Rajeev Samant | University of Alabama at Birmingham (original) (raw)

Papers by Rajeev Samant

International Journal of Breast Cancer, 2012

The microenvironment at the site of tumor metastasis plays a key role in determining the fate of ... more The microenvironment at the site of tumor metastasis plays a key role in determining the fate of the metastasizing tumor cells. This ultimately has a direct impact on the progression of cancer. Bone is the preferred site of metastasis of breast cancer. Painful, debilitating osteolytic lesions are formed as a result of crosstalk between breast cancer cells and cells in the bone, predominantly the osteoblasts and osteoclasts. In this paper, we have discussed the temporal and spatial role of hedgehog (Hh) signaling in influencing the fate of metastatic breast cancer cells in bone. By virtue of its secreted ligands, the Hh pathway is capable of homotypic and heterotypic signaling and consequently altering the microenvironment in the bone. We also have put into perspective the therapeutic implications of using Hh inhibitors to prevent and/or treat bone metastases of breast cancer.

Cancer Metastasis — Biology and Treatment, 2005

... 7. Ohtaki T., Shintani Y., Honda S., Matsumoto H., Hori A., Kanehashi K., Torao Y., Kumano S.... more ... 7. Ohtaki T., Shintani Y., Honda S., Matsumoto H., Hori A., Kanehashi K., Torao Y., Kumano S., Takatsu Y., Matsuda Y., Ishibashi Y ... Saunders MM, Seraj MJ, Li ZY, Zhou ZY, Winter CR, Welch DR and Donahue HJ Breast cancer metastatic potential correlates with a breakdown in ...

International Journal of Breast Cancer, 2012

PLoS ONE, 2012

The remodeling process in bone yields numerous cytokines and chemokines that mediate crosstalk be... more The remodeling process in bone yields numerous cytokines and chemokines that mediate crosstalk between osteoblasts and osteoclasts and also serve to attract and support metastatic tumor cells. The metastatic tumor cells disturb the equilibrium in bone that manifests as skeletal complications. The Hedgehog (Hh) pathway plays an important role in skeletogenesis. We hypothesized that the Hh pathway mediates an interaction between tumor cells and osteoblasts and influences osteoblast differentiation in response to tumor cells. We have determined that breast tumor cells have an activated Hh pathway characterized by upregulation of the ligand, IHH and transcription factor GLI1. Breast cancer cells interact with osteoblasts and cause an enhanced differentiation of pre-osteoblasts to osteoblasts that express increased levels of the osteoclastogenesis factors, RANKL and PTHrP. There is sustained expression of osteoclast-promoting factors, RANKL and PTHrP, even after the osteoblast differentiation ceases and apoptosis sets in. Moreover, tumor cells that are deficient in Hh signaling are compromised in their ability to induce osteoblast differentiation and consequently are inefficient in causing osteolysis. The stimulation of osteoblast differentiation sets the stage for osteoclast differentiation and overall promotes osteolysis. Thus, in the process of developing newer therapeutic strategies against breast cancer metastasis to bone it would worthwhile to keep in mind the role of the Hh pathway in osteoblast differentiation in an otherwise predominant osteolytic phenomenon.

Cancer Metastasis — Biology and Treatment, 2002

Cancer Metastasis – Biology and Treatment, 2007

Metastasis of breast cancer is a complex event involving coordinated cross-talk of several protei... more Metastasis of breast cancer is a complex event involving coordinated cross-talk of several proteins. Genes that control the resultant metastasis can be broadly classified as metastasis promoter genes (MPGs) and metastasis suppressor genes (MSGs). There is an explosion of information in the studies that focus on these genes; however, thus far, a very few of them are actually tested clinically

Utilizing gene microarray profiling of melanoma samples, we have recently identified a novel gene... more Utilizing gene microarray profiling of melanoma samples, we have recently identified a novel gene overexpressed in both thick primary and metastatic melanomas. This gene, progestagen-associated endometrial protein (PAEP), has never before been implicated in the oncogenic processes of melanoma, with its true function in oncogenesis and tumour progression relatively unknown. Overexpression of the PAEP gene in freshly procured thick primary and metastatic melanoma samples (58%) and daughter cell lines (77%) is confirmed by quantitative RT-PCR, immunohistochemistry, Western blotting and mass spectrometric analysis. We suggest that PAEP gene overexpression is involved with melanoma tumour progression as well as an aggressive phenotype. Transfection of melanoma cells with PAEP small interfering RNA (siRNA) reveals a significant decrease in soft agar colony formation and a marked inhibition of both cell migration and cell invasion. Furthermore, we establish stable melanoma transfectants via PAEP lentiviral small hairpin RNA (shRNA), examine their growth characteristics in a murine xenograft model and reveal that tumour growth is significantly inhibited in two separate melanoma cell lines. Our data strongly implicate the PAEP gene as a tumour growth promoter with oncogenic properties and a potential therapeutic target for patients with advanced melanoma.

