The bone marrow cells containing progenitors of the various blood cell lineages are cultured in a semisolid matrix (for example, ReachBio’s ColonyGEL™) with appropriate cytokines to induce the progenitor cells to divide and differentiate into morphologically distinct colonies of mature cell types. The addition of test molecules to this system can be used to detect compound-induced changes in both number of colonies formed (quantitative) or colony size and morphology (qualitative), both of which infer toxicity (Figure 1).
By using combinations of cytokines that allow the simultaneous growth of both myeloid (granulocytic) and erythroid (red blood) cell colonies, it can be determined if a compound’s toxicity is restricted to a specific lineage or if the damage is more generalized. Another significant advantage of this assay system is that it allows for the simultaneous testing of multiple compounds, and analyses of colony numbers can predict whether compounds in combination will have additive or synergistic toxicity to the bone marrow compartment.
This may be of particular importance when developing new therapeutics for patients who are likely to have been heavily pretreated with traditional chemotherapeutic compounds (unintentional combination therapy) or new therapeutics that are designed to be used with other compounds to enhance the efficacy of both (intentional combination therapy).
Although many animal models (in vitro and in vivo) are useful tools for toxicity assessment, studies have revealed significant differences between human, dog, rat, and mouse hematopoietic progenitor cells with regard to their sensitivities to certain pharmaceuticals.
In particular, there are some classes of compounds where a curative dose (blood levels) in mice against a human tumor xenograft may not be achievable in patients due to a higher sensitivity of normal bone marrow cells in humans than in mice. For these reasons, some groups have looked at ratios of the human and mouse values for myeloid progenitor (CFU-GM) growth for specific drug classes (the camptothecins, for example).
The European Centre for the Validation of Alternative Methods recently validated the CFU-GM assay and suggested that through the use of the ratio between mouse and human IC90 CFU-GM values as well as the maximum tolerated dose (MTD) of the compound in mice, the MTD in patients could be predicted.