Maty Tzukerman - Academia.edu (original) (raw)
Papers by Maty Tzukerman
Molecular Biology of the Cell, Dec 1, 2000
Three different cell differentiation experimental model systems (human embryonic stem cells, mous... more Three different cell differentiation experimental model systems (human embryonic stem cells, mouse F9 cells, and human HL-60 promyelocytic cells) were used to determine the relationship between the reduction in telomerase activity after differentiation and the regulation of the promoter for the hTERT gene. Promoter constructs of three different lengths were subcloned into the PGL3-basic luciferase reporter vector. In all three experimental systems, all three promoter constructs drove high levels of reporter activity in the nondifferentiated state, with a marked and time-dependent reduction after the induction of differentiation. In all cases, the smallest core promoter construct (283 nt upstream of the ATG) gave the highest activity. Electrophoretic mobility shift assays revealed transcription factor binding to two E-box domains within the core promoter. There was also a marked time-dependent reduction in this binding with differentiation. In addition, a distinct and novel element was identified within the core promoter, which also underwent time-dependent reduction in transcription factor binding with differentiation. Sitedirected mutagenesis of this novel element revealed a correlation between transcription factor binding and promoter activity. Taken together, the results indicate that regulation of overall telomerase activity with differentiation is mediated at least in part at the level of the TERT promoter and provides new information regarding details of the regulatory interactions that are involved in this process.
Journal of clinical & experimental pathology, Aug 31, 2013
Humana Press eBooks, Sep 7, 2006
There is no available experimental system wherein human cancer cells can be grown in the context ... more There is no available experimental system wherein human cancer cells can be grown in the context of a mixed population of normal differentiated human cells for testing biological aspects of cancer cell growth (tumor cell invasion, angiogenesis) or response to anti-cancer therapies. Human embryonic stem cells when implanted into immunocompromised mice develop teratomas containing complex structures, comprising differentiated cell types representing the major germline-derived lineages. We sought to determine whether human cancer cells would grow within such teratomas and display properties associated with malignancy such as invasiveness and recruitment of blood vessels. Ovarian cancer cells (HEY), stably expressing an H2A-GFP fusion protein, which allows tracking of tumor cells, were injected into mature teratomas and developed into tumors. The growth, proliferation capacity, invasion, and induction of blood vessel formation were examined. We propose using the novel experimental platform we have described, consisting of human tumor cells growing within a human cellular microenvironment derived from human embryonic stem cells, to develop a preclinical model for investigating and manipulating the stromal response in tumor cell growth, as an additional tool in cancer research.
PubMed, Jul 15, 2001
The telomerase RNA-protein complex responsible for maintenance of telomeric DNA at chromosome end... more The telomerase RNA-protein complex responsible for maintenance of telomeric DNA at chromosome ends, is usually inactive in most primary somatic human cells, but is specifically activated with in vitro immortalization and during tumorigenesis. Although expression of the RNA component of telomerase appears to be constitutive, the expression pattern of human telomerase reverse transcriptase (hTERT), the catalytic subunit of telomerase, is correlated with measured enzyme activity. In particular, a >80% concordance has been reported between telomerase activity and hTERT mRNA expression in ovarian tumors. Accordingly, to learn more about the mechanism regulating hTERT gene expression in ovarian carcinoma, we have performed a detailed analysis of the 5'-flanking promoter region of the hTERT gene. We have reported previously the isolation and analysis of a 5.8-kb genomic fragment containing the human hTERT gene promoter (M. Tzukerman et al., Mol. Biol. Cell, 11: 4381-4391, 2000). Deletion analysis of this promoter was carried out using transient transfection of promoter-reporter constructs in four different telomerase-expressing, ovarian carcinoma-derived cell lines, the tumorigenic properties of which have been characterized, and was compared with telomerase-negative primary human fibroblasts and nontransformed ovarian epithelial cells. These assays have shown that the hTERT promoter is inactive in telomerase-negative cells and is active in telomerase-positive cell lines. A core promoter of 283 bp upstream of the transcription initiation site (TI) was found to be sufficient for maximum promoter activity, suggesting the presence of inhibitory elements within the larger promoter sequence. Gel shift analysis of the core promoter using nuclear extracts from the ovarian and control cell lines revealed specific transcription factor binding using extracts from telomerase-positive cells. Among the binding elements, we identified two E-boxes (CACGTG) as well as a novel element (MT-box), which we identified recently in a number of differentiation systems. Site-directed mutagenesis was used to introduce mutations into this novel transcription factor binding element. These mutations significantly affect the transcriptional activity of hTERT promoter in a cell type-specific manner and suggest that the transcription factors that bind to the E-box and the novel element cooperatively function as major determinants of hTERT expression and telomerase activity in ovarian cancer. Further comparison of promoter activity, telomerase activity, and telomere length among the different ovarian cancer cells indicated that a threshold level of telomerase activity is apparently sufficient to protect telomere integrity and permit the immortal state of the different ovarian cancer cell lines.
PDF File - 11199K, Co-localization of TVM expression with hCD31 expression.
