Publications by authors named "Saraswati Sukumar"

153 Publications

HEYL Regulates Neoangiogenesis Through Overexpression in Both Breast Tumor Epithelium and Endothelium.

Front Oncol 2020 15;10:581459. Epub 2021 Jan 15.

Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD, United States.

Blocking tumor angiogenesis is an appealing therapeutic strategy, but to date, success has been elusive. We previously identified HEYL, a downstream target of Notch signaling, as an overexpressed gene in both breast cancer cells and as a tumor endothelial marker, suggesting that HEYL overexpression in both compartments may contribute to neoangiogenesis. Carcinomas arising in double transgenic Her2-neu/HeyL mice showed higher tumor vessel density and significantly faster growth than tumors in parental Her2/neu mice. Providing mechanistic insight, microarray-based mRNA profiling of HS578T-tet-off-HEYL human breast cancer cells revealed upregulation of several angiogenic factors including CXCL1/2/3 upon HEYL expression, which was validated by RT-qPCR and protein array analysis. Upregulation of the cytokines CXCL1/2/3 occurred through direct binding of HEYL to their promoter sequences. We found that vessel growth and migration of human vascular endothelial cells (HUVECs) was promoted by conditioned medium from HS578T-tet-off-HEYL carcinoma cells, but was blocked by neutralizing antibodies against CXCL1/2/3. Supporting these findings, suppressing HEYL expression using shRNA in MDA-MB-231 cells significantly reduced tumor growth. In addition, suppressing the action of proangiogenic cytokines induced by HEYL using a small molecule inhibitor of the CXCl1/2/3 receptor, CXCR2, in combination with the anti-VEGF monoclonal antibody, bevacizumab, significantly reduced tumor growth of MDA-MB-231 xenografts. Thus, HEYL expression in tumor epithelium has a profound effect on the vascular microenvironment in promoting neoangiogenesis. Furthermore, we show that lack of HEYL expression in endothelial cells leads to defects in neoangiogenesis, both under normal physiological conditions and in cancer. Thus, HeyL-/- mice showed impaired vessel outgrowth in the neonatal retina, while the growth of mammary tumor cells E0771 was retarded in syngeneic HeyL-/- mice compared to wild type C57/Bl6 mice. Blocking HEYL's angiogenesis-promoting function in both tumor cells and tumor-associated endothelium may enhance efficacy of therapy targeting the tumor vasculature in breast cancer.
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http://dx.doi.org/10.3389/fonc.2020.581459DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7845423PMC
January 2021

HOX genes and the NF-κB pathway: A convergence of developmental biology, inflammation and cancer biology.

Biochim Biophys Acta Rev Cancer 2020 12 10;1874(2):188450. Epub 2020 Oct 10.

Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD, United States of America. Electronic address:

The roles of HOX transcription factors as oncogenes and tumor suppressor genes, and the NF-KB pathway in chronic inflammation, both leading to cancer are well-established. HOX transcription factors are members of an evolutionarily conserved family of proteins required for anteroposterior body axis patterning during embryonic development, and are often dysregulated in cancer. The NF-KB pathway aids inflammation and immunity but it is also important during embryonic development. It is frequently activated in both solid and hematological malignancies. NF-KB and HOX proteins can influence each other through mutual transcriptional regulation, protein-protein interactions, and regulation of upstream and downstream interactors. These interactions have important implications both in homeostasis and in disease. In this review, we summarize the role of HOX proteins in regulating inflammation in homeostasis and disease- with a particular emphasis on cancer. We also describe the relationship between HOX genes and the NF-KB pathway, and discuss potential therapeutic strategies.
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http://dx.doi.org/10.1016/j.bbcan.2020.188450DOI Listing
December 2020

Breast-Specific Epigenetic Regulation of DeltaNp73 and Its Role in DNA-Damage-Response of -Mutated Human Mammary Epithelial Cells.

Cancers (Basel) 2020 Aug 21;12(9). Epub 2020 Aug 21.

Department of Oncology, Kaplan Medical Center, Rehovot and the Faculty of Medicine, The Hebrew University, Jerusalem 7661041, Israel.

The function of BRCA1/2 proteins is essential for maintaining genomic integrity in all cell types. However, why women who carry deleterious germline mutations in face an extremely high risk of developing breast and ovarian cancers specifically has remained an enigma. We propose that breast-specific epigenetic modifications, which regulate tissue differentiation, could team up with BRCA deficiency and affect tissue susceptibility to cancer. In earlier work, we compared genome-wide methylation profiles of various normal epithelial tissues and identified breast-specific methylated gene promoter regions. Here, we focused on deltaNp73, the truncated isoform of p73, which possesses antiapoptotic and pro-oncogenic functions. We showed that the promoter of is unmethylated in normal human breast epithelium and methylated in various other normal epithelial tissues and cell types. Accordingly, deltaNp73 was markedly induced by DNA damage in human mammary epithelial cells (HMECs) but not in other epithelial cell types. Moreover, the induction of deltaNp73 protected HMECs from DNA damage-induced cell death, and this effect was more substantial in HMECs from mutation carriers. Notably, when BRCA1 was knocked down in MCF10A, a non-malignant breast epithelial cell line, both deltaNp73 induction and its protective effect from cell death were augmented upon DNA damage. Interestingly, deltaNp73 induction also resulted in inhibition of BRCA1 and BRCA2 expression following DNA damage. In conclusion, breast-specific induction of deltaNp73 promotes survival of BRCA1-deficient mammary epithelial cells upon DNA damage. This might result in the accumulation of genomic alterations and allow the outgrowth of breast cancers. These findings indicate deltaNp73 as a potential modifier of breast cancer susceptibility in mutation carriers and may stimulate novel strategies of prevention and treatment for these high-risk women.
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http://dx.doi.org/10.3390/cancers12092367DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7564633PMC
August 2020

Intraductal Therapy in Breast Cancer: Current Status and Future Prospective.

J Mammary Gland Biol Neoplasia 2020 06 23;25(2):133-143. Epub 2020 Jun 23.

Department of Breast and Thyroid Surgery, Renmin Hospital of Wuhan University, No 238 Jiefang Road, Wuchang District, Wuhan, Hubei, 430060, People's Republic of China.

