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Published in final edited form as: Cancer Lett. 2015 Jul 30;368(1):79–87. doi: 10.1016/j.canlet.2015.07.032

Small molecule inhibitor of the bone morphogenetic protein pathway DMH1 reduces ovarian cancer cell growth

Laura D Hover 1, Christian D Young 2, Neil E Bhola 2, Andrew J Wilson 1,3, Dineo Khabele 1,3, Charles C Hong 4,5, Harold L Moses 6, Philip Owens 6
PMCID: PMC4554828  NIHMSID: NIHMS712461  PMID: 26235139

Abstract

The bone morphogenetic protein (BMP) pathway belonging to the Transforming Growth Factor beta (TGFβ) family of secreted cytokines/growth factors is an important regulator of cancer. BMP ligands have been shown to play both tumor suppressive and promoting roles in human cancers. We have found that BMP ligands are amplified in human ovarian cancers and that BMP receptor expression correlates with poor progression-free-survival (PFS). Furthermore, active BMP signaling has been observed in human ovarian cancer tissue. We also determined that ovarian cancer cell lines have active BMP signaling in a cell autonomous fashion. Inhibition of BMP signaling with a small molecule receptor kinase antagonist is effective at reducing ovarian tumor sphere growth. Furthermore, BMP inhibition can enhance sensitivity to Cisplatin treatment and regulates gene expression involved in platinum resistance in ovarian cancer. Overall, these studies suggest targeting the BMP pathway as a novel source to enhance chemo-sensitivity in ovarian cancer.

Keywords: BMP, BMPR1a, DMH1, ovarian cancer, platinum resistance

1. Introduction

Ovarian cancer is the leading cause of cancer deaths from gynecological malignancies [1]. It is estimated that in 2015, 21,290 women will be diagnosed with ovarian cancer and over 14,000 women will die from ovarian cancer [1; 2]. The five year survival rate is dismal at 45.6% and despite advances in treatment, the survival rates have not improved [3]. Ovarian cancer remains deadly due to its invasive nature leading to metastasis, late diagnosis, tumor heterogeneity and resistance to chemotherapy. Recently, the bone morphogenetic protein (BMP) pathway has been discovered to play critical roles during tumor formation and progression in ovarian cancers [4].

The BMP pathway is a complex director of development and part of the transforming growth factor beta (TGFβ) family. BMP ligands, which are secreted by many cell types,bind to type I and type II serine/threonine kinase receptors. Upon ligand binding, type I receptors phosphorylate Smads 1, 5 and 8(mouse)/9(human), which then bind with Smad4. The complex is then translocated to the nucleus to regulate transcription of target genes. Canonical BMP target genes in response to ligand stimulation are ID1 and the inhibitory Smad, Smad6, which functions in a negative feedback manner thus tightly regulating BMP signaling [5; 6; 7].

BMP signaling was originally characterized as tumor suppressive. Similar to TGFβ, in cancer cells, the role of BMP signaling is becoming increasingly complex showing dual tumor suppressive and promoting roles. The basis of BMP tumor suppression originated with the discovery that patients with mutations in BMPR1a effectively inhibiting BMP signaling developed Juvenile Polyposis Syndrome associated hamartomas. Concordantly, mice with targeted deletion of BMPR1a in skin develop similar hamartomatous lesions [8; 9; 10; 11; 12]. When the type II BMP receptor, BMPR2, is expressed as a dominant negative in a mouse model of breast cancer, it enhances tumor metastasis [13]. Many studies have shown that treatment of normal and cancerous cells with BMP ligands reduces cell proliferation and growth and, similar to TGFβ treatment, induces transcription of p21/27/57 to repress the MYC oncogene [14; 15; 16; 17]. In addition it has been shown that treatment of cells with BMP ligand antagonists Noggin and Coco can lead to increased cell proliferation and promote metastasis [18; 19]. Recently, it has been shown that breast cancer cells show enhanced cell migration and invasion when treated with BMP ligands [20; 21]. In addition, when BMP ligands are overexpressed in various cancer cells such as prostate cancer, they can demonstrate tumor-promoting phenotypes such as increased invasion and metastasis [22]. Recent reviews have highlighted these dual roles for BMPs in cancer [23; 24].

In ovarian cancer, BMP signaling has been shown to have dual tumor suppressive and promoting roles as well. For example, treatment of ovarian cancer cells with recombinant BMP-2 has an anti-proliferative effect [14] and constitutively active BMPR1a receptor activity limits metastasis of ovarian cancer cells through decreased adhesion [25]. Additionally, the BMP receptors BMPR1a and BMPR1b have redundant roles that can suppress ovarian tumor formation [26]. Conversely, BMP tumor promoting activity has been demonstrated in both ovarian tumor cells and the surrounding microenvironment [27; 28]. BMP stimulation can enhance the proliferation of ovarian cancer cells [4; 29; 30; 31]. Furthermore, BMP signaling has been shown to increase ovarian cancer cell motility and migration via EMT and other pathways such as AKT [32; 33; 34]. Ovarian cancer is known to be heterogeneous at the cellular and molecular level and it remains unknown, which contexts dictate BMP function as tumor suppressive or promoting. We find that BMP activity is endogenously high in human ovarian cancers and in our model of ovarian cancer, BMP inhibition restricts growth and cooperates with standard platinum based chemotherapy.

2. Materials and Methods

2.1 Cell culture and tumor sphere assay

Cells were grown in DMEM medium with 10% Fetal Bovine Serum (FBS) and Triple antibiotic/antimycotic (Life Technologies). Cells grown as spheres were placed in ultra-low adhesion culture 6-well plates (Corning) and after five days treated with drug and allowed to grow for five additional days. Sphere media was comprised of DMEM/F12 with triple antibiotic/antimycotic, 20ng/ml EGF and 1× B27 supplement (Life Technologies). Spheres were analyzed using the Gelcount automated colony counting system. Equal sized masks were used in analysis and performed in triplicate. Routine mycoplasma testing was performed and cells were treated for 24 hours prophylactically with ciprofloxin for 24 hours (GeneHunter). DMH1 was used at 20uM for all experiments and Cisplatin was used at 1uM, and equal quantities of DMSO were used as vehicle control.

2.2 Immunohistochemistry

A Tissue microarray comprised of 209 samples from patients diagnosed with ovarian cancer was constructed at Vanderbilt University Medical Center (VUMC) as previously described [35]. All samples consisted of tumor tissue of various grades and histologies.