Supplementary Table S2 from Niche-Dependent Tumorigenic Capacity of Malignant Ovarian Ascites-Der... more Supplementary Table S2 from Niche-Dependent Tumorigenic Capacity of Malignant Ovarian Ascites-Derived Cancer Cell Subpopulations
PDF File - 9042K, TVM antibodies do not cross-react with murine tumor vessels.
There is currently no experimental system available wherein human cancer cells can be grown in th... more There is currently no experimental system available wherein human cancer cells can be grown in the context of a mixed population of normal differentiated human cells for testing biological aspects of cancer cell growth (tumor cell invasion, angiogenesis), or response to anticancer therapies. Human embryonic stem cells when implanted into immunocompromised mice develop teratomas containing complex structures, comprising differentiated cell types representing the major germ-line derived lineages. We sought to determine whether human cancer cells would grow within such teratomas and display properties associated with malignancy, such as invasiveness and recruitment of blood vessels. Ovarian cancer cells (HEY), stably expressing an H2A-GFP fusion protein, which allows tracking of tumor cells, were injected into mature teratomas and developed into tumors. The growth, proliferative capacity, invasion, and induction of blood vessel formation were examined. We propose using this novel experimental platform to develop a preclinical model for investigating and manipulating the stromal response in tumor cell growth as an additional tool in cancer research. Keywords: Angiogenesis; Human embryonic stem cells; Immunodeficient SCID/beige mouse; Teratoma; Tumorigenesis; Tumor microenvironment; Vasculogenesis
Journal of Biological Chemistry, Apr 1, 2010
Sirt1, a NAD-dependent protein deacetylase, is reported to regulate intracellular metabolism and ... more Sirt1, a NAD-dependent protein deacetylase, is reported to regulate intracellular metabolism and attenuate reactive oxidative species (ROS)-induced apoptosis leading to longevity and acute stress resistance. We created transgenic (TG) mice with kidney-specific overexpression of Sirt1 using the promoter sodium-phosphate cotransporter IIa (Npt2) driven specifically in proximal tubules and investigated the kidney-specific role of Sirt1 in the protection against acute kidney injury (AKI). We also elucidated the role of number or function of peroxisome and mitochondria in mediating the mechanisms for renal protective effects of Sirt1 in AKI. Cisplatin-induced AKI decreased the number and function of peroxisomes as well as mitochondria and led to increased local levels of ROS production and renal tubular apoptotic cells. TG mice treated with cisplatin mitigated AKI, local ROS, and renal tubular apoptotic tubular cells. Consistent with these results, TG mice treated with cisplatin also exhibited recovery of peroxisome number and function, as well as rescued mitochondrial function; however, mitochondrial number was not recovered. Immunoelectron microscopic findings consistently demonstrated that the decrease in peroxisome number by cisplatin in wild type mice was restored in transgenic mice. In HK-2 cells, a cultured proximal tubule cell line, overexpression of Sirt1 rescued the cisplatin-induced cell apoptosis through the restoration of peroxisome number, although the mitochondria number was not restored. These results indicate that Sirt1 overexpression in proximal tubules rescues cisplatin-induced AKI by maintaining peroxisomes number and function, concomitant up-regulation of catalase, and elimination of renal ROS levels. Renal Sirt1 can be a potential therapeutic target for the treatment of AKI.
Molecular Endocrinology, Mar 1, 1991
The identification of hormone response elements in the promoter regions of hormonally regulated g... more The identification of hormone response elements in the promoter regions of hormonally regulated genes has revealed a striking similarity between the estrogen response element (ERE) and a palindromic thyroid hormone response element (TRE) derived from the GH gene promoter. In addition, this TRE was described as a strong retinoic acid receptor response element for all three subtypes: a, 0, and 7. We show here that the TRE in the absence of thyroid hormone receptor (TR) behaves similarly to imperfect EREs, which can synergize to mediate a strong estrogen-dependent activation of transcription. However, in the presence of TR, but the absence of T 3 , activation of the TRE constructs by estrogen receptor (ER) is inhibited. In vitro, ER and TR were found to bind to the TRE in the absence and presence of their respective ligands; however, TRs form a more stable complex with the TRE than does ER. To examine whether repression of ER activity on the TRE constructs by TR was due to heterodimer formation, we employed truncated TR mutants (tTR) that lacked the DNA-binding domain, but contained the ligand-binding/dimerization domain. The tTRs were shown to be efficient inhibitors of TR, but not of ER. Thus, inhibition of ER activity on TREs by TRs does not result from heterodimer formation. We discuss a mechanism in which TRs, in the absence of thyroid hormone, control TRE activation by related receptors by preventing their access to the TRE. This mechanism can greatly enhance the fidelity of the ligand-specific response from a TRE.