With our improved understanding of the biological behavior of breast cancer, minimally invasive intervention is urgently needed for personalized treatment of early disease. Intraductal therapy is one such minimally invasive approach. With the help of appropriate tools, technologies using the intraductal means of entering the ducts may be used both to diagnose and treat lesions in the mammary duct system with less trauma and at the same time avoid systemic toxicity. Traditional agents such as those targeting pathways, endocrine therapy, immunotherapy, or gene therapy can be used alone or combined with other new technologies, such as nanomaterials, through the intraductal route. Additionally, relevant mammary tumor models in rodents which reflect changes in the tumor microenvironment will help deepen our understanding of their biological behavior and heterogeneity. This article reviews the current status and future prospects of intraductal therapy in breast cancer, with emphasis on ductal carcinoma in situ.
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http://dx.doi.org/10.1007/s10911-020-09453-4DOI Listing
June 2020

Multiple roles of HOX proteins in Metastasis: Let me count the ways.

Cancer Metastasis Rev 2020 09;39(3):661-679

Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Department of Oncology, Johns Hopkins University School of Medicine, 1650 Orleans Street, CRB1-143, Baltimore, MD, 21231, USA.

Knowledge of the role of HOX proteins in cancer has been steadily accumulating in the last 25 years. They are encoded by 39 HOX genes arranged in 4 distinct clusters, and have unique and redundant function in all types of cancers. Many HOX genes behave as oncogenic transcriptional factors regulating multiple pathways that are critical to malignant progression in a variety of tumors. Some HOX proteins have dual roles that are tumor-site specific, displaying both oncogenic and tumor suppressor function. The focus of this review is on how HOX proteins contribute to growth or suppression of metastasis. The review will cover HOX protein function in the critical aspects of epithelial-mesenchymal transition, in cancer stem cell sustenance and in therapy resistance, manifested as distant metastasis. The emerging role of adiposity in both initiation and progression of metastasis is described. Defining the role of HOX genes in the metastatic process has identified candidates for targeted cancer therapies that may combat the metastatic process. We will discuss potential therapeutic opportunities, particularly in pathways influenced by HOX proteins.
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http://dx.doi.org/10.1007/s10555-020-09908-4DOI Listing
September 2020

Unpredicted central inversion in a sgRNA flanked by inverted repeats.

Mol Biol Rep 2020 Aug 18;47(8):6375-6378. Epub 2020 May 18.

Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA.

In genome engineering, sgRNAs define the genomic target to be modified in CRISPR/Cas9 system. Either for single gene editing or genome-wide screens, sgRNAs are cloned into plasmid vectors. During our performance of CRISPR/Cas9 gene knock out, we found that the central part of a sgRNA was inverted after transformation into Escherichia coli. Interestingly, the inverted portion was found to be flanked by inverted repeats, and sealing of nicks inside the plasmid could correct the inversion. This type of sgRNA recombination completely changed its original sequence and should be noted during sgRNA design and performance of CRISPR/Cas9.
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http://dx.doi.org/10.1007/s11033-020-05524-1DOI Listing
August 2020

Nanoparticle interactions with immune cells dominate tumor retention and induce T cell-mediated tumor suppression in models of breast cancer.

Sci Adv 2020 03 25;6(13):eaay1601. Epub 2020 Mar 25.

Department of Radiation Oncology and Molecular Radiation Sciences, School of Medicine, Johns Hopkins University, Baltimore, MD 21231, USA.

The factors that influence nanoparticle fate in vivo following systemic delivery remain an area of intense interest. Of particular interest is whether labeling with a cancer-specific antibody ligand ("active targeting") is superior to its unlabeled counterpart ("passive targeting"). Using models of breast cancer in three immune variants of mice, we demonstrate that intratumor retention of antibody-labeled nanoparticles was determined by tumor-associated dendritic cells, neutrophils, monocytes, and macrophages and not by antibody-antigen interactions. Systemic exposure to either nanoparticle type induced an immune response leading to CD8 T cell infiltration and tumor growth delay that was independent of antibody therapeutic activity. These results suggest that antitumor immune responses can be induced by systemic exposure to nanoparticles without requiring a therapeutic payload. We conclude that immune status of the host and microenvironment of solid tumors are critical variables for studies in cancer nanomedicine and that nanoparticle technology may harbor potential for cancer immunotherapy.
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http://dx.doi.org/10.1126/sciadv.aay1601DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7096167PMC
March 2020

Targeted nanopore sequencing with Cas9-guided adapter ligation.

Nat Biotechnol 2020 04 10;38(4):433-438. Epub 2020 Feb 10.

Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, USA.

Despite recent improvements in sequencing methods, there remains a need for assays that provide high sequencing depth and comprehensive variant detection. Current methods are limited by the loss of native modifications, short read length, high input requirements, low yield or long protocols. In the present study, we describe nanopore Cas9-targeted sequencing (nCATS), an enrichment strategy that uses targeted cleavage of chromosomal DNA with Cas9 to ligate adapters for nanopore sequencing. We show that nCATS can simultaneously assess haplotype-resolved single-nucleotide variants, structural variations and CpG methylation. We apply nCATS to four cell lines, to a cell-line-derived xenograft, and to normal and paired tumor/normal primary human breast tissue. Median sequencing coverage was 675× using a MinION flow cell and 34× using the smaller Flongle flow cell. The nCATS sequencing requires only ~3 μg of genomic DNA and can target a large number of loci in a single reaction. The method will facilitate the use of long-read sequencing in research and in the clinic.
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http://dx.doi.org/10.1038/s41587-020-0407-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7145730PMC
April 2020

DNA methylation markers predict recurrence-free interval in triple-negative breast cancer.

NPJ Breast Cancer 2020 31;6. Epub 2020 Jan 31.

1Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD 21205 USA.

