inhibitors, and AKT inhibitors are the most advanced in the clinic. They are approved for the treatment of several carcinomas.     

Targeting of mTORC2 prevents cell migration and promotes apoptosis in breast cancer

Most of breast cancers are resistant to mammalian target of rapamycin complex 1 (mTORC1) inhibitors rapamycin and rapalogs. Recent studies indicate mTORC2 is emerging as a promising cancer therapeutic target. In this study, we compared the inhibitory effects of targeting mTORC1 with mTORC2 on a variety of breast cancer cell lines and xenograft. We demonstrated that inhibition of mTORC1/2 by mTOR kinase inhibitors PP242 and OSI-027 effectively suppress phosphorylation of Akt (S473) and breast cancer cell proliferation. Targeting of mTORC2 either by kinase inhibitors or rictor knockdown, but not inhibition of mTORC1 either by rapamycin or raptor knockdown promotes serum starvation- or cisplatin-induced apoptosis. Furthermore, targeting of mTORC2 but not mTORC1 efficiently prevent breast cancer cell migration. Most importantly, in vivo administration of PP242 but not rapamycin as single agent effectively prevents breast tumor growth and induces apoptosis in xenograft. Our data suggest that agents that inhibit mTORC2 may have advantages over selective mTORC1 inhibitors in the treatment of breast cancers. Given that mTOR kinase inhibitors are in clinical trials, this study provides a strong rationale for testing the use of mTOR kinase inhibitors or combination of mTOR kinase inhibitors and cisplatin in the clinic.     

Dual inhibition of the mTORC1 and mTORC2 signaling pathways is a promising therapeutic target for adult T‐cell leukemia

Adult T‐cell leukemia (ATL) has a poor prognosis as a result of severe immunosuppression and rapid tumor progression with resistance to conventional chemotherapy. Recent integrated‐genome analysis has revealed mutations in many genes involved in the T‐cell signaling pathway, suggesting that the aberration of this pathway is an important factor in ATL pathogenesis and ATL‐cell proliferation. We screened a siRNA library to examine signaling‐pathway functionality and found that the PI3K/Akt/mTOR pathway is critical to ATL‐cell proliferation. We therefore investigated the effect of mammalian target of rapamycin (mTOR) inhibitors, including the dual inhibitors PP242 and AZD8055 and the mTORC1 inhibitors rapamycin and everolimus, on human T‐cell leukemia virus type 1 (HTLV‐1)‐infected‐cell and ATL‐cell lines. Both dual inhibitors inhibited the proliferation of all tested cell lines by inducing G1‐phase cell‐cycle arrest and subsequent cell apoptosis, whereas the effects of the 2 mTORC1 inhibitors were limited, as they did not induce cell apoptosis. In the ATL‐cell lines and in the primary ATL samples, both dual inhibitors inhibited phosphorylation of AKT at serine‐473, a target of mTORC2, as well as that of S6K, whereas the mTORC1 inhibitors only inhibited mTORC1. Furthermore, AZD8055 more significantly inhibited the in vivo growth of the ATL‐cell xenografts than did everolimus. These results indicate that the PI3K/mTOR pathway is critical to ATL‐cell proliferation and might thus be a new therapeutic target in ATL.     

Targeting Ras-PI3K/mTOR pathway and the predictive biomarkers in endometrial cancer