Methods in Enzymology, 1990
ABSTRACT
PubMed, Dec 1, 1991
Retinoids such as retinoic acid (RA) are potent anti-arthritic and anti-neoplastic agents. We inv... more Retinoids such as retinoic acid (RA) are potent anti-arthritic and anti-neoplastic agents. We investigated the mechanism by which RA inhibits induction of collagenase gene transcription by inflammatory mediators, tumor promoters, and proto-oncogenes. We found that the RA receptors (RARs) are potent inhibitors of AP-1 activity generated either by cJun homodimers or cJun/cFos heterodimers. In addition, both cJun and cFos can inhibit RAR activity. In vitro experiments suggested that this inhibition is due to an interaction between RAR and AP-1 proteins that results in mutual loss of DNA-binding activity. The RARs need not bind to the AP-1 site, neither does AP-1 bind to RA response elements. An understanding of this antagonism between the RAR and AP-1 might help to elucidate the anti-neoplastic and anti-arthritic effects of RA as well as its effects on cell differentiation and proliferation.
PubMed, Dec 1, 2003
Extract: There has been a growing awarenes in recent years that tumorigenesis properties are mark... more Extract: There has been a growing awarenes in recent years that tumorigenesis properties are markedly affected by the surrounding tissue microenvironment at both primary and metastatic sites. Thus, it has been shown that tumor progression is associated with extensive remodeling of adjacent tissues to provide a supportive microenvironment for cancer cell proliferation, migration, invasion, and the formation of blood vessels required for supporting cancer growth. As an example, human prostate carcinoma-associated fibroblasts can promote tumorigenic transformation in initiated human prostate epithelial cells. Appreciation of the importance of the stromal response has led to the development of novel anti-cancer therapeutic agents targeted to frustrate stromal response factors, which support progressive tumor growth. Targets for investigation have included proteases, heparanase and other enzymes expressed by cancer cells or by adjacent stromal cells, which degrade extracellular matrix components and facilitate the release of cytokines and growth factors which stimulate angiogenesis, or support the growth and invasion of cancer cells. A great deal of attention has been directed at the development of anti-angiogenic molecules in particular, some of which have reached clinical trials.
PubMed, Jun 1, 2002
The recent characterization of MHC-displayed tumor-associated antigensthat recognize effector cel... more The recent characterization of MHC-displayed tumor-associated antigensthat recognize effector cells of the immune system has created new perspectives for cancer therapy. Antibodies that recognize these tumor-associated MHC-peptide complexes with the same specificity as the T-cell antigen receptor will therefore be valuable tools for immunotherapy as well as for studying antigen presentation in human cancers. Most tumor-associated antigens are expressed in only one or a few tumor types; however, recently specific T-cell epitopes derived from the telomerase catalytic subunit (hTERT) that are widely expressed in many cancers were identified and shown to be recognized by CTLs derived from cancer patients. We selected a large nonimmune repertoire of phage Fab antibodies on recombinant human class I HLA-A2 complexes displaying two distinct antigenic T-cell epitopes derived from hTERT. We isolated a surprisingly large panel of high-affinity human recombinant Fab antibodies that exhibited peptide-specific, MHC-restricted binding characteristics of T cells. The analyzed Fabs not only recognize the cognate MHC-peptide complex in a recombinant soluble form but also the native complex as displayed on the surface of antigen-presenting cells and hTERT-expressing tumor cells. These findings demonstrate for the first time the ability to transform the unique fine specificity but low intrinsic affinity of TCRs on T cells into high-affinity soluble antibody molecules endowed with a T-cell antigen receptor-like specificity. These molecules may prove to be very important and widely applicable for monitoring the expression of specific MHC-peptide complexes on the surface of tumor and immune cells, for structure-function studies of TCR-peptide-MHC interactions, as well as for developing new targeting agents for immunotherapy.
Solid tumor complexity emanates from the requirement of a supportive microenvironment that provid... more Solid tumor complexity emanates from the requirement of a supportive microenvironment that provides a compatible network of interactions between heterogeneous cancer cells and various tumor supporting cells. In previous studies we have established a model for studying tumorigenic processes where cancer cells can directly interact with a human microenvironment based on the potential of human embryonic stem cells (hESC) to generate in vivo teratoma tissue. We utilized the hESC - derived experimental platform to advantage as a supportive tumor microenvironment to demonstrate that the balance between self-renewal and tumorigenic differentiation differs strikingly among cancer cell subpopulations (CCSPs), and importantly is dynamically dependent on the tumor microenvironment as a crucial determinant of tumor growth properties. Six clonally expanded CCSPs derived from the ovarian clear cell carcinoma of a single tumor were used to demonstrate striking intratumoral heterogeneity that is also affected by the tumor microenvironment. While only four out of the six CCSPs developed into tumors in a conventional xenograft model, all of the CCSPs robustly generated tumors in the hESC-derived cellular microenvironment, highlighting the potential experimental utility of the hESC-based model in supporting tumor promotion. Moreover, each of the six CCSPs displays a different level of morphologic and tumorigenic differentiation, wherein growth in the hESC-derived microenvironment favors growth of CD44+/ALDH+ pockets of self-renewing cells that sustain tumor growth through a process of tumorigenic differentiation into CD44-/ALDH- derivatives. These