We lack tools to risk-stratify triple-negative breast cancer (TNBC). Our goal was to develop molecular tools to predict disease recurrence. Methylation array analysis was performed on 110 samples treated by locoregional therapy obtained from institutional cohorts. Discovered marker sets were then tested by Kaplan-Meier analyses in a prospectively collected TNBC cohort of 49 samples from the no-chemotherapy arms of IBCSG trials VIII and IX, and by logistic regression in a chemotherapy-treated cohort of 121 TNBCs from combined IBCSG trials and institutional repositories. High methylation was associated with shorter recurrence-free interval in the no-chemotherapy arm of the IBCSG studies, as well as in the chemotherapy-treated patients within the combined institutional and IBCSG chemotherapy cohorts (100 marker panel,  = 0.002; 30 marker panel,  = 0.05). Chromosome 19 sites were enriched among these loci. In conclusion, our hypermethylation signatures identify increased recurrence risk independent of whether patients receive chemotherapy.
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http://dx.doi.org/10.1038/s41523-020-0145-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6994477PMC
January 2020

Discovery of a Potent GLUT Inhibitor from a Library of Rapafucins by Using 3D Microarrays.

Angew Chem Int Ed Engl 2019 11 31;58(48):17158-17162. Epub 2019 Oct 31.

Department of Pharmacology and Molecular Sciences, The SJ Yan and HJ Mao Laboratory of Chemical Biology, Johns Hopkins University School of Medicine, Room 516, Hunterian Building, 725 N. Wolfe Street, Baltimore, MD, USA.

Glucose transporters play an essential role in cancer cell proliferation and survival and have been pursued as promising cancer drug targets. Using microarrays of a library of new macrocycles known as rapafucins, which were inspired by the natural product rapamycin, we screened for new inhibitors of GLUT1. We identified multiple hits from the rapafucin 3D microarray and confirmed one hit as a bona fide GLUT1 ligand, which we named rapaglutin A (RgA). We demonstrate that RgA is a potent inhibitor of GLUT1 as well as GLUT3 and GLUT4, with an IC value of low nanomolar for GLUT1. RgA was found to inhibit glucose uptake, leading to a decrease in cellular ATP synthesis, activation of AMP-dependent kinase, inhibition of mTOR signaling, and induction of cell-cycle arrest and apoptosis in cancer cells. Moreover, RgA was capable of inhibiting tumor xenografts in vivo without obvious side effects. RgA could thus be a new chemical tool to study GLUT function and a promising lead for developing anticancer drugs.
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http://dx.doi.org/10.1002/anie.201905578DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6861656PMC
November 2019

Perturbed myoepithelial cell differentiation in BRCA mutation carriers and in ductal carcinoma in situ.

Nat Commun 2019 09 13;10(1):4182. Epub 2019 Sep 13.

Department of Medical Oncology, Dana-Farber Cancer Institute Boston, Boston, MA, 02215, USA.

Myoepithelial cells play key roles in normal mammary gland development and in limiting pre-invasive to invasive breast tumor progression, yet their differentiation and perturbation in ductal carcinoma in situ (DCIS) are poorly understood. Here, we investigated myoepithelial cells in normal breast tissues of BRCA1 and BRCA2 germline mutation carriers and in non-carrier controls, and in sporadic DCIS. We found that in the normal breast of non-carriers, myoepithelial cells frequently co-express the p63 and TCF7 transcription factors and that p63 and TCF7 show overlapping chromatin peaks associated with differentiated myoepithelium-specific genes. In contrast, in normal breast tissues of BRCA1 mutation carriers the frequency of p63TCF7 myoepithelial cells is significantly decreased and p63 and TCF7 chromatin peaks do not overlap. These myoepithelial perturbations in normal breast tissues of BRCA1 germline mutation carriers may play a role in their higher risk of breast cancer. The fraction of p63TCF7 myoepithelial cells is also significantly decreased in DCIS, which may be associated with invasive progression.
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http://dx.doi.org/10.1038/s41467-019-12125-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6744561PMC
September 2019

DNA Methylation Markers for Breast Cancer Detection in the Developing World.

Clin Cancer Res 2019 11 12;25(21):6357-6367. Epub 2019 Jul 12.

Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, Maryland.

Purpose: An unmet need in low-resource countries is an automated breast cancer detection assay to prioritize women who should undergo core breast biopsy and pathologic review. Therefore, we sought to identify and validate a panel of methylated DNA markers to discriminate between cancer and benign breast lesions using cells obtained by fine-needle aspiration (FNA). Two case-control studies were conducted comparing cancer and benign breast tissue identified from clinical repositories in the United States, China, and South Africa for marker selection/training ( = 226) and testing ( = 246). Twenty-five methylated markers were assayed by Quantitative Multiplex-Methylation-Specific PCR (QM-MSP) to select and test a cancer-specific panel. Next, a pilot study was conducted on archival FNAs (49 benign, 24 invasive) from women with mammographically suspicious lesions using a newly developed, 5-hour, quantitative, automated cartridge system. We calculated sensitivity, specificity, and area under the receiver-operating characteristic curve (AUC) compared with histopathology for the marker panel.

Results: In the discovery cohort, 10 of 25 markers were selected that were highly methylated in breast cancer compared with benign tissues by QM-MSP. In the independent test cohort, this panel yielded an AUC of 0.937 (95% CI = 0.900-0.970). In the FNA pilot, we achieved an AUC of 0.960 (95% CI = 0.883-1.0) using the automated cartridge system.

Conclusions: We developed and piloted a fast and accurate methylation marker-based automated cartridge system to detect breast cancer in FNA samples. This quick ancillary test has the potential to prioritize cancer over benign tissues for expedited pathologic evaluation in poorly resourced countries.
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http://dx.doi.org/10.1158/1078-0432.CCR-18-3277DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6825533PMC
November 2019

Validation of a low-cost, carbon dioxide-based cryoablation system for percutaneous tumor ablation.

PLoS One 2019 3;14(7):e0207107. Epub 2019 Jul 3.

Department of Biomedical Engineering, Johns Hopkins University, Baltimore, Maryland, United States of America.

