Breast Cancer Disparity Among African American Women: Insights Powered by Sanger Sequencing

Written by Caroline McMahon and Cleta D’Sa

Breast cancer is a leading cause of cancer death among African American women, with those under the age of 45 most likely to develop breast cancer, and those of all ages most likely to die from breast cancer.¹ Data show that African American women in Memphis, Tennessee, suffer disproportionately from higher breast cancer mortality rates compared to non-Hispanic white women.^2,3,4 Considering recent reports of higher breast cancer mortality rates among women in Memphis, compared to 49 of the largest US cities,^2,3 could genetic variants and environmental exposures be contributing to this disparity? Athena Starlard-Davenport, PhD, was determined to find the answer and has made examining the devastating effects of breast cancer in African American women her life’s work.

Growing up in a single-parent household, Starlard-Davenport was surrounded by women. Her close bond with these women significantly influenced an interest in pursuing breast cancer research from early on; Starlard-Davenport began this work in graduate school in 2002, and received her Ph.D. in Biochemistry and Molecular Biology from the University of Arkansas for Medical Sciences in Little Rock. During that time, she joined a lab where she focused on drug-metabolizing enzymes and how certain drugs metabolize in the liver.

As the disease began to hit closer to home with the passing of family and friends, and eventually her aunt-in-law, Starlard-Davenport became even more interested in understanding the role of metabolizing enzymes because they also metabolize estrogen. And as is already known, prolonged exposure to estrogen increases the risk of breast cancer. Her goal then was to look at particular metabolizing enzymes called UDP-glucuronosyltransferases (UGTs) and characterize them in normal and breast cancer tissues from African American and white women. A study came out showing that a promoter polymorphism in one of the UGTs that is majorly responsible for metabolizing both parent estrogens and estrogen metabolites might be associated with premenopausal breast cancer.⁵ That mutation is actually a TA repeat polymorphism in the promoter, and so it’s been well-studied in individuals who have many of these TA repeats in the TATA box region of the promoter. When a person has fewer TA repeats, the enzyme is more active in metabolizing certain drugs.⁶

The study also showed that individuals who have many of these TA repeats have less metabolism of estrogen, and so it’s thought that if an individual has these repeats, they are more likely to have an accumulation of estrogen in the body, which might predispose them to breast cancer. Says Starlard-Davenport, “With samples collected throughout the Memphis community from African American women, I was just curious to see, based upon my hypothesis, if women have this TA repeat polymorphism, then maybe they might be at higher risk of breast cancer. And so that’s exactly what we did.”

Enter Sanger sequencing

Starlard-Davenport and her team designed primers around the TA repeat region so that they could amplify it by PCR. Genotypes of the TA repeat promoter polymorphism were determined by bidirectional sequencing using the Sanger method with forward or reverse primers to amplify a 351 bp region containing the polymorphic region. “From the results,” says Starlard-Davenport, “we were able to stratify women based upon whether or not they were cases or controls and see if there was an association with breast cancer based upon premenopausal status and postmenopausal status. We found that women with this TA repeat polymorphism were more likely to have breast cancer and it was more pronounced in women [who] were premenopausal.”⁷

Sanger sequencing played an important role in this discovery. According to Starlard-Davenport, “Although high-throughput next-generation sequencing (HT-NGS) technologies [have] led to the identification of genetic variations associated with various diseases and inherited disorders, Sanger sequencing is the gold standard for sequencing single genes, including small gene regions. By using Sanger sequencing to analyze the DNA fragment that contains the UGT promoter polymorphism, we do not need to sequence the entire human genome, which is very costly and time consuming when considering the additional bioinformatic support necessary to clean, analyze, and interpret large amounts of sequencing data. Additionally, unlike HT-NGS, with Sanger sequencing, we do not have to worry about how to interpret and manage incidental findings from our sequencing data⁸ such as variants of unknown significance that have not been characterized.”

The future of breast cancer research

Starlard-Davenport believes that women who have the TA repeat polymorphism could be checked and could then perhaps benefit from avoiding hormone replacement therapy, which includes certain estrogen-containing compounds that might cause an accumulation of estrogen in the body and could increase their risk of developing breast cancer.

