Adapting high-throughput screening methods and assays for biocontainment laboratories

Abstract

High-throughput screening (HTS) has been integrated into the drug discovery process, and multiple assay formats have been widely used in many different disease areas but with limited focus on infectious agents. In recent years, there has been an increase in the number of HTS campaigns using infectious wild-type pathogens rather than surrogates or biochemical pathogen-derived targets. Concurrently, enhanced emerging pathogen surveillance and increased human mobility have resulted in an increase in the emergence and dissemination of infectious human pathogens with serious public health, economic, and social implications at global levels. Adapting the HTS drug discovery process to biocontainment laboratories to develop new drugs for these previously uncharacterized and highly pathogenic agents is now feasible, but HTS at higher biosafety levels (BSL) presents a number of unique challenges. HTS has been conducted with multiple bacterial and viral pathogens at both BSL-2 and BSL-3, and pilot screens have recently been extended to BSL-4 environments for both Nipah and Ebola viruses. These recent successful efforts demonstrate that HTS can be safely conducted at the highest levels of biological containment. This review outlines the specific issues that must be considered in the execution of an HTS drug discovery program for high-containment pathogens. We present an overview of the requirements for HTS in high-level biocontainment laboratories.

Fig. 1.

The optimization of high-throughput screening (HTS) liquid handler size. (A) Shows the relative size of the Biomek 2000 that was used in early infectious disease screening in biosafety level BSL-2 laboratories. Later efforts that were adapted to BSL-3 and -4 laboratories made use of smaller noncontact dispensers, such as the Matrix WellMate (B).

Fig. 2.

Assay plate layout. Compound library plates contain 320 compounds in columns 3–22 rows A–P as indicated in yellow (A). Columns 1, 2, 23, and 24 are empty to accommodate assay controls. For antiviral screening in the BSL-3, compounds are transferred to the assay plates and DMSO is added to the control wells. Cell controls are added to column 1 and 2 with one dispenser, and cells plus virus mixed together are added to columns 3–24 with another dispenser (A). For screening in the BSL-4, this process was simplified further so that all components could be added with one dispenser in a single pass. This format is illustrated in (B). One line of the dispense head is used to dispense cells into the cell control wells (row A, B), while the remaining lines, which are in a separate reservoir, contain cells and virus mixed together, which are dispensed to the remainder of the plate that includes test compounds and virus control wells (B).

Fig. 3.

The amount of personal protective equipment (PPE) increases with BSL. An operator at a class II biosafety cabinet is shown at BSL-2, BSL-3/3+, and BSL-4. The differences in PPE (described in Table 1) are shown for each level. The operator is shown manipulating a small-footprint noncontact liquid dispenser.

Fig. 4.

The optimized HTS workflow for BSL-3/-4 containment laboratories. Six operational steps are outlined, separated by performance in low-containment (BSL-1/-2) or high-containment (BSL-3/-4). These steps are designed to minimize plate handling, operator workload, and equipment dedication in high-containment laboratories.