Breast cancer rates are rising in low- and middle-income countries (LMICs), yet there is a lack of accessible and cost-effective treatment. As a result, the cancer burden and death rates are highest in LMICs. In an effort to meet this need, our work presents the design and feasibility of a low-cost cryoablation system using widely-available carbon dioxide as the only consumable. This system uses an 8-gauge outer-diameter needle and Joule-Thomson expansion to percutaneously necrose tissue with cryoablation. Bench top experiments characterized temperature dynamics in ultrasound gel demonstrated that isotherms greater than 2 cm were formed. Further, this system was applied to mammary tumors in an in vivo rat model and necrosis was verified by histopathology. Finally, freezing capacity under a large heat load was assessed with an in vivo porcine study, where volumes of necrosis greater than 1.5 cm in diameter confirmed by histopathology were induced in a highly perfused liver after two 7-minute freeze cycles. These results demonstrate the feasibility of a carbon-dioxide based cryoablation system for improving solid tumor treatment options in resource-constrained environments.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0207107PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6608927PMC
February 2020

Intraductal fulvestrant for therapy of ERα-positive ductal carcinoma in situ of the breast: a preclinical study.

Carcinogenesis 2019 07;40(7):903-913

Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD, USA.

Mammographic screening for breast cancer has led to increased detection of ductal carcinoma in situ (DCIS) and a reappraisal of the necessity of aggressive treatment with their attendant toxicities for a preneoplastic lesion. Fulvestrant, a selective estrogen receptor degrader, is very effective in the treatment of estrogen receptor positive (ER+) breast cancer, but delivery by the painful intramuscular (i.m) route is limiting. We hypothesized that intraductal (i.duc) administration of fulvestrant will provide a direct, safe and effective treatment for DCIS. Mice bearing mammary ductal xenografts of ER+, luciferase-tagged MCF-7 breast cancer cells were administered vehicle or fulvestrant i.m or i.duc. I.duc MCF-7-luc tumors in mice treated with fulvestrant i.duc or i.m grew significantly slower than vehicle control. Whole mount analysis and histopathology showed that i.duc fulvestrant achieved significantly larger cancer-free areas. Western blot analysis showed reduced levels of estrogen receptor alpha (ERα) and its downstream targets, c-Myc and Cyclin D1, and increased levels of ERβ, which is known to inhibit ERα function. Immunohistochemical analysis of tumor sections showed that Ki67 and ERα protein levels decreased by 3-fold, and neoangiogenesis was inhibited by i.duc fulvestrant treatment. I.duc fulvestrant also reduced outgrowth of ERα+, autochthonous N-methyl-N-nitrosourea-induced mammary tumors in rats. Overall, we have shown that i.duc fulvestrant was significantly more effective than, or equivalent in action to i.m fulvestrant in two preclinical models of breast cancer. These studies provide evidence for a novel and safe route for fulvestrant therapy of DCIS and prevention of breast cancer. This preclinical study provides a strong basis for conducting clinical trials for DCIS and early breast cancer.
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http://dx.doi.org/10.1093/carcin/bgz084DOI Listing
July 2019

A biologic scaffold-associated type 2 immune microenvironment inhibits tumor formation and synergizes with checkpoint immunotherapy.

Sci Transl Med 2019 01;11(477)

Translational Tissue Engineering Center, Baltimore, MD 21231, USA.

Biomaterials in regenerative medicine are designed to mimic and modulate tissue environments to promote repair. Biologic scaffolds (derived from decellularized tissue extracellular matrix) promote a wound-healing (proregenerative) immune phenotype and are used clinically to treat tissue loss, including in the context of tumor resection. It is unknown whether a biomaterial microenvironment that encourages tissue formation may also promote tumor development. We implanted a urinary bladder matrix (UBM) scaffold, which is used clinically for wound management, with syngeneic cancer cell lines in mice to study how wound-healing immune responses affect tumor formation and sensitivity to immune checkpoint blockade. The UBM scaffold created an immune microenvironment that inhibited B16-F10 melanoma tumor formation in a CD4 T cell-dependent and macrophage-dependent manner. In-depth immune characterization revealed an activated type 2-like immune response that was distinct from the classical tumor microenvironment, including activated type 2 T helper T cells, a unique macrophage phenotype, eosinophil infiltration, angiogenic factors, and complement. Tumor growth inhibition by PD-1 and PD-L1 checkpoint blockade was potentiated in the UBM scaffold immune microenvironment. Engineering the local tumor microenvironment to promote a type 2 wound-healing immune signature may serve as a therapeutic target to improve immunotherapy efficacy.
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http://dx.doi.org/10.1126/scitranslmed.aat7973DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7254933PMC
January 2019

Induction of cell cycle arrest and inflammatory genes by combined treatment with epigenetic, differentiating, and chemotherapeutic agents in triple-negative breast cancer.

Breast Cancer Res 2018 11 28;20(1):145. Epub 2018 Nov 28.

Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD, USA.

Background: A combination of entinostat, all-trans retinoic acid, and doxorubicin (EAD) induces cell death and differentiation and causes significant regression of xenografts of triple-negative breast cancer (TNBC).

Methods: We investigated the mechanisms underlying the antitumor effects of each component of the EAD combination therapy by high-throughput gene expression profiling of drug-treated cells.

Results: Microarray analysis showed that entinostat and doxorubicin (ED) altered expression of genes related to growth arrest, inflammation, and differentiation. ED downregulated MYC, E2F, and G2M cell cycle genes. Accordingly, entinostat sensitized the cells to doxorubicin-induced growth arrest at G2. ED induced interferon genes, which correlated with breast tumors containing a higher proportion of tumor-infiltrating lymphocytes. ED also increased the expression of immune checkpoint agonists and cancer testis antigens. Analysis of TNBC xenografts showed that EAD enhanced the inflammation score in nude mice. Among the genes differentially regulated between the EAD and ED groups, an all-trans retinoic acid (ATRA)-regulated gene, DHRS3, was induced in EAD-treated xenografts. DHRS3 was expressed at lower levels in human TNBC metastases compared to normal breast or primary tumors. High expression of ED-induced growth arrest and inflammatory genes was associated with better prognosis in TNBC patients.

Conclusions: Entinostat potentiated doxorubicin-mediated cell death and the combination induced inflammatory signatures. The ED-induced immunomodulation may improve immunotherapy. Addition of ATRA to ED may potentiate inflammation and contribute to TNBC regression.
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http://dx.doi.org/10.1186/s13058-018-1068-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6263070PMC
November 2018

Breast Hormone Concentrations in Random Fine-Needle Aspirates of Healthy Women Associate with Cytological Atypia and Gene Methylation.

