A team of bioengineers, molecular biologists, and clinicians used a novel rare cell-sorter to isolate breast cancer cells from the blood of patients, with the aim of identifying the most effective drugs to treat each individual tumor. Circulating tumor cells (CTCs) were isolated and grown in the laboratory for extensive genetic analysis, which enabled the identification and testing of the most effective cancer-killing drugs for those tumors.
The ability to perform such genetic analysis in the laboratory paves the way for providing the most effective treatment, not only initially, but throughout the course of the disease, as mutating tumors become resistant to certain drugs, but susceptible to others.
CTCs are tumor cells that are shed from primary tumors in the body and are carried through the circulation. For a number of years now, researchers have worked to develop technologies to capture and perform genetic analysis on these cells to learn about their growth characteristics and molecular evolution. Led by senior authors Daniel Haber, MD, PhD, and Shyamala Maheswaran, PhD, of Massachusetts General Hospital Cancer Center at Harvard Medical School; and Mehmet Toner, PhD, Harvard Center for Bioengineering in Medicine, the research team, who developed a microfluidic chip called the CTC-iChip, used it to isolate the minute numbers of tumor cells circulating in the blood. The work is reported in the July 11 issue of Science.
The study design incorporated several crucial technologies necessary for successfully using CTCs to gain accurate information about the tumors from which they originated. The CTC-iChip is unique in that it does not use cancer cell markers on the surface of the CTCs to identify cells for capture. This is important because these markers change as the cancer progresses, so that captured cells obtained by using such markers may only represent a subset of cells shed by the tumor. Instead the iChip efficiently removes the normal blood cells, leaving behind viable CTCs that represent cells shed from both the primary tumor as well as metastatic tumor deposits – tumor nodules arising in distant organs due to the deposition of CTCs.
The other key advance in the study was the development of a cell culture system that allowed the CTCs to successfully grow in the laboratory. The expansion of the CTCs in cell culture is critical for having enough cells for genetic analysis and subsequent testing of anti-cancer drugs and drug combinations that target the newly evolved mutations. After much trial and error the group found that the cells could be successfully grown and expanded when cultured as suspended spheres of cells rather than when attached as a monolayer to the bottom of the cell culture plate. Importantly, with this new culture technique, the cells did not mutate over time while in culture -- a common problem when growing cells in the laboratory.
The researchers used the iChip to isolate CTCs from the blood of 36 breast cancer patients. Cell lines were successfully established from the CTCs of six of these patients. Genetic analysis of the cell lines were compared with biopsies from the parent tumor to determine whether the metastatic tumors had evolved over time. Cultures derived from the same patient at multiple time points were compared to verify that culture conditions did not result in genetic changes in the CTCs. Standard care at Massachusetts General Hospital involves screening for a variety of mutations in just 25 genes. In contrast, the CTC cell lines enabled a far more extensive mutational analysis, including screening for mutations in 1,000 cancer genes.
Armed with such a comprehensive genetic analysis, the researchers then tested CTC lines for sensitivity to panels of single medications and medication combinations, based on knowledge of the susceptibility of various cancer mutations to certain drugs. The aim of these experiments was to identify which medications worked best on the tumor cells of each individual patient.
The test results indicated that tumors from several patients responded to therapy with medications commonly used for tumors carrying the mutations identified with the standard 25-gene analysis. However, several of the tumor samples did not respond to such treatments, but were responsive to different combinations of medications identified through the more extensive 1,000 gene screening made possible by isolation of the CTCs. Therefore, this proof of concept study successfully demonstrated that this approach has the potential to identify a wider range of genetic mutations, enabling treatments that successfully target the specific mutations in each patient’s tumor.
The work is a significant first step towards “precision medicine” in oncology where treatments are tailored to the drug sensitivity patterns in individual patients. Furthermore, the system offers the opportunity to adjust treatments throughout the course of disease based on evolving tumor mutation profiles. By repeated sampling of CTCs throughout the course of a patient’s disease, medications can be adjusted as individual tumors become resistant to certain medications but susceptible to others.
Tiffani Lash, PhD., the NIBIB Program Director for Micro- and Nano-System Technologies, elaborates on the significance of the research. “With the hope of improving cancer therapies, NIBIB has supported the development of the sophisticated technologies necessary to capture and analyze CTCs. The ability to grow the CTCs in the laboratory and test them for cancer drug susceptibility is a major step towards providing targeted therapies with the real potential to significantly improve patient outcomes.”