Faculty Publications

Thyroid tumors arising from the follicular cells often harbor mutations leading to the constitutive activation of the PI3K and Ras signaling cascades. However, it is still unclear what their respective contribution to the neoplastic process is, as well as to what extent they interact. We have used mice harboring a Kras oncogenic mutation and a Pten deletion targeted to the thyroid epithelium to address in vivo these questions. Here, we show that although each of these two pathways, alone, :is unable to transform thyroid follicular cells, their simultaneous activation is highly oncogenic, leading to invasive and metastatic follicular carcinomas. In particular, phosphatidylinositol-3-kinase (PI3K) activation suppressed Kras-initiated feedback signals that uncouple mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinase (MEK) and ERK activation, thus stunting MAPK activity; in addition, PI3K and Kras cooperated to drastically up-regulate cyclin D1 mRNA levels. Finally, combined pharmacologic inhibition of PI3K and MAPK completely inhibited the growth of double-mutant cancer cell lines, providing a compelling rationale for the dual targeting of these pathways in thyroid cancer. [Cancer Res 2009;69(8):3689-94]

The oncogene v-akt was isolated from a retrovirus that induced murine thymic lymphomas. Transgenic mice expressing a constitutively activated form of the cellular homologue Akt2 specifically in immature T cells develop spontaneous thymic lymphomas. We hypothesized that tumors from these mice might exhibit oncogenic chromosomal rearrangements that cooperate with activated Akt2 in lymphomagenesis. Cytogenetic analysis revealed a recurrent clonal inversion of chromosome 6, inv(6), in thymic lymphomas from multiple transgenic founder lines, including one line in which 15 of 15 primary tumors exhibited this same rearrangement. Combined fluorescence in situ hybridization, PCR, and DNA sequence analyses showed that the distal inv(6) breakpoint resides at the T-cell receptor beta chain locus, Tcrb. The proximal breakpoint maps to a region near a locus comprising the linked homeobox/transcription factor genes Dlx5 and Dlx6. Expression analysis of genes translocated to the vicinity of the Tcrb enhancer revealed that Dlx5 and Dlx6 are overexpressed in tumors exhibiting the inv(6). Experimental overexpression of Dlx5 in mammalian cells resulted in enhanced cell proliferation and increased colony formation, and clonogenic assays revealed cooperativity when both Dlx5 and activated Akt2 were coexpressed. In addition, DLX5, but not DLX6, was found to be abundantly expressed in three of seven human T-cell lymphomas tested. These findings suggest that the Dlx5 can act as an oncogene by cooperating with Akt2 to promote lymphomagenesis.

Thyroid-stimulating hormone (TSH) has long been recognized as the major proliferative and functional stimulus for thyroid follicular cells. TSH receptor (TSHR) engagement stimulates the production of cyclic AMP and the subsequent activation of downstream effector molecules, including protein kinase A, S6K1, and Rap1, whereas the role of the RAS and phosphatidylinositol-3-kinase signaling cascades downstream of TSHR is still controversial. Despite the abundance of candidates, it is still unclear which of these pathways represent(s) the key mitogenic output of TSH-initiated signaling. We have used an in vivo model of goitrogenesis to dissect the contribution of these pathways to TSH-induced thyrocyte proliferation and thyroid hyperplasia. We show that the in vivo proliferative response to chronic TSHR stimulation relies heavily on the activation of the mTOR/S6K1 axis, and that mTOR inhibition during goitrogenic stimulation abrogates the hyperplastic but not the hypertrophic thyrocyte responses to TSH, thus functionally uncoupling these two processes. Strikingly, goitrogenesis was not associated with an increase in AKT phosphorylation levels, underlining the existence of an AKT-independent pathway leading to mTOR activation upon TSH stimulation. [Cancer Res 2007;67(17):8002-6].

Endometrial carcinoma, a common malignancy of the female genital tract, is composed of a number of tumor types with different light-microscopic features, molecular genetic alterations, and prognoses. In addition, hormonal influences significantly impact growth regulatory pathways and interact with genetic alterations in the pathogenesis of at least some types of endometrial carcinorna. These factors have complicated the analyses of endometrial carcinoma, but over the past decades, awareness of the different types of endometrial carcinoma, in addition to careful clinicopathological studies, molecular analyses, and animal studies of the biological underpinnings of the different tumor types, has increased. We present the current understanding of endometrial carcinoma, from a molecular vantage point, highlighting what are presently thought to be the fundamental pathways involved in the development and progression of the major types of endometrial carcinorna.

Loss of PTEN is the earliest detectable genetic lesion in the endometrioid subtype of endometrial cancer (EEC), a tumor thought to be associated with an increase in unopposed estrogen activity. Pten(+)/(-) mice develop endometrial neoplastic lesions with full penetrance, despite having normal estrogen levels. We have utilized oligonucleoticle arrays to identify the alterations in gene expression patterns associated with loss of Pten and consequent neoplastic transformation of the endometrium. We show that 487 and 330 genes are substantially up- and downregulated, respectively, in Pten(+/-) mice. Several genes whose expression levels are impacted by loss of Pten are associated with pathways and functions that are relevant to the transformation and progression processes. Strikingly, we found that the expression levels of over 100 genes known to be regulated by estrogen receptor alpha (ER alpha) are also altered in the neoplastic uterus from Pten(+)/(-) mice, thus mimicking a h! yperestrogenic environment. These results provide in vivo evidence supporting the hypothesis that loss of Pten and subsequent Akt activation result in the activation of several ER alpha-dependent pathways that, mimicking increased estrogen signaling, may play a pivotal role in the neoplastic process.