The Ras-PI3K (phosphatidylinositol-3-kinase)/mTOR (mammalian Target of Rapamycin) pathway is frequently activated in various types of cancers. A number of inhibitors targeting the PI3K/mTOR pathway and MAPK pathway (another Ras effector pathway) are under development. PI3K/AKT activating mutations, including mutations in PTEN (50%), PIK3CA (30%), and K-Ras (20%), are frequently observed in endometrial cancer. A coexistence of these mutations is also commonly observed. We classified 13 endometrial cancer cell lines into three groups according to their mutational status in these genes: Group A (n=9); K-Ras wild-type and PTEN mutant, Group B (n=2); K-Ras mutant, and Group C (n=2) without any mutations in K-Ras, PTEN or PIK3CA. We determined the effects a dual PI3K/mTOR inhibitor (Inhibitor P) on these cell lines. MTT assay revealed that all the nine cell lines in Group A were sensitive to the inhibitor P (IC50<100 nM), whereas the other four cell lines in Group B or C were less sensitive to it(IC50>100 nM). Daily oral administration of inhibitor P showed anti-tumor effects in the mice bearing Group A tumors. Our data suggest that dual inhibition of the PI3K/mTOR is a promising molecular-targeted therapeutic for certain endometrial cancers, and that the mutational status of K-Ras and PI3K pathway-related genes, like PTEN and PIK3CA, could be useful for predicting sensitivities to such agents.     

An integrative genomic and proteomic analysis of PIK3CA, PTEN, and AKT mutations in breast cancer

Phosphatidylinositol 3-kinase (PI3K)/AKT pathway aberrations are common in cancer. By applying mass spectroscopy-based sequencing and reverse-phase protein arrays to 547 human breast cancers and 41 cell lines, we determined the subtype specificity and signaling effects of PIK3CA, AKT, and PTEN mutations and the effects of PIK3CA mutations on responsiveness to PI3K inhibition in vitro and on outcome after adjuvant tamoxifen. PIK3CA mutations were more common in hormone receptor-positive (34.5%) and HER2-positive (22.7%) than in basal-like tumors (8.3%). AKT1 (1.4%) and PTEN (2.3%) mutations were restricted to hormone receptor-positive cancers. Unlike AKT1 mutations that were absent from cell lines, PIK3CA (39%) and PTEN (20%) mutations were more common in cell lines than tumors, suggesting a selection for these but not AKT1 mutations during adaptation to culture. PIK3CA mutations did not have a significant effect on outcome after adjuvant tamoxifen therapy in 157 hormone receptor-positive breast cancer patients. PIK3CA mutations, in comparison with PTEN loss and AKT1 mutations, were associated with significantly less and inconsistent activation of AKT and of downstream PI3K/AKT signaling in tumors and cell lines. PTEN loss and PIK3CA mutation were frequently concordant, suggesting different contributions to pathophysiology. PTEN loss rendered cells significantly more sensitive to growth inhibition by the PI3K inhibitor LY294002 than did PIK3CA mutations. Thus, PI3K pathway aberrations likely play a distinct role in the pathogenesis of different breast cancer subtypes. The specific aberration present may have implications for the selection of PI3K-targeted therapies in hormone receptor-positive breast cancer.     

Autophagy limits the cytotoxic effects of the AKT inhibitor AZ7328 in human bladder cancer cells

Background: Mutations that activate the PI3K/AKT/mTOR pathway are relatively common in urothelial (bladder) cancers, but how these pathway mutations affect AKT dependency is not known. We characterized the relationship between AKT pathway mutational status and sensitivity to the effects of the selective AKT kinase inhibitor AZ7328 using a panel of 12 well-characterized human bladder cancer cell lines.Methods: Sequenome DNA sequencing was performed to identify mutations in a panel of 12 urothelial cancer cell lines. Drug-induced proliferative inhibition and apoptosis were quantified using MTT assays and propidium iodide staining with FACS analyses. Protein activation via phosphorylation was measured by immunoblotting. Autophagy was measured by LC3 immunofluorescence and immunoblotting.Results: AZ7328 inhibited proliferation and AKT substrate phosphorylation in a concentration-dependent manner but had minimal effects on apoptosis. Proliferative inhibition correlated loosely with the presence of activating PIK3CA mutations and was strengthened in combination with the mTOR inhibitor rapamycin. AZ7328 induced autophagy in some of the lines, and in the cells exposed to a combination of AZ7328 and chemical autophagy inhibitors apoptosis was induced.Conclusions: The cytostatic effects of AZ7328 correlate with PIK3CA mutations and are greatly enhanced by dual pathway inhibition using an mTOR inhibitor. Furthermore, AZ7328 can interact with autophagy inhibitors to induce apoptosis in some cell lines. Overall, our results support the further evaluation of combinations of PI3K/AKT/mTOR pathway and autophagy inhibitors in pre-clinical in vivo models and ultimately in patients with PIK3CA mutant bladder cancers.     

