One of the earliest examples of a vaccine biomarker assay still in use is the polio antibody neutralization assay, which evaluates the immunogenicity of different polio vaccines. This assay is used to support concomitant vaccine clinical trials and developing world implementation studies.
At the just-concluded Cambridge Healthtech “Biomarker Discovery Summit” in Philadelphia, Mark T. Esser, Ph.D., senior research fellow, vaccines and biologics research at Merck, noted that vaccine biomarker assays have been widely used for many years to evaluate both the immunogenicity and the efficacy of vaccines in clinical trials and postmarketing studies.
A successful example of using a vaccine biomarker assay in the development of a vaccine, Dr. Esser said, is the serum bactericidal assay developed for Neisseria meningitidis type C. The results from this biomarker assay were used to license a Meningitis C conjugate vaccine in the U.K., without performing a clinical efficacy study.
Dr. Esser’s talk focused on vaccines, biomarkers, and correlates of protection. Multiplexed antibody biomarker assays use either spotting array technologies or microsphere array technologies to monitor the antibody response to multiple antigens simultaneously.
Merck’s HPV serology assay is based on Luminex’ xMAP technology, which uses multicolored microspheres and the principles of flow cytometry. The technology enables users to perform up to 100 biomarker assays simultaneously on a single sample, Dr. Esser noted. The Merck HPV serology assay simultaneously measures the immune antibody response to neutralizing epitopes on several virus-like particle (VLP) types. Multiplexing an assay allows a generation of more results, quickly and cost effectively compared to running traditional single-test biomarker assays.
Correlates of Protection
The FDA defines a correlate of protection as a laboratory parameter that has been shown, from adequate and well-controlled clinical trials, to be associated with protection from clinical disease, Dr. Esser continued.
Correlates of protection exist for some older vaccines such as Recombivax™ and Energix® for heptatitis B, for which the internationally agreed upon correlate of protection is 10 ml of anti-Hep B surface antigen antibodies. For newer vaccines, such as Gardasil™ for the prevention of cervical cancer, and Rotateq™ for the prevention of rotavirus-induced diarrhea, correlates of protection have not yet been determined, he noted.
Several different biomarker assays were used in the development of Gardisil™. Virus neutralization assays, multiplexed Luminex-based antibody assays, T cell immunology assays, and molecular assays for detecting HPV type-specific infections were all developed to support both pre-clinical animal studies and Phase I–III clinical trials, Dr. Esser explained.
Early in development, serology assays showed that a quadrivalent HPV type 6, 11, 16, and 18 VLP vaccine formulated on Merck’s aluminum adjuvant was immunogenic and elicited T cell responses and both serum and mucosal antibodies in animal models. In clinical development, an HPV 6, 11, 16, and 18 multiplexed, competitive Luminex® Immunoassay was used to select the optimal formulation in dose ranging studies, to show the long-term duration of antibodies following vaccination and the presence of immune memory.
Molecular HPV genotyping assays were also used in clinical development to determine whether cervical or genital lesions were caused by HPV 6, 11, 16, 18, or nonvaccine HPV types. More importantly, the immunology assay was used in an immunobridging study in 9–15 year-old females. The results indicated that Gardasil was safe and immunogenic in this population. The antibody biomarker data were included in the new drug application for Gardasil in adolescent females.
In her presentation, Nandini Raghavan, Ph.D., the principal biostatistician in the nonclinical biostatistics department at Johnson & Johnson Pharmaceutical Research & Development, described the development of a gene-expression based signature to predict nongenotoxic carcinogenicity (NGTC) with high accuracy using 24-hour microarray experiments on rats. Dr. Raghavan pointed out that this is especially critical, since short-term assays for nongenotoxic carcinogenicity—which is commonly observed in long-term rodent studies—have proven difficult to develop. Prime objectives of the study were development, validation, and standardization of biomarkers for safety evaluation.
Dr. Raghavan noted that high-throughput genomic technologies are being introduced at a rapid pace, and that these require sophisticated analysis strategies.
“This represents a paradigm shift and data analysis software tools cannot be used,” she stated. “We’re interrogating 35,000 sequences at a time, so the chance of picking up false positives is quite high. Overfitting is a significant problem, leading to nonreproducible results.”