Principal Investigator: Andrew Lewis, MD
Office / Division / Lab: OVRR / DVP / LDV
Vaccines are an essential public health tool for controlling viral diseases. Viral vaccines are produced in living cells. The cells that are used for vaccine manufacture are called cell substrates. The development of safe and effective viral vaccines requires that these substrates and the other substances used in the production of the vaccines must be as safe as possible.
There are a variety of types of cell substrates, including cells from eggs and cells from mammals grown in culture. Some cells used as cell substrates are immortalized (transformed), that is, they continually multiply so the culture never dies out. Cells from these cultures of transformed cells have the potential to form tumors in animals (tumor-forming cell substrates).
The major challenge to the safety of vaccines manufactured in transformed cell substrates is contamination with genetic material (DNA) from the cell substrate that might trigger the growth of tumors in the vaccine recipient; or the genetic material might encode infectious agents including cancer-causing viruses. However, transformed cell substrates are important for the development of vaccines for HIV/AIDS, vaccines against annual and pandemic influenza (such substrates are used in Europe to make influenza vaccines) and for vaccines to protect against agents of bioterrorism. FDA reviewers must evaluate the safety issues posed by all cell substrates used in the manufacture of viral vaccines.
Regulatory evaluation of transformed cell substrates could be improved by a better understanding of the processes involved in cell transformation. Thus, our laboratory is attempting to understand mammalian cell transformation, how transformed cells develop the ability to form tumors, and how these tumors actually develop. Our laboratory is also developing ways to evaluate the possible risks that might be associated with the genetic material (DNA) from tumor-forming cell substrates. DNA from these substrates poses two risks: 1) the possibility of transferring cancer-causing activity to vaccines, and 2) the possibility of transferring infectious microorganisms to vaccines.
To extend our work to include all aspects of the risks associated with tumor-forming cell substrates, we are developing ways to 1) determine whether DNA from such cells can be a source of cancer-causing activity and infectious microorganisms; and 2) evaluate the impact of addition of enzymes during manufacturing that would cause DNA degradation (breakdown) on the relative risks associated with these processes.
Our laboratory is currently focused on two projects: 1) characterizing the tumorigenic phenotypes expressed by neoplastically transformed, cell-substrate reagents, and 2) evaluating the oncogenicity/infectivity of DNA from neoplastic cell substrates.
Establishing quantitative methods for defining tumorigenicity as a means of characterizing the tumorigenic phenotypes of cell-substrate reagents to be used for vaccine manufacture: We are using quantitative, (dose-response) tumorigenicity assays in immuno-incompetent strains of mice to characterize the ability to form tumors (tumorigenic phenotype) expressed by transformed cell substrates and to evaluate mechanisms of tumor formation in animals. As part of our efforts to further understand neoplastic transformation, our studies of neoplastic cell tumorigenicity are providing the reagents that we use to study the patterns of miRNA expression associated with the neoplastic processes during the passage of mammalian cells in tissue culture.
Evaluating the oncogenic activity posed by DNA from neoplastic cell substrates: Cellular H-ras and c-myc oncogenes are oncogenic in mice when injected together in different plamids or cloned together in the same plasmid. Less than a picogram of single plasmid DNA containing both oncogenes can induce tumors. We use these plasmid-mouse models to develop assays to assess the possible oncogenic activity associated with the DNA of neoplastic cell substrates. We use similar models to evaluate the impact of DNA degradation and removal on oncogenic activity and the infectivity of cell DNA-containing retroviral genomes.
Vaccine: X 2019 Apr 11;(1):100004
Responsiveness to basement membrane extract as a possible trait for tumorigenicity characterization.
Murata H, Omeir R, Tu W, Lanning L, Phy K, Foseh G, Lewis AM Jr, Peden K
Vaccine 2017 Oct 4;35(41):5503-9
Assessment of potential miRNA biomarkers of VERO-cell tumorigenicity in a new line (AGMK1-9T7) of African green monkey kidney cells.
Teferedegne B, Rotroff DM, Macauley J, Foseh G, Lewis G, Motsinger-Rief A, Lewis AM Jr
Chromosome Res 2015 Dec;23(4):663-80
A novel canine kidney cell line model for the evaluation of neoplastic development: karyotype evolution associated with spontaneous immortalization and tumorigenicity.
Omeir R, Thomas R, Teferedegne B, Williams C, Foseh G, Macauley J, Brinster L, Beren J, Peden K, Breen M, Lewis AM Jr
Vaccine 2015 Dec 16;33(51):7254-61
RT-qPCR-based microneutralization assay for human cytomegalovirus using fibroblasts and epithelial cells.
Wang X, Peden K, Murata H