Biomedical subjects

S Maheswaran

Publications and source records attributed to S Maheswaran.

At least 19 recordsLinked to original sources

PubMed2005

Mullerian inhibiting substance suppresses tumor growth in the C3(1)T antigen transgenic mouse mammary carcinoma model.

Mullerian inhibiting substance (MIS) inhibits breast cancer cell growth in vitro. To extend the use of MIS to treat breast cancer, it is essential to test the responsiveness of mammary tumor growth to MIS in vivo. Mammary tumors arising in the C3(1) T antigen mouse model expressed the MIS type II receptor, and MIS in vitro inhibited the growth of cells derived from tumors. Administration of MIS to mice was associated with a lower number of palpable mammary tumors compared with vehicle-treated mice (P=0.048), and the mean mammary tumor weight in the MIS-treated group was significantly lower compared with the control group (P=0.029). Analysis of proliferating cell nuclear antigen (PCNA) expression and caspase-3 cleavage in tumors revealed that exposure to MIS was associated with decreased proliferation and increased apoptosis, respectively, and was not caused by a decline in T antigen expression. The effect of MIS on tumor growth was also evaluated on xenografted human breast cancer cell line MDA-MB-468, which is estrogen receptor- and retinoblastoma-negative and expresses mutant p53, and thus complements the C3(1)Tag mouse mammary tumors that do not express estrogen receptor and have functional inactivation of retinoblastoma and p53. In agreement with results observed in the transgenic mice, MIS decreased the rate of MDA-MB-468 tumor growth and the gain in mean tumor volume in severe combined immunodeficient mice compared with vehicle-treated controls (P=0.004). These results suggest that MIS can suppress the growth of mammary tumors in vivo.

Animals↗

PubMed2004

Transforming Growth Factor-beta superfamily: evaluation as breast cancer biomarkers and preventive agents.

The Transforming Growth Factor-beta (TGFbeta) superfamily of cytokines is comprised of a number of structurally-related, secreted polypeptides that regulate a multitude of cellular processes including proliferation, differentiation and neoplastic transformation. These growth regulatory molecules induce ligand-mediated hetero-oligomerization of distinct type II and type I serine/threonine kinase receptors that transmit signals predominantly through receptor-activated Smad proteins but also induce Smad-independent pathways. Ligands, receptors and intracellular mediators of signaling initiated by members of the TGFbeta family are expressed in the mammary gland and disruption of these pathways may contribute to the development and progression of human breast cancer. Since many facets of TGFbeta and breast cancer have been recently reviewed in several articles, except for discussion of recent developments on some aspects of TGFbeta, the major focus of this review will be on the role of activins, inhibins, BMPs, nodal and MIS-signaling in breast cancer with emphasis on their utility as potential diagnostic, prognostic and therapeutic targets.

Activins↗

PubMed2003

Synthesis, spectra, dioxygen affinity and antifungal activity of Ru(III) Schiff base complexes.

The reaction of ruthenium(III) complexes, [RuX(3)(EPh(3))(3)] (E=As, X=Cl or Br; E=P, X=Cl) and [RuBr(3)(PPh(3))(2)(CH(3)OH)] with bidendate Schiff base ligands derived by condensing salicylaldehyde with methylamine (Hsalmet), cyclohexylamine (Hsalchx), 2-aminopyridine (Hsalampy) have been carried out. The complexes were characterized by analytical and spectral studies (IR, electronic and EPR) and are formulated as [RuX(EPh(3))(LL')(2)] (where LL'=monobasic bidentate Schiff base ligand; E=P or As, X=Cl or Br). An octahedral geometry has been tentatively proposed for the new complexes. Dioxygen affinity of some of the Ru(III) Schiff base complexes was studied by cyclic voltammetry. The representative Schiff bases and their complexes were tested in vitro to evaluate their activity against fungi, namely, Aspergillus flavus (A. flavus) and Fusarium species.

Antifungal Agents↗

PubMed2003

Computational models of oral and craniofacial development, growth, and repair.

