Ken Nephew Lab

The major areas of research in Dr. Kenneth P. Nephew’s laboratory are:

1.      Cancer epigenetics, DNA methylation and chromatin remodeling.

2.      Epigenetic-based therapies.

3.      Integrated Cancer Biology

4.      Breast cancer and antiestrogen resistance.

5.      Estrogen receptor signaling and receptor degradation.

6.      Ubiquitin and ubiquitin-like  pathways.

7.      Cancer gene therapy- transcriptional targeting of ovarian cancer and thyroid cancer.

Areas 1-3  Aberrant DNA methylation is a hallmark of most cancers, and the association between increased CpG island methylation and inactivation of genes with known roles in tumor development and progression has been demonstrated for essentially all human cancers, including ovarian.  In addition, genes known to be involved in drug sensitivity can become methylated in ovarian tumors.  Using a microarray based approach called differential methylation hybridization (DMH), developed by collaborator Tim Huang, we determined that a higher level of methylation is associated with early disease recurrence following chemotherapy.  Furthermore, we identified a select group of CpG island loci potentially useful as epigenetic markers for predicting treatment outcome for ovarian cancer patients.  Thus, epigenetic changes appear to play a role in chemotherapy response in ovarian cancer patients, and DNA methylation patterns of ovarian tumors may independently predict response or progression free survival of patients.  In addition, our work and the work of others suggest that DNA methylation is reversible in ovarian cancer cells.  Thus, we are pursuing the clinical development of epigenetic therapies for this disease.  In the laboratory, we are combining pharmacological reversal of aberrant methylation with standard chemotherapy.  We hope to take this approach in the clinic and improve the outcome for recurrent ovarian cancer patients. 

Area 3. Interrogating Epigenetic Changes in Cancer Genomes:  Predicting Drug Resistance in Cancer Genomes by DNA Methylation Profiling. We will utilize mathematical models to explore DNA methylation in cancer cells.  We will test the hypothesis that in both intrinsic and acquired drug-resistance, increased methylation density in CpG island loci occurs during ovarian cancer progression and can further be selected for during chemotherapy.  This aberrant DNA methylation leads to eventual silencing of genes that regulate drug responses and apoptotic signaling networks and contributes to the insensitivity of ovarian cancer cells to chemotherapy and poor treatment outcomes.

Areas 4-6. Antiestrogens such as tamoxifen are commonly used as adjuvant therapy for women with ERa-positive breast cancer.  Most breast cancers eventually develop resistance to tamoxifen, and another antiestrogen named Fulvestrant (ICI 182,780; also known clinically as Faslodex) is used as a second line therapy.   Fulvestrant inhibits breast cancer cell growth by causing receptor degradation, we are  investigating the role of ubiquitination and proteasomal degradation of ERa in this process.  Our work further suggests that disruptions in ubiquitin-associated pathways may contribute to the development of antiestrogen-resistance in human breast cancer, and emerging treatments for breast cancer include drugs that target estrogen receptor degradation and the proteasome. 

Area 7.  Limitations of current ovarian cancer gene therapies include lack of specificity and transduction of normal tissues.  One strategy toward overcoming these limitations is to direct gene therapy specifically to ovarian cancer cells by using tissue/tumor specific promoters.   The whey acidic protein HE4 is frequently overexpressed in ovarian cancer, suggesting that the HE4 promoter is highly transcriptionally active in the disease.  We isolated the HE4 promoter and used it to drive specific reporter gene expression in epithelial ovarian cancer cell lines.  Along with our colleagues at IUSM, we are continuing to develop this promoter and others for use in transcriptional targeting in ovarian cancer gene therapy

Active Grants

R01 CA85289 (PI:  Kenneth P. Nephew, Ph.D)

Agency: National Institutes of Health/National Cancer Institute

Title: DNA Methylation and Ovarian Cancer

Co-Investigator: Tim H.-M. Huang, Ph.D.

RSG TBE-104125 (PI: Kenneth P. Nephew, Ph.D.)