Tumor spheres were formalin fixed, placed into Histogel (Harlan Scientific), were embedded and sectioned at 5 µM and dewaxed in xylene and rehydrated in alcohol with citrate antigen retrieval as previously described [13]. Standard Mayer's hematoxylin and eosin (H&E) was performed. The sections were immunostained using antibodies for Cleaved Caspase-3 (Cell Signaling Cat#9661, 1:200) and Ki-67 (Novoastra Cat#ki67-mm1, 1:200) Paraffin derived sections were counterstained with hematoxylin (Vector Labs QS) and mounted with Cytoseal. Quantification of IHC and IF was performed using NIH ImageJ as previously described [36].

2.3 RNA isolation, cDNA synthesis, qPCR and primer selection

RNA isolation was performed with RNeasy purification including DNAseI treatment (Qiagen). Equal amounts of RNA were synthesized into cDNA using the VILO cDNA synthesis kit (Invitrogen). LuminoCt (Sigma) 2× SYBR mastermix was combined with 1 µM of both a forward and reverse primer sequence (primer sequences are listed in Supplemental Table 1) into 20 µl reactions and cycled for 95°–10 s to 60° for 30 s for 40 cycles followed by a melting curve. BioRad CFX96 was used and instrument provided software was used to determine relative normalized expression to GAPDH expression.

2.4 Western blot

Total protein was isolated using Complete LysisM Buffer (Roche). Protein was diluted to equal concentrations and equally loaded on 10% polyacrylamide gels prior to transfer to a nitrocellulose membrane. Protein concentration was determined using micro plate BCA assay (Pierce). Blots were incubated overnight with Smad1 (Cell Signaling Cat#6944 1:1000), pSmad1/5 (Cell Signaling Cat#9516 1:1000), and Actin (Sigma Cat#A2066 1:4000) antibodies. HRP-conjugated secondary antibodies were used to visualize band intensity via x-ray film exposure using ECL western substrate (Perkin Elmer).

2.5 Human breast cancer databases and statistical analysis

For analysis of the TCGA data set, we used the cBio portal (http://www.cbio.portal.org/) [31; 37]. We queried the TCGA provisional serous ovarian dataset which was comprised of 594 patients and 609 tumors and the CCLE dataset which contained 50 ovarian cancer cell lines. Human gene symbols were queried and accessed on 11 March 2015. RNA expression cutoff was adjusted to 2.0. Analysis of gene expression correlating with RFS was performed using the kmplotter (http://kmplot.com). Human gene symbols were entered into ovarian (GSE14764), and JetSet probe selection was used to determine optimal representative microarray probe [38]. Automatic cutoff scores were selected during queries and PFS were selected. Statistical analysis was performed using Excel (Microsoft, Redmond, WA, USA), Prism (Graphpad, La Jolla, CA, USA), and FlowJo (TreeStar, Ashland, OR, USA) software. Statistical significance was deemed for any comparison where P<0.05.

3. Results

3.1 BMP expression is amplified and active in ovarian cancer and correlates with poor relapse free survival

The BMP pathway has many ligands and their expression in cancer has only recently been addressed. We queried The Cancer Genome Atlas (TCGA) using the cBIO.org portal from Memorial Sloan Kettering Cancer Center and specifically selected the serous ovarian cancer data set. We found that BMP ligands were rarely deleted or downregulated and were commonly upregulated, with BMP7 upregulation or amplification in 17% of patients with serous ovarian cancer (Figure 1A). The datasets provided from the TCGA are from whole tumor tissue samples and can represent gene expression from the tumor microenvironment in addition to the tumor cells. To assess BMP signaling specifically in tumor cells, we analyzed BMP ligand expression in ovarian cancer cell lines, which have been profiled in the Cancer Cell Line Encyclopedia (CCLE) and are available through the cBIO.org portal. Upon analysis of the cell lines, we found that most cell lines showed amplification or upregulation of BMP ligands. BMP7 was the most frequently altered, upregulated or amplified in 32% of the ovarian cancer cell lines (Figure S1A). In addition to investigating BMP ligands, we looked at the expression of TGFβ family receptors and co-receptors in ovarian cancer patients in the TCGA data set and found all receptors to be altered in only 10% or less of patients. The majority of these alterations were amplifications or upregulations (Figure S1B).

Figure 1. BMPs are widely expressed in human ovarian cancer and BMP receptors correlate with poor survival.

Figure 1

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A. Oncoprint from cBIO.org indicating the percent change in BMP ligands expressed in the provisional TCGA human serous ovarian cancer dataset. B. Progression Free Survival (PFS) based on gene expression of BMP receptors. C. IHC for pSmad1/5/9 in primary human ovarian cancers grade 1 and grade 3. Black arrows indicate positive staining. Scale bars indicate 50µm.

Next, we looked at the association between gene expression of BMP signaling molecules and survival using kmplot.com. The gene expression of two type I receptors, ACVR1 (also known as ALK2) and BMPR1a (also known as ALK3) is correlated with poor progression free survival (PFS) in human ovarian cancer (Figure 1B). To examine active BMP signaling within ovarian cancer, we analyzed the phosphorylation of Smads 1/5/9 by immunohistochemistry using a TMA comprised of 209 samples of ovarian cancer of various grades, stages and histologies. Our results revealed heterogeneous, yet positive staining throughout all grades and stages of ovarian cancers. p-Smad1/5/9 was expressed in both the tumor epithelium and the surrounding stroma (Figure 1C).

3.2 Human ovarian cell lines display cell endogenous BMP activity that when inhibited decreases sphere size

We investigated three commonly used human ovarian cancer cell lines, OVCAR3, OVCAR8 and SKOV3 and found that all untreated cells displayed active BMP signaling indicated by phosphorylation of Smads1,5 and 9 in a cell autonomous fashion (Figure 2A). Using DMH1, a small molecule kinase inhibitor for the type I BMP receptors, at 20uM for 24 hours we could reduce the canonical phosphorylation of Smads 1,5 and 9. Interestingly, we found that acute treatment of these cells with Cisplatin at 1uM for 24 hours also resulted in a reduction of BMP Smad phosphorylation, and when combined with DMH1 eliminated the remaining detectable phosphorylated Smads 1,5 and 9 (Figure 2A). When ovarian cancer metastasizes, it can form spheroids that can invade and travel to distant sites [39; 40]. To mimic this phenomenon in vitro, cancer cells can be grown as spheroids using ultra-low adhesion cell culture plates. We grew OVCAR3, OVCAR8 and SKOV3 cells in spheroid conditions for ten days and found that OVCAR8 had the most efficient sphere forming ability and produced the largest number of spheres from an initial seeding of 50,000 cells (Figure S2A). OVCAR8 cells were also the largest in size and made well-defined spheres within ten days of growth (Figure S2B). Interestingly, when we closely examined the spheres, we noticed that OVCAR3 cells were mostly hollow and created cyst-like structures (Figure S2B). SKOV3 cells formed the fewest number of spheres and tended to grow in aggregates and made poor cell-cell connections (Figure S2B).