This paper illustrates how biological and clinical problems stimulate research in biomedical informatics and how such research contributes to their solution. The computational models described use techniques from Logic Programming, Machine Learning, Computer Vision, and Biomathematics. They address problems in the development, growth, and repair of oral and craniofacial tissues arising in cell biology, clinical genetics, and dentistry. At the micro-level, the dynamic interaction of cells in the oral epithelium is modeled. At the macro-level, models are constructed of either the craniofacial shape of an individual or the craniofacial shape differences within and between healthy and congenitally abnormal populations. In between, in terms of scale, there are models of normal dentition and the use of computerized expert knowledge to guide the design of dental prostheses used to restore function in partially edentulous patients.

Adult↗

PubMed2001

BRCA1 and GADD45 mediated G2/M cell cycle arrest in response to antimicrotubule agents.

BRCA1 is a tumour suppressor gene implicated in the predisposition to early onset breast and ovarian cancer. We have generated cell lines with inducible expression of BRCA1 to evaluate its role in mediating the cellular response to various chemotherapeutic drugs commonly used in the treatment of breast and ovarian cancer. Induction of BRCA1 in the presence of Taxol and Vincristine resulted in a dramatic increase in cell death; an effect that was preceded by an acute arrest at the G2/M phase of the cell cycle and which correlated with BRCA1 mediated induction of GADD45. A proportion of the arrested cells were blocked in mitosis suggesting activation of both a G2 and a mitotic spindle checkpoint. In contrast, no specific interaction was observed between BRCA1 induction and treatment of cells with a range of DNA damaging agents including Cisplatin and Adriamycin. Inducible expression of GADD45 in the presence of Taxol induced both G2 and mitotic arrest in these cells consistent with a role for GADD45 in contributing to these effects. Our results support a role for both BRCA1 and GADD45 in selectively regulating a G2/M checkpoint in response to antimicrotubule agents and raise the possibility that their expression levels in cells may contribute to the toxicity observed with these compounds.

Antineoplastic Agents↗

PubMed2001

Mullerian inhibiting substance regulates NFkappaB signaling and growth of mammary epithelial cells in vivo.

Müllerian inhibiting substance (MIS) inhibits breast cancer cell growth in vitro through interference with cell cycle progression and induction of apoptosis, a process associated with NFkappaB activation and up-regulation of one of its important target genes, IEX-1S (Segev, D. L., Ha, T., Tran, T. T., Kenneally, M., Harkin, P., Jung, M., MacLaughlin, D. T., Donahoe, P. K., and Maheswaran, S. (2000) J. Biol. Chem. 275, 28371-28379). Here we demonstrate that MIS activates the NFkappaB signaling cascade, induces IEX-1S mRNA, and inhibits the growth of MCF10A, an immortalized human breast epithelial cell line with characteristics of normal cells. In vivo, an inverse correlation was found to exist between various stages of mammary growth and MIS type II receptor expression. Receptor mRNA significantly diminished during puberty, when the ductal system branches and invades the adipose stroma and during the expansive growth at lactation, but it was up-regulated during involution, a time of regression and apoptosis. Peripartum variations in MIS type II receptor expression correlated with NFkappaB activation and IEX-1S mRNA expression. Administration of MIS to female mice induced NFkappaB DNA binding and IEX-1S mRNA expression in the breast. Furthermore, exposure to MIS in vivo increased apoptosis in the mouse mammary ductal epithelium. Thus, MIS may function as an endogenous hormonal regulator of NFkappaB signaling and growth in the breast.

Animals↗

PubMed2001

Müllerian inhibiting substance: an instructive developmental hormone with diagnostic and possible therapeutic applications.

