Biomarkers’ Role in COVID Research
Pharmacodynamic (PD) immune response biomarkers in vaccine studies and beyond
Biomarkers are playing a pivotal role both in guiding improvements in the clinical management of patients with COVID-19 and in world-wide vaccine development efforts. In a recent pre-print (‘COVID-19 Biomarkers in research’), Gogate et al sought to summarise the current state of COVID-19 biomarker knowledge. Prompted by the lack of a single resource, they suggest there is an urgent need within the research community for an integrated COVID-19 Biomarker Knowledgebase.
Using high school and undergraduate student volunteers, they crowdsourced a literature review and summarized the output into a list of 138 corroborated biomarkers. Their manually-collated resource found that the biomarkers most frequently cited related to the immune system (e.g. TNF-a, and IP-10), coagulopathies (e.g D-dimer) or established cancer biomarkers (e.g. ACE2 and IL-6).
The FDA-NIH Biomarker Working Group (FNBWG) recognises a number of different biomarker categories, namely; Diagnostic, Monitoring, Prognostic, Predictive, Pharmacodynamic/Response, Susceptibility/Risk and Safety. The COVID-19 biomarkers identified by the literature review of Gogate et al were classified into these different categories and thus have different roles in the clinical management of patients (for example, diagnosing the condition or predicting the likely disease severity). Whilst biomarkers used to inform routine patient management are run in hospital laboratories close to the point of care, in the context of vaccine clinical trials certain specialized biomarker tests (such as the quantification of antigen-specific T cells) may be included which require the use of more geographically distant, central immunology laboratories. In this scenario, the added complexities of sample shipment and logistics must be considered.
As the name suggests, Pharmacodynamic (or ‘Response’) Biomarkers are employed to measure the response of the body – in this case, the immune system - to a candidate drug or vaccine. PD biomarkers are typically included as exploratory endpoints in clinical trials and do not directly guide patient management. These endpoints are run in support of the primary aims of the early phase trial, which will often be to establish the safety profile of the new vaccine, with the ultimate ‘gold standard’ being the demonstration of clinical efficacy whereby immunised subjects are protected from subsequent infection or challenge. The exploratory, PD biomarker analysis normally requires that blood samples are obtained from trial subjects at various timepoints pre- and post-treatment, then analysed and compared to determine whether the desired immune response has been induced in vivo. This serves as ‘Proof of Mechanism’ confirmation for the proposed vaccine.
Broadly speaking, immune responses are commonly analysed at the protein/secreted soluble factor level or the cellular level. Where these tests are not available in local laboratories, assessment of soluble factors such as circulating antibodies (as evidence of humoral/ B cell immunity) and inflammatory cytokines can be performed on frozen serum or plasma. Sample freezing means that these assay types lend themselves to shipping from geographically distant sites and batching of samples for PD biomarker analysis (subject to appropriate assay validation). The assessment of T cell immunity induced by a vaccine can be measured by readouts such as flow cytometry and ELISpot analysis. Where direct ex vivo measurement of immune cell subsets is sufficient, flow cytometry of whole blood samples can be performed.
Delays in blood draw-to-processing due to shipment can be overcome by the use of fixatives, once it has been established that antigenic epitopes withstand this process. However, the situation for functional assays – such as any antigen-specific T cell assessments – is more complex. In this case, a period of ex vivo activation is often required to stimulate T cell function, meaning that prior sample fixation will not be tolerated. Consequently, other solutions must be investigated. These include the storage of peripheral blood mononuclear cells (PBMC) at laboratories local to the clinical centre, prior to frozen shipment to the immunology lab for activation and analysis post-thawing. Alternatively, activation of the samples may be performed in the local lab before the shipment step (or possible fixation or freezing steps) occurs.
Pre-analytical variation begins as soon as the blood sample has left the patient’s arm. An understanding of the immune response being modulated and the possible stability issues affecting analyte or cell types being measured helps to inform the experimental strategy and to de-risk sample handling solutions. Even for exploratory endpoints, reasonable efforts need to be made to establish that the proposed sample handling and experimental approaches generate data which is fit for the intended purpose.
PD biomarkers are one of the recognized sub-types of biomarkers. Whilst not directly in patient management, they can be employed as exploratory endpoints to demonstrate that a vaccine is inducing the desired antigen-specific immune responses after dosing in the context of clinical trials. Meaningful results can be generated even where shipping to geographically distant, specialized immunology labs is required, as long as logistics considerations are addressed during the validation phase.