Development of tuberculosis imaging probes based on sugars and proteins

Thesis

Tuberculosis (TB) is one of the oldest known human diseases, caused by Mycobacterium tuberculosis. The techniques currently employed for diagnosis are generally insensitive and non-specific. Positron emission tomography (PET) combined with computed tomography (CT) is used extensively in clinics and is among the advanced molecular imaging modalities for disease diagnosis and for monitoring the response to treatment.

In this thesis we described new probes based on sugars and proteins. We used an optimised biocatalytic approach for the synthesis of 2-fluoro-2-deoxy-D-trehalose (FDT). We scaled up synthesis for pre-clinical studies giving strong consideration to the expression host for enzymes. The use of rare biocatalytic methods employed in the synthesis was unique for the development of a radioisotope probe.

We then sought to develop an LC-MS based method for the detection of FDT in vitro and in vivo. Toxicity of the synthesised sugar compound was assessed in two different animal models. FDT administered to naïve marmosets labeled very little tissue in the lung, but when the animals were infected with M. tuberculosis, the tubercular lesions took up the radiolabelled FDT and [¹⁸F] FDT monomycolate was isolated from tubercular lesions indicating that the probe reached the site of bacterial incorporation. In addition, FDT labelling of tubercular lesions was reduced with 4-drug standard TB treatment in marmosets. This work has permitted the development of specific PET imaging probe for TB diagnosis towards the clinic.

We demonstrated that a connvergent protein synthesis using a "tag-and-modify" approach allowed the creation of homogenous, modified CCL2 variants bearing dual labels. We incorporated an unnatural amino acid azidohomoalanine (Aha) in the wild type CCL2 to obtain modified variants. Through initial work we have shown the successful modification of various potential PET imaging probes for TB diagnosis.

Authors: Khan, RMN

• DOI: 10.5287/ora-zg0g11qpx
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: Azidohomoalanine; enzymatic sugar synthesis; PET; FDT; Imaging; Tuberculosis; protein chemistry; CCL2; biocatalyst
• Subjects: Radiochemistry; Chemistry, Organic; Chemical Biology