Targeting the phosphatidylinositol 3-kinase signaling pathway in breast cancer

The phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) network plays a key regulatory function in cell survival, proliferation, migration, metabolism, angiogenesis, and apoptosis. Genetic aberrations found at different levels, either with activation of oncogenes or inactivation of tumor suppressors, make this pathway one of the most commonly disrupted in human breast cancer. The PI3K-dependent phosphorylation and activation of the serine/threonine kinase AKT is a key activator of cell survival mechanisms. The activation of the oncogene PIK3CA and the loss of regulators of AKT including the tumor suppressor gene PTEN are mutations commonly found in breast tumors. AKT relieves the negative regulation of mTOR to activate protein synthesis and cell proliferation through S6K and 4EBP1. The common activation of the PI3K pathway in breast cancer has led to the development of compounds targeting the effector mechanisms of the pathway including selective and pan-PI3K/pan-AKT inhibitors, rapamycin analogs for mTOR inhibition, and TOR-catalytic subunit inhibitors. The influences of other oncogenic pathways such as Ras-Raf-Mek on the PI3K pathway and the known feedback mechanisms of activation have prompted the use of compounds with broader effect at multiple levels and rational combination strategies to obtain a more potent antitumor activity and possibly a meaningful clinical effect. Here, we review the biology of the network, its role in the development and progression of breast cancer, and the evaluation of targeted therapies in clinical trials.     

The PI3K/AKT/mTOR Signaling Pathway: Implications in the Treatment of Breast Cancer

Epidemiologic and experimental studies support a key role of the phosphatidyl inositol 3-kinase/AKT/mammalian target of rapamycin (PI3K/AKT/mTOR) pathway in the biology of human cancers. Alterations resulting in activation of PI3K/Akt/mTOR signaling are perhaps the most frequent events observed in solid tumors, including breast cancer, and contribute to neoplastic transformation. The PI3K/mTOR pathway can be activated by overproduction of growth factors or chemokines, loss of phosphatase and tensin homolog (PTEN) expression, or by mutations in growth factor receptors Ras, PTEN, or PI3K itself. Activation of this pathway contributes to cell cycle proliferation, growth, cell cycle entry, survival, cell motility, protein synthesis, and glucose metabolism, all important aspects of tumorigenesis. The most common genetic aberrations in breast cancer are activating somatic missense mutations in the gene encoding the p110a (PIK3CA) subunit of PI3K. The PTEN gene is often hypermethylated or decreased in expression, through as yet unclear mechanisms, in breast cancer. Studies have shown that PI3K/PTEN/AKT pathway modulation is implicated in HER2/neu-tumorigenesis and in response to the HER2-targeting antibody trastuzumab. Components of the pathway are regulated by feed-back and cross-talk to other signaling cascades and appear to be implicated with drug resistance. Over the past few years, a number of components of this signaling cascade have been the subject of intense drug-discovery activities. Rapamycin analogs have already been shown to have antitumor efficacy in some tumor types. Newer-generation PI3K, AKT, and mTOR inhibitors have shown significant promise preclinically and are now in clinical trials. This article summarizes the progress made in the elucidation of the pathway, clinical implications in pathology of breast cancer, and reviews novel drugs targeting this pathway for cancer treatment, particularly inhibitors of PI3K, AKT, and mTOR, currently undergoing clinical trials. Potential combination strategies, safety concerns, and resistance mechanisms for this new generation of anticancer agents are also discussed.     