Figure 2. Human ovarian cell lines have active BMP signaling.

Figure 2

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A. Western blot for three human ovarian cancer cell lines OVCAR3, OVCAR8 and SKOV3 reveals active BMP signaling by the presence of phospho-Smad1/5/9. Cells were treated with DMH1 (20µM), Cisplatin (1µM), or both for 24 hours as indicated by a plus symbol. B. OVCAR8 cells grown in tumor sphere conditions for ten days were treated with DMH1 (20µM), Cisplatin (1µM), or both on day 5 after seeding. C. Tumor spheres embedded in histogel were stained for H&E. Error bars indicate SEM. *p=<0.05, **p=<0.01. Scale bars indicate 100µm.

We continued with a focus on OVCAR8 cells because they showed the greatest ability to be able to form spheres and spheroid cells have been associated with increased tumor cell aggressiveness [41]. We sought to see if BMP inhibition and Cisplatin treatment could affect the growth of tumor spheres. Using ultra-low-adherent culture 6-well culture plates we seeded 50,000 OVCAR8 cells in triplicate for five days. After five days, small spheres had begun to form and we treated the tumor spheres once with DMH1 (20µM), Cisplatin (1µM), or both for five additional days. We measured sphere volume and diameter and following tumor sphere measurements we Formalin-Fixed Paraffin Embedded (FFPE) spheres into Histogel and perform Hematoxylin and Eosin (H&E) staining (Figure 2B, C). We found that DMH1 was sufficient to inhibit the growth of tumor spheres by total volume and sphere size (Figure 2B). Interestingly, Cisplatin treatment alone did not affect sphere size or volume; however in combination with DMH1 we observed a significant reduction in sphere size and volume (Figure 2B).

3.3 Cisplatin resistant cell lines respond to BMP inhibition

Subsequently we wanted to examine BMP inhibition in the context of chemotherapy resistance. NCI-RES is a chemo-resistant cell line derived from OVCAR8 cells [42]. We repeated our treatments of DMH1 and Cisplatin on OVCAR8 cells and NCI-RES cells grown as spheres and observed that treatment with DMH1 as a single agent was able to inhibit sphere size in OVCAR8 cells, but not in NCI-RES cells. However, when used in combination, treatment with DMH1 (20uM) and Cisplatin (1µM) for five days resulted in a significant reduction in sphere volume and size in NCI-RES resistant cells (Figure 3A). To investigate if the changes in sphere volume and size were due to proliferation, we performed IHC for the proliferation marker Ki-67 on FFPE tumor spheres from the various treatment groups after ten days of growth in suspension (Figure 3B). We found that Ki-67 positive staining was significantly decreased in response to combination therapy of DMH1 and Cisplatin in the OVCAR8 cells (Figure S3A). However, there was no significant change in proliferation, measured by the presence of Ki-67, after combination therapy in the NCI-RES cells. We next investigated cell death in the spheres in response to treatment by analyzing the expression of cleaved Caspase-3 by IHC (Figure 3C). We found no significant changes in the expression of cleaved Caspase-3 in any tumor spheres in response to treatment (Figure S3B).

Figure 3. Cisplatin resistant cell lines respond to BMP inhibition.

Figure 3

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A. OVCAR8 (white bars) and NCI-RES (black bars) cells were grown as tumor spheres and were treated after five days of culture with DMH1 (20µM), Cisplatin (1µM), or both for five days. B. Tumor spheres were FFPE and IHC for proliferation marker Ki-67 was performed. C. IHC for apoptotic marker cleaved Caspase-3 was performed on FFPE tumor spheres. Error bars indicate SEM. *p=<0.05, **p=<0.01. Scale bars indicate 100µm.

3.4 BMP inhibition regulates genes associated with Cisplatin resistance

The regulation of gene expression has recently been associated with resistance in ovarian cancer cells. Recent large genomic studies have begun to identify gene expression patterns which are altered in ovarian Cisplatin resistant cells [43; 44; 45]. We treated OVCAR8 and NCI-RES cells with acute 24-hour treatment of DMH1 (20µM), Cisplatin (1µM), or both and isolated RNA. Next we analyzed the panel of genes shown to be up or downregulated in chemo-resistant ovarian cancer cells by Li et al by qPCR. We found that BMP inhibition regulates the expression of 4 genes shown to be associated with resistance: ROBO2, JAG1, CY1B1 and HAPLN1.. ROBO2 and JAG1, which were shown to be downregulated in Cisplatin resistant ovarian cancer cells, were significantly increased in response to combination therapy of DMH1 and Cisplatin in both Cisplatin sensitive OVCAR8 cells as well as insensitive NCI-RES cells [43]. ROBO2 mRNA was significantly increased only when treated with both DMH1 and Cisplatin. JAG1 however was significantly upregulated after DMH1 treatment alone or in combination with Cisplatin (Figure 4A). Previously CY1B1 and HAPLN1 were shown to be upregulated in Cisplatin resistant cells [43]. We found that CYP1B1 was reduced with single agent DMH1 treatment in both OVCAR8 and NCI-RES cells. HAPLN1 expression showed a trending increase in expression with single agent exposure to either DMH1 or Cisplatin, but when combined, gene expression was decreased significantly in both OVCAR8 and NCI-RES cells (Figure 4B).

Figure 4. BMP inhibition regulates gene expression associated with ovarian Cisplatin resistance.

Figure 4

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mRNA was collected from OVCAR8 (white bars) and NCI-RES (black bars) cells treated with DMH1 (20µM), Cisplatin (1µM), or both for 24 hours. to measure expression of genes A. qPCR analysis of ROBO2 and JAG1 which are downregulated in Cisplatin resistant cells. B. qPCR analysis of CYP1B1 and HAPLN1 which are upregulated in Cisplatin resistant cells. mRNA is normalized to GAPDH levels and relative to DMSO treated cells. Error bars indicate SEM. *p=<0.05, **p=<0.01.