derivative cells display microenvironment-dependent plasticity with the capacity to restore self-renewal and CD44 expression. Gene expression profiles of two specific CCSPs grown in vitro and then injected either as intramuscular or as intrateratoma tumors were examined. This gene expression array analysis revealed that the hESC-based model supports the capacity of distinct CCSPs to generate tumors, and maintains CSC properties, in contrast to the conventional direct tumor xenograft platform. Significant gene ontologies correlates for each group of tumors showed greater complexity in the interaction between the tumor cells and the hESC-derived cellular microenvironment as opposed to the xenotransplantation model. In addition, these results demonstrated that the murine microenvironment is incapable to discriminate between different cancer cell populations of a single tumor and therefore excessively simplify tumor progression events. Taken together, the hESC-based in vivo model renders intratumoral heterogeneity in the self-renewal and tumorigenic differentiation amenable to biological analysis as well as anticancer therapy testing. Citation Format: Sagi Abelson, Yeela Shamai, Liron Berger, Irena Reiter, Roni Shouval, Karl Skorecki, Maty Tzukerman. Microenvironment-dependent intratumoral heterogeneity in the self-renewal and tumorigenic differentiation of ovarian cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2641. doi:10.1158/1538-7445.AM2013-2641
Intratumoral heterogeneity challenges existing paradigms for anti-cancer therapy. We have previou... more Intratumoral heterogeneity challenges existing paradigms for anti-cancer therapy. We have previously demonstrated that the human embryonic stem cells (hESC)-derived cellular microenvironment in immunocompromised mice, enables functional distinction of heterogeneous tumor cells, including cells which do not grow into a tumor in conventional direct tumor xenograft platform. We have identified and characterized six cancer cell subpopulations each clonally expanded from a single cell, derived from human ovarian clear cell carcinoma of a single tumor, to demonstrate striking intratumoral phenotypic heterogeneity that is dynamically dependent on the tumor growth microenvironment. These cancer cell subpopulations, characterized as cancer stem cell subpopulations, faithfully recapitulate the full spectrum of histological phenotypic heterogeneity known for human ovarian clear cell carcinoma. Each of the six subpopulations displays a different level of morphologic and tumorigenic differentiation wherein growth in the hESC-derived microenvironment favors growth of CD44+/aldehyde dehydrogenase positive pockets of self-renewing cells that sustain tumor growth through a process of tumorigenic differentiation into CD44-/aldehyde dehydrogenase negative derivatives. Strikingly, these derivative cells display microenvironment-dependent plasticity with the capacity to restore self-renewal markers and CD44 expression. We delineated the distinct gene expression and epigenetic profiles of two such subpopulations, representing extremes of phenotypic heterogeneity in terms of niche-dependent self-renewal and tumorigenic differentiation. By combining Gene Set Enrichment, Gene Ontology and Pathway-focused array analyses with methylation status and secretome analyses, we propose a suite of robust differences in tumor self-renewal and differentiation pathways, and interaction with stromal cells in the tumor microenvironment that underlay the striking intratumoral phenotypic heterogeneity which characterized this and other solid tumor malignancies. Citation Format: Maty Tzukerman, Sagi Abelson, Yeela Shamai, Liron Berger, Karl Skorecki. Intratumoral heterogeneity of ovarian cancer stem cells and their interactions with the tumor microenvironment. [abstract]. In: Abstracts: AACR Special Conference on Cellular Heterogeneity in the Tumor Microenvironment; 2014 Feb 26-Mar 1; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(1 Suppl):Abstract nr B43. doi:10.1158/1538-7445.CHTME14-B43
Cell regulation, Jul 1, 1991
Gene regulation by thyroid hormones is mediated through multiple nuclear receptors. Only some of ... more Gene regulation by thyroid hormones is mediated through multiple nuclear receptors. Only some of these thyroid hormone receptor (TR) isoforms become transcriptional enhancers in the presence of the thyroid hormone T3. Here we analyze the regulatory function of the human TRa2 isoform. This protein does not bind T3 and is not a transcriptional activator of thyroid hormone-responsive elements (TRE). Transfected TRa2 functions as a constitutive repressor of the transcriptional activators TRal and TR,81 but also represses heterologous receptors, including the retinoic acid receptor and the estrogen receptor, which can activate TRE-controlled genes. TRa2 protein showed strongly reduced DNA binding to a palindromic TRE when compared with the active TRs. Hybrid receptor analysis revealed that the special properties of the TRa2 protein, including its repressor function and DNA binding characteristics, are intrinsic properties of its carboxyterminus and can be transferred to other receptors. Although it has been shown that the active TRs can act as repressors and silencers due to their strong DNA binding in the absence of hormone, our data show that TRa2 is unlikely to inhibit TRs and other receptors through a competitive DNA binding mechanism. Antibody gel shift experiments suggest that repression by TRa2 might result from interaction with active receptors. Thus, the receptorlike TRa2 isoform differs from typical nuclear receptors in its DNA-binding and ligand-binding properties and appears to regulate the activity of other receptors via protein-protein interaction.
Blood, Jul 19, 2012
leukemia cases, > 1 lineage was observed at relapse, indicating that diverse mechanisms can promo... more leukemia cases, > 1 lineage was observed at relapse, indicating that diverse mechanisms can promote relapse in the same patient. In conclusion, diverse relapse mechanisms can be observed by systematic reconstruction of cell lineage trees of patients with leukemia. (Blood.