Sorafenib enhances the therapeutic efficacy of rapamycin in colorectal cancers harboring oncogenic KRAS and PIK3CA

Activation of phosphatidylinositol 3-kinase (PI3K)/Akt signaling is associated with tumorigenesis and metastasis of colorectal cancer (CRC). The mammalian target of rapamycin (mTOR) kinase, a downstream effector of PI3K/Akt signaling, regulates tumorigenesis and metastasis of CRCs, indicating that mTOR inhibition may have therapeutic potential. Notwithstanding, many cancers, including CRC, demonstrate resistance to the antitumorigenic effects of rapamycin. In this study, we show that inhibition of mTORC1 with rapamycin leads to feedback activation of PI3K/Akt and Ras-MAPK signaling, resulting in cell survival and possible contribution to rapamycin resistance. Combination with the multikinase inhibitor, sorafenib, abrogates rapamycin-induced activation of PI3K/Akt and Ras-MAPK signaling pathways. Combination of rapamycin with sorafenib synergistically inhibits proliferation of CRC cells. CRCs harboring coexistent KRAS and PIK3CA mutations are partially sensitive to either rapamycin or sorafenib monotherapy, but highly sensitive to combination treatment with rapamycin and sorafenib. Combination with sorafenib enhances therapeutic efficacy of rapamycin on induction of apoptosis and inhibition of cell-cycle progression, migration and invasion of CRCs. We demonstrate efficacy and safety of concomitant treatment with rapamycin and sorafenib at inhibiting growth of xenografts from CRC cells with coexistent mutations in KRAS and PIK3CA. The efficacy and tolerability of combined treatment with rapamycin and sorafenib provides rationale for use in treating CRC patients, particularly those with tumors harboring coexistent KRAS and PIK3CA mutations.     

非小细胞肺癌组织中表皮生长因子受体磷脂酰肌醇3激酶蛋白激酶B1及哺乳动物雷帕霉素靶蛋白mRNA表达及其临床意义

目的探讨磷脂酰肌醇3激酶/蛋白激酶B1/哺乳动物雷帕霉素靶蛋白(PI3K/Akt1/m TOR)通路与表皮生长因子受体(EGFR)基因的关系,及其PI3K/Akt1/m TOR通路导致EGFR-酪氨酸激酶抑制剂(TKI)耐药的具体机制。方法选择135例未经过放化疗或靶向药物治疗的非小细胞肺癌(NSCLC)患者的肿瘤组织标本,检测其EGFR、PIK3CA、Akt1和m TOR的mRNA表达水平,并进行相关性分析。结果 EGFR与Akt1、m TOR的表达水平呈正相关(r分别为0.342、0.320和0.281,均P<0.01),与PIK3CA基因无关。结论 EGFR基因表达与Akt1和m TOR相关,与PIK3CA无关,提示EGFR基因可能直接激活Akt及其下游通路,而不经过PI3K。     

PI3K independent activation of mTORC1 as a target in lapatinib-resistant ERBB2+ breast cancer cells

Therapies targeting the ERBB2 receptor, including the kinase inhibitor lapatinib (Tykerb, GlaxoSmithKline), have improved clinical outcome for women with ERBB2-amplified breast cancer. However, acquired resistance to lapatinib remains a significant clinical problem, and the mechanisms governing resistance remain poorly understood. We sought to define molecular alterations that confer an acquired lapatinib resistance phenotype in ER?/ERBB2+ human breast cancer cells. ERBB2-amplified SKBR3 breast cancer cells were rendered resistant to lapatinib via culture in increasing concentrations of the drug, and molecular changes associated with a resistant phenotype were interrogated using a collaborative enzyme-enhanced immunoassay platform and immunoblotting techniques for detection of phosphorylated signaling cascade proteins. Interestingly, despite apparent inactivation of the PI3K/AKT signaling pathway, resistant cells exhibited constitutive activation of mammalian target of rapamycin complex 1 (mTORC1) and were highly sensitive to mTOR inhibition with rapamycin and the dual PI3K/mTOR inhibitor NVP-BEZ235. These data demonstrate a role for downstream activation of mTORC1 in the absence of molecular alterations leading to PI3K/AKT hyperactivation as a potential mechanism of lapatinib resistance in this model of ERBB2+ breast cancer and support the rationale of combination or sequential therapy using ERBB2 and mTOR-targeting molecules to prevent or target resistance to lapatinib. Moreover, our data suggest that assessment of mTOR substrate phosphorylation (i.e., S6) may serve as a more robust biomarker to predict sensitivity to mTOR inhibitors in the context of lapatinib resistance than PI3K mutations, loss of PTEN and p-AKT levels.     