Introduction: Breast cancer is one of the most common malignancies affecting women in the United ... more Introduction: Breast cancer is one of the most common malignancies affecting women in the United States and Europe. Approximately three out of every four women with breast cancer develop metastases in bone which, in turn, diminishes quality of life. The α v β 3 integrin has previously been implicated in multiple aspects of tumor progression, metastasis and osteoclast bone resorption. Therefore, we hypothesized that the α v β 3 -selective inhibitor, S247, would decrease the development of osteolytic breast cancer metastases. Materials and methods: Cells were treated in vitro with S247 and assessed for viability and adhesion to matrix components. Athymic mice received intracardiac (left ventricle) injections of human MDA-MB-435 breast carcinoma cells expressing enhanced green-fluorescent protein.

The exact cellular and molecular mechanisms involved in melanoma tumorigenesis remain obscure. Pr... more The exact cellular and molecular mechanisms involved in melanoma tumorigenesis remain obscure. Previous gene expression profiling analyses performed upon NHEM and human melanoma samples identified WFDC1 as one of the most frequently down-regulated genes. Here we further showed that NHEM readily express WFDC1 but expression is reduced or completely lost in 80% of the patients-derived melanoma cell lines and tissue samples examined. Furthermore, we show that promoter hypermethylation accounts for the silencing of the WFDC1 gene in 20% of the melanoma cell lines examined. The over-expression of WFDC1 in two metastatic melanoma cell lines, A375 and LOX, resulted in a significant delay of tumor growth in a murine xenograft model, despite a non-significant difference in tumor cell growth in vitro. Gene expression microarray analysis and further expression validation suggests that the Dickkopf-1 (Dkk1) gene is up-regulated in WFDC1 over-expressing cell lines, suggesting that the tumor suppressive function of WFDC1 may be partially a result of up-regulated Dkk1 gene expression, which is known to be a potent inhibitor of the Wnt signaling pathway.

Cancer letters, Jan 10, 2014

22 Available online xxxx 23 Keywords: 24 Niclosamide 25 FDA-approved drug 26 Multi-targeted thera... more 22 Available online xxxx 23 Keywords: 24 Niclosamide 25 FDA-approved drug 26 Multi-targeted therapy 27 Drug discovery 28 Cancer stem cells 29 3 0 a b s t r a c t 31 The rapid development of new anticancer drugs that are safe and effective is a common goal shared by 32 basic scientists, clinicians and patients. The current review discusses one such agent, namely niclosamide, 33 which has been used in the clinic for the treatment of intestinal parasite infections. Recent studies repeat-34 edly identified niclosamide as a potential anticancer agent by various high-throughput screening cam-35 paigns. Niclosamide not only inhibits the Wnt/b-catenin, mTORC1, STAT3, NF-jB and Notch signaling 36 pathways, but also targets mitochondria in cancer cells to induce cell cycle arrest, growth inhibition 37 and apoptosis. A number of studies have established the anticancer activities of niclosamide in both 38 in vitro and in vivo models. Moreover, the inhibitory effects of niclosamide on cancer stem cells provide 39

Molecular cancer therapeutics, 2014

Basal-like breast cancers (BLBC) are poorly differentiated and display aggressive clinical behavi... more Basal-like breast cancers (BLBC) are poorly differentiated and display aggressive clinical behavior. These tumors become resistant to cytotoxic agents, and tumor relapse has been attributed to the presence of cancer stem cells (CSC). One of the pathways involved in CSC regulation is the Wnt/b-catenin signaling pathway. LRP6, a Wnt ligand receptor, is one of the critical elements of this pathway and could potentially be an excellent therapeutic target. Niclosamide has been shown to inhibit the Wnt/b-catenin signaling pathway by causing degradation of LRP6. TRA-8, a monoclonal antibody specific to TRAIL death receptor 5, is cytotoxic to BLBC cell lines and their CSC-enriched populations. The goal of this study was to examine whether niclosamide is cytotoxic to BLBCs, specifically the CSC population, and if in combination with TRA-8 could produce increased cytotoxicity. Aldehyde dehydrogenase (ALDH) is a known marker of CSCs. By testing BLBC cells for ALDH expression by flow cytometry, we were able to isolate a nonadherent population of cells that have high ALDH expression. Niclosamide showed cytotoxicity against these nonadherent ALDH-expressing cells in addition to adherent cells from four BLBC cell lines: 2LMP, SUM159, HCC1187, and HCC1143. Niclosamide treatment produced reduced levels of LRP6 and b-catenin, which is a downstream Wnt/b-catenin signaling protein. The combination of TRA-8 and niclosamide produced additive cytotoxicity and a reduction in Wnt/b-catenin activity. Niclosamide in combination with TRA-8 suppressed growth of 2LMP orthotopic tumor xenografts. These results suggest that niclosamide or congeners of this agent may be useful for the treatment of BLBC. Mol Cancer Ther; 13(4); 800-11. Ó2014 AACR.