Dr. Alfred Jost pioneered the field of reproductive endocrinology with his seminal observation that two hormones produced by the testes are required for the male embryo to develop a normal internal reproductive tract. T induces the Wolffian ducts to differentiate into epididymides, vasa deferens, and seminal vesicles. Müllerian inhibiting substance (MIS) causes regression of the Müllerian ducts, which in its absence would normally develop into the Fallopian tubes, uterus, and upper vagina as is observed in female embryos. This review will summarize our current understanding of molecular mechanisms underlying the function of MIS both as a fetal gonadal hormone that causes Müllerian duct regression and as an adult hormone, the roles for which are currently being investigated, i.e., inhibition of steroidogenesis, germ cell development, and cancer. We will also address the regulation of MIS expression as one of the first genes expressed after the commitment of the bipotential gonads to differentiate into testes under the influence of SRY, the gene on the sex-determining region of the Y chromosome. We will discuss what is known regarding MIS signal transduction, which as with other members of the TGFbeta family of growth and differentiation factors, occurs through a heteromeric complex of single transmembrane serine/threonine kinase receptors to effect downstream signaling events, including Smad, nuclear factor-kappaB, beta-catenin, and p16 activation. Finally, we will assess the clinical relevance of studying MIS in patients with persistent Müllerian duct syndrome and our efforts to determine the therapeutic value of MIS for patients with ovarian and other MIS receptor-expressing cancers.

Aging↗

PubMed2000

Mullerian inhibiting substance inhibits ovarian cell growth through an Rb-independent mechanism.

Müllerian inhibiting substance (MIS), a transforming growth factor-beta family member, causes regression of the Müllerian duct in male embryos. MIS overexpression in transgenic mice ablates the ovary, and MIS inhibits the growth of ovarian cancer cell lines in vitro, suggesting a key role for this hormone in postnatal development of the ovary. This report describes a mechanism for MIS-mediated growth inhibition in both a human epithelial ovarian cancer cell line and a cell line derived from normal ovarian surface epithelium, which is the origin of human epithelial ovarian cancers. MIS-treated cells accumulated in the G(1) phase of the cell cycle and subsequently underwent apoptosis. MIS up-regulated the cyclin-dependent kinase inhibitor p16 through an MIS type II receptor-mediated mechanism and inhibited growth in the absence of detectable or inactive Rb protein. Prolonged treatment with MIS down-regulated the Rb-related protein p130 and increased the Rb family-regulated transcription factor E2F1, overexpression of which inhibited growth. These findings demonstrate that p16 is required for MIS-mediated growth inhibition in ovarian epithelial cells and tumor cells and suggest that up-regulation of E2F1 also plays a role in this process.

Animals↗

PubMed2000

Mullerian inhibiting substance inhibits breast cancer cell growth through an NFkappa B-mediated pathway.

Müllerian inhibiting substance (MIS), a member of the transforming growth factor-beta superfamily, induces regression of the Müllerian duct in male embryos. In this report, we demonstrate MIS type II receptor expression in normal breast tissue and in human breast cancer cell lines, breast fibroadenoma, and ductal adenocarcinomas. MIS inhibited the growth of both estrogen receptor (ER)-positive T47D and ER-negative MDA-MB-231 breast cancer cell lines, suggesting a broader range of target tissues for MIS action. Inhibition of growth was manifested by an increase in the fraction of cells in the G(1) phase of the cell cycle and induction of apoptosis. Treatment of breast cancer cells with MIS activated the NFkappaB pathway and selectively up-regulated the immediate early gene IEX-1S, which, when overexpressed, inhibited breast cancer cell growth. Dominant negative IkappaBalpha expression ablated both MIS-mediated induction of IEX-1S and inhibition of growth, indicating that activation of the NFkappaB signaling pathway was required for these processes. These results identify the NFkappaB-mediated signaling pathway and a target gene for MIS action and suggest a putative role for the MIS ligand and its downstream interactors in the treatment of ER-positive as well as negative breast cancers.

Animals↗

PubMed1999

Induction of GADD45 and JNK/SAPK-dependent apoptosis following inducible expression of BRCA1.

