Cell lines are one of cancer researchers’ most powerful tools for studying the disease that accounts for one in four deaths in the US. Cell-based studies using these lines allow scientists to probe cancer’s molecular mechanisms: how it starts and spreads, and what molecules fuel or suppress it. Biologists create these cell lines by searching through donated cancer cells from individual patients to find cells that can be immortalized, meaning they can be grown again and again in petri dishes and in mice.
While the most famous cell line, HeLa, came from an African-American patient, Henrietta Lacks, the majority of cancer cell lines are derived from patients with European ancestry. Yet researchers have increasingly observed differences in how certain cancers behave at the biological level in various ethnic groups, highlighting the need for ethnically diverse cell lines to probe the molecular basis for these differences.
In the past, scientists assigned an ethnicity to a cell line through a doctor’s observation or a self-report by the patient. Now, a multiinstitutional team of researchers has analyzed the genetic ancestry of 15 prostate, breast, and cervical cancer cell lines and found that several lines labeled as mixed or black/African-American had been misclassified (Cancer Epidemiol. Biomarkers Prev. 2019, DOI: 10.1158/1055-9965.EPI-18-1132). The authors say the results should motivate scientists to confirm the identity of cell lines they use in experiments.
The team used a set of 105 ancestry genetic biomarkers—specific DNA base differences commonly found in people of a certain ancestry—to determine the percentage of each cell line’s West African, Native American, and European ancestry. The team reports that HeLa cells contain 66% West African ancestry, which is slightly lower than the average of about 80% West African ancestry for US born African-Americans. The scientists also found that one prostate cancer cell line called 22Rv1 had 99% European ancestry though it had previously been determined to have mixed ancestry using a small panel of biomarkers. (Prostate 2017, DOI: 10.1002/pros.23437)
But the main misclassification was in a prostate cancer cell line called E006AA-hT, which is labeled and sold by vendors as African-American, but actually carries 91% European ancestry.
Prostate cancer disproportionately affects black men, whose mortality rate from the disease is twice that of the rate for white men, according to the Centers for Disease Control and Prevention. Experts say this disparity is a result of societal variables, such as socioeconomic status and access to health care, as well as biological factors. For example, prostate cancer tends to be associated with more inflammation and to be more aggressive in African-American men than in men of European ancestry, suggesting a biological difference. Yet only two of the dozen most common commercially-available prostate cell lines are sold as African-American; the rest are sold as having European ancestry.
It’s upsetting to have so few African-American cell lines to start with and then to find that those resources were misclassified, says study coauthor Rick A. Kittles, a cancer and health disparities researcher at the City of Hope.
Study coauthor K. Sean Kimbro of North Carolina Central University says resolving this disparity becomes particularly important as cancer treatment moves toward precision medicine, essentially targeting specific genes that drive disease, some of which may differ between ethnic populations. The lack of cell lines from different ethnic groups makes the science biased against non-European ancestry groups, Kimbro says. “Not necessarily intentionally but just by default because the cell lines don’t include those individuals,” Kimbro says.
Kittles says cell line misclassifications are pretty common and may happen either through mix-ups in the lab or, in the case of ancestry, by relying only on people’s observations of patients. “Unless you ask [patients] about their family history and then also type these ancestry markers, there’s no real way to be 100% sure,” he says.
The E006AA cell line originated from cells donated by a 50-year-old African American man who was a patient at Louisiana State University-Health Sciences Center, according to the 2004 publication that originally established the cell line (Prostate 2004, DOI: 10.1002/pros.20053). Scientists slightly modified the cells in 2014 to produce the highly tumorigenic variant, E006AA-hT, that can grow in mice (Int. J. Biol. Sci. 2014, DOI: 10.7150/ijbs.9406). C&EN could not reach corresponding author Shahriah Koochekpour, last listed as a researcher at the Roswell Park Comprehensive Cancer Center, or any of the either paper’s authors for comment on the new ancestry results by press time.
Koochekpour and colleagues deposited the line at the American Type Culture Collection, a common resource for biological researchers, where it is currently sold as an African-American cell line. ATCC says they don’t perform genetic ancestry testing on their cell lines and rely on the scientists depositing the samples for that information. “In this case, we will work with the depositor to confirm the ethnicity of the deposited cell line and if we confirm that the information originally reported needs to be revised, ATCC will take all necessary steps to correct the situation,” the company says.
The authors of the new study tested a sample of E006AA-hT purchased from ATCC and three separate sources of the E006AA cell line and found more than 90% European ancestry in all cases.
More than a dozen published research papers have used either E006AA or E006AA-hT cells, including several of which used the cell line specifically to investigate the biology of prostate cancer in African-American men.
Omar Franco, a cancer researcher at NorthShore University HealthSystem, bought E006AA-hT through ATCC and used the cells in a study published online in September 2018 (Cancer Res. 2018, DOI: 10.1158/0008-5472.CAN-17-3810). Franco says he first heard about the line’s misclassification in November when a friend texted him after seeing Kittles present the ancestry data at a scientific meeting. Franco says the cells’ ancestry doesn’t directly impact the conclusions of his study. The study investigated the progression of prostate cancer in African-American men, but it focused on the effect of fibroblasts, cells involved in inflammation, on the prostate cancer cells. He also used the other commercially available African-American cell line, MDA-PCa-2b, which the new study confirms as African-American with 86% West African ancestry.
C&EN contacted the journals Scientific Reports and the Prostate, which published several papers that may be impacted by the new study’s findings, for comment. A representative of Scientific Reports tells C&EN, “when issues are raised with the Scientific Reports editorial team about papers we have published, we look into them carefully on a case-by-case basis. Once such processes have concluded and we have the necessary information to make an informed decision, we will take action where appropriate to ensure the accuracy of the scientific record. For confidentiality reasons, we are unable to comment further at this time.” the Prostate did not respond by press time.
Franco and several other cancer researchers told C&EN that they will stop using E006AA-hT as an African-American cell line. But some point out that the cells could still be useful as a general prostate cancer cell line. “We’re still searching for good lines,” Franco says. “Hopefully somebody will make some.”
Nishadi Rajapakse, a biologist and program director at the National Institute on Minority Health and Health Disparities says that as part of the agency’s mission to reduce health disparities, they’ve funded efforts to improve diversity and representation of underrepresented groups in biomedical research. Studies like this one are important, she says, because they encourage researchers to verify their cell lines’ ancestry.
Tuskegee University’s Clayton Yates’ lab is one of several groups working to increase the number of African-American cell line models. His lab recently developed an African-American prostate cancer cell line called RC-77T/E that was validated in the new study, but is not yet commercially available. Yates says poor diversity in cell lines stems from several factors, including the difficulty of immortalizing prostate cells. Also, cell lines often have been collected at major research institutions that don’t always have diverse patient populations and African-Americans’ historical mistreatment in the US has generated an inherent lack of trust of institutions. These barriers can be overcome, he says, with the help of researchers and clinicians that represent the diversity within communities.