Cancer genomics & proteomics

have been used for the discovery of genes and pathways involved in melanoma and other types of ca... more have been used for the discovery of genes and pathways involved in melanoma and other types of cancer. However, in many cases, the results from various tumor models failed to be validated successfully in clinical studies. Limited information is available on how closely these models reflect the in vivo physiological conditions. In this study, a comprehensive genomics approach was used to systematically compare the expression patterns of snap frozen samples obtained from patients with primary melanoma, lymph node metastasis, and distant metastases, and compare these patterns to those of their corresponding cell lines and tumor xenografts in nude mice. The GE Healthcare 20k human genome array was used and the expression data was normalized and analyzed using GeneSpring 7.2 software. Based on the expression analysis, the correlation rate between the snap frozen primary patient samples vs. derived cell lines was 66%, with 1687 differentially expressed genes. The correlation rate between the snap frozen primary patient samples and the tumor xenografts was 75%, with 1,374 differentially expressed genes, and the correlation rate comparing tumor xenografts to derived cell lines ranged between 58% and 84%. These results demonstrated significant gene expression differences between tumor materials with different in vitro and in vivo growth microenvironments. Such studies can help us to distinguish between genes up-or downregulated as a result of the microenvironment and those stably expressed independently of the tumor milieu. With the extensive use of cell lines and xenografts in cancer research, the information obtained using our approach may help to better interpret results generated from different tumor models by understanding common differences, as well as similarities at the gene expression level, information that may have important practical and biological implications.

Encyclopedic Reference of Cancer, 2001

Oncology, 2010

Breast cancer metastasis suppressor 1 (BRMS1) has been shown to functionally reduce the metastati... more Breast cancer metastasis suppressor 1 (BRMS1) has been shown to functionally reduce the metastatic potential of melanoma. We also previously reported that BRMS1 negatively regulates the expression of the oncoprotein osteopontin (OPN). This study was carried out to assess the clinical relevance of BRMS1 and OPN in melanoma. Epigenetic regulation of BRMS1 was assessed by treating clinically derived melanoma cell lines with the demethylating agent 5-aza-2'-deoxycytidine (DAC) and the histone deacetylase inhibitor trichostatin A (TSA), followed by sodium bisulfite modification and methylation-specific PCR. Assessments of BRMS1 and OPN levels were performed using immunoblotting, quantitative real-time RT-PCR or reporter assays. RNA silencing was employed to abrogate the expression of OPN in melanoma-derived cell lines. The in vivo relevance of our findings was determined with experiments using athymic nude mice. The reduced expression of BRMS1 in surgically excised melanoma specimens correlated with increased OPN expression during the progression from primary to metastatic melanoma. Treatment with DAC and TSA elevated BRMS1 levels, but caused an inconsistent change in OPN gene expression. Abrogating the expression of OPN in BRMS1-deficient metastatic melanoma-derived cell lines retarded the growth of melanoma tumor xenografts in athymic nude mice. While treatment with DAC and TSA may not be a universally applicable treatment alternative in melanoma, silencing the expression of OPN in metastatic melanomas that have lost expression of BRMS1 is a potential option for therapeutic intervention.

Matrix Biology, 2014

Osteopontin (OPN) is a multifunctional cytokine that impacts cell proliferation, survival, drug r... more Osteopontin (OPN) is a multifunctional cytokine that impacts cell proliferation, survival, drug resistance, invasion, and stem like behavior. Due to its critical involvement in regulating cellular functions, its aberrant expression and/or splicing is functionally responsible for undesirable alterations in disease pathologies, specifically cancer. It is implicated in promoting invasive and metastatic progression of many carcinomas. Due to its autocrine and paracrine activities OPN has been shown to be a crucial mediator of cellular cross talk and an influential factor in the tumor microenvironment. OPN has been implicated as a prognostic and diagnostic marker for several cancer types. It has also been explored as a possible target for treatment. In this article we hope to provide a broad perspective on the importance of OPN in the pathophysiology of cancer.