Molecular Biology of the Cell, Dec 1, 2000
Three different cell differentiation experimental model systems (human embryonic stem cells, mous... more Three different cell differentiation experimental model systems (human embryonic stem cells, mouse F9 cells, and human HL-60 promyelocytic cells) were used to determine the relationship between the reduction in telomerase activity after differentiation and the regulation of the promoter for the hTERT gene. Promoter constructs of three different lengths were subcloned into the PGL3-basic luciferase reporter vector. In all three experimental systems, all three promoter constructs drove high levels of reporter activity in the nondifferentiated state, with a marked and time-dependent reduction after the induction of differentiation. In all cases, the smallest core promoter construct (283 nt upstream of the ATG) gave the highest activity. Electrophoretic mobility shift assays revealed transcription factor binding to two E-box domains within the core promoter. There was also a marked time-dependent reduction in this binding with differentiation. In addition, a distinct and novel element was identified within the core promoter, which also underwent time-dependent reduction in transcription factor binding with differentiation. Sitedirected mutagenesis of this novel element revealed a correlation between transcription factor binding and promoter activity. Taken together, the results indicate that regulation of overall telomerase activity with differentiation is mediated at least in part at the level of the TERT promoter and provides new information regarding details of the regulatory interactions that are involved in this process.
Journal of clinical & experimental pathology, Aug 31, 2013
Humana Press eBooks, Sep 7, 2006
There is no available experimental system wherein human cancer cells can be grown in the context ... more There is no available experimental system wherein human cancer cells can be grown in the context of a mixed population of normal differentiated human cells for testing biological aspects of cancer cell growth (tumor cell invasion, angiogenesis) or response to anti-cancer therapies. Human embryonic stem cells when implanted into immunocompromised mice develop teratomas containing complex structures, comprising differentiated cell types representing the major germline-derived lineages. We sought to determine whether human cancer cells would grow within such teratomas and display properties associated with malignancy such as invasiveness and recruitment of blood vessels. Ovarian cancer cells (HEY), stably expressing an H2A-GFP fusion protein, which allows tracking of tumor cells, were injected into mature teratomas and developed into tumors. The growth, proliferation capacity, invasion, and induction of blood vessel formation were examined. We propose using the novel experimental platform we have described, consisting of human tumor cells growing within a human cellular microenvironment derived from human embryonic stem cells, to develop a preclinical model for investigating and manipulating the stromal response in tumor cell growth, as an additional tool in cancer research.
PubMed, Jul 15, 2001
The telomerase RNA-protein complex responsible for maintenance of telomeric DNA at chromosome end... more The telomerase RNA-protein complex responsible for maintenance of telomeric DNA at chromosome ends, is usually inactive in most primary somatic human cells, but is specifically activated with in vitro immortalization and during tumorigenesis. Although expression of the RNA component of telomerase appears to be constitutive, the expression pattern of human telomerase reverse transcriptase (hTERT), the catalytic subunit of telomerase, is correlated with measured enzyme activity. In particular, a >80% concordance has been reported between telomerase activity and hTERT mRNA expression in ovarian tumors. Accordingly, to learn more about the mechanism regulating hTERT gene expression in ovarian carcinoma, we have performed a detailed analysis of the 5'-flanking promoter region of the hTERT gene. We have reported previously the isolation and analysis of a 5.8-kb genomic fragment containing the human hTERT gene promoter (M. Tzukerman et al., Mol. Biol. Cell, 11: 4381-4391, 2000). Deletion analysis of this promoter was carried out using transient transfection of promoter-reporter constructs in four different telomerase-expressing, ovarian carcinoma-derived cell lines, the tumorigenic properties of which have been characterized, and was compared with telomerase-negative primary human fibroblasts and nontransformed ovarian epithelial cells. These assays have shown that the hTERT promoter is inactive in telomerase-negative cells and is active in telomerase-positive cell lines. A core promoter of 283 bp upstream of the transcription initiation site (TI) was found to be sufficient for maximum promoter activity, suggesting the presence of inhibitory elements within the larger promoter sequence. Gel shift analysis of the core promoter using nuclear extracts from the ovarian and control cell lines revealed specific transcription factor binding using extracts from telomerase-positive cells. Among the binding elements, we identified two E-boxes (CACGTG) as well as a novel element (MT-box), which we identified recently in a number of differentiation systems. Site-directed mutagenesis was used to introduce mutations into this novel transcription factor binding element. These mutations significantly affect the transcriptional activity of hTERT promoter in a cell type-specific manner and suggest that the transcription factors that bind to the E-box and the novel element cooperatively function as major determinants of hTERT expression and telomerase activity in ovarian cancer. Further comparison of promoter activity, telomerase activity, and telomere length among the different ovarian cancer cells indicated that a threshold level of telomerase activity is apparently sufficient to protect telomere integrity and permit the immortal state of the different ovarian cancer cell lines.
PDF File - 11199K, Co-localization of TVM expression with hCD31 expression.