Agency:  American Cancer Society

Title: Regulation of Estrogen Receptor Function by the Ubiquitin-Like NEDD8 Pathway

BC10839 (PI:  Kenneth P. Nephew, Ph.D)            

Agency: United States Army, Department of Defense,

Title: Role of the Neddylation Enzyme Uba3, a New Estrogen Receptor Corepressor, in Breast Cancer (IDEA Award)

BC010402 (PI:  Kenneth P. Nephew, Ph.D)          

Agency: United States Army, Department of Defense

Title: Role of the Neddylation Enzyme Uba3, a New Estrogen Receptor Corepressor, in Breast Cancer, Career Development Award

Biomedical Research Pilot Funding Application (PI: Kenneth P. Nephew, Ph.D)

Agency: Indiana University School of Medicine

Title: Role of the NEDD8 Pathway in Mammary Gland Development and Tumor Formation

Pilot Project Grant, IU Cancer Center (PI; Co-PI is Ross Weatherman, Ph.D., Dept of Medicinal Chemistry, Purdue Cancer Center)

         Title: Chemical Probes of the Mechanism Action of Antiestrogen Action in Breast Cancer

Structure-Activity Relationships of NEDD8-Induced Estrogen Receptor Degradation by Breast Cancer Therapeutics

P50 CA113001 (PI:  Tim Huang, Ph.D.)                  

Agency: National Institutes of Health/National Cancer Institute

Integrated Cancer Biology Program

Title of the Program Project:  Interrogating Epigenetic Changes in Cancer Genomes

Representative Publications

Ahluwalia A, Yan P, Hurteau JA, Bigsby RM, Jung SH, Huang T, Nephew KP. DNA methylation and ovarian cancer. I: Analysis of CpG island hypermethylation in human ovarian cancer using differential methylation hybridization. Gynecol Oncol, 82:261-268, 2001.

Ahluwalia A, Hurteau JA, Bigsby RM, Nephew KP. DNA methylation and ovarian cancer. II: Expression of DNA methyltransferases in ovarian cancer cell lines and normal ovarian epithelial cells. Gynecol Oncol, 82:299-304, 2001.

Fan M, Long X, Bailey JA, Reed CA, Osborne E, Bigsby RM, Nephew KP. The activating enzyme of NEDD8 inhibits steroid receptor function. Mol Endocrinol, 16:315-330, 2002.

Wei S, Chen C, Shi H, Yan P, Harnsomburana J, Shyu C, Nephew KP, Brown R, Huang T. Methylation microarray analysis: late stage ovarian carcinomas distinguish disease-free survival in patients. Clin Cancer Res, 8:2246-52, 2002.

Nephew KP, Huang T. Epigenetic gene silencing in cancer initiation and progression. Cancer Lett,190:125-133, 2003.

Fan M, Bigsby RM, Nephew KP. The NEDD8 pathway is required for proteasome mediated degradation of human estrogen receptor-a and essential for the antiproliferation activity of ICI 182,780 in ER-positive breast cancer cells. Mol Endocrinol, 17:356-365, 2003. (cover article)

Shi H, Wei SH, Leu Y-W, Rahmatpanah F, Liu JC, Yan PS, Nephew KP, Huang THM. Triple analysis of the cancer epigenome: an integrated microarray system for assessing gene expression, DNA methylation and histone acetylation. Cancer  Res, 63:2164-2171, 2003. (cover article)

Berry NB, Cho YM, Harrington MA, Foley J, Williams SD, Nephew KP. Transcriptional targeting in ovarian cancer cells using the HE4 promoter. Gynecologic Oncol, 92:896-904, 2004.

Fan M, Nakshatri H, Nephew KP.Inhibiting proteasomal proteolysis sustains estrogen receptor-a activation

Mol Endocrinol, 18:2603-15, 2004

Balch C, Huang TH-M, Brown R, Nephew KP.  The epigenetics of ovarian cancer drug resistance and resensitization American Obstet Gynecol, 191:1552-72, 2004

Leu YW, Yan PS, Fan W, Jin VX, Liu CJ, Curran EM, Welshons WV, Wei HS, Davuluri RV, Plass C, Nephew KP, Huang TH-M. Loss of estrogen signaling triggers epigenetic silencing of its downstream targets Cancer Res, 64:8184-8192, 2004

Jin VX, Leu YW, Fan M, Liyanarachchi S, Sun H, Nephew KP, Huang TH-M, Davuluri RV. Identifying estrogen receptor-α target genes using integrated computational genomics and chromatin immunoprecipitation microarray.  Nucleic Acids Res, 32:6627-6635, 2004

Li, L, Shi H, Yiannoutsos C, Huang TH, Nephew KP.  Epigenetic hypothesis tests for methylation and acetylation in a triple microarray system.  J. Computational Biol (in press)

Balch C, Montgomery JS, Paik H-I, Kim SH, Huang T H-M, Nephew KP.  Emerging epigenetic therapies and biomarkers for cancer. Front Biosci (in press)