Biology subjects

McMahon, M. T.

Publications and source records attributed to McMahon, M. T..

3 recordsLinked to original sources

bioRxiv2025

Biocatalytic Construction of a CEST MRI Nucleoside Probe: Synthesis and Evaluation of 5-Methyl-5,6-dihydrothymidine

Magnetic Resonance Imaging (MRI) is a cornerstone of modern clinical diagnostics, often enhanced by contrast agents. Traditionally, these agents are chemically synthesized, which can involve complex, costly, and environmentally unfriendly processes. Here, we report a novel biocatalytic approach for the efficient, safe, and eco-friendly synthesis of 5-methyl-5,6-dihydrothymidine (5-MDHT), a potent Chemical Exchange Saturation Transfer (CEST) MRI probe for imaging in vivo expression of the Herpes Simplex Virus Type-1 Thymidine Kinase (HSV1-TK) reporter gene. We demonstrate that 5-MDHT can be biosynthesized via one- or two-step enzymatic reactions using human purine nucleoside phosphorylase (hPNPase) and the SgvMVAV SAM-dependent methyltransferase. hPNPase catalyzed the base-exchange reaction with catalytic efficiencies (kcat/KM) between 138-316 s-1 M-1, while SgvMVAV methylation of 5,6-dihydrothymidine yielded 5-MDHT with a catalytic efficiency of 26 s-1 M-1. Molecular dynamics simulations supported the enzymatic binding and selectivity observed experimentally. The resulting 5-MDHT was validated using CEST-MRI, showing a distinct exchangeable imino proton signal at 5.3 ppm. These findings highlight the chemo- and regioselectivity of the biocatalysts and establish biocatalysis as a viable platform for producing clinically relevant MRI contrast agents.

biochemistry↗

bioRxiv2024

The Proton Resonance Enhancement for CEST imaging and Shift Exchange (PRECISE) family of RF pulse shapes for Chemical Exchange Saturation Transfer MRI

PurposeTo optimize a 100 msec pulse for producing CEST MRI contrast and evaluate in mice. MethodsA gradient ascent algorithm was employed to generate a family of 100 point, 100 msec pulses for use in CEST pulse trains ( PRECISE). Gradient ascent optimizations were performed for exchange rates (kca) = 500 s-1, 1,500 s-1, 2,500 s-1, 3,500 s-1 and 4,500 s-1 and offsets ({Delta}{omega}) = 9.6, 7.8, 4.2 and 2.0 ppm. 7 PRECISE pulse shapes were tested on an 11.7 T scanner using a phantom containing three representative CEST agents with peak saturation B1 = 4 T. The pulse producing the most contrast in phantoms was then evaluated for CEST MRI pH mapping of the kidneys in healthy mice after iopamidol administration. ResultsThe most promising pulse in terms of contrast performance across all three phantoms was the 9.6 ppm, 2500 s-1 optimized pulse with [~]2.7 x improvement over Gaussian and [~]1.3xs over Fermi pulses. This pulse also displayed a large improvement in contrast over the Gaussian pulse after administration of iopamidol in live mice. ConclusionA new 100 msec pulse was developed based on gradient ascent optimizations which produced better contrast compared to standard Gaussian and Fermi pulses in phantoms. This shape also showed a substantial improvement for CEST MRI pH mapping in live mice over the Gaussian shape and appears promising for a wide range of CEST applications.

bioengineering↗

bioRxiv2020

Redesigned Reporter Gene for Improved Proton Exchange-based Molecular MRI Contrast

Reporter gene imaging allows for non-invasive monitoring of molecular processes in living cells, providing insights on the mechanisms underlying pathology and therapy. A lysine-rich protein (LRP) chemical exchange saturation transfer (CEST) MRI reporter gene has previously been developed and used to image tumor cells, cardiac viral gene transfer, and oncolytic virotherapy. However, the highly repetitive nature of the LRP reporter gene sequence leads to DNA recombination events and the expression of a range of truncated LRP protein fragments, thereby greatly limiting the CEST sensitivity. Here we report the use of a redesigned LRP reporter (rdLRP), aimed to provide excellent stability and CEST sensitivity. The rdLRP contains no DNA repeats or GC rich regions and 30% less positively charged amino-acids. RT-PCR of cell lysates transfected with rdLRP demonstrated a stable reporter gene with a single distinct band corresponding to full-length DNA. A distinct increase in CEST-MRI contrast was obtained in cell lysates of rdLRP transfected cells and in in vivo LRP expressing mouse brain tumors (p=0.0275, n=10).

bioengineering↗