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  5. Development of Quantitative Assays to Evaluate the Safety of Cell Substrates and Vaccines
  1. Biologics Research Projects

Development of Quantitative Assays to Evaluate the Safety of Cell Substrates and Vaccines

Principal Investigator: Keith Peden, PhD
Office / Division / Lab: OVRR / DVP / LDV

General Overview

Viral vaccines are produced in animal cells called "cell substrates”. Therefore, the characteristics of those cell substrates directly affect the quality of the vaccine. Our division in CBER regulates vaccines against diseases caused by viruses, and our program is developing new tools that could be used to evaluate the safety of cell substrates used to produce viral vaccines.

There are a variety of vaccines that protect the public against viral diseases; these diseases include polio, influenza, measles, mumps, rubella, and smallpox. These vaccines, which are made in monkey, chick, or human cell substrates, are safe and effective. However, not all vaccines can be manufactured in these cell substrates because they may not be able to grow, thus making them unsuitable for producing certain other vaccines, such as those against HIV/AIDS, or against newly emerging infectious organisms (e.g., Ebola virus and SARS).

The reason that there are only a limited variety of cell substrates is that scientists in the 1950s and 1960s agreed that cell lines established from human tumors should not be used to make viral vaccines. The rationale for this decision was based on the concern that if a cell substrate were derived from a human tumor, or was shown in an experiment to be able to form a tumor in an animal (i.e., was tumorigenic), then components from those cells could be present in vaccines manufactured in them. Such vaccines might then induce cancer or other diseases in recipients of the vaccines.

However, the growth of some viruses requires the use of tumorigenic cells. Therefore, there is renewed interest in using these cells to develop new vaccines. Over the past several decades researchers have learned much about how cancer develops, and that allows FDA to reopen the question about the safety concerns and address them in a more mechanistic and data-driven manner.

Our laboratory is developing new approaches, using rodent models to investigate whether the use of tumorigenic cells or cells derived from human or animal tumors for vaccine production pose safety concerns to the recipients of the vaccines.

We believe that the risk of using tumorigenic cells to make viral vaccines would increase either by the presence of disease-causing microorganisms (adventitious agents), or from the DNA of the cell. Therefore, we have begun to address each of these concerns. This work requires the development of sensitive animal and cell-culture based assays.

Our work will provide critical data to help the vaccine community evaluate whether it is safe to end the 40-year ban on using tumorigenic cells to make viral vaccines.

Scientific Overview

The major concerns with the use of novel neoplastic cell substrates are the potential presence of adventitious agents and the unavoidable presence of residual cellular DNA in vaccines.

Therefore, our laboratory is developing assays to evaluate the safety of novel cell substrates. We have divided this work among five projects: 1) assessing the in vivo oncogenicity of cell-substrate DNA; 2) developing in vitro assays to quantify the elimination of biological activity of DNA; 3) determining whether the tumorigenicity of a cell substrate affects the safety of vaccine manufactured in them; 4) developing methods to detect adventitious agents; and 5) developing methods to detect neutralizing antibodies to viruses.

The variety of cell substrates used to manufacture licensed vaccines is limited to primary avian or monkey cells, diploid cells, and one continuous cell line, Vero. This repertoire is insufficient for the production of the next generation of vaccines.

All of the mammalian cell substrates being evaluated are neoplastic, since they are immortal, and some are tumorigenic. The fear that components from the production-cell substrate could induce cancer in vaccine recipients was the main reason that tumorigenic cells were proscribed for vaccine manufacture for over 40 years. This proscription was partly due to the inability to identify the risk factors; yet even after they were identified, there was a lack of assays that could quantify the risk from these factors.

Projects 1 and 2 have been the main focus of our work. Because DNA can be oncogenic or infectious, we must consider both activities. As part of our studies to assess the risk from residual cell-substrate DNA in vaccines, we are developing assays to quantify the biological activities of DNA and thus enable us to estimate the risks DNA poses. Our risk estimates will be conservative, since they will be based on the most sensitive assays available.

We have also assessed methods for inactivating the biological activities of DNA and estimated subsequent risk reduction. In addition, we determined that the biological activity of DNA can be reduced more than 10e7, a reduction that has been accepted as sufficient by the Vaccine and Related Biological Products Advisory Committee to permit vaccines manufactured in neoplastic-cell substrates to move into clinical trials.

A broader issue is whether the tumorigenicity of a cell substrate affects the safety of a vaccine manufactured in it. While there is a perception that a tumorigenic cell represents a safety concern, there are few data available to answer this question.

One of the goals of Project 3 is to evaluate whether the tumorigenicity of a cell has relevance to safety or is a characteristic of the cell that needs to be documented. In other words, if the cell line can be documented to be free from adventitious agents, and the residual cellular DNA in the product manufactured in that cell line is of an amount and size that is considered acceptable, should the fact that a cell line is tumorigenic preclude its use as a cell substrate for vaccine manufacture?

An additional goal of this work is to identify biomarkers for the acquisition of a tumorigenic phenotype by cell substrates. Such biomarkers would both reduce the cost to sponsors and the use of animals.


NPJ Vaccines 2020 Jun 18;5:52
WHO informal consultation on the guidelines for evaluation of the quality, safety, and efficacy of DNA vaccines, Geneva, Switzerland, December 2019.
Sheets R, Kang HN, Meyer H, Knezevic I, Sheets R, Kang HN, Meyer H, Knezevic I, Abwao E, Alali M, Aprea P, Bae C, Blades CDRZ, Boyer J, Broderick KE, Duffy P, Farnsworth A, Gangakhedkar J, Gutsch D, Hafiz RA, Jackson N, Kaslow D, Khan AS, Ledgerwood J, Liu MA, Maslow J, Nkansah E, Park Y, Patel A, Peden K, Racine T, Rose N, Roy P, Song M, Wei W

Vaccine 2020 Feb 18;38(8):1869-80
The science of vaccine safety: summary of meeting at Wellcome Trust.
Plotkin SA, Offit PA, DeStefano F, Larson HJ, Arora NK, Zuber PLF, Fombonne E, Sejvar J, Lambert PH, Hviid A, Halsey N, Garcon N, Peden K, Pollard AJ, Markowitz LE, Glanz J

NPJ Vaccines 2019 Oct 14;4:42
Harmonization of Zika neutralization assays by using the WHO International Standard for anti-Zika virus antibody.
Mattiuzzo G, Knezevic I, Hassall M, Ashall J, Myhill S, Faulkner V, Hockley J, Rigsby P, Wilkinson DE, Page M, collaborative study participants

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):5481-6
Development of a micro-neutralization assay for ebolaviruses using a replication-competent vesicular stomatitis hybrid virus and a quantitative PCR readout.
Lee SS, Phy K, Peden K, Sheng-Fowler L

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

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


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