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|Thesis Advisor||Bell, Thomas W|
|Date of Issue||2021|
|Description||Cyclotriazadisulfonamide (CADA) prevents HIV entry into host cells by down-modulating CD4 receptor expression. The model for binding the CADA compounds involves a dominant dipole- dipole interaction between one of the side arms and a glutamine residue (Q15) in the hydrophobic (H) region of the CD4 signal peptide. There are three different regions in the CADA molecule that may actively bind with the signal peptide. These three regions are the isobutylene head group, the arenesulfonyl sidearms, and the tail group. Different modifications were previously done to these regions of CADA analogs to investigate structure-activity relationships (SAR). This study aims to modify the tail and the sidearm of the CADA compounds to find the structure-activity relationship for the down-modulation of sortilin, human-CD4, and ACE2.Interestingly, the signal peptide-containing protein sortilin was identified as a new target for CADA. Sortilin is a transmembrane protein similar to hCD4 that shuttles between the cell surface. Both cellular expression and in vitro co-translational translocation of sortilin were significantly reduced by CADA, similar to hCD4. My research aim is to target the signaling sequence of sortilin and ACE2 using newly developed CADA analogs. A negatively charged aspartate in the sortilin sequence at position 35 may play an essential role in binding with the carboxylic acid tail in SG analogs. A glutamine residue is also present at position 25 in the hydrophobic (H) region, which may play a similar role to Q15 in the hCD4 signal peptide. Similarly, the cationic arginine residue at positions 36 could play an essential role in binding. The strategy is to design CADA analogs that can bind with the signal sequence of sortilin to down- modulate its expression by forming a salt bridge with the amino acid residues present in the sortilin sequence. In this study, the current CADA analog’s benzyl tail is replaced with a long chain carboxylic acid tail to synthesized 14 different new CADA analogs. Among the synthesized 14 ii new analogs, SG001, SG003, SG010, SG006, and SG008 have an ester functional group at the tail region, while SG002, SG004, and SG011 has a carboxylic acid tail. In addition to this, the debenzylated analogs SG005 and SG012 are synthesized as an intermediate. We propose that the carboxylic acid group form an ion pair and/or hydrogen bond with glutamine Q25 in the signal sequence of sortilin and increase drug potency for sortilin down-modulation. We also hypothesized the carboxylic acid tail is essential. It may form a salt bridge with arginine 36 or a hydrogen bond interaction with aspartic acid 35, passing the signal peptidase cleavage site between serine 33 and glutamine 34. The signal peptide-containing protein ACE2 is also a new target for CADA. ACE2 is a monocarboxypeptidase, a part of the renin-angiotensin-aldosterone system (RAAS). ACE2 is responsible for converting Ang II to Ang (1-7), which has some beneficial effects on the human body. However, ACE2 is a receptor for SARS-CoV2 to enter the human alveolar cell. So, the drug that can downmodulate the expression of ACE2 could be potential therapeutics for SARS-CoV2. This study involves screening of newly synthesized SG analogs in Calu-3 cells to see the down- modulation of ACE2 using the peptide Mca-APK-Dnp as a fluorescence substrate. The screening result shows that SG analogs with long hydrophobic ester tail are more potent than the carboxylic acid tail analogs.|
|Title||Synthesis of CADA Analogs for Down-Modulation of Sortilin, CD4 and ACE2|
|Degree Level||Master's Degree|