Cancer Prev Res (Phila) 2018 09 28;11(9):557-568. Epub 2018 Jun 28.

Department of Surgery, Northwestern University Feinberg School of Medicine, Chicago, Illinois.

Sex steroid hormones contribute to breast cancer development, but data on concentrations of these within breast tissue are limited. We performed simultaneous multiparameter measurement of breast sex steroids, breast epithelial cytology, and DNA methylation in 119 healthy women (54 pre- and 65 postmenopausal) without a history of breast cancer. Random fine-needle aspiration (rFNA) of the breast was performed simultaneously with blood collection. Breast samples were analyzed by LC/MS-MS for estrone, estradiol, progesterone, androstenedione, and testosterone. Blood samples were assayed for estradiol and progesterone by immunoassay. Cytomorphology was classified using the Masood Score, and DNA methylation of eight genes was analyzed using quantitative multiplexed methylation-specific PCR, and expressed as the cumulative methylation index (CMI). Serum and breast concentrations of estradiol and progesterone showed significant correlation (Spearman = 0.34, = 0.001 and = 0.69, < 0.0006, respectively). Progesterone concentration was significantly higher in the premenopausal breast ( < 0.0008), and showed a luteal surge. Breast estrone and estradiol concentrations did not differ significantly by menopause, but androstenedione concentration was higher in the breasts of postmenopausal women ( = 0.026 and = 0.208). Breast androgens were significantly correlated with breast density (Spearman = 0.27, = 0.02 for testosterone) and CMI (Spearman = 0.3, = 0.038 for androstenedione). Our data indicate that future larger studies of breast steroid hormones along with other parameters are feasible. Significant associations of breast androgen concentrations with breast density and gene methylation warrant future study. .
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http://dx.doi.org/10.1158/1940-6207.CAPR-17-0323DOI Listing
September 2018

Quantitative phosphoproteomic analysis reveals reciprocal activation of receptor tyrosine kinases between cancer epithelial cells and stromal fibroblasts.

Clin Proteomics 2018 15;15:21. Epub 2018 Jun 15.

1Department of Biological Chemistry, Johns Hopkins University, Baltimore, MD USA.

Background: Cancer-associated fibroblasts (CAFs) are one of the most important components of tumor stroma and play a key role in modulating tumor growth. However, a mechanistic understanding of how CAFs communicate with tumor cells to promote their proliferation and invasion is far from complete. A major reason for this is that most current techniques and model systems do not capture the complexity of signal transduction that occurs between CAFs and tumor cells.

Methods: In this study, we employed a stable isotope labeling with amino acids in cell culture (SILAC) strategy to label invasive breast cancer cells, MDA-MB-231, and breast cancer patient-derived CAF this has already been defined above cells. We used an antibody-based phosphotyrosine peptide enrichment method coupled to LC-MS/MS to catalog and quantify tyrosine phosphorylation-mediated signal transduction events induced by the bidirectional communication between patient-derived CAFs and tumor cells.

Results: We discovered that distinct signaling events were activated in CAFs and in tumor epithelial cells during the crosstalk between these two cell types. We identified reciprocal activation of a number of receptor tyrosine kinases including EGFR, FGFR1 and EPHA2 induced by this bidirectional communication.

Conclusions: Our study not only provides insights into the mechanisms of the interaction between CAFs and tumor cells, but the model system described here could be used as a prototype for analysis of intercellular communication in many different tumor microenvironments.
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http://dx.doi.org/10.1186/s12014-018-9197-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6003199PMC
June 2018

Mutational profiles of breast cancer metastases from a rapid autopsy series reveal multiple evolutionary trajectories.

JCI Insight 2017 12 21;2(24). Epub 2017 Dec 21.

Department of Oncology, The Sidney Kimmel Comprehensive Cancer Center, and.

Heterogeneity within and among tumors in a metastatic cancer patient is a well-established phenomenon that may confound treatment and accurate prognosis. Here, we used whole-exome sequencing to survey metastatic breast cancer tumors from 5 patients in a rapid autopsy program to construct the origin and genetic development of metastases. Metastases were obtained from 5 breast cancer patients using a rapid autopsy protocol and subjected to whole-exome sequencing. Metastases were evaluated for sharing of somatic mutations, correlation of copy number variation and loss of heterozygosity, and genetic similarity scores. Pathological features of the patients' disease were assessed by immunohistochemical analyses. Our data support a monoclonal origin of metastasis in 3 cases, but in 2 cases, metastases arose from at least 2 distinct subclones in the primary tumor. In the latter 2 cases, the primary tumor presented with mixed histologic and pathologic features, suggesting early divergent evolution within the primary tumor with maintenance of metastatic capability in multiple lineages. We used genetic and histopathological evidence to demonstrate that metastases can be derived from a single or multiple independent clones within a primary tumor. This underscores the complexity of breast cancer clonal evolution and has implications for how best to determine and implement therapies for early- and late-stage disease.
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http://dx.doi.org/10.1172/jci.insight.96896DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5752302PMC
December 2017

Quantitation of DNA Methylation by Quantitative Multiplex Methylation-Specific PCR (QM-MSP) Assay.

Methods Mol Biol 2018 ;1708:473-496

Breast and Ovarian Cancer Program, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, 1650 Orleans St., CRB1, RM 143, Baltimore, MD, 21321, USA.