Laboratory Investigation, 2012

DNAJB6 is a constitutively expressed member of the HSP40 family. It has been described as a negat... more DNAJB6 is a constitutively expressed member of the HSP40 family. It has been described as a negative regulator of breast tumor progression and a regulator of epithelial phenotype. Expression of DNAJB6 is reported to be compromised with tumor progression. However, factors responsible for its down-regulation are still undefined. We used a knowledge-based screen for identifying miRNAs capable of targeting DNAJB6. In this work, we present our findings that hsa-miR-632 (miR-632) targets the coding region of DNAJB6. Invasive and metastatic breast cancer cells express high levels of miR-632 compared to mammary epithelial cells. Analysis of RNA from breast tumor specimens reveals inverse expression patterns of DNAJB6 transcript and miR-632. In response to exogenous miR-632 expression, DNAJB6 protein levels are down regulated and the resultant cell population shows significantly increased invasive ability. Silencing endogenous miR-632 abrogates invasive ability of breast cancer cells and promotes epithelial like characteristics noted by E-cadherin expression with concomitant decrease in mesenchymal markers such as Zeb2 and Slug. Thus, miR-632 is a potentially important epigenetic regulator of DNAJB6 which contributes to the down regulation of DNAJB6 and plays a supportive role in malignant progression.

Journal of Cellular and Molecular Medicine, 2009

The growth of cancer cells as multicellular spheroids has frequently been reported to mimic the i... more The growth of cancer cells as multicellular spheroids has frequently been reported to mimic the in vivo tumour architecture and physiology and has been utilized to study antitumour drugs. In order to determine the distinctive characteristics of the spheroid-derived cells compared to the corresponding monolayer-derived cells, we enriched multicellular spheroid-forming subpopulations of cells from three human breast cancer cell lines (MCF7, MCF10AT and MCF10DCIS.com). These spheroid-derived cells were injected into female athymic nude mice to assess their tumorigenic potential and were profiled for their characteristic miRNA signature. We discovered that the sphe-

International Journal of Cancer, 2014

Hedgehog (Hh) signaling regulates embryonic patterning and organ morphogenesis. It is also involv... more Hedgehog (Hh) signaling regulates embryonic patterning and organ morphogenesis. It is also involved in regeneration and repair of tissues. Aberrant Hh pathway activation is a feature of many human malignancies. Classical Hh signaling is activated by Hh ligands that can signal in an autocrine or paracrine manner generating a tumor-stromal crosstalk. In contrast to canonical Hh signaling that culminates in the activation of GLI transcription factors, "noncanonical" Hh signaling does not involve GLI transcriptional activity. Several Hh pathway inhibitors have progressed to clinical trials, where the outcomes have not been very encouraging in many solid tumors. Here we discuss the likely role of "nonclassical" Hh-GLI signaling that is activated by growth factors and cytokines from the tumor and/or its microenvironment; these uncouple Hh signaling from the vital regulatory protein Smoothened, and result in the activation of GLI. While efforts are being made to target tumor-intrinsic Hh targets, it is imperative to acknowledge the role of the complex molecular networks and crosstalk between different components of the tumor microenvironment that can result in the emergence of resistance to conventional Hh therapy. These considerations have an important bearing on appreciating the need to mitigate the effects of tumor microenvironment to combat resistance to Hh inhibitors.

International Journal of Cancer, 2004

Our previous studies demonstrate that introduction of a ϳ70 cM region (now estimated at 63.75 Mb ... more Our previous studies demonstrate that introduction of a ϳ70 cM region (now estimated at 63.75 Mb by the Human Genome Project) of human chromosome 12 into the highly metastatic Dunning rat prostate cancer cell line AT6.1 results in >30-fold (>90%) reduction in the number of overt metastases in spontaneous metastasis assays. We report the further localization and biological characterization of the metastasis-suppressor activity encoded by a reduced region of chromosome 12. To localize this metastasis-suppressor activity, a panel of AT6.1 microcell hybrids that retain varying portions of human chromosome 12 was constructed and subjected to sequence-tagged site (STS)-based PCR analysis and assessment of in vivo metastatic ability. Data from these complementary approaches localized the metastasis-suppressor activity to a ϳ4.2 Mb portion of human chromosome 12q24.3 comprised of 3 separate regions. Reverse transcriptase-polymerase chain reaction (RT-PCR) and immunoblotting were used for differential expression analyses to identify which characterized genes, predicted gene sequences and expressed sequence tags (ESTs) within this region could be responsible for the observed metastasis suppression. Comprehensive in vivo studies showed that suppressed AT6.1-12 hybrids that retain the metastasis-suppressor region on 12q24.3 are capable of arriving at the secondary site, but are not able to persist there. Thus, unlike other metastasis-suppressor genes characterized to date, the metastasis-suppressor gene encoded by this region appears to utilize a different biologic mechanism to suppress the growth of overt metastases at the secondary site.