PTEN deficiency is associated with reduced sensitivity to mTOR inhibitor in human bladder cancer through the unhampered feedback loop driving PI3K/Akt activation

Background:

Preclinical studies have shown that PTEN loss enhances sensitivity to mammalian target of Rapamycin (mTOR) inhibitors because of facilitated PI3K (phosphatidylinositol-3 kinase)/Akt activation and consecutive stimulation of the mTOR pathway. In patients with advanced transitional cell carcinoma (TCC) treated with the mTOR inhibitor everolimus, PTEN loss was, however, associated with resistance to treatment.

Methods:

Transitional cell carcinoma specimens, human bladder cancer cells and derived mouse xenografts were used to evaluate how the PTEN status influences the activity of mTOR inhibitors.

Results:

Transitional cell carcinoma patients with a shorter progression-free survival under everolimus exhibited PTEN deficiency and increased Akt activation. Moreover, PTEN-deficient bladder cancer cells were less sensitive to rapamycin than cells expressing wild-type PTEN, and rapamycin strikingly induced Akt activation in the absence of functional PTEN. Inhibition of Akt activation by the PI3K inhibitor wortmannin interrupted this rapamycin-induced feedback loop, thereby enhancing the antiproliferative effects of the mTOR inhibitor both in vitro and in vivo.

Conclusion:

Facilitation of Akt activation upon PTEN loss can have a more prominent role in driving the feedback loop in response to mTOR inhibition than in promoting the mTOR pathway. These data support the use of both PI3K and mTOR inhibitors to treat urothelial carcinoma, in particular in the absence of functional PTEN.     

Targeting phosphatidylinositol 3 kinase (PI3K)-Akt beyond rapalogs

The activation of the phosphatidylinositol 3 kinase (PI3K)-Akt pathway is a known causal mechanism of oncogenesis and resistance to cancer treatments. The process of PI3K-Akt pathway activation is complex and includes receptor tyrosine kinase(RTK) activation, PIK3CA mutations, loss of phosphatase and tensin homolog (PTEN), Akt mutations, tuberous sclerosis complex (TSC) mutations, and Ras homologue enriched in brain (RHEB) gene amplifications. The blockage of mammalian target of rapamycin (mTOR), the key downstream pathway protein, has been successful in selected cancer types, with mTOR-targeting agents available for clinical use. Other novel drugs blocking this pathway such as PI3K inhibitors, Akt inhibitors and PDK-1 inhibitors are currently only available for investigational use, but have shown promise as cancer therapies in both preclinical and early phase clinical studies. The newer generations of these inhibitors are more specific and have improved potency and safety. The combinations of targeted treatments against this pathway, blocking multiple different steps, are under preliminary investigation. Further research is needed to identify the biomarkers that predict treatment response and resistance in order to optimize personalized medicine.     