“It would be good to be able to get more samples from women just to confirm this in a larger study, to see if indeed that this mutation is associated with breast cancer,” says Starlard-Davenport. “If we could confirm that and just have ways where we can get women tested, for example, like with the Precision Medicine Initiative, just to be able to determine that if someone has a certain mutation, maybe they should not, for example, be taking birth control pills that contain estrogen.”

In January 2016, Starlard-Davenport joined the faculty at the University of Tennessee Health Science Center (UTHSC) as an Assistant Professor in the Department of Genetics, Genomics, and Informatics in the College of Medicine, where she continues her research and is currently pursuing another hypothesis: individuals who have breast cancer may have a different microbiome, microbiota in the breast, compared to women without breast cancer.

Starlard-Davenport’s early research results show that African American women have a distinct microbiome in their breast cancer tumors, making them more vulnerable to breast cancer’s effects.⁹ While it has become her mission to discover the reason, she knows her research will be difficult and asserts, “Breast cancer is not one disease. There are many tumor subtypes that strike pre-menopausal vs. post-menopausal women differently.” She adds that having the opportunity to get outside of the lab and actually shadow a surgical oncologist has enabled her to see and hear first-hand the very difficult journey experienced by women with breast cancer, which has served to further motivate her mission to help future generations determine the cause of, and one day prevent, breast cancer.

Starlard-Davenport firmly believes that with hard work and determination, you can achieve your goals, and she has certainly proven it: she was the first African American student to receive a Ph.D. in the Biochemistry and Molecular Biology program at the University of Arkansas for Medical Sciences in Little Rock in 2007, and she took her qualifying exams while pregnant. She is also involved in sickle cell disease research, studying microRNA as a potential targeted therapy for inducing levels of fetal hemoglobin in patients with sickle cell disease.

Explore Sanger sequencing applications, instrumentation, reagents, consumables, and software designed to respond to the unlimited potential of scientific inquiry.

References

1. Desantis, C.E., Ma, J et al. (2017) “Breast cancer statistics, 2017, racial disparity in mortality by state,” CA Cancer J Clin, 67(6), pp. 439–448. doi:10.3322/caac.21412
2. Hunt, B.R., Whitman, S. et al. (2014) “Increasing Black:White disparities in breast cancer mortality in the 50 largest cities in the United States,” Cancer Epidemiol, 38(2), pp. 118–123. doi:10.1016/j.canep.2013.09.009
3. Vidal, G., Bursac, Z. et al. (2017) “Racial disparities in survival outcomes by breast tumor subtype among African American women in Memphis, Tennessee,” Cancer Med, 6(7), pp. 1776–1786. doi:10.1002/cam4.1117
4. Whitman, S., Orsi, J. et al. (2012) “The racial disparity in breast cancer mortality in the 25 largest cities in the Unites States,” Cancer Epidemiol, 36(2), pp. e147–151. doi:10.106/j.canep.2011.10.012
5. Guillemette, C., Millikan, R.C. et al. (2000) “Genetic polymorphisms in uridine diphospho-glucuronosyltransferase 1A1 and association with breast cancer among African Americans,” Cancer Res, 60(4), pp. 960–966.
6. Guillemette, C., De Vivo, I. et al. (2001) “Association of genetic polymorphisms in UGT1A1 with breast cancer and hormone levels,” Cancer Epidemiol Biomarkers Prev, 10(6), pp. 711–714.
7. Smith, A., Cropp, C. et al. (2018) “Prevalence of the UGT1A1*28 promoter polymorphism and breast cancer risk among African American women in Memphis, TN,” Cancer Health Disparities, 2, pp. e1–e12. doi:10.9777/chd.2018.10025
8. Di Resta, C., Galbiati, S. et al. (2018) “Next-generation sequencing approach for the diagnosis of human diseases: Open challenges and new opportunities,” EJIFCC, 29(1), pp. 4–14. PMID: 29765282 PMCID: PMC5949614
9. Smith, A., Pierre, J.F. et al. (2019) “Distinct microbial communities that differ by race, stage, or breast-tumor subtype in breast tissues of non-Hispanic Black and non-Hispanic White women,” Sci Rep, 9(1), 11940. doi:10.1038/s41598-019-48348-1