Screening for small molecule activators of mitochondrial enzyme aldehyde dehydrogenase 1B1

Abstract

Aldehydes are known as a class of highly reactive molecules and one of the most important drivers of redox damage in living cells. Aldehyde levels are mainly controlled by enzymes from Aldehyde Dehydrogenase superfamily (ALDH), of which ALDH2 is the most well characterized member. The main acetaldehyde metabolizing enzyme is mainly known by a mutation (E540K, named ALDH2*2) present in about 40% of the east Asian population, which show a lower ALDH2 enzymatic activity (nearly 5% of the wild-type activity) and is associated with very severe deleterious phenotypes, such as the Asian flush reaction, increased cardiovascular disease morbimortality and cancer occurrence. In 2008, Daria Mochly-Rosen's lab at Stanford University developed a small molecule activator of aldehyde dehydrogenase 2, termed Alda-1, which increases ALDH2*2 activity by 11-fold. Since then, Alda-1 has been applied to a myriad of experimental disease models and tested in humans as well (phase 2 clinical trial).Sharing the mitochondrial environment with ALDH2 lies ALDH1B1, a lesser studied member of the ALDH family. Such as ALDH2, ALDH1B1 has several high prevalent cataloged non-synonymous mutations, but their consequence to enzyme activity and to the individual's biology are still elusive. Some evidence found in literature links a few genotypes to cardiovascular diseases, cancers, and alcohol intolerance, suggesting that ALDH1B1 may have an important role in these diseases and that some of these mutations may negatively affect its catalytic activity. Thus, we decided to characterize the biochemical and biological consequences of the seven most frequent non-synonymous mutations in ALDH1B1. Our preliminary results show that out the 7 target mutations, two of them (193fs and 388fs) yield a completely unstable protein when expressed in both bacteria (E. coli) and immortalized carcinoma cells (A375 cells); and other two (253v and 470a) display impaired catalytic activity. Moreover, we discovered that ALDH1B1 and ALDH2 are capable of physically associate to each other forming heterotetramers (chimeras), indicating that physical interaction between ALDH1B1 and ALDH2 monomers might either propagate or counteract inadvertent genetic variations in ALDH1B1 and ALDH2. Finally, we provide evidence that Alda-1 (and its derivates) increases catalytic activity of ALDH2:ALDH1B1 chimeras, but not ALDH1B1, depicting a lack of ALDH1B1 activators available for tests and treatments. The current application for internship abroad is a joint effort between Dr. Julio C.B. Ferreira and Dr. Daria Mochly-Rosen labs (Department of Chemical and Systems Biology Operations- Stanford University) with the primary objective as discovering a new molecule capable of activating ALDH1B1 enzyme. Noteworthy, the experimental strategy applied here will be the same strategy successfully used by the same research group for the discovery of Alda-1. This new molecule will be extremely useful on unraveling potential pathophysiological roles of ALDH1B1 and possibly being developed into a drug in the future. (AU)