Validation of the canonical BMP response genes, ID1 and SMAD6, revealed that only treatments including DMH1 directly decrease the BMP response (Figure S4A). In previous reports of genes linked to Cisplatin resistance in ovarian cancer cells, there were two BMP signaling molecules that were found to be downregulated, BAMBI and CHRDL1 [43]. We performed qPCR for these genes and found that Cisplatin treatment decreased BAMBI mRNA levels in OVCAR8 cells, yet combined treatment in NCI-RES cells led to an increase in BAMBI gene expression (Figure S4B). CHRDL1 expression was only significantly decreased in response to Cisplatin in the OVCAR8 cells and was not significantly altered in response to BMP inhibition in either cell line (Figure S4B).

4. Discussion

The BMP pathway is crucial to normal ovarian function and has been shown to regulate hormones, ovulation and oocyte development [46]. We and others have recently shown BMP signaling to be active in and regulate ovarian cancer cells as well. It has previously been shown that BMP ligands and signaling is increased in ovarian cancer cells in comparison to normal ovarian cells [46; 47]. Similar to previous studies, here we show that BMP ligands and receptors are overexpressed in more than 50% of ovarian cancer tumors and ovarian cancer cell lines (Figure 1, Supplementary Figure 1) and is endogenously active in ovarian cancer cell lines (Figure 2). Additionally we found that BMP signaling is active in ovarian cancer samples across multiple grades, stages, histology and in both chemo-sensitive and resistant cells (Figure 1).

The BMP pathway has been historically thought of as tumor suppressive, but as a large amount of genetic data from human patients becomes more available, BMPs have emerged as possible tumor promoters. Recently the BMP ligands, BMP-2 and BMP-4, have been shown to have prognostic association with a poor outcome for ovarian cancer patients [48; 49; 50]. We chose to look at the receptor partners for BMP signaling ACVR1 (ALK2) and BMPR1a (ALK3), which we determined are also correlated with poor outcome (Figure 1B). Recent work looking at the downstream mediator of canonical signaling, Smad5, has also been implicated in poor survival in ovarian cancer patients [4].

Inhibition of the BMP pathway has only recently been investigated as a putative target in cancer therapies. BMP small molecule inhibitors have radically progressed from toxic non-selective/specific inhibitors such as Dorsomorphin and LDN-193189 to more selective and specific inhibitors such as DMH1 [51; 52; 53; 54; 55]. Other groups have used Dorsomorphin and LDN-193189 for successful outcomes in many cancer types such as Lung, Prostate, Breast and acute lymphoblastic leukemia (ALL) [56; 57; 58; 59]. We have previously reported the successful use of DMH1 in metastatic breast cancer mouse model both on the tumor cells as well as the surrounding tumor microenvironment [60]. Here we show that as a neoadjuvant, DMH1 enhanced acute low doses of Cisplatin and reduced ovarian cancer sphere growth.

One of the primary assays we used to assess the efficacy of DMH1 was the sphere assay. One of the unique characteristics of ovarian cancer cells is their ability to form spheres during the process of metastasis [33, 34]. Therefore, these spheres are crucial to investigate as they play a primary role in disease progression. As shown in Figure 3, BMP inhibition reduced sphere size yet not proliferation or apoptosis, indicating that sphere growth is an important and independent factor to consider, specifically in ovarian cancer studies.

Most notably, DMH1 was able to enhance the response of chemo-resistant cells to treatment. Currently the standard of treatment for ovarian cancer patients is to receive a combination therapy of platinum and taxane based treatments. Resistance to platinum agents, such as Cisplatin, remains a significant source of morbidity [43; 61] and overcoming resistance is a key goal in ovarian cancer treatment [62; 63; 64; 65]. The discovery of new targets mediating resistance and overcoming or reversing resistance to chemotherapy has begun to emerge [35; 44; 66; 67; 68]. Recent gene expression studies have linked not only the BMP pathway but also the Epithelial to Mesenchymal Transition (EMT) process to chemoresistance [43; 61; 69; 70]. The BMP pathway plays an important role in EMT as it not only promotes the reverse process MET, but also antagonizes TGFβ directed EMT [71; 72; 73].

In our studies, we found that dual therapy of DMH1 and Cisplatin altered sphere size and gene expression of chemo-resistant cells in a distinct manner from single-agent therapies (Figure 3A, Figure 4). Sphere size of these cells was only significantly decreased in response to dual treatment. Interestingly, inhibition of BMP signaling in NCI-RES cells was able to upregulate genes that were shown to be downregulated during chemo-resistance and downregulate genes shown to be upregulated during chemo-resistance (Figure 4) indicating the ability of BMP inhibition to affect drug resistance. We saw unique effects of adjuvant therapy specifically on the expression of ROBO2 and HAPLN1 highlighting the distinct, synergistic effect of dual therapy on resistant cells (Figure 4). This may also indicate that ROBO2 and HAPLN1 could act as markers of overcoming resistance due to the fact that only combined significantly affects the gene expression. Our findings suggest that the addition of DMH1 to the current chemotherapeutic regimens may enhance the ability of chemo-resistant cell lines to respond.

We are optimistic that based on our findings, the BMP/TGFβ pathway is a targetable pathway in all ovarian cancer stages, grades and histology and that inhibition of the BMP pathway could positively affect patient response to the current chemotherapeutic agents used. More broadly, our findings show that as large datasets become available, we will be able to more accurately define pathways and signaling molecules that are important to each cancer subtype. As a result we will be able to effectively combine multiple drugs, making substantial improvements to the current therapies. Ultimately we hope that our findings will set the stage for advanced studies with additional cell lines and drug treatments to elucidate the role of BMP signaling in ovarian cancer cells and chemo-resistance.

Supplementary Material

1
2
3

Highlights.

Bone morphogenetic protein (BMP) ligands are overexpressed in ovarian cancer.

BMP receptors increased expression correlate with poor progression-free-survival (PFS).

Small molecule BMP receptor inhibitor DMH1 reduces ovarian cancer cell growth.

DMH1 enhances effect of cisplatin chemotherapy and in cisplatin resistant cell lines.

Acknowledgements

This work was supported by the Department of Defense-Breast Cancer Research Program postdoctoral fellowship awards BC087501 (to P.O.); and National Institutes of Health Grants R01 CA085492 and R01 CA102162, the Robert J. and Helen C. Kleberg Foundation and the T.J. Martell Foundation to HLM. CCH receives support from NIH grant HL10440 and VA Merit grant BX000771. The VUMC Flow Cytometry Shared Resource is supported by the Vanderbilt Ingram Cancer Center (P30 CA68485) and the Vanderbilt Digestive Disease Research Center (DK058404). We would like to acknowledge assistance from Vanderbilt Translational Pathology Shared Resource (TPSR), which is funded by the NIH/NCI Vanderbilt Cancer Center Support Grant (2P30 CA068485-14).