PI3K/PTEN/Akt pathway status affects the sensitivity of high-grade glioma cell cultures to the insulin-like growth factor-1 receptor inhibitor NVP-AEW541

IGF-1 receptor signaling contributes to the growth of many solid tumors, including glioblastoma. This study analyzed the sensitivity of 8 glioblastoma cultures to the IGF-1 receptor inhibitor NVP-AEW541. Growth reduction, caused by a combination of antiproliferative and proapoptotic effects, varied between 20% and 100%. Growth-inhibitory effects of IGF-1 receptor siRNA were also demonstrated in 2 of the cultures. Activating mutations in PIK3CA were found in 2 cultures, and 2 other cultures displayed ligand-independent Akt phosphorylation. Growth inhibition was significantly reduced in cultures with PIK3CA mutations or ligand-independent Akt phosphorylation. PTEN siRNA experiments supported the notion that the status of the PI3K/PTEN/Akt pathway is involved in determining NVP-AEW541 sensitivity. Combination treatments with either PI3 kinase or mTOR inhibitors together with NVP-AEW541 were performed. These experiments demonstrated the effects of NVP-AEW541 in cells not responding to mono-treatment with the IGF-1 receptor inhibitor, when used together with either of the 2 other inhibitors. Together, the studies support continued clinical development of IGF-1 receptor antagonists for glioblastomas and identify links between PI3K/PTEN/Akt status and sensitivity to mono-treatment with NVP-AEW541. Furthermore, the studies suggest that NVP-AEW541 is also active together with PI3 kinase and mTOR inhibitors in cultures with a dysregulated PI3K/PTEN/Akt pathway. These studies should assist in future clinical development of IGF-1 receptor antagonists for glioblastoma and other tumors.     

PI3K inhibitors in trastuzumab-resistant HER2-positive breast cancer cells with PI3K pathway alterations

The activation of the PI3K signaling pathway resulting from genetic alterations induces carcinogenesis and resistance to anticancer therapies. Breast cancer is a major malignancy that is associated with dysregulation of the PI3K signaling pathway. PIK3CA mutations and PTEN loss occur in every subtype of breast cancer. PI3K inhibitors are being evaluated in breast cancer after the success of an alpha isoform-specific PI3K inhibitor in estrogen receptor (ER)-positive/HER2-negative metastatic breast cancer. Some preclinical data indicate the potential for PI3K/mTOR targeting in combination with trastuzumab for HER2-positive breast cancer with or without expression of the estrogen receptor. However, the role of this therapy in HER2-positive breast cancer with PIK3CA mutations and/or PTEN loss remains unclear. We examined three HER2-positive, ER-negative breast cancer cell lines to determine the efficacy of a novel alpha isoform-specific PI3K inhibitor in combination with trastuzumab. The results indicated that this combination was effective in PIK3CA-mutant or PTEN-deficient breast cancer cells by inducing apoptosis and inhibiting the expression of downstream proteins. PTEN loss by siRNA modulation in parental HER2-positive cancer cells with PI3K signaling pathway alterations could not confer resistance to alpelisib or GDC-0077 plus trastuzumab. We selected the CK-MB-1 cell line without alterations in the PI3K pathway to demonstrate that PI3K inhibitors plus trastuzumab represented a biomarker-specific treatment. In vivo effects of alpelisib plus trastuzumab were tested and confirmed in a mouse model, showing the combination strategy offered the best opportunity to achieve tumor volume reduction. With known safety profiles, this cytotoxic chemotherapy-free regimen warrants further attention as a biomarker-driven strategy for treating HER2-positive breast cancer.     

Reduced PTEN expression in breast cancer cells confers susceptibility to inhibitors of the PI3 kinase/Akt pathway.