The breast cancer susceptibility gene BRCA1 encodes a protein implicated in the cellular response to DNA damage, with postulated roles in homologous recombination as well as transcriptional regulation. To identify downstream target genes, we established cell lines with tightly regulated inducible expression of BRCA1. High-density oligonucleotide arrays were used to analyze gene expression profiles at various times following BRCA1 induction. A major BRCA1 target is the DNA damage-responsive gene GADD45. Induction of BRCA1 triggers apoptosis through activation of c-Jun N-terminal kinase/stress-activated protein kinase (JNK/SAPK), a signaling pathway potentially linked to GADD45 gene family members. The p53-independent induction of GADD45 by BRCA1 and its activation of JNK/SAPK suggest a pathway for BRCA1-induced apoptosis.

Apoptosis↗

PubMed1999

Human ovarian cancer, cell lines, and primary ascites cells express the human Mullerian inhibiting substance (MIS) type II receptor, bind, and are responsive to MIS.

Six human ovarian cancer cell lines and samples of ascites cells isolated from 27 patients with stage III or IV ovarian papillary serous cystadenocarcinoma were studied individually to test whether recombinant human Mullerian inhibiting substance (rhMIS) acts via its receptor. To do these experiments, we scaled up production of rhMIS and labeled it successfully with biotin for binding studies, cloned the human MIS type II receptor for mRNA detection, and raised antibodies to an extracellular domain peptide for protein detection. These probes were first tested on the human ovarian cancer cell lines and then applied to primary ovarian ascites cells. rhMIS inhibited colony growth of five of six cell lines that expressed the human MIS type II receptor mRNA by Northern analysis while not inhibiting receptor-negative COS cells. Flow cytometry performed on MIS-sensitive ovarian cancer cell lines demonstrated specific and saturable binding of rhMIS (Kd = 10.2 nM). Ascites cells from 15 of 27 or 56% of patients tested bound biotinylated MIS (MIS-biotin) and, of the 11 that grew in soft agarose, 9 of 11 or 82% showed statistically significant inhibition of colony formation. Of the 15 patients who bound biotinylated MIS, mRNA was available for analysis from 9, and 8 of 9 expressed MIS type II receptor mRNA by reverse transcription-PCR, showing a statistically significant correlation, compared with binding, by chi2 analysis (P = 0.025). Solid ovarian cancers were positive for the MIS type II receptor protein by immunohistochemical staining, which colocalized with staining for antibody to CA-125 (OC-125). Thus, the detection of the MIS type I receptor by flow cytometry may be a useful predictor of therapeutic response to MIS and may be a modality to rapidly choose patients with late-stage ovarian cancer for treatment with MIS.

Adult↗

PubMed1998

E1B 55K sequesters WT1 along with p53 within a cytoplasmic body in adenovirus-transformed kidney cells.

WT1 encodes a tumor suppressor that is expressed in cells of the developing kidney and is inactivated in Wilms tumor, a pediatric kidney cancer. The adenovirus E1B 55K gene product contributes to the transformation of primary baby rat kidney (BRK) cells by binding and inactivating the product of the p53 tumor suppressor. We have previously demonstrated that WT1 and p53 are present within a protein complex in vivo. We now show that WT1 is physically associated with E1B 55K in adenovirus-transformed cells, an interaction that is mediated by the first two zinc fingers of WT1. Immunodepletion of p53 abrogates the coimmunoprecipitation of E1B 55K and WT1, consistent with the presence of a trimeric protein complex containing these three proteins. In the presence of E1B 55K, WT1 which is normally localized in the nucleus, is retained within a very high molecular weight complex and sequestered in the characteristic perinuclear cytoplasmic body that contains E1B 55K and p53. Expression of E1B 55K in osteosarcoma cells that undergo apoptosis following expression of WT1 inhibits WT1-mediated cell death. We conclude that E1B 55K may target WT1 along with p53, resulting in the functional inactivation of both tumor suppressor gene products by this viral oncoprotein.

Adenoviridae↗

PubMed1998

Inhibition of cellular proliferation by the Wilms tumor suppressor WT1 requires association with the inducible chaperone Hsp70.