Role of the Funding Source:

The funding provided for these studies had no involvement in the study design, collection of data, analysis and interpretation of the data. The funding sources also did not participate in the writing of the report and the choice of where to submit the study for publication.

Footnotes

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Disclosure of Potential Conflicts of Interest:

The authors have no conflicts of interest to declare.

References

  • 1.Kurman RJ, Shih Ie M. The origin and pathogenesis of epithelial ovarian cancer: a proposed unifying theory. Am J Surg Pathol. 2010;34:433–443. doi: 10.1097/PAS.0b013e3181cf3d79. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 2.Siegel RL, Miller KD, Jemal A. Cancer statistics, 2015. CA Cancer J Clin. 2015;65:5–29. doi: 10.3322/caac.21254. [DOI] [PubMed] [Google Scholar]
  • 3.SEER Stat Fact Sheets: Ovary Cancer (1992–2012. NCI. 2015 [Google Scholar]
  • 4.Peng J, Yoshioka Y, Mandai M, Matsumura N, Baba T, Yamaguchi K, Hamanishi J, Kharma B, Murakami R, Abiko K, Murphy SK, Konishi I. The BMP signaling pathway leads to enhanced proliferation in serous ovarian cancer-a potential therapeutic target. Mol Carcinog. 2015 doi: 10.1002/mc.22283. [DOI] [PubMed] [Google Scholar]
  • 5.Miyazono K, Kamiya Y, Morikawa M. Bone morphogenetic protein receptors and signal transduction. J Biochem. 2010;147:35–51. doi: 10.1093/jb/mvp148. [DOI] [PubMed] [Google Scholar]
  • 6.Chen D, Zhao M, Mundy GR. Bone morphogenetic proteins. Growth Factors. 2004;22:233–241. doi: 10.1080/08977190412331279890. [DOI] [PubMed] [Google Scholar]
  • 7.Singh A, Morris RJ. The Yin and Yang of bone morphogenetic proteins in cancer. Cytokine & growth factor reviews. 2010;21:299–313. doi: 10.1016/j.cytogfr.2010.06.003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 8.Howe JR, Bair JL, Sayed MG, Anderson ME, Mitros FA, Petersen GM, Velculescu VE, Traverso G, Vogelstein B. Germline mutations of the gene encoding bone morphogenetic protein receptor 1A in juvenile polyposis. Nat Genet. 2001;28:184–187. doi: 10.1038/88919. [DOI] [PubMed] [Google Scholar]
  • 9.Kobielak K, Pasolli HA, Alonso L, Polak L, Fuchs E. Defining BMP functions in the hair follicle by conditional ablation of BMP receptor IA. J Cell Biol. 2003;163:609–623. doi: 10.1083/jcb.200309042. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10.Andl T, Ahn K, Kairo A, Chu EY, Wine-Lee L, Reddy ST, Croft NJ, Cebra-Thomas JA, Metzger D, Chambon P, Lyons KM, Mishina Y, Seykora JT, Crenshaw EB, 3rd, Millar SE. Epithelial Bmpr1a regulates differentiation and proliferation in postnatal hair follicles and is essential for tooth development. Development. 2004;131:2257–2268. doi: 10.1242/dev.01125. [DOI] [PubMed] [Google Scholar]
  • 11.Ming Kwan K, Li AG, Wang XJ, Wurst W, Behringer RR. Essential roles of BMPR-IA signaling in differentiation and growth of hair follicles and in skin tumorigenesis. Genesis. 2004;39:10–25. doi: 10.1002/gene.20021. [DOI] [PubMed] [Google Scholar]
  • 12.Yuhki M, Yamada M, Kawano M, Iwasato T, Itohara S, Yoshida H, Ogawa M, Mishina Y. BMPR1A signaling is necessary for hair follicle cycling and hair shaft differentiation in mice. Development. 2004;131:1825–1833. doi: 10.1242/dev.01079. [DOI] [PubMed] [Google Scholar]
  • 13.Owens P, Pickup MW, Novitskiy SV, Chytil A, Gorska AE, Aakre ME, West J, Moses HL. Disruption of bone morphogenetic protein receptor 2 (BMPR2) in mammary tumors promotes metastases through cell autonomous and paracrine mediators. Proc Natl Acad Sci U S A. 2012;109:2814–2819. doi: 10.1073/pnas.1101139108. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Soda H, Raymond E, Sharma S, Lawrence R, Cerna C, Gomez L, Timony GA, Von Hoff DD, Izbicka E. Antiproliferative effects of recombinant human bone morphogenetic protein-2 on human tumor colony-forming units. Anticancer Drugs. 1998;9:327–331. doi: 10.1097/00001813-199804000-00006. [DOI] [PubMed] [Google Scholar]
  • 15.Ghosh-Choudhury N, Ghosh-Choudhury G, Celeste A, Ghosh PM, Moyer M, Abboud SL, Kreisberg J. Bone morphogenetic protein-2 induces cyclin kinase inhibitor p21 and hypophosphorylation of retinoblastoma protein in estradiol-treated MCF-7 human breast cancer cells. Biochim Biophys Acta. 2000;1497:186–196. doi: 10.1016/s0167-4889(00)00060-4. [DOI] [PubMed] [Google Scholar]
  • 16.Ghosh-Choudhury N, Woodruff K, Qi W, Celeste A, Abboud SL, Ghosh Choudhury G. Bone morphogenetic protein-2 blocks MDA MB 231 human breast cancer cell proliferation by inhibiting cyclin-dependent kinase-mediated retinoblastoma protein phosphorylation. Biochem Biophys Res Commun. 2000;272:705–711. doi: 10.1006/bbrc.2000.2844. [DOI] [PubMed] [Google Scholar]
  • 17.Pouliot F, Labrie C. Role of Smad1 and Smad4 proteins in the induction of p21WAF1, Cip1 during bone morphogenetic protein-induced growth arrest in human breast cancer cells. J Endocrinol. 2002;172:187–198. doi: 10.1677/joe.0.1720187. [DOI] [PubMed] [Google Scholar]
  • 18.Kulessa H, Turk G, Hogan BL. Inhibition of Bmp signaling affects growth and differentiation in the anagen hair follicle. The EMBO journal. 2000;19:6664–6674. doi: 10.1093/emboj/19.24.6664. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 19.Gao H, Chakraborty G, Lee-Lim AP, Mo Q, Decker M, Vonica A, Shen R, Brogi E, Brivanlou AH, Giancotti FG. The BMP inhibitor Coco reactivates breast cancer cells at lung metastatic sites. Cell. 2012;150:764–779. doi: 10.1016/j.cell.2012.06.035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20.Ketolainen JM, Alarmo EL, Tuominen VJ, Kallioniemi A. Parallel inhibition of cell growth and induction of cell migration and invasion in breast cancer cells by bone morphogenetic protein 4. Breast Cancer Res Treat. 2010;124:377–386. doi: 10.1007/s10549-010-0808-0. [DOI] [PubMed] [Google Scholar]
