SINEUP™ can be used to up-regulate protein expression for any target mRNA. The gene encoding the protein can be endogenous or non-endogenous, and in any mammalian cell. SINEUP™ increases the efficiency of translation but does not affect mRNA levels. Therefore, SINEUP™ only works in cells where a gene is already being expressed. This targeting mechanism and lack of activity in cells where a gene is quiescent provide a very high degree of cell specificity, even in mixed cell populations.
SINEUP™ utilizes a plasmid construct, pDUAL, which can be transiently or stably transfected into cells. The SINEUP™ construct contains a H1 promoter which drives the expression of the SINEUP™ sequence. This is non-coding and contains two elements separated by a short spacer. The first of these two elements is the binding domain (BD), a short sequence (around 40 nucleotides) complementary to the target gene. The second element is an effector domain (ED) which contains sequences which (similar to IRES elements) recruits ribosomes to the target mRNA.
Importantly, SINEUP™ is not a gene editing technology, and the target gene operon is completely unaltered. Increases in protein levels are typically 1.5-fold, but increases as much as 10-fold have been observed. Although relatively small, these effects are sufficient to generate measurable phenotypic effects. The hundreds of haploinsufficiency disorders and syndromes associated with trisomy illustrate the long-term effects of subtle changes in expression levels.