The Wilms tumor suppressor WT1 encodes a zinc finger transcription factor that is expressed in glomerular podocytes during a narrow window in kidney development. By immunoprecipitation and protein microsequencing analysis, we have identified a major cellular protein associated with endogenous WT1 to be the inducible chaperone Hsp70. WT1 and Hsp70 are physically associated in embryonic rat kidney cells, in primary Wilms tumor specimens and in cultured cells with inducible expression of WT1. Colocalization of WT1 and Hsp70 is evident within podocytes of the developing kidney, and Hsp70 is recruited to the characteristic subnuclear clusters that contain WT1. The amino-terminal transactivation domain of WT1 is required for binding to Hsp70, and expression of that domain itself is sufficient to induce expression of Hsp70 through the heat shock element (HSE). Substitution of a heterologous Hsp70-binding domain derived from human DNAJ is sufficient to restore the functional properties of a WT1 protein with an amino-terminal deletion, an effect that is abrogated by a point mutation in DNAJ that reduces binding to Hsp70. These observations indicate that Hsp70 is an important cofactor for the function of WT1, and suggest a potential role for this chaperone during kidney differentiation.

Amino Acid Sequence↗

PubMed1997

Induction of p21 by the Wilms' tumor suppressor gene WT1.

WT1 encodes a zinc finger transcription factor that is expressed in the developing kidney and the inactivation of which leads to Wilms' tumor, a pediatric kidney cancer. We have recently shown that inducible expression of WT1 in osteosarcoma cells triggers programmed cell death, an effect that is associated with transcriptional repression of the endogenous epidermal growth factor receptor. We now show that WT1-mediated apoptosis is preceded by induction of the cyclin-dependent kinase inhibitor p21, associated with G1 phase arrest. This effect is only demonstrated by WT1 isoforms with an intact DNA binding domain, and it is associated with increased expression of endogenous p21 mRNA. WT1-mediated induction of p21 is independent of p53, another tumor suppressor gene known to regulate p21 expression. In the kidney, p21 is expressed in differentiating glomerular podocytes along with WT1. We conclude that induction of p21 expression may contribute to WT1-dependent differentiation pathways in the kidney and potentially to the function of WT1 as a tumor suppressor gene.

Anti-Bacterial Agents↗

PubMed1997

The EWS-WT1 translocation product induces PDGFA in desmoplastic small round-cell tumour.

Chromosomal translocations resulting in chimaeric transcription factors underlie specific malignancies, but few authentic target genes regulated by these fusion proteins have been identified. Desmoplastic small round-cell tumour (DSRT) is a multiphenotypic primitive tumour characterized by massive reactive fibrosis surrounding nests of tumour cells. The t(11;22)(p13;q12) chromosomal translocation that defines DSRT produces a chimaeric protein containing the potential transactivation domain of the Ewing-sarcoma protein (EWS) fused to zinc fingers 2-4 of the Wilms tumour suppressor and transcriptional repressor WT1 (refs 2,3). By analogy with other EWS fusion products, the EWS-WT1 chimaera may encode a transcriptional activator whose target genes overlap with those repressed by WT1 (ref. 4). To characterize its functional properties, we generated osteosarcoma cell lines with tightly regulated inducible expression of EWS-WT1. Expression of EWS-WT1 induced the expression of endogenous platelet-derived growth factor-A (PDGFA), a potent secreted mitogen and chemoattractant whose promoter contains the many potential WT1-binding sites. Native PDGFA was not regulated by wild-type WT1, indicating a difference in target gene specificity between this tumour suppressor and its oncogenic derivative. PDGFA was expressed within tumour cells in primary DSRT specimens, but it was absent in Wilms tumours expressing WT1 and Ewing sarcomas with an EWS-Fli translocation. We conclude that the oncogenic fusion of EWS to WT1 in DSRT results in the induction of PDGFA, a potent fibroblast growth factor that contributes to the characteristic reactive fibrosis associated with this unique tumour.

Binding Sites↗

PubMed1996

Functional properties of WT1.