The defining feature of the Quantitative Multiplex Methylation-Specific PCR (QM-MSP) method to sensitively quantify DNA methylation is the two-step PCR approach for a multiplexed analysis of a panel of up to 12 genes in clinical samples with minimal quantities of DNA. In the first step, for up to 12 genes tested, one pair of gene-specific primers (forward and reverse) amplifies the methylated and unmethylated copies of the same gene simultaneously and in multiplex, in one PCR reaction. This methylation-independent amplification step produces amplicons of up to 10 copies per μL after 36 cycles of PCR. In the second step, the amplicons of the first reaction (STEP 1) are quantified with a standard curve using real-time PCR and two independent fluorophores to detect methylated/unmethylated DNA of each gene in the same well (e.g., 6FAM and VIC). One methylated copy is detectable in 100,000 reference gene copies. Methylation is reported on a continuous scale. For the gene panel, the highest level of normal DNA methylation above which a sample would be called positive is derived by using Receiver Operating Characteristic (ROC), maximizing assay specificity and sensitivity to distinguish between normal/benign versus tumor DNA. QM-MSP can be applied to clinical samples of fresh or fixed ductal cells, ductal fluid, nipple fluid, fine needle aspirates, core biopsies, and tumor tissue sections.
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http://dx.doi.org/10.1007/978-1-4939-7481-8_24DOI Listing
July 2018

Tumor and serum DNA methylation in women receiving preoperative chemotherapy with or without vorinostat in TBCRC008.

Breast Cancer Res Treat 2018 01 16;167(1):107-116. Epub 2017 Sep 16.

Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins School of Medicine, 1650 Orleans St., Baltimore, MD, 21287, USA.

Background: Methylated gene markers have shown promise in predicting breast cancer outcomes and treatment response. We evaluated whether baseline and changes in tissue and serum methylation levels would predict pathological complete response (pCR) in patients with HER2-negative early breast cancer undergoing preoperative chemotherapy.

Methods: The TBCRC008 trial investigated pCR following 12 weeks of preoperative carboplatin and albumin-bound paclitaxel + vorinostat/placebo (n = 62). We measured methylation of a 10-gene panel by quantitative multiplex methylation-specific polymerase chain reaction and expressed results as cumulative methylation index (CMI). We evaluated association between CMI level [baseline, day 15 (D15), and change] and pCR using univariate and multivariable logistic regression models controlling for treatment and hormone receptor (HR) status, and performed exploratory subgroup analyses.

Results: In univariate analysis, one log unit increase in tissue CMI levels at D15 was associated with 40% lower chance of obtaining pCR (odds ratio, OR 0.60, 95% CI 0.37-0.97; p = 0.037). Subgroup analyses suggested a significant association between tissue D15 CMI levels and pCR in vorinostat-treated [OR 0.44 (0.20, 0.93), p = 0.03], but not placebo-treated patients.

Conclusion: In this study investigating the predictive roles of tissue and serum CMI levels in patients with early breast cancer for the first time, we demonstrate that high D15 tissue CMI levels may predict poor response. Larger studies and improved analytical procedures to detect methylated gene markers in early stage breast cancer are needed. TBCRC008 is registered on ClinicalTrials.gov (NCT00616967).
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http://dx.doi.org/10.1007/s10549-017-4503-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5793219PMC
January 2018

Inhibition of platelet function using liposomal nanoparticles blocks tumor metastasis.

Theranostics 2017 26;7(5):1062-1071. Epub 2017 Feb 26.

CAS Key Laboratory for Biomedical Effects of Nanomaterials & Nanosafety, CAS Center for Excellence in Nanoscience, National Center for Nanoscience and Technology, Beijing 100190, China.

Extensive evidence has shown that platelets support tumor metastatic progression by inducing epithelial-mesenchymal transition of cancer cells and by shielding circulating tumor cells from immune-mediated elimination. Therefore, blocking platelet function represents a potential new avenue for therapy focused on eliminating metastasis. Here we show that liposomal nanoparticles bearing the tumor-homing pentapeptide CREKA (Cys-Arg-Glu-Lys-Ala) can deliver a platelet inhibitor, ticagrelor, into tumor tissues to specifically inhibit tumor-associated platelets. The drug-loaded nanoparticles (CREKA-Lipo-T) efficiently blocked the platelet-induced acquisition of an invasive phenotype by tumor cells and inhibited platelet-tumor cell interaction . Intravenously administered CREKA-Lipo-T effectively targeted tumors within 24 h, and inhibited tumor metastasis without overt side effects. Thus, the CREKA-Lipo formulation provides a simple strategy for the efficient delivery of anti-metastatic drugs and shows considerable promise as a platform for novel cancer therapeutics.
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http://dx.doi.org/10.7150/thno.17908DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5399576PMC
January 2018

Inhibitors of STAT3, β-catenin, and IGF-1R sensitize mouse PIK3CA-mutant breast cancer to PI3K inhibitors.

Mol Oncol 2017 05 6;11(5):552-566. Epub 2017 Apr 6.

Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, MD, USA.

Although mutations in the phosphoinositide 3-kinase catalytic subunit (PIK3CA) are common in breast cancer, PI3K inhibitors alone have shown modest efficacy. We sought to identify additional pathways altered in PIK3CA-mutant tumors that might be targeted in combination with PI3K inhibitors. We generated two transgenic mouse models expressing the human PIK3CA-H1047R- and the -E545K hotspot-mutant genes in the mammary gland and evaluated their effects on development and tumor formation. Molecular analysis identified pathways altered in these mutant tumors, which were also targeted in multiple cell lines derived from the PIK3CA tumors. Finally, public databases were analyzed to determine whether novel pathways identified in the mouse tumors were altered in human tumors harboring mutant PIK3CA. Mutant mice showed increased branching and delayed involution of the mammary gland compared to parental FVB/N mice. Mammary tumors arose in 30% of the MMTV-PIK3CA-H1047R and in 13% of -E545K mice. Compared to MMTV-Her-2 transgenic mouse mammary tumors, H1047R tumors showed increased upregulation of Wnt/β-catenin/Axin2, hepatocyte growth factor (Hgf)/Stat3, insulin-like growth factor 2 (Igf-2), and Igf-1R pathways. Inhibitors of STAT3, β-catenin, and IGF-1R sensitized H1047R-derived mouse tumor cells and PIK3CA-H1047R overexpressing human HS578T breast cancer cells to the cytotoxic effects of PI3K inhibitors. Analysis of The Cancer Genome Atlas database showed that, unlike primary PIK3CA-wild-type and HER-2 breast carcinomas, PIK3CA-mutant tumors display increased expression of AXIN2, HGF, STAT3, IGF-1, and IGF-2 mRNA and activation of AKT, IGF1-MTOR, and WNT canonical signaling pathways. Drugs targeting additional pathways that are altered in PIK3CA-mutant tumors may improve treatment regimens using PI3K inhibitors alone.
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http://dx.doi.org/10.1002/1878-0261.12053DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5527464PMC
May 2017

The non-receptor tyrosine kinase TNK2/ACK1 is a novel therapeutic target in triple negative breast cancer.

