Date on Master's Thesis/Doctoral Dissertation


Document Type

Doctoral Dissertation

Degree Name

Ph. D.



Degree Program

Chemistry, PhD

Committee Chair

Nantz, Michael

Committee Member

Handa, Sanchin

Committee Member

Ramezanipour, Farshid

Committee Member

Shapiro, Bruce

Author's Keywords

PS-750-M; oxime ether lipid; gene silencing; lung cancer; RNA interference; boronic acid-containg cationic lipids


RNA-based therapeutics is a rapidly expanding field due to its enormous potential for treatment of diseases through knockdown of genes or expression of therapeutic proteins. However, due to the overall negative-charge of an RNA molecule, RNA-based therapeutics must rely on delivery systems to overcome the various biological barriers for ultimate release of an RNA payload into the cytosol. Over the past three decades, the development of lipid-based RNA delivery systems, especially lipid nanoparticles (LNPs), have been comprehensively studied due to their unique properties. LNPs represent the most widely used delivery systems for RNA-based therapeutics, as evidenced by the clinical approvals of three LNP-RNA formulations: patisiran (ONPATTRO®), and BNT162b2 and mRNA-1273, which are LNP-encapsulated mRNA-based vaccines for prevention of COVID-19. Cationic lipids are also considered suitable candidates for RNAi therapeutics delivery. To this end, the present thesis work centers on examining liposome formulations of oxime ether (OELs) containing hydroxylated head groups in animal studies to validate their suitability as in vivo RNA carriers.

This thesis work also examines the effects of adding a boron-containing lipid to OEL4-based formulations. Conjugation chemistry, particularly oximation, is a critical component for the development of these lipids. Oximation involves the reaction of an aminooxy group with a carbonyl group of an aldehyde or ketone to form an oxime ether. The versatility of oximation is highlighted and exploited in this thesis. In addition, as part of my initial studies on micelles, the synthesis and synthetic applications of a novel, environmentally benign surfactant developed to form micelles containing polar cores is also described.

This thesis is divided into two parts. Part I focuses on the synthesis and applications of PS-750-M, a surfactant developed to enable organic synthesis transformations in water. Accordingly, Chapter 1 reviews the historical development of micellar catalytic processes.

Chapter 2 details the synthesis of the surfactant PS-750-M and its application in Suzuki-Miyaura couplings of unactivated quinoline systems. PS-750-M contains a proline-based linker between its hydrophobic and hydrophilic regions that imparts polarity to the micellar core, which is presumed to play a significant metal-coordinating role in the micellar Suzuki-Miyaura coupling reactions. PS-750-M functions as a green solvent that mimics toxic dipolar-aprotic solvents, such as DMF, DMAc, NMP, and 1, 4-dioxane. The micellar reaction medium derived from PS-750-M is demonstrated to be recyclable.

Part II focuses on the use of hydroxylated oxime ether lipids (OELs) as delivery agents for RNAi therapeutics. We first explore the in vivo utility of OELs. We found that OEL4 liposomes formulated using 3 mol% DSPE-PEG350 accumulate in tumors at highest efficiency as compared to other formulations and exhibited significant increases in tumor to liver ratios, indicative of improved delivery selectivity. The PEGylated liposomes also showed a statistically significant luciferase signal reduction in tumors as compared to an untreated mice control group.

Chapter 3 reviews the structural features of cationic lipids in siRNA/DNA therapy and contrasts it with novel features of cationic OELs. A modified synthesis of OELs that proceeds in 7 steps in overall 13% yield, constituting a 4-fold improvement over the previously reported synthesis, is also presented. The formulation and results of in vitro and in vivo transfection studies using OEL4 are presented in Chapter 4. Our studies show that the surface modification of OEL4 was essential for tumor accumulation in mice bearing human lung cancer xenografts. The surface-modified OEL4 formulations were developed by inclusion of distearoylphosphatidylethanolamine (DSPE) bound to varying lengths of poly(ethylene glycol) (PEG) polymer. The results of in vivo studies on mice bearing human lung cancer (A549-luc2) determined the optimal OEL4 formulation for delivery of an RNA payload to be the formulation OEL4:DOPE:DSPE-PEG350 (48.5:48.5:3). Chapter 5 discusses the synthesis of boron-containing lipids and preliminary results of boron-assisted RNA transfection studies using both oxime ether- and boron lipid-derived formulations. The results of in vitro transfection studies using oxime ether- and boron lipid-derived formulations established that inclusion of the novel boronic acid-containing cationic lipid DMDBH to an OEL4 lipoplex formulation enhances the gene silencing of capability of OEL4 by about 20%. The action of DMDBH in this regard remains unknown; however, nmr studies of OEL4 and DMDBH suggest dimerization of DMDBH and OEL4, which could increase the strength of the interaction with RNA and increase lipoplex stability. Chapter 6 presents detailed experimental procedures of the chemistry and biology described in Chapters 2, 3, 4 and 5. Supporting spectroscopic data of synthesized compounds are also included in Chapter 6.

Available for download on Wednesday, August 10, 2022