International Journal of Breast Cancer, 2012

The microenvironment at the site of tumor metastasis plays a key role in determining the fate of ... more The microenvironment at the site of tumor metastasis plays a key role in determining the fate of the metastasizing tumor cells. This ultimately has a direct impact on the progression of cancer. Bone is the preferred site of metastasis of breast cancer. Painful, debilitating osteolytic lesions are formed as a result of crosstalk between breast cancer cells and cells in the bone, predominantly the osteoblasts and osteoclasts. In this paper, we have discussed the temporal and spatial role of hedgehog (Hh) signaling in influencing the fate of metastatic breast cancer cells in bone. By virtue of its secreted ligands, the Hh pathway is capable of homotypic and heterotypic signaling and consequently altering the microenvironment in the bone. We also have put into perspective the therapeutic implications of using Hh inhibitors to prevent and/or treat bone metastases of breast cancer.

Cancer Metastasis — Biology and Treatment, 2005

... 7. Ohtaki T., Shintani Y., Honda S., Matsumoto H., Hori A., Kanehashi K., Torao Y., Kumano S.... more ... 7. Ohtaki T., Shintani Y., Honda S., Matsumoto H., Hori A., Kanehashi K., Torao Y., Kumano S., Takatsu Y., Matsuda Y., Ishibashi Y ... Saunders MM, Seraj MJ, Li ZY, Zhou ZY, Winter CR, Welch DR and Donahue HJ Breast cancer metastatic potential correlates with a breakdown in ...

International Journal of Breast Cancer, 2012

PLoS ONE, 2012

The remodeling process in bone yields numerous cytokines and chemokines that mediate crosstalk be... more The remodeling process in bone yields numerous cytokines and chemokines that mediate crosstalk between osteoblasts and osteoclasts and also serve to attract and support metastatic tumor cells. The metastatic tumor cells disturb the equilibrium in bone that manifests as skeletal complications. The Hedgehog (Hh) pathway plays an important role in skeletogenesis. We hypothesized that the Hh pathway mediates an interaction between tumor cells and osteoblasts and influences osteoblast differentiation in response to tumor cells. We have determined that breast tumor cells have an activated Hh pathway characterized by upregulation of the ligand, IHH and transcription factor GLI1. Breast cancer cells interact with osteoblasts and cause an enhanced differentiation of pre-osteoblasts to osteoblasts that express increased levels of the osteoclastogenesis factors, RANKL and PTHrP. There is sustained expression of osteoclast-promoting factors, RANKL and PTHrP, even after the osteoblast differentiation ceases and apoptosis sets in. Moreover, tumor cells that are deficient in Hh signaling are compromised in their ability to induce osteoblast differentiation and consequently are inefficient in causing osteolysis. The stimulation of osteoblast differentiation sets the stage for osteoclast differentiation and overall promotes osteolysis. Thus, in the process of developing newer therapeutic strategies against breast cancer metastasis to bone it would worthwhile to keep in mind the role of the Hh pathway in osteoblast differentiation in an otherwise predominant osteolytic phenomenon.

Cancer Metastasis — Biology and Treatment, 2002

Cancer Metastasis – Biology and Treatment, 2007

Metastasis of breast cancer is a complex event involving coordinated cross-talk of several protei... more Metastasis of breast cancer is a complex event involving coordinated cross-talk of several proteins. Genes that control the resultant metastasis can be broadly classified as metastasis promoter genes (MPGs) and metastasis suppressor genes (MSGs). There is an explosion of information in the studies that focus on these genes; however, thus far, a very few of them are actually tested clinically

Utilizing gene microarray profiling of melanoma samples, we have recently identified a novel gene... more Utilizing gene microarray profiling of melanoma samples, we have recently identified a novel gene overexpressed in both thick primary and metastatic melanomas. This gene, progestagen-associated endometrial protein (PAEP), has never before been implicated in the oncogenic processes of melanoma, with its true function in oncogenesis and tumour progression relatively unknown. Overexpression of the PAEP gene in freshly procured thick primary and metastatic melanoma samples (58%) and daughter cell lines (77%) is confirmed by quantitative RT-PCR, immunohistochemistry, Western blotting and mass spectrometric analysis. We suggest that PAEP gene overexpression is involved with melanoma tumour progression as well as an aggressive phenotype. Transfection of melanoma cells with PAEP small interfering RNA (siRNA) reveals a significant decrease in soft agar colony formation and a marked inhibition of both cell migration and cell invasion. Furthermore, we establish stable melanoma transfectants via PAEP lentiviral small hairpin RNA (shRNA), examine their growth characteristics in a murine xenograft model and reveal that tumour growth is significantly inhibited in two separate melanoma cell lines. Our data strongly implicate the PAEP gene as a tumour growth promoter with oncogenic properties and a potential therapeutic target for patients with advanced melanoma.