The PTEN protein is a lipid phosphatase with putative tumor suppressing abilities, including inhibition of the PI3K/Akt signaling pathway. Inactivating mutations or deletions of the PTEN gene, which result in hyper-activation of the PI3K/Akt signaling pathway, are increasingly being reported in human malignancies, including breast cancer, and have been related to features of poor prognosis and resistance to chemotherapy and hormone therapy. Prior studies in different tumor models have shown that, under conditions of PTEN deficiency, the PI3K/Akt signaling pathway becomes a fundamental proliferative and survival pathway, and that pharmacological inhibition of this pathway results in tumor growth inhibition. This study aimed to explore further this hypothesis in breast cancer cells. To this end, we have determined the growth response to inhibition of the PI3K/Akt signaling pathway in a series of breast cancer cell lines with different PTEN levels. The PTEN-negative cell line displayed greater sensitivity to the growth inhibitory effects of the PI3K inhibitor, LY294002 and rapamycin, an inhibitor of the PI3K/Akt downstream mediator mTOR, compared with the PTEN-positive cell lines. To determine whether or not these differences in response are specifically due to effects of PTEN, we developed a series of cell lines with reduced PTEN protein expression compared with the parental cell line. These reduced PTEN cells demonstrated an increased sensitivity to the anti-proliferative effects induced by LY294002 and rapamycin compared with the parental cells, which corresponded to alterations in cell cycle response. These findings indicate that inhibitors of mTOR, some of which are already in clinical development (CCI-779, an ester of rapamycin), have the potential to be effective in the treatment of breast cancer patients with PTEN-negative tumors and should be evaluated in this setting.     

PIK3CA mutations correlate with hormone receptors, node metastasis, and ERBB2, and are mutually exclusive with PTEN loss in human breast carcinoma

Deregulation of the phosphatidylinositol 3-kinase (PI3K) pathway either through loss of PTEN or mutation of the catalytic subunit alpha of PI3K (PIK3CA) occurs frequently in human cancer. We identified PIK3CA mutations in 26% of 342 human breast tumor samples and cell lines at about equal frequency in tumor stages I to IV. To investigate the relationship between PTEN and PIK3CA, we generated a cohort of tumors that had lost PTEN expression and compared it with a matched control set that had retained PTEN. A highly significant association between PIK3CA mutations and retention of PTEN protein expression was observed. In addition, PIK3CA mutations were associated with expression of estrogen and progesterone receptors (ER/PR), lymph node metastasis, and ERBB2 overexpression. The fact that PIK3CA mutations and PTEN loss are nearly mutually exclusive implies that deregulated phosphatidylinositol-3,4,5-triphosphate (PIP(3)) is critical for tumorigenesis in a significant fraction of breast cancers and that loss of PIP(3) homeostasis by abrogation of either PIK3CA or PTEN relieves selective pressure for targeting of the other gene. The correlation of PIK3CA mutation to ER/PR-positive tumors and PTEN loss to ER/PR-negative tumors argues for disparate branches of tumor evolution. Furthermore, the association between ERBB2 overexpression and PIK3CA mutation implies that more than one input activating the PI3K/AKT pathway may be required to overcome intact PTEN. Thus, mutation of PIK3CA is frequent, occurs early in carcinoma development, and has prognostic and therapeutic implications.     

PIK3CA mutations,phosphatase and tensin homolog,human epidermal growth factor receptor 2, and insulin-like growth factor 1 receptor and adjuvant tamoxifen resistance in postmenopausal breast cancer patients

Introduction

Inhibitors of the phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/AKT/mTOR) pathway can overcome endocrine resistance in estrogen receptor (ER) α-positive breast cancer, but companion diagnostics indicating PI3K/AKT/mTOR activation and consequently endocrine resistance are lacking. PIK3CA mutations frequently occur in ERα-positive breast cancer and result in PI3K/AKT/mTOR activation in vitro. Nevertheless, the prognostic and treatment-predictive value of these mutations in ERα-positive breast cancer is contradictive. We tested the clinical validity of PIK3CA mutations and other canonic pathway drivers to predict intrinsic resistance to adjuvant tamoxifen. In addition, we tested the association between these drivers and downstream activated proteins.

Methods

Primary tumors from 563 ERα-positive postmenopausal patients, randomized between adjuvant tamoxifen (1 to 3 years) versus observation were recollected. PIK3CA hotspot mutations in exon 9 and exon 20 were assessed with Sequenom Mass Spectometry. Immunohistochemistry was performed for human epidermal growth factor receptor 2 (HER2), phosphatase and tensin homolog (PTEN), and insulin-like growth factor 1 receptor (IGF-1R). We tested the association between these molecular alterations and downstream activated proteins (like phospho-protein kinase B (p-AKT), phospho-mammalian target of rapamycin (p-mTOR), p-ERK1/2, and p-p70S6K). Recurrence-free interval improvement with tamoxifen versus control was assessed according to the presence or absence of canonic pathway drivers, by using Cox proportional hazard models, including a test for interaction.