  • 21.Katsuno Y, Hanyu A, Kanda H, Ishikawa Y, Akiyama F, Iwase T, Ogata E, Ehata S, Miyazono K, Imamura T. Bone morphogenetic protein signaling enhances invasion and bone metastasis of breast cancer cells through Smad pathway. Oncogene. 2008;27:6322–6333. doi: 10.1038/onc.2008.232. [DOI] [PubMed] [Google Scholar]
  • 22.Morrissey C, Brown LG, Pitts TE, Vessella RL, Corey E. Bone morphogenetic protein 7 is expressed in prostate cancer metastases and its effects on prostate tumor cells depend on cell phenotype and the tumor microenvironment. Neoplasia. 2010;12:192–205. doi: 10.1593/neo.91836. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 23.Ehata S, Yokoyama Y, Takahashi K, Miyazono K. Bi-directional roles of bone morphogenetic proteins in cancer: Another molecular Jekyll and Hyde? Pathol Int. 2013;63:287–296. doi: 10.1111/pin.12067. [DOI] [PubMed] [Google Scholar]
  • 24.Alarmo EL, Kallioniemi A. Bone morphogenetic proteins in breast cancer: dual role in tumourigenesis? Endocr Relat Cancer. 2010;17:R123–R139. doi: 10.1677/ERC-09-0273. [DOI] [PubMed] [Google Scholar]
  • 25.Shepherd TG, Mujoomdar ML, Nachtigal MW. Constitutive activation of BMP signalling abrogates experimental metastasis of OVCA429 cells via reduced cell adhesion. J Ovarian Res. 2010;3:5. doi: 10.1186/1757-2215-3-5. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 26.Edson MA, Nalam RL, Clementi C, Franco HL, Demayo FJ, Lyons KM, Pangas SA, Matzuk MM. Granulosa cell-expressed BMPR1A and BMPR1B have unique functions in regulating fertility but act redundantly to suppress ovarian tumor development. Mol Endocrinol. 2010;24:1251–1266. doi: 10.1210/me.2009-0461. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 27.McLean K, Gong Y, Choi Y, Deng N, Yang K, Bai S, Cabrera L, Keller E, McCauley L, Cho KR, Buckanovich RJ. Human ovarian carcinoma-associated mesenchymal stem cells regulate cancer stem cells and tumorigenesis via altered BMP production. J Clin Invest. 2011;121:3206–3219. doi: 10.1172/JCI45273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 28.Ma W, Ma J, Xu J, Qiao C, Branscum A, Cardenas A, Baron AT, Schwartz P, Maihle NJ, Huang Y. Lin28 regulates BMP4 and functions with Oct4 to affect ovarian tumor microenvironment. Cell Cycle. 2013;12:88–97. doi: 10.4161/cc.23028. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 29.Herrera B, van Dinther M, Ten Dijke P, Inman GJ. Autocrine bone morphogenetic protein-9 signals through activin receptor-like kinase-2/Smad1/Smad4 to promote ovarian cancer cell proliferation. Cancer Res. 2009;69:9254–9262. doi: 10.1158/0008-5472.CAN-09-2912. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 30.Tsai CL, Tsai CN, Lin CY, Chen HW, Lee YS, Chao A, Wang TH, Wang HS, Lai CH. Secreted stress-induced phosphoprotein 1 activates the ALK2-SMAD signaling pathways and promotes cell proliferation of ovarian cancer cells. Cell Rep. 2012;2:283–293. doi: 10.1016/j.celrep.2012.07.002. [DOI] [PubMed] [Google Scholar]
  • 31.Peart TM, Correa RJ, Valdes YR, Dimattia GE, Shepherd TG. BMP signalling controls the malignant potential of ascites-derived human epithelial ovarian cancer spheroids via AKT kinase activation. Clin Exp Metastasis. 2012;29:293–313. doi: 10.1007/s10585-011-9451-3. [DOI] [PubMed] [Google Scholar]
  • 32.Moll F, Millet C, Noel D, Orsetti B, Bardin A, Katsaros D, Jorgensen C, Garcia M, Theillet C, Pujol P, Francois V. Chordin is underexpressed in ovarian tumors and reduces tumor cell motility. FASEB J. 2006;20:240–250. doi: 10.1096/fj.05-4126com. [DOI] [PubMed] [Google Scholar]
  • 33.Theriault BL, Shepherd TG, Mujoomdar ML, Nachtigal MW. BMP4 induces EMT and Rho GTPase activation in human ovarian cancer cells. Carcinogenesis. 2007;28:1153–1162. doi: 10.1093/carcin/bgm015. [DOI] [PubMed] [Google Scholar]
  • 34.Shepherd TG, Theriault BL, Nachtigal MW. Autocrine BMP4 signalling regulates ID3 proto-oncogene expression in human ovarian cancer cells. Gene. 2008;414:95–105. doi: 10.1016/j.gene.2008.02.015. [DOI] [PubMed] [Google Scholar]
  • 35.Wilson AJ, Liu AY, Roland J, Adebayo OB, Fletcher SA, Slaughter JC, Saskowski J, Crispens MA, Jones HW, 3rd, James S, Fadare O, Khabele D. TR3 modulates platinum resistance in ovarian cancer. Cancer Res. 2013;73:4758–4769. doi: 10.1158/0008-5472.CAN-12-4560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36.Owens P, Engelking E, Han G, Haeger SM, Wang XJ. Epidermal Smad4 deletion results in aberrant wound healing. Am J Pathol. 2010;176:122–133. doi: 10.2353/ajpath.2010.090081. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 37.Cerami E, Gao J, Dogrusoz U, Gross BE, Sumer SO, Aksoy BA, Jacobsen A, Byrne CJ, Heuer ML, Larsson E, Antipin Y, Reva B, Goldberg AP, Sander C, Schultz N. The cBio cancer genomics portal: an open platform for exploring multidimensional cancer genomics data. Cancer Discov. 2012;2:401–404. doi: 10.1158/2159-8290.CD-12-0095. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 38.Gyorffy B, Lanczky A, Eklund AC, Denkert C, Budczies J, Li Q, Szallasi Z. An online survival analysis tool to rapidly assess the effect of 22,277 genes on breast cancer prognosis using microarray data of 1,809 patients. Breast Cancer Res Treat. 2010;123:725–731. doi: 10.1007/s10549-009-0674-9. [DOI] [PubMed] [Google Scholar]