Introduction: Breast cancer is one of the most common malignancies affecting women in the United ... more Introduction: Breast cancer is one of the most common malignancies affecting women in the United States and Europe. Approximately three out of every four women with breast cancer develop metastases in bone which, in turn, diminishes quality of life. The α v β 3 integrin has previously been implicated in multiple aspects of tumor progression, metastasis and osteoclast bone resorption. Therefore, we hypothesized that the α v β 3 -selective inhibitor, S247, would decrease the development of osteolytic breast cancer metastases. Materials and methods: Cells were treated in vitro with S247 and assessed for viability and adhesion to matrix components. Athymic mice received intracardiac (left ventricle) injections of human MDA-MB-435 breast carcinoma cells expressing enhanced green-fluorescent protein.

The exact cellular and molecular mechanisms involved in melanoma tumorigenesis remain obscure. Pr... more The exact cellular and molecular mechanisms involved in melanoma tumorigenesis remain obscure. Previous gene expression profiling analyses performed upon NHEM and human melanoma samples identified WFDC1 as one of the most frequently down-regulated genes. Here we further showed that NHEM readily express WFDC1 but expression is reduced or completely lost in 80% of the patients-derived melanoma cell lines and tissue samples examined. Furthermore, we show that promoter hypermethylation accounts for the silencing of the WFDC1 gene in 20% of the melanoma cell lines examined. The over-expression of WFDC1 in two metastatic melanoma cell lines, A375 and LOX, resulted in a significant delay of tumor growth in a murine xenograft model, despite a non-significant difference in tumor cell growth in vitro. Gene expression microarray analysis and further expression validation suggests that the Dickkopf-1 (Dkk1) gene is up-regulated in WFDC1 over-expressing cell lines, suggesting that the tumor suppressive function of WFDC1 may be partially a result of up-regulated Dkk1 gene expression, which is known to be a potent inhibitor of the Wnt signaling pathway.

Cancer letters, Jan 10, 2014

22 Available online xxxx 23 Keywords: 24 Niclosamide 25 FDA-approved drug 26 Multi-targeted thera... more 22 Available online xxxx 23 Keywords: 24 Niclosamide 25 FDA-approved drug 26 Multi-targeted therapy 27 Drug discovery 28 Cancer stem cells 29 3 0 a b s t r a c t 31 The rapid development of new anticancer drugs that are safe and effective is a common goal shared by 32 basic scientists, clinicians and patients. The current review discusses one such agent, namely niclosamide, 33 which has been used in the clinic for the treatment of intestinal parasite infections. Recent studies repeat-34 edly identified niclosamide as a potential anticancer agent by various high-throughput screening cam-35 paigns. Niclosamide not only inhibits the Wnt/b-catenin, mTORC1, STAT3, NF-jB and Notch signaling 36 pathways, but also targets mitochondria in cancer cells to induce cell cycle arrest, growth inhibition 37 and apoptosis. A number of studies have established the anticancer activities of niclosamide in both 38 in vitro and in vivo models. Moreover, the inhibitory effects of niclosamide on cancer stem cells provide 39

Molecular cancer therapeutics, 2014

Basal-like breast cancers (BLBC) are poorly differentiated and display aggressive clinical behavi... more Basal-like breast cancers (BLBC) are poorly differentiated and display aggressive clinical behavior. These tumors become resistant to cytotoxic agents, and tumor relapse has been attributed to the presence of cancer stem cells (CSC). One of the pathways involved in CSC regulation is the Wnt/b-catenin signaling pathway. LRP6, a Wnt ligand receptor, is one of the critical elements of this pathway and could potentially be an excellent therapeutic target. Niclosamide has been shown to inhibit the Wnt/b-catenin signaling pathway by causing degradation of LRP6. TRA-8, a monoclonal antibody specific to TRAIL death receptor 5, is cytotoxic to BLBC cell lines and their CSC-enriched populations. The goal of this study was to examine whether niclosamide is cytotoxic to BLBCs, specifically the CSC population, and if in combination with TRA-8 could produce increased cytotoxicity. Aldehyde dehydrogenase (ALDH) is a known marker of CSCs. By testing BLBC cells for ALDH expression by flow cytometry, we were able to isolate a nonadherent population of cells that have high ALDH expression. Niclosamide showed cytotoxicity against these nonadherent ALDH-expressing cells in addition to adherent cells from four BLBC cell lines: 2LMP, SUM159, HCC1187, and HCC1143. Niclosamide treatment produced reduced levels of LRP6 and b-catenin, which is a downstream Wnt/b-catenin signaling protein. The combination of TRA-8 and niclosamide produced additive cytotoxicity and a reduction in Wnt/b-catenin activity. Niclosamide in combination with TRA-8 suppressed growth of 2LMP orthotopic tumor xenografts. These results suggest that niclosamide or congeners of this agent may be useful for the treatment of BLBC. Mol Cancer Ther; 13(4); 800-11. Ó2014 AACR.