Results

PIK3CA mutations (both exon 9 and exon 20) were associated with low tumor grade. An enrichment of PIK3CA exon 20 mutations was observed in progesterone receptor- positive tumors. PIK3CA exon 20 mutations were not associated with downstream-activated proteins. No significant interaction between PIK3CA mutations or any of the other canonic pathway drivers and tamoxifen-treatment benefit was found.

Conclusion

PIK3CA mutations do not have clinical validity to predict intrinsic resistance to adjuvant tamoxifen and may therefore be unsuitable as companion diagnostic for PI3K/AKT/mTOR inhibitors in ERα- positive, postmenopausal, early breast cancer patients.     

Frequent alterations of the PI3K/AKT/mTOR pathways in hereditary nonpolyposis colorectal cancer

The phosphatidylinositol 3-kinases-AKT-mammalian target of rapamycin pathway (PI3K/AKT/mTOR) is central in colorectal tumors. Data on its role in hereditary cancers are, however, scarce and we therefore characterized mutations in PIK3CA and KRAS, and expression of PIK3CA, phosphorylated AKT, and PTEN in colorectal cancers linked to hereditary nonpolyposis colorectal cancer (HNPCC). Sequencing was used to identify mutations in PIK3CA, a real-time PCR-based method to identify KRAS mutations, and immunohistochemical staining was used to evaluate the expression of PIK3CA, phosphorylated AKT and PTEN in 58 HNPCC-associated colorectal cancers. Derangements of at least one of the PI3K/AKT/mTOR components analyzed were found in 51/58 (88%) tumors. Mutations in PIK3CA and KRAS were identified in 14 and 31% of the tumors respectively. Overexpression of PIK3CA and phosphorylated AKT occurred in 59 and 75% and were strongly associated (P = 0.005). Reduced/lost PTEN expression was found in 63% of the tumors. Though HNPCC-associated colorectal cancers show simple genetic profiles with few chromosomal alterations, we demonstrate frequent and repeated targeting of the PI3K/AKT/mTOR pathway, which suggests that therapeutic strategies directed at this pathway are likely to be beneficial also in HNPCC.     

Involvement of Akt-1 and mTOR in sensitivity of breast cancer to targeted therapy

Elucidating the response of breast cancer cells to chemotherapeutic and hormonal based drugs is clearly important as these are frequently used therapeutic approaches. A signaling pathway often involved in chemo- and hormonal-resistance is the Ras/PI3K/PTEN/Akt/mTOR cascade. In the studies presented in this report, we have examined the effects of constitutive activation of Akt on the sensitivity of MCF-7 breast cancer cells to chemotherapeutic- and hormonal-based drugs as well as mTOR inhibitors. MCF-7 cells which expressed a constitutively-activated Akt-1 gene [ΔAkt-1(CA)] were more resistant to doxorubicin, etoposide and 4-OH-tamoxifen (4HT) than cells lacking ΔAkt-1(CA). Cells which expressed ΔAkt-1(CA) were hypersensitive to the mTOR inhibitor rapamycin. Furthermore, rapamycin lowered the IC₅₀s for doxorubicin, etoposide and 4HT in the cells which expressed ΔAkt-1(CA), demonstrating a potential improved method for treating certain breast cancers which have deregulated PI3K/PTEN/Akt/mTOR signaling. Understanding how breast cancers respond to chemo- and hormonal-based therapies and the mechanisms by which they can become drug resistant may enhance our ability to treat breast cancer. These results also document the potential importance of knowledge of the mutations present in certain cancers which may permit more effective therapies.