Oncotarget 2017 01;8(2):2971-2983

Department of Biological Chemistry, Johns Hopkins University School of Medicine Baltimore, MD 21205, U.S.A.

Breast cancer is the most prevalent cancer in women worldwide. About 15-20% of all breast cancers do not express estrogen receptor, progesterone receptor or HER2 receptor and hence are collectively classified as triple negative breast cancer (TNBC). These tumors are often relatively aggressive when compared to other types of breast cancer, and this issue is compounded by the lack of effective targeted therapy. In our previous phosphoproteomic profiling effort, we identified the non-receptor tyrosine kinase TNK2 as activated in a majority of aggressive TNBC cell lines. In the current study, we show that high expression of TNK2 in breast cancer cell lines correlates with high proliferation, invasion and colony forming ability. We demonstrate that knockdown of TNK2 expression can substantially suppress the invasiveness and proliferation advantage of TNBC cells in vitro and tumor formation in xenograft mouse models. Moreover, inhibition of TNK2 with small molecule inhibitor (R)-9bMS significantly compromised TNBC proliferation.Finally, we find that high levels of TNK2 expression in high-grade basal-like breast cancers correlates significantly with poorer patient outcome. Taken together, our study suggests that TNK2 is a novel potential therapeutic target for the treatment of TNBC.
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http://dx.doi.org/10.18632/oncotarget.13579DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5356856PMC
January 2017

Monitoring of Serum DNA Methylation as an Early Independent Marker of Response and Survival in Metastatic Breast Cancer: TBCRC 005 Prospective Biomarker Study.

J Clin Oncol 2017 Mar 21;35(7):751-758. Epub 2016 Nov 21.

Kala Visvanathan, Johns Hopkins University School of Medicine and Bloomberg School of Public Health; MaryJo S. Fackler, Zhe Zhang, Zoila A. Lopez-Bujanda, Stacie C. Jeter, Lori J. Sokoll, Leslie M. Cope, Christopher B. Umbricht, David M. Euhus, Saraswati Sukumar, and Antonio C. Wolff, Johns Hopkins University School of Medicine, Baltimore, MD; Elizabeth Garrett-Mayer, Medical University of South Carolina, Charleston, SC; Andres Forero, University of Alabama at Birmingham, Birmingham, AL; Anna M. Storniolo, Indiana University, Bloomington, IN; Rita Nanda, University of Chicago, Chicago, IL; Nancy U. Lin, Dana-Farber Cancer Institute, Boston, MA; Lisa A. Carey, University of North Carolina, Chapel Hill, NC; and James N. Ingle, Mayo Clinic, Rochester, MN.

Purpose Epigenetic alterations measured in blood may help guide breast cancer treatment. The multisite prospective study TBCRC 005 was conducted to examine the ability of a novel panel of cell-free DNA methylation markers to predict survival outcomes in metastatic breast cancer (MBC) using a new quantitative multiplex assay (cMethDNA). Patients and Methods Ten genes were tested in duplicate serum samples from 141 women at baseline, at week 4, and at first restaging. A cumulative methylation index (CMI) was generated on the basis of six of the 10 genes tested. Methylation cut points were selected to maximize the log-rank statistic, and cross-validation was used to obtain unbiased point estimates. Logistic regression or Cox proportional hazard models were used to test associations between the CMI and progression-free survival (PFS), overall survival (OS), and disease status at first restaging. The added value of the CMI in predicting survival outcomes was evaluated and compared with circulating tumor cells (CellSearch). Results Median PFS and OS were significantly shorter in women with a high CMI (PFS, 2.1 months; OS, 12.3 months) versus a low CMI (PFS, 5.8 months; OS, 21.7 months). In multivariable models, among women with MBC, a high versus low CMI at week 4 was independently associated with worse PFS (hazard ratio, 1.79; 95% CI, 1.23 to 2.60; P = .002) and OS (hazard ratio, 1.75; 95% CI, 1.21 to 2.54; P = .003). An increase in the CMI from baseline to week 4 was associated with worse PFS ( P < .001) and progressive disease at first restaging ( P < .001). Week 4 CMI was a strong predictor of PFS, even in the presence of circulating tumor cells ( P = .004). Conclusion Methylation of this gene panel is a strong predictor of survival outcomes in MBC and may have clinical usefulness in risk stratification and disease monitoring.
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http://dx.doi.org/10.1200/JCO.2015.66.2080DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5455421PMC
March 2017

Guidelines for the selection of functional assays to evaluate the hallmarks of cancer.

Biochim Biophys Acta 2016 12 11;1866(2):300-319. Epub 2016 Oct 11.

MTA TTK Lendület Cancer Biomarker Research Group, Magyar tudósok körútja 2, H-1117 Budapest, Hungary; 2nd Department of Pediatrics, Semmelweis University, H-1094 Budapest, Hungary. Electronic address:

The hallmarks of cancer capture the most essential phenotypic characteristics of malignant transformation and progression. Although numerous factors involved in this multi-step process are still unknown to date, an ever-increasing number of mutated/altered candidate genes are being identified within large-scale cancer genomic projects. Therefore, investigators need to be aware of available and appropriate techniques capable of determining characteristic features of each hallmark. We review the methods tailored to experimental cancer researchers to evaluate cell proliferation, programmed cell death, replicative immortality, induction of angiogenesis, invasion and metastasis, genome instability, and reprogramming of energy metabolism. Selecting the ideal method is based on the investigator's goals, available equipment and also on financial constraints. Multiplexing strategies enable a more in-depth data collection from a single experiment - obtaining several results from a single procedure reduces variability and saves time and relative cost, leading to more robust conclusions compared to a single end point measurement. Each hallmark possesses characteristics that can be analyzed by immunoblot, RT-PCR, immunocytochemistry, immunoprecipitation, RNA microarray or RNA-seq. In general, flow cytometry, fluorescence microscopy, and multiwell readers are extremely versatile tools and, with proper sample preparation, allow the detection of a vast number of hallmark features. Finally, we also provide a list of hallmark-specific genes to be measured in transcriptome-level studies. Although our list is not exhaustive, we provide a snapshot of the most widely used methods, with an emphasis on methods enabling the simultaneous evaluation of multiple hallmark features.
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http://dx.doi.org/10.1016/j.bbcan.2016.10.002DOI Listing
December 2016

The dual role of FOXF2 in regulation of DNA replication and the epithelial-mesenchymal transition in breast cancer progression.

