Summary of research presentation by Adrian Krainer (Cold Spring Harbor Laboratory, NY) at the 2005 FightSMA conference

The SMN2 gene is an excellent therapeutic target because it has the potential to produce normal SMN protein. However, due to a process called RNA splicing, a very small, but very important, piece of the SMN2 gene is not made into protein. Briefly, the DNA that encodes the SMN2 gene produces a pre-mRNA transcript. This is essentially an exact duplicate of the DNA but has now been copied into a slightly different molecular form called RNA. RNA is what is used to make protein (DNA cannot be used for this process), however, only a small percentage of the actual pre-mRNA contains the information that is used in the production of a protein. As an example, this pre-mRNA could be equated to a book that has 20 chapters, however, the information that is important for a particular recipe is found in chapters 1, 4, 9, and 20, the remaining chapters are simply junk and can be discarded. This is essentially what happens in RNA splicing. The important regions that are the instructions for making a particular protein are encoded in “exons” and the sequences in between are called “introns.” The cell can find the exons, bring them together, and remove the intron sequences, forming an RNA that is ready to make a protein. Unfortunately, in the case of SMN2, there is a mistake that tells that cell to throw out the final “chapter” or specifically, exon 7. Dr. Krainer has previously very elegantly shown that the basis for this error in assembling the SMN RNA is due to a disruption in a binding site for a protein that tells the cell’s machinery to include exon 7 in the final RNA.

The protein that is responsible for telling the cell that SMN exon 7 is an actual exon (and important “chapter”) is called SF2/ASF. This protein has two parts: 1) a large region that binds a specific RNA sequence such as the one present in SMN1 exon 7; and 2) a smaller section that is an “activator” domain. The activator domain is the portion of the protein that is directly involved making sure that SMN exon 7 is included in the final spliced RNA. The SF2/ASF protein and others are essential for producing the full-length SMN protein, however, it is highly unlikely that a therapy for SMA (or any other disorder) would involve treating a cell with a full-sized splicing factor such as SF2/ASF. To this end, Dr. Krainer and colleagues have developed small novel synthetic molecules that are designed to stimulate SMN2 expression. These molecules are not drugs per se, rather they are rationally designed molecules that are conceptually similar to SF2/ASF. The molecules are comprised of an RNA-binding domain and a potent activation domain, however these domains are significantly smaller than the similar domains found in SF2/ASF.

A critical distinction between the normal SF2/ASF protein in the novel small molecules is that the “RNA binding domain” of the small molecules is an anti-sense molecule. Anti-sense technology is based upon the biochemical nature of nucleic acids. Briefly, DNA and RNA are comprised of small building blocks called bases. There are only four bases (G, A, T, C; U substitutes for T in RNA sequences) and there are rules for how the bases interact with one another. For example, in double-stranded DNA, G forms a bond with C and T (or U in RNA) forms a bond with A. Based upon these principles, Dr. Krainer and colleagues have develop short nucleic acid molecules that are the “complement” to various regions throughout SMN exon 7 and they have systematically analyzed anti-sense molecules that bind to SMN exon 7 to identify the ones that result in the highest levels of full-length (good) SMN. This has been performed in collaboration with ISIS pharmaceuticals, a company that specializes in anti-sense technology. This approach has identified 3 anti-sense molecules that appear to change the splicing pattern of SMN2, resulting in high levels of full-length SMN from the SMN2 gene.
Read the original summary