WT1 encodes a zinc finger transcription factor that is inactivated in a subset of Wilms' tumors. We have recently shown that introduction of wild-type WT1 into a Wilms' tumor-derived cell line, RM1, results in growth suppression, consistent with its function as a tumor suppressor gene. WT1-mediated growth suppression was also observed in other cells derived from embryonal tumors, including two osteosarcoma cell lines, U2OS and Saos-2, notable for the respective presence or absence of wild-type p53. To further characterize the functional properties of WT1, multiple U2OS and Saos-2 cell lines were established, expressing either wild-type WT1 splicing variants or naturally occurring mutants under control of a tightly regulated tetracycline repressable promoter. Induction of WT1 in these cells resulted in programmed cell death. This effect was preferentially mediated by WT1 isoform B (encoding alternative splice I, lacking alternative splice II "KTS"), and it was independent of p53, occurring in both U2OS and Saos-2 cells. WT1-mediated apoptosis was associated with transcriptional repression of the epidermal growth factor receptor (EGFR) and reduced synthesis of endogenous EGFR protein synthesis. Constitutive expression of EGFR abrogated WT1-mediated cell death. We conclude that wild-type WT1 can induce apoptosis in embryonal cancer cells, presumably through the withdrawal of required growth factor survival signals, and that EGFR is a physiological target gene for WT1.

Alternative Splicing↗

PubMed1995

Truncated WT1 mutants alter the subnuclear localization of the wild-type protein.

WT1 encodes a zinc-finger protein, expressed as distinct isoforms, that is inactivated in a subset of Wilms tumors. Both constitutional and somatic mutations disrupting the DNA-binding domain of WT1 result in a potentially dominant-negative phenotype. In generating inducible cell lines expressing wild-type isoforms of WT1 and WT1 mutants, we observed dramatic differences in the subnuclear localization of the induced proteins. The WT1 isoform that binds with high affinity to a defined DNA target, WT1(-KTS), was diffusely localized throughout the nucleus. In contrast, expression of an alternative splicing variant with reduced DNA binding affinity, WT1 (+KTS), or WT1 mutants with a disrupted zinc-finger domain resulted in a speckled pattern of expression within the nucleus. Although similar in appearance, the localization of WT1 variants to subnuclear clusters was clearly distinct from that of the essential splicing factor SC35, suggesting that WT1 is not directly involved in pre-mRNA splicing. Localization to subnuclear clusters required the N terminus of WT1, and coexpression of a truncated WT1 mutant and wild-type WT1(-KTS) resulted in their physical association, the redistribution of WT1(-KTS) from a diffuse to a speckled pattern, and the inhibition of its transactivational activity. These observations suggest that different WT1 isoforms and WT1 mutants have distinct subnuclear compartments. Dominant-negative WT1 proteins physically associate with wild-type WT1 in vivo and may result in its sequestration within subnuclear structures.

Binding Sites↗

PubMed1995

WT1 suppresses synthesis of the epidermal growth factor receptor and induces apoptosis.

The Wilms tumor suppressor gene WT1 encodes a developmentally regulated transcription factor that is mutated in a subset of embryonal tumors. To test its functional properties, we developed osteosarcoma cell lines expressing WT1 under an inducible tetracycline-regulated promoter. Induction of WT1 resulted in programmed cell death. This effect, which was differentially mediated by the alternative splicing variants of WT1, was independent of p53. WT1-mediated apoptosis was associated with reduced synthesis of the epidermal growth factor receptor (EGFR), but not of other postulated WT1-target genes, and it was abrogated by constitutive expression of EGFR. WT1 repressed transcription from the EGFR promoter, binding to two TC-rich repeat sequences. In the developing kidney, EGFR expression in renal precursor cells declined with the onset of WT1 expression. Repression of EGFR and induction of apoptosis by mechanism that may contribute to its critical role in normal kidney development and to the immortalization of tumor cells with inactivated WT1 alleles.

Alternative Splicing↗