Supplementary Table S2 from Niche-Dependent Tumorigenic Capacity of Malignant Ovarian Ascites-Der... more Supplementary Table S2 from Niche-Dependent Tumorigenic Capacity of Malignant Ovarian Ascites-Derived Cancer Cell Subpopulations
PDF File - 9042K, TVM antibodies do not cross-react with murine tumor vessels.
There is currently no experimental system available wherein human cancer cells can be grown in th... more There is currently no experimental system available wherein human cancer cells can be grown in the context of a mixed population of normal differentiated human cells for testing biological aspects of cancer cell growth (tumor cell invasion, angiogenesis), or response to anticancer therapies. Human embryonic stem cells when implanted into immunocompromised mice develop teratomas containing complex structures, comprising differentiated cell types representing the major germ-line derived lineages. We sought to determine whether human cancer cells would grow within such teratomas and display properties associated with malignancy, such as invasiveness and recruitment of blood vessels. Ovarian cancer cells (HEY), stably expressing an H2A-GFP fusion protein, which allows tracking of tumor cells, were injected into mature teratomas and developed into tumors. The growth, proliferative capacity, invasion, and induction of blood vessel formation were examined. We propose using this novel experimental platform to develop a preclinical model for investigating and manipulating the stromal response in tumor cell growth as an additional tool in cancer research. Keywords: Angiogenesis; Human embryonic stem cells; Immunodeficient SCID/beige mouse; Teratoma; Tumorigenesis; Tumor microenvironment; Vasculogenesis
Journal of Biological Chemistry, Apr 1, 2010
Sirt1, a NAD-dependent protein deacetylase, is reported to regulate intracellular metabolism and ... more Sirt1, a NAD-dependent protein deacetylase, is reported to regulate intracellular metabolism and attenuate reactive oxidative species (ROS)-induced apoptosis leading to longevity and acute stress resistance. We created transgenic (TG) mice with kidney-specific overexpression of Sirt1 using the promoter sodium-phosphate cotransporter IIa (Npt2) driven specifically in proximal tubules and investigated the kidney-specific role of Sirt1 in the protection against acute kidney injury (AKI). We also elucidated the role of number or function of peroxisome and mitochondria in mediating the mechanisms for renal protective effects of Sirt1 in AKI. Cisplatin-induced AKI decreased the number and function of peroxisomes as well as mitochondria and led to increased local levels of ROS production and renal tubular apoptotic cells. TG mice treated with cisplatin mitigated AKI, local ROS, and renal tubular apoptotic tubular cells. Consistent with these results, TG mice treated with cisplatin also exhibited recovery of peroxisome number and function, as well as rescued mitochondrial function; however, mitochondrial number was not recovered. Immunoelectron microscopic findings consistently demonstrated that the decrease in peroxisome number by cisplatin in wild type mice was restored in transgenic mice. In HK-2 cells, a cultured proximal tubule cell line, overexpression of Sirt1 rescued the cisplatin-induced cell apoptosis through the restoration of peroxisome number, although the mitochondria number was not restored. These results indicate that Sirt1 overexpression in proximal tubules rescues cisplatin-induced AKI by maintaining peroxisomes number and function, concomitant up-regulation of catalase, and elimination of renal ROS levels. Renal Sirt1 can be a potential therapeutic target for the treatment of AKI.
Molecular Endocrinology, Mar 1, 1991
The identification of hormone response elements in the promoter regions of hormonally regulated g... more The identification of hormone response elements in the promoter regions of hormonally regulated genes has revealed a striking similarity between the estrogen response element (ERE) and a palindromic thyroid hormone response element (TRE) derived from the GH gene promoter. In addition, this TRE was described as a strong retinoic acid receptor response element for all three subtypes: a, 0, and 7. We show here that the TRE in the absence of thyroid hormone receptor (TR) behaves similarly to imperfect EREs, which can synergize to mediate a strong estrogen-dependent activation of transcription. However, in the presence of TR, but the absence of T 3 , activation of the TRE constructs by estrogen receptor (ER) is inhibited. In vitro, ER and TR were found to bind to the TRE in the absence and presence of their respective ligands; however, TRs form a more stable complex with the TRE than does ER. To examine whether repression of ER activity on the TRE constructs by TR was due to heterodimer formation, we employed truncated TR mutants (tTR) that lacked the DNA-binding domain, but contained the ligand-binding/dimerization domain. The tTRs were shown to be efficient inhibitors of TR, but not of ER. Thus, inhibition of ER activity on TREs by TRs does not result from heterodimer formation. We discuss a mechanism in which TRs, in the absence of thyroid hormone, control TRE activation by related receptors by preventing their access to the TRE. This mechanism can greatly enhance the fidelity of the ligand-specific response from a TRE.