  • 39.Burleson KM, Boente MP, Pambuccian SE, Skubitz AP. Disaggregation and invasion of ovarian carcinoma ascites spheroids. J Transl Med. 2006;4:6. doi: 10.1186/1479-5876-4-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 40.Shield K, Ackland ML, Ahmed N, Rice GE. Multicellular spheroids in ovarian cancer metastases: Biology and pathology. Gynecol Oncol. 2009;113:143–148. doi: 10.1016/j.ygyno.2008.11.032. [DOI] [PubMed] [Google Scholar]
  • 41.Liao J, Qian F, Tchabo N, Mhawech-Fauceglia P, Beck A, Qian Z, Wang X, Huss WJ, Lele SB, Morrison CD, Odunsi K. Ovarian cancer spheroid cells with stem cell-like properties contribute to tumor generation, metastasis an d chemotherapy resistance through hypoxia-resistant metabolism. PLoS One. 2014;9 doi: 10.1371/journal.pone.0084941. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 42.Liscovitch M, Ravid D. A case study in misidentification of cancer cell lines: MCF-7/AdrR cells (re-designated NCI/ADR-RES) are derived from OVCAR-8 human ovarian carcinoma cells. Cancer Lett. 2007;245:350–352. doi: 10.1016/j.canlet.2006.01.013. [DOI] [PubMed] [Google Scholar]
  • 43.Li J, Wood WH, 3rd, Becker KG, Weeraratna AT, Morin PJ. Gene expression response to cisplatin treatment in drug-sensitive and drug-resistant ovarian cancer cells. Oncogene. 2007;26:2860–2872. doi: 10.1038/sj.onc.1210086. [DOI] [PubMed] [Google Scholar]
  • 44.Januchowski R, Zawierucha P, Andrzejewska M, Rucinski M, Zabel M. Microarray-based detection and expression analysis of ABC and SLC transporters in drug-resistant ovarian cancer cell lines. Biomed Pharmacother. 2013;67:240–245. doi: 10.1016/j.biopha.2012.11.011. [DOI] [PubMed] [Google Scholar]
  • 45.Sherman-Baust CA, Becker KG, Wood Iii WH, Zhang Y, Morin PJ. Gene expression and pathway analysis of ovarian cancer cells selected for resistance to cisplatin, paclitaxel, or doxorubicin. J Ovarian Res. 2011;4:1757–2215. doi: 10.1186/1757-2215-4-21. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 46.Shepherd TG, Nachtigal MW. Identification of a putative autocrine bone morphogenetic protein-signaling pathway in human ovarian surface epithelium and ovarian cancer cells. Endocrinology. 2003;144:3306–3314. doi: 10.1210/en.2003-0185. [DOI] [PubMed] [Google Scholar]
  • 47.Le Page C, Ouellet V, Madore J, Ren F, Hudson TJ, Tonin PN, Provencher DM, Mes-Masson AM. Gene expression profiling of primary cultures of ovarian epithelial cells identifies novel molecular classifiers of ovarian cancer. Br J Cancer. 2006;94:436–445. doi: 10.1038/sj.bjc.6602933. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 48.Le Page C, Puiffe ML, Meunier L, Zietarska M, de Ladurantaye M, Tonin PN, Provencher D, Mes-Masson AM. BMP-2 signaling in ovarian cancer and its association with poor prognosis. J Ovarian Res. 2009;2:4. doi: 10.1186/1757-2215-2-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 49.Ma Y, Ma L, Guo Q, Zhang S. Expression of bone morphogenetic protein-2 and its receptors in epithelial ovarian cancer and their influence on the prognosis of ovarian cancer patients. J Exp Clin Cancer Res. 2010;29:85. doi: 10.1186/1756-9966-29-85. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 50.Laatio L, Myllynen P, Serpi R, Rysa J, Ilves M, Lappi-Blanco E, Ruskoaho H, Vahakangas K, Puistola U. BMP-4 expression has prognostic significance in advanced serous ovarian carcinoma and is affected by cisplatin in OVCAR-3 cells. Tumour Biol. 2011;32:985–995. doi: 10.1007/s13277-011-0200-7. [DOI] [PubMed] [Google Scholar]
  • 51.Hong CC, Yu PB. Applications of small molecule BMP inhibitors in physiology and disease. Cytokine & growth factor reviews. 2009;20:409–418. doi: 10.1016/j.cytogfr.2009.10.021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 52.Hao J, Ho JN, Lewis JA, Karim KA, Daniels RN, Gentry PR, Hopkins CR, Lindsley CW, Hong CC. In vivo structure-activity relationship study of dorsomorphin analogues identifies selective VEGF and BMP inhibitors. ACS Chem Biol. 2010;5:245–253. doi: 10.1021/cb9002865. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 53.Ao A, Hao J, Hopkins CR, Hong CC. DMH1, a novel BMP small molecule inhibitor, increases cardiomyocyte progenitors and promotes cardiac differentiation in mouse embryonic stem cells. PLoS One. 2012;7:e41627. doi: 10.1371/journal.pone.0041627. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 54.Neely MD, Litt MJ, Tidball AM, Li GG, Aboud AA, Hopkins CR, Chamberlin R, Hong CC, Ess KC, Bowman AB. DMH1, a highly selective small molecule BMP inhibitor promotes neurogenesis of hiPSCs: comparison of PAX6 and SOX1 expression during neural induction. ACS Chem Neurosci. 2012;3:482–491. doi: 10.1021/cn300029t. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 55.Engers DW, Frist AY, Lindsley CW, Hong CC, Hopkins CR. Synthesis and structure-activity relationships of a novel and selective bone morphogenetic protein receptor (BMP) inhibitor derived from the pyrazolo[1.5-a] pyrimidine scaffold of dorsomorphin: the discovery of ML347 as an ALK2 versus ALK3 selective MLPCN probe. Bioorg Med Chem Lett. 2013;23:3248–3252. doi: 10.1016/j.bmcl.2013.03.113. