Production of synthetic retinoids to control cell differentiation
A major challenge in stem cell biology is the ability to control the development of cells and tissues in a predictable way. Biotechnologists require molecules that induce reliable and reproducible biological activity resulting in consistent modes of cell differentiation.
Retinoids are naturally occurring derivatives of vitamin A, which play a major role in mammalian development, and are commonly used to induce the differentiation of cultured stem cells in the laboratory. However, it is not widely appreciated that such molecules, including the commonly used all trans form of retinoic acid, degrade readily. This degradation is difficult to avoid during the routine use of these molecules in cell culture and results in the formation of isomers that effect cell differentiation in alternative ways compared to the intact parent molecule. This in turn introduces a potential source of variation in the control of cell differentiation.
Molecular structure of all-trans retinoic acid (ATRA), a retinoid derivative most regularly used to induce the differentiation of cultured cells. Naturally occurring retinoids, including ATRA, 9 cis retinoic acid and 13 cis retinoic acid, can be described as chromophores since they selectively absorb light. Such molecules are essentially composed of three structurally distinct regions: a hydrophobic end, a polyene linker and an acidic group. The polyene linker in naturally occurring retinoids is highly conjugated and it is this region that gives it the ability to absorb light (at a frequency of 300 -400 nm depending on the solvent). It is due to this feature that these molecules are particularly susceptible to photoisomerisation and can degrade into a mixture of different retinoic acid isomers. The resulting concentration of retinoid levels has also been shown to decrease markedly over time in culture and this could be a consequence of both their degradation and metabolism. Moreover, ATRA is temperature sensitive and is known to oxidise readily.
To address this problem, we have designed and synthesised a small collection of well defined synthetic analogues of retinoic acid that have significantly improved chemical and physical stability and mimic the biological activity of retinoid derivatives currently used to induce cell differentiation. Retinoids are polyene-containing natural products and it is the combination of olefin geometries for these types of compound that has a profound effect on biological processes. We are developing methods for the stereo controlled synthesis of polyene building blocks from which we will assemble retinoid related polyenes for biological testing.
Overview of the retinoid development programme
The reagents we have produced so far possess demonstrated ability to modulate the differentiation of stem cells in a manner resembling naturally occurring retinoids. This programme of research is ongoing and will produce a range of well characterised small molecules that have proven ability to regulate cell development. With the benefit of improved stability, the use of these reagents will reduce cellular heterogeneity in cultures of differentiating cells. Stable, synthetic modulators of cell differentiation offer distinct advantages over existing technology and will be of significant commercial value.
We have developed a small library of well defined synthetic analogues of retinoic acid that have significantly improved chemical and physical stability and mimic the biological activity of retinoid derivatives currently used to induce cell differentiation. In the example above, Molecule A induced human pluripotent stem cells to form epithelial cells, whereas Molecule B induced the same cells to differentiate and produce neurons.