Cancer genomics & proteomics

have been used for the discovery of genes and pathways involved in melanoma and other types of ca... more have been used for the discovery of genes and pathways involved in melanoma and other types of cancer. However, in many cases, the results from various tumor models failed to be validated successfully in clinical studies. Limited information is available on how closely these models reflect the in vivo physiological conditions. In this study, a comprehensive genomics approach was used to systematically compare the expression patterns of snap frozen samples obtained from patients with primary melanoma, lymph node metastasis, and distant metastases, and compare these patterns to those of their corresponding cell lines and tumor xenografts in nude mice. The GE Healthcare 20k human genome array was used and the expression data was normalized and analyzed using GeneSpring 7.2 software. Based on the expression analysis, the correlation rate between the snap frozen primary patient samples vs. derived cell lines was 66%, with 1687 differentially expressed genes. The correlation rate between the snap frozen primary patient samples and the tumor xenografts was 75%, with 1,374 differentially expressed genes, and the correlation rate comparing tumor xenografts to derived cell lines ranged between 58% and 84%. These results demonstrated significant gene expression differences between tumor materials with different in vitro and in vivo growth microenvironments. Such studies can help us to distinguish between genes up-or downregulated as a result of the microenvironment and those stably expressed independently of the tumor milieu. With the extensive use of cell lines and xenografts in cancer research, the information obtained using our approach may help to better interpret results generated from different tumor models by understanding common differences, as well as similarities at the gene expression level, information that may have important practical and biological implications.

Encyclopedic Reference of Cancer, 2001

Oncology, 2010

Breast cancer metastasis suppressor 1 (BRMS1) has been shown to functionally reduce the metastati... more Breast cancer metastasis suppressor 1 (BRMS1) has been shown to functionally reduce the metastatic potential of melanoma. We also previously reported that BRMS1 negatively regulates the expression of the oncoprotein osteopontin (OPN). This study was carried out to assess the clinical relevance of BRMS1 and OPN in melanoma. Epigenetic regulation of BRMS1 was assessed by treating clinically derived melanoma cell lines with the demethylating agent 5-aza-2'-deoxycytidine (DAC) and the histone deacetylase inhibitor trichostatin A (TSA), followed by sodium bisulfite modification and methylation-specific PCR. Assessments of BRMS1 and OPN levels were performed using immunoblotting, quantitative real-time RT-PCR or reporter assays. RNA silencing was employed to abrogate the expression of OPN in melanoma-derived cell lines. The in vivo relevance of our findings was determined with experiments using athymic nude mice. The reduced expression of BRMS1 in surgically excised melanoma specimens correlated with increased OPN expression during the progression from primary to metastatic melanoma. Treatment with DAC and TSA elevated BRMS1 levels, but caused an inconsistent change in OPN gene expression. Abrogating the expression of OPN in BRMS1-deficient metastatic melanoma-derived cell lines retarded the growth of melanoma tumor xenografts in athymic nude mice. While treatment with DAC and TSA may not be a universally applicable treatment alternative in melanoma, silencing the expression of OPN in metastatic melanomas that have lost expression of BRMS1 is a potential option for therapeutic intervention.

Matrix Biology, 2014

Osteopontin (OPN) is a multifunctional cytokine that impacts cell proliferation, survival, drug r... more Osteopontin (OPN) is a multifunctional cytokine that impacts cell proliferation, survival, drug resistance, invasion, and stem like behavior. Due to its critical involvement in regulating cellular functions, its aberrant expression and/or splicing is functionally responsible for undesirable alterations in disease pathologies, specifically cancer. It is implicated in promoting invasive and metastatic progression of many carcinomas. Due to its autocrine and paracrine activities OPN has been shown to be a crucial mediator of cellular cross talk and an influential factor in the tumor microenvironment. OPN has been implicated as a prognostic and diagnostic marker for several cancer types. It has also been explored as a possible target for treatment. In this article we hope to provide a broad perspective on the importance of OPN in the pathophysiology of cancer.