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 56.Langenfeld E, Hong CC, Lanke G, Langenfeld J. Bone morphogenetic protein type I receptor antagonists decrease growth and induce cell death of lung cancer cell lines. PloS one. 2013;8:e61256. doi: 10.1371/journal.pone.0061256. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 57.Nishimori H, Ehata S, Suzuki HI, Katsuno Y, Miyazono K. Prostate cancer cells and bone stromal cells mutually interact with each other through bone morphogenetic protein-mediated signals. J Biol Chem. 2012;287:20037–20046. doi: 10.1074/jbc.M112.353094. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 58.Balboni AL, Hutchinson JA, DeCastro AJ, Cherukuri P, Liby K, Sporn MB, Schwartz GN, Wells WA, Sempere LF, Yu PB, DiRenzo J. DeltaNp63alpha-mediated activation of bone morphogenetic protein signaling governs stem cell activity and plasticity in normal and malignant mammary epithelial cells. Cancer research. 2013;73:1020–1030. doi: 10.1158/0008-5472.CAN-12-2862. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 59.Robson NC, Hidalgo L, McAlpine T, Wei H, Martinez VG, Entrena A, Melen GJ, MacDonald AS, Phythian-Adams A, Sacedon R, Maraskovsky E, Cebon J, Ramirez M, Vicente A, Varas A. Optimal effector functions in human natural killer cells rely upon autocrine bone morphogenetic protein signaling. Cancer Res. 2014;74:5019–5031. doi: 10.1158/0008-5472.CAN-13-2845. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 60.Owens P, Pickup MW, Novitskiy SV, Giltnane JM, Gorska AE, Hopkins CR, Hong CC, Moses HL. Inhibition of BMP signaling suppresses metastasis in mammary cancer. Oncogene. 2014 doi: 10.1038/onc.2014.189. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 61.Miow QH, Tan TZ, Ye J, Lau JA, Yokomizo T, Thiery JP, Mori S. Epithelial-mesenchymal status renders differential responses to cisplatin in ovarian cancer. Oncogene. 2014 doi: 10.1038/onc.2014.136. [DOI] [PubMed] [Google Scholar]
  • 62.Davis A, Tinker AV, Friedlander M. "Platinum resistant" ovarian cancer: what is it, who to treat and how to measure benefit? Gynecol Oncol. 2014;133:624–631. doi: 10.1016/j.ygyno.2014.02.038. [DOI] [PubMed] [Google Scholar]
  • 63.Fraser M, Leung B, Jahani-Asl A, Yan X, Thompson WE, Tsang BK. Chemoresistance in human ovarian cancer: the role of apoptotic regulators. Reprod Biol Endocrinol. 2003;1:66. doi: 10.1186/1477-7827-1-66. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 64.Siddik ZH. Cisplatin: mode of cytotoxic action and molecular basis of resistance. Oncogene. 2003;22:7265–7279. doi: 10.1038/sj.onc.1206933. [DOI] [PubMed] [Google Scholar]
  • 65.Ali AY, Farrand L, Kim JY, Byun S, Suh JY, Lee HJ, Tsang BK. Molecular determinants of ovarian cancer chemoresistance: new insights into an old conundrum. Ann N Y Acad Sci. 2012;1271:58–67. doi: 10.1111/j.1749-6632.2012.06734.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 66.Bowden NA. Nucleotide excision repair: why is it not used to predict response to platinum-based chemotherapy? Cancer Lett. 2014;346:163–171. doi: 10.1016/j.canlet.2014.01.005. [DOI] [PubMed] [Google Scholar]
  • 67.Pavan S, Olivero M, Cora D, Di Renzo MF. IRF-1 expression is induced by cisplatin in ovarian cancer cells and limits drug effectiveness. Eur J Cancer. 2013;49:964–973. doi: 10.1016/j.ejca.2012.09.024. [DOI] [PubMed] [Google Scholar]
  • 68.Helleman J, Jansen MP, Burger C, van der Burg ME, Berns EM. Integrated genomics of chemotherapy resistant ovarian cancer: a role for extracellular matrix, TGFbeta and regulating microRNAs. Int J Biochem Cell Biol. 2010;42:25–30. doi: 10.1016/j.biocel.2009.10.016. [DOI] [PubMed] [Google Scholar]
  • 69.Marchini S, Fruscio R, Clivio L, Beltrame L, Porcu L, Fuso Nerini I, Cavalieri D, Chiorino G, Cattoretti G, Mangioni C, Milani R, Torri V, Romualdi C, Zambelli A, Romano M, Signorelli M, di Giandomenico S, D'Incalci M. Resistance to platinum-based chemotherapy is associated with epithelial to mesenchymal transition in epithelial ovarian cancer. Eur J Cancer. 2013;49:520–530. doi: 10.1016/j.ejca.2012.06.026. [DOI] [PubMed] [Google Scholar]
  • 70.Baribeau S, Chaudhry P, Parent S, Asselin E. Resveratrol inhibits cisplatin-induced epithelial-to-mesenchymal transition in ovarian cancer cell lines. PLoS One. 2014;9:e86987. doi: 10.1371/journal.pone.0086987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 71.Scheel C, Eaton EN, Li SH, Chaffer CL, Reinhardt F, Kah KJ, Bell G, Guo W, Rubin J, Richardson AL, Weinberg RA. Paracrine and autocrine signals induce and maintain mesenchymal and stem cell states in the breast. Cell. 2011;145:926–940. doi: 10.1016/j.cell.2011.04.029. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 72.Samavarchi-Tehrani P, Golipour A, David L, Sung HK, Beyer TA, Datti A, Woltjen K, Nagy A, Wrana JL. Functional genomics reveals a BMP-driven mesenchymal-to-epithelial transition in the initiation of somatic cell reprogramming. Cell Stem Cell. 2010;7:64–77. doi: 10.1016/j.stem.2010.04.015. [DOI] [PubMed] [Google Scholar]
  • 73.Buijs JT, Henriquez NV, van Overveld PG, van der Horst G, ten Dijke P, van der Pluijm G. TGF-beta and BMP7 interactions in tumour progression and bone metastasis. Clin Exp Metastasis. 2007;24:609–617. doi: 10.1007/s10585-007-9118-2. [DOI] [PubMed] [Google Scholar]

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Supplementary Materials

1
NIHMS712461-supplement-1.pdf (1.1MB, pdf)
2
NIHMS712461-supplement-2.xlsx (46.8KB, xlsx)
3
NIHMS712461-supplement-3.pdf (321.6KB, pdf)

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