Cell Signal 2016 10 1;28(10):1502-19. Epub 2016 Jul 1.

Breast Cancer Program, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD, USA. Electronic address:

Dysregulation of Forkhead-box (FOX) transcription factors is linked to cancers of numerous tissue types. Here, we report that FOXF2 is frequently silenced in luminal-type and HER2-positive breast cancers, but is overexpressed in basal-like breast cancers; thus, FOXF2 appears to play distinct roles in different breast cancer subtypes. Inactivation of FOXF2 in luminal-type and HER2-positive breast cancers is attributable to epigenetic silencing. Silencing of FOXF2 is associated with poor prognosis in luminal-type breast cancer. Ectopic expression of FOXF2 in luminal and HER2-positive breast cancer cells suppresses their tumorigenic properties in vitro and in vivo via inhibition of the CDK2-RB-E2F cascade. The in vivo function of FOXF2 is to maintain the stringency of DNA replication, and its loss triggers dysregulation of DNA replication, which in turn activates the p53 checkpoint pathway. Besides its role in cell cycle regulation, FOXF2 is functionally required for mobility and epithelial-to-mesenchymal transition (EMT) of normal breast epithelial cells. In basal-like breast cancer cells, the cell-cycle function of FOXF2 is impaired. However, the EMT function of FOXF2 is still required for mobility, invasiveness and anchorage-independent growth of basal-like breast cancer cells. Our gene expression profiling studies demonstrate that FOXF2 regulates the expression of genes implicated in cell cycle and EMT regulation. Moreover, FOXF2 is highly co-expressed with basal- and metastasis-related genes in breast cancer. These findings suggest that FOXF2 has a dual role in breast tumorigenesis and functions as either a tumor suppressor or an oncogene depending on the breast tumor subtype.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5056599PMC
http://dx.doi.org/10.1016/j.cellsig.2016.06.021DOI Listing
October 2016

HOXC10 Expression Supports the Development of Chemotherapy Resistance by Fine Tuning DNA Repair in Breast Cancer Cells.

Cancer Res 2016 08 14;76(15):4443-56. Epub 2016 Jun 14.

Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, Maryland.

Development of drug resistance is a major factor limiting the continued success of cancer chemotherapy. To overcome drug resistance, understanding the underlying mechanism(s) is essential. We found that HOXC10 is overexpressed in primary carcinomas of the breast, and even more significantly in distant metastasis arising after failed chemotherapy. High HOXC10 expression correlates with shorter recurrence-free and overall survival in patients with estrogen receptor-negative breast cancer undergoing chemotherapy. We found that HOXC10 promotes survival in cells treated with doxorubicin, paclitaxel, or carboplatin by suppressing apoptosis and upregulating NF-κB Overexpressed HOXC10 increases S-phase-specific DNA damage repair by homologous recombination (HR) and checkpoint recovery in cells at three important phases. For double-strand break repair, HOXC10 recruits HR proteins at sites of DNA damage. It enhances resection and lastly, it resolves stalled replication forks, leading to initiation of DNA replication following DNA damage. We show that HOXC10 facilitates, but is not directly involved in DNA damage repair mediated by HR. HOXC10 achieves integration of these functions by binding to, and activating cyclin-dependent kinase, CDK7, which regulates transcription by phosphorylating the carboxy-terminal domain of RNA polymerase II. Consistent with these findings, inhibitors of CDK7 reverse HOXC10-mediated drug resistance in cultured cells. Blocking HOXC10 function, therefore, presents a promising new strategy to overcome chemotherapy resistance in breast cancer. Cancer Res; 76(15); 4443-56. ©2016 AACR.
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http://dx.doi.org/10.1158/0008-5472.CAN-16-0774DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4970943PMC
August 2016

Gene Methylation and Cytological Atypia in Random Fine-Needle Aspirates for Assessment of Breast Cancer Risk.

Cancer Prev Res (Phila) 2016 Aug 3;9(8):673-682. Epub 2016 Jun 3.

Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins School of Medicine, Baltimore, MD.

Methods to determine individualized breast cancer risk lack sufficient sensitivity to select women most likely to benefit from preventive strategies. Alterations in DNA methylation occur early in breast cancer. We hypothesized that cancer-specific methylation markers could enhance breast cancer risk assessment. We evaluated 380 women without a history of breast cancer. We determined their menopausal status or menstrual cycle phase, risk of developing breast cancer (Gail model), and breast density and obtained random fine-needle aspiration (rFNA) samples for assessment of cytopathology and cumulative methylation index (CMI). Eight methylated gene markers were identified through whole-genome methylation analysis and included novel and previously established breast cancer detection genes. We performed correlative and multivariate linear regression analyses to evaluate DNA methylation of a gene panel as a function of clinical factors associated with breast cancer risk. CMI and individual gene methylation were independent of age, menopausal status or menstrual phase, lifetime Gail risk score, and breast density. CMI and individual gene methylation for the eight genes increased significantly (P < 0.001) with increasing cytological atypia. The findings were verified with multivariate analyses correcting for age, log (Gail), log (percent density), rFNA cell number, and body mass index. Our results demonstrate a significant association between cytological atypia and high CMI, which does not vary with menstrual phase or menopause and is independent of Gail risk and mammographic density. Thus, CMI is an excellent candidate breast cancer risk biomarker, warranting larger prospective studies to establish its utility for cancer risk assessment. Cancer Prev Res; 9(8); 673-82. ©2016 AACR.
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http://dx.doi.org/10.1158/1940-6207.CAPR-15-0377DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4970896PMC
August 2016