Laboratory Investigation, 2012

DNAJB6 is a constitutively expressed member of the HSP40 family. It has been described as a negat... more DNAJB6 is a constitutively expressed member of the HSP40 family. It has been described as a negative regulator of breast tumor progression and a regulator of epithelial phenotype. Expression of DNAJB6 is reported to be compromised with tumor progression. However, factors responsible for its down-regulation are still undefined. We used a knowledge-based screen for identifying miRNAs capable of targeting DNAJB6. In this work, we present our findings that hsa-miR-632 (miR-632) targets the coding region of DNAJB6. Invasive and metastatic breast cancer cells express high levels of miR-632 compared to mammary epithelial cells. Analysis of RNA from breast tumor specimens reveals inverse expression patterns of DNAJB6 transcript and miR-632. In response to exogenous miR-632 expression, DNAJB6 protein levels are down regulated and the resultant cell population shows significantly increased invasive ability. Silencing endogenous miR-632 abrogates invasive ability of breast cancer cells and promotes epithelial like characteristics noted by E-cadherin expression with concomitant decrease in mesenchymal markers such as Zeb2 and Slug. Thus, miR-632 is a potentially important epigenetic regulator of DNAJB6 which contributes to the down regulation of DNAJB6 and plays a supportive role in malignant progression.

Journal of Cellular and Molecular Medicine, 2009

The growth of cancer cells as multicellular spheroids has frequently been reported to mimic the i... more The growth of cancer cells as multicellular spheroids has frequently been reported to mimic the in vivo tumour architecture and physiology and has been utilized to study antitumour drugs. In order to determine the distinctive characteristics of the spheroid-derived cells compared to the corresponding monolayer-derived cells, we enriched multicellular spheroid-forming subpopulations of cells from three human breast cancer cell lines (MCF7, MCF10AT and MCF10DCIS.com). These spheroid-derived cells were injected into female athymic nude mice to assess their tumorigenic potential and were profiled for their characteristic miRNA signature. We discovered that the sphe-

International Journal of Cancer, 2014

Hedgehog (Hh) signaling regulates embryonic patterning and organ morphogenesis. It is also involv... more Hedgehog (Hh) signaling regulates embryonic patterning and organ morphogenesis. It is also involved in regeneration and repair of tissues. Aberrant Hh pathway activation is a feature of many human malignancies. Classical Hh signaling is activated by Hh ligands that can signal in an autocrine or paracrine manner generating a tumor-stromal crosstalk. In contrast to canonical Hh signaling that culminates in the activation of GLI transcription factors, "noncanonical" Hh signaling does not involve GLI transcriptional activity. Several Hh pathway inhibitors have progressed to clinical trials, where the outcomes have not been very encouraging in many solid tumors. Here we discuss the likely role of "nonclassical" Hh-GLI signaling that is activated by growth factors and cytokines from the tumor and/or its microenvironment; these uncouple Hh signaling from the vital regulatory protein Smoothened, and result in the activation of GLI. While efforts are being made to target tumor-intrinsic Hh targets, it is imperative to acknowledge the role of the complex molecular networks and crosstalk between different components of the tumor microenvironment that can result in the emergence of resistance to conventional Hh therapy. These considerations have an important bearing on appreciating the need to mitigate the effects of tumor microenvironment to combat resistance to Hh inhibitors.

International Journal of Cancer, 2004

Our previous studies demonstrate that introduction of a ϳ70 cM region (now estimated at 63.75 Mb ... more Our previous studies demonstrate that introduction of a ϳ70 cM region (now estimated at 63.75 Mb by the Human Genome Project) of human chromosome 12 into the highly metastatic Dunning rat prostate cancer cell line AT6.1 results in >30-fold (>90%) reduction in the number of overt metastases in spontaneous metastasis assays. We report the further localization and biological characterization of the metastasis-suppressor activity encoded by a reduced region of chromosome 12. To localize this metastasis-suppressor activity, a panel of AT6.1 microcell hybrids that retain varying portions of human chromosome 12 was constructed and subjected to sequence-tagged site (STS)-based PCR analysis and assessment of in vivo metastatic ability. Data from these complementary approaches localized the metastasis-suppressor activity to a ϳ4.2 Mb portion of human chromosome 12q24.3 comprised of 3 separate regions. Reverse transcriptase-polymerase chain reaction (RT-PCR) and immunoblotting were used for differential expression analyses to identify which characterized genes, predicted gene sequences and expressed sequence tags (ESTs) within this region could be responsible for the observed metastasis suppression. Comprehensive in vivo studies showed that suppressed AT6.1-12 hybrids that retain the metastasis-suppressor region on 12q24.3 are capable of arriving at the secondary site, but are not able to persist there. Thus, unlike other metastasis-suppressor genes characterized to date, the metastasis-suppressor gene encoded by this region appears to utilize a different biologic mechanism to suppress the growth of overt metastases at the secondary site.