Context: TGFBR1*6A is a common polymorphis of the type I transforming growth factor ? receptor (TGFBR1). Epidemiological studies suggest that TGFBR1*6A may act as a tumor susceptibility allele. How TGFBR1*6A contributes to cancer development is largely unknown. Objectives: To determine whether TGFBR1*6A is somatically acquired by primary tumors and metastases during cancer development and whether the 3-amino acid deletion that differentiates TGFBR1*6A from TGFBR1 is part of the mature receptor or part of the signal sequence and to investigate TGFBR1*6A signaling in cancer cells. Design, Setting, and Patients: Tumor and germline tissues from 531 patients with a diagnosis of head and neck, colorectal, or breast cancer recruited from 3 centers in the United States and from 1 center in Spain from June 1, 1994, through June 30, 2004. In vitro translation assays, MCF-7 breast cancer cells stably transfected with TGFBR1*6A, TGFBR1, or the vector alone, DLD-1 colorectal cancer cells that endogenously carry TGFBR1*6A, and SW48 colorectal cancer cells that do not carry TGFBR1*6A. Main Outcome Measures: TGFBR1*6A somatic acquisition in cancer. Determination of the amino terminus of the mature TGFBR1*6A and TGFBR1 receptors. Determination of TGF-?-dependent cell proliferation. Results: TGFBR1*6A was somatically acquired in 13 of 44 (29.5%) colorectal cancer metastases, in 4 of 157 (2.5%) of colorectal tumors, in 4 of 226 (1.8%) head and neck primary tumors, and in none of the 104 patients with breast cancer. TGFBR1*6A somatic acquisition is not associated with loss of heterozygosity, microsatellite instability, or a mutator phenotype. The signal sequences of TGFBR1 and TGFBR1*6A are cleaved at the same site resulting in identical mature receptors. TGFBR1*6A may switch TGF-? growth inhibitory signals into growth stimulatory signals in MCF-7 breast cancer cells and in DLD-1 colorectal cancer cells. Conclusions: TGFBR1*6A is somatically acquired in 29.5% of liver metastases from colorectal cancer and may bestow cancer cells with a growth advantage in the presence of TGF-?. The functional consequences of this conversion appear to be mediated by the TGFBR1*6A signal sequence rather than by the mature receptor. The results highlight a new facet of TGF-? signaling in cancer and suggest that TGFBR1*6A may represent a potential therapeutic target in cancer. ©2005 American Medical Association. All rights reserved.

Activation of AKT and overexpression of fatty acid synthase (FAS) are frequently obsd. in human ovarian cancer. To explore a possible connection between AKT and FAS, immunohistochem. analyses were conducted on an ovarian cancer tissue microarray, which revealed a significant correlation between phosphorylated AKT (phospho-AKT) and expression of FAS. To investigate the relationship between phospho-AKT and FAS in vitro, a variety of expts. employing a specific phosphatidylinositol 3-OH kinase (PI3K) inhibitor (LY294002), inducible PTEN expression in PTEN-null cells, or AKT1 siRNA demonstrated that phosphatidylinositol-3 kinase (PI3K)/AKT signaling modulates FAS expression. In contrast, inhibition of FAS activity by the drug C75 resulted in downregulation of phospho-AKT and increased cell death. To explore the functional relationship between phospho-AKT and FAS, the authors used SKOV3, C200, and OVCAR10 ovarian carcinoma cells, which have constitutively active AKT, and OVCAR5 cells, which have very low basal phospho-AKT levels. Treatment with LY294002 abolished AKT activity and potentiated apoptosis induced by FAS inhibitors cerulenin or C75 only in cells with constitutively active AKT, suggesting that constitutive activation of AKT protects against FAS inhibitor-induced cell death. Furthermore, inhibition of FAS activity by cerulenin or C75 resulted in downregulation of phospho-AKT, which preceded the induction of apoptosis. To investigate the relationship between phospho-AKT and FAS in vivo, severe combined immunodeficient mice injected i.p. with SKOV3 cells were treated with C75. Growth of SKOV3 xenografts was markedly inhibited by C75. Anal. of the levels of phospho-AKT and FAS in C75-treated tumors revealed concordant downregulation of phospho-AKT and FAS. Collectively, the authors' findings are consistent with a working model in which AKT activation regulates FAS expression, at least in part, whereas FAS activity modulates AKT activation. [on SciFinder (R)]

Objective. The lack of an endometrial epithelium-specific promoter has slowed down the development of technically advanced mouse models of endometrial cancer. The aim of this study was to test whether direct in vivo adenoviral-mediated gene delivery can be used to circumvent this problem. Methods. Adenoviruses expressing the LacZ reporter gene or the Cre recombinase were injected into the left horn of the mouse uterus. Histochemistry and immunohistochemistry were used to detect expression of the reporter gene as well as targeted deletion of a floxed allele. Results. Our data demonstrate that in vivo direct injection of adenoviruses can efficiently target the endometrium in the mouse, specifically transducing genes to the glandular epithelial component. Conclusions. This approach will allow the generation of more refined and genetically defined mouse models of endometrial cancer. Endometrial gland-specific transient expression of recombinases, such as Cre, may thus be employed to delete engineered alleles of tumor suppressor genes and to activate the expression of latent oncogenes. (C) 2004 Elsevier Inc. All rights reserved.