Research ProGrAMs

Targeting Essential Enzymes in Mycobacterium Species

We are investigating various pathways related to trehalose, a disaccharide and essential metabolite in Mycobacterium tuberculosis (Mtb). Several enzymes involved in trehalose utilization are essential for the organism’s survival, making them promising targets for therapeutic intervention.

Our research focuses on developing probe molecules to elucidate the detailed molecular interactions between these enzymes and their substrates. For instance, we are using carbohydrates and glycomimetics as components of substrate analogs and transition state analogs to target key enzymes, including:

Mtb Antigen 85s (Ag85s): Ag85s utilize trehalose monomycolate (TMM) to construct essential components of the mycobacterial cell wall.

GlgE: This enzyme is involved in alpha-glucan and capsule synthesis, both essential for Mtb viability and pathogenicity.

OtsB2: OtsB2 plays a critical role in producing trehalose from trehalose-6-phosphate. It is a promising antimicrobial drug target due to its essential function in Mtb metabolism.

Additionally, we have identified novel inhibitors of polyketide synthase Pks13, an enzyme responsible for both mycolic acid synthesis and the formation of TMM. In addition we have reported on the total synthesis of new puupehenone derivatives (terpenoid-based natural products) which are curiosly more active on dormant Mtb over actively replicating Mtb and also have potentent activity agaist Clostridioides difficile which can be a devistating infection in the elderly.

These molecular probes and inhibitors hold significant potential for uncovering new enzyme inhibitors that could be developed into treatments for multidrug-resistant Mtb, addressing a growing global health threat.

Figure 1. Trehalose utilization pathways — **Figure 1.** Trehalose utilization pathways

**Program 2. Oligosaccharide-based Immunotherapeutics. ** Carbohydrates found in naturally occurring glycoconjugates often serve as critical markers for cancer and infectious diseases, making them compelling targets for vaccine development. One notable example are the oligosaccharides that compose the lipopolysaccharide (LPS) of Pseudomonas aeruginosa, a significant ESKAPE pathogen.

Our research focuses on synthesizing homogeneous oligosaccharides derived from LPS for use in glycoconjugate vaccines. For instance, Figure 2 illustrates a conjugatable fragment from the core domain of *Pseudomonas aeruginosa *LPS.

In addition, we are developing innovative strategies to enhance vaccine immunogenicity by leveraging endogenous anti-L-rhamnose antibodies naturally present in humans. Our studies have also led to the synthesis, isolation, and formulation of new adjuvant combinations. By incorporating L-rhamnosides as antibody-recruiting molecules (ARMs) or integrating novel adjuvants formulations into vaccine designs, we aim to optimize immune responses.

See Figure 3 for an illustration of this concept.

Figure 3. Synthesis of a tetrasaccharide component of the lipopolysaccharide of P. aeruginosa. — **Figure 2.** Synthesis of a tetrasaccharide component of the lipopolysaccharide of *P. aeruginosa*.