Methods in Enzymology, 1990
ABSTRACT
PubMed, Dec 1, 1991
Retinoids such as retinoic acid (RA) are potent anti-arthritic and anti-neoplastic agents. We inv... more Retinoids such as retinoic acid (RA) are potent anti-arthritic and anti-neoplastic agents. We investigated the mechanism by which RA inhibits induction of collagenase gene transcription by inflammatory mediators, tumor promoters, and proto-oncogenes. We found that the RA receptors (RARs) are potent inhibitors of AP-1 activity generated either by cJun homodimers or cJun/cFos heterodimers. In addition, both cJun and cFos can inhibit RAR activity. In vitro experiments suggested that this inhibition is due to an interaction between RAR and AP-1 proteins that results in mutual loss of DNA-binding activity. The RARs need not bind to the AP-1 site, neither does AP-1 bind to RA response elements. An understanding of this antagonism between the RAR and AP-1 might help to elucidate the anti-neoplastic and anti-arthritic effects of RA as well as its effects on cell differentiation and proliferation.
PubMed, Dec 1, 2003
Extract: There has been a growing awarenes in recent years that tumorigenesis properties are mark... more Extract: There has been a growing awarenes in recent years that tumorigenesis properties are markedly affected by the surrounding tissue microenvironment at both primary and metastatic sites. Thus, it has been shown that tumor progression is associated with extensive remodeling of adjacent tissues to provide a supportive microenvironment for cancer cell proliferation, migration, invasion, and the formation of blood vessels required for supporting cancer growth. As an example, human prostate carcinoma-associated fibroblasts can promote tumorigenic transformation in initiated human prostate epithelial cells. Appreciation of the importance of the stromal response has led to the development of novel anti-cancer therapeutic agents targeted to frustrate stromal response factors, which support progressive tumor growth. Targets for investigation have included proteases, heparanase and other enzymes expressed by cancer cells or by adjacent stromal cells, which degrade extracellular matrix components and facilitate the release of cytokines and growth factors which stimulate angiogenesis, or support the growth and invasion of cancer cells. A great deal of attention has been directed at the development of anti-angiogenic molecules in particular, some of which have reached clinical trials.
PubMed, Jun 1, 2002
The recent characterization of MHC-displayed tumor-associated antigensthat recognize effector cel... more The recent characterization of MHC-displayed tumor-associated antigensthat recognize effector cells of the immune system has created new perspectives for cancer therapy. Antibodies that recognize these tumor-associated MHC-peptide complexes with the same specificity as the T-cell antigen receptor will therefore be valuable tools for immunotherapy as well as for studying antigen presentation in human cancers. Most tumor-associated antigens are expressed in only one or a few tumor types; however, recently specific T-cell epitopes derived from the telomerase catalytic subunit (hTERT) that are widely expressed in many cancers were identified and shown to be recognized by CTLs derived from cancer patients. We selected a large nonimmune repertoire of phage Fab antibodies on recombinant human class I HLA-A2 complexes displaying two distinct antigenic T-cell epitopes derived from hTERT. We isolated a surprisingly large panel of high-affinity human recombinant Fab antibodies that exhibited peptide-specific, MHC-restricted binding characteristics of T cells. The analyzed Fabs not only recognize the cognate MHC-peptide complex in a recombinant soluble form but also the native complex as displayed on the surface of antigen-presenting cells and hTERT-expressing tumor cells. These findings demonstrate for the first time the ability to transform the unique fine specificity but low intrinsic affinity of TCRs on T cells into high-affinity soluble antibody molecules endowed with a T-cell antigen receptor-like specificity. These molecules may prove to be very important and widely applicable for monitoring the expression of specific MHC-peptide complexes on the surface of tumor and immune cells, for structure-function studies of TCR-peptide-MHC interactions, as well as for developing new targeting agents for immunotherapy.
Solid tumor complexity emanates from the requirement of a supportive microenvironment that provid... more Solid tumor complexity emanates from the requirement of a supportive microenvironment that provides a compatible network of interactions between heterogeneous cancer cells and various tumor supporting cells. In previous studies we have established a model for studying tumorigenic processes where cancer cells can directly interact with a human microenvironment based on the potential of human embryonic stem cells (hESC) to generate in vivo teratoma tissue. We utilized the hESC - derived experimental platform to advantage as a supportive tumor microenvironment to demonstrate that the balance between self-renewal and tumorigenic differentiation differs strikingly among cancer cell subpopulations (CCSPs), and importantly is dynamically dependent on the tumor microenvironment as a crucial determinant of tumor growth properties. Six clonally expanded CCSPs derived from the ovarian clear cell carcinoma of a single tumor were used to demonstrate striking intratumoral heterogeneity that is also affected by the tumor microenvironment. While only four out of the six CCSPs developed into tumors in a conventional xenograft model, all of the CCSPs robustly generated tumors in the hESC-derived cellular microenvironment, highlighting the potential experimental utility of the hESC-based model in supporting tumor promotion. Moreover, each of the six CCSPs displays a different level of morphologic and tumorigenic differentiation, wherein growth in the hESC-derived microenvironment favors growth of CD44+/ALDH+ pockets of self-renewing cells that sustain tumor growth through a process of tumorigenic differentiation into CD44-/ALDH- derivatives. These derivative cells display microenvironment-dependent plasticity with the capacity to restore self-renewal and CD44 expression. Gene expression profiles of two specific CCSPs grown in vitro and then injected either as intramuscular or as intrateratoma tumors were examined. This gene expression array analysis revealed that the hESC-based model supports the capacity of distinct CCSPs to generate tumors, and maintains CSC properties, in contrast to the conventional direct tumor xenograft platform. Significant gene ontologies correlates for each group of tumors showed greater complexity in the interaction between the tumor cells and the hESC-derived cellular microenvironment as opposed to the xenotransplantation model. In addition, these results demonstrated that the murine microenvironment is incapable to discriminate between different cancer cell populations of a single tumor and therefore excessively simplify tumor progression events. Taken together, the hESC-based in vivo model renders intratumoral heterogeneity in the self-renewal and tumorigenic differentiation amenable to biological analysis as well as anticancer therapy testing. Citation Format: Sagi Abelson, Yeela Shamai, Liron Berger, Irena Reiter, Roni Shouval, Karl Skorecki, Maty Tzukerman. Microenvironment-dependent intratumoral heterogeneity in the self-renewal and tumorigenic differentiation of ovarian cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2641. doi:10.1158/1538-7445.AM2013-2641
Intratumoral heterogeneity challenges existing paradigms for anti-cancer therapy. We have previou... more Intratumoral heterogeneity challenges existing paradigms for anti-cancer therapy. We have previously demonstrated that the human embryonic stem cells (hESC)-derived cellular microenvironment in immunocompromised mice, enables functional distinction of heterogeneous tumor cells, including cells which do not grow into a tumor in conventional direct tumor xenograft platform. We have identified and characterized six cancer cell subpopulations each clonally expanded from a single cell, derived from human ovarian clear cell carcinoma of a single tumor, to demonstrate striking intratumoral phenotypic heterogeneity that is dynamically dependent on the tumor growth microenvironment. These cancer cell subpopulations, characterized as cancer stem cell subpopulations, faithfully recapitulate the full spectrum of histological phenotypic heterogeneity known for human ovarian clear cell carcinoma. Each of the six subpopulations displays a different level of morphologic and tumorigenic differentiation wherein growth in the hESC-derived microenvironment favors growth of CD44+/aldehyde dehydrogenase positive pockets of self-renewing cells that sustain tumor growth through a process of tumorigenic differentiation into CD44-/aldehyde dehydrogenase negative derivatives. Strikingly, these derivative cells display microenvironment-dependent plasticity with the capacity to restore self-renewal markers and CD44 expression. We delineated the distinct gene expression and epigenetic profiles of two such subpopulations, representing extremes of phenotypic heterogeneity in terms of niche-dependent self-renewal and tumorigenic differentiation. By combining Gene Set Enrichment, Gene Ontology and Pathway-focused array analyses with methylation status and secretome analyses, we propose a suite of robust differences in tumor self-renewal and differentiation pathways, and interaction with stromal cells in the tumor microenvironment that underlay the striking intratumoral phenotypic heterogeneity which characterized this and other solid tumor malignancies. Citation Format: Maty Tzukerman, Sagi Abelson, Yeela Shamai, Liron Berger, Karl Skorecki. Intratumoral heterogeneity of ovarian cancer stem cells and their interactions with the tumor microenvironment. [abstract]. In: Abstracts: AACR Special Conference on Cellular Heterogeneity in the Tumor Microenvironment; 2014 Feb 26-Mar 1; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(1 Suppl):Abstract nr B43. doi:10.1158/1538-7445.CHTME14-B43
Cell regulation, Jul 1, 1991
Gene regulation by thyroid hormones is mediated through multiple nuclear receptors. Only some of ... more Gene regulation by thyroid hormones is mediated through multiple nuclear receptors. Only some of these thyroid hormone receptor (TR) isoforms become transcriptional enhancers in the presence of the thyroid hormone T3. Here we analyze the regulatory function of the human TRa2 isoform. This protein does not bind T3 and is not a transcriptional activator of thyroid hormone-responsive elements (TRE). Transfected TRa2 functions as a constitutive repressor of the transcriptional activators TRal and TR,81 but also represses heterologous receptors, including the retinoic acid receptor and the estrogen receptor, which can activate TRE-controlled genes. TRa2 protein showed strongly reduced DNA binding to a palindromic TRE when compared with the active TRs. Hybrid receptor analysis revealed that the special properties of the TRa2 protein, including its repressor function and DNA binding characteristics, are intrinsic properties of its carboxyterminus and can be transferred to other receptors. Although it has been shown that the active TRs can act as repressors and silencers due to their strong DNA binding in the absence of hormone, our data show that TRa2 is unlikely to inhibit TRs and other receptors through a competitive DNA binding mechanism. Antibody gel shift experiments suggest that repression by TRa2 might result from interaction with active receptors. Thus, the receptorlike TRa2 isoform differs from typical nuclear receptors in its DNA-binding and ligand-binding properties and appears to regulate the activity of other receptors via protein-protein interaction.
Blood, Jul 19, 2012
leukemia cases, > 1 lineage was observed at relapse, indicating that diverse mechanisms can promo... more leukemia cases, > 1 lineage was observed at relapse, indicating that diverse mechanisms can promote relapse in the same patient. In conclusion, diverse relapse mechanisms can be observed by systematic reconstruction of cell lineage trees of patients with leukemia. (Blood.