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Preclinical PET animal studies require immobilization of the animal, typically accomplished through the administration of anesthesia, which may affect the radiotracer biodistribution. The use of 18F point sources attached to the rat head is one of the most promising methods for motion compensation in awake rat PET studies. However, the presence of radioactive markers may degrade image quality. In this study, we aimed to investigate the most favorable conditions for preclinical PET studies using awake rats with attached point sources. Firstly, we investigate the optimal activity conditions for the markers and rat-injected tracer using Monte Carlo simulations to determine the parameters of maximum detectability without compromising image quality. Additionally, we scrutinize the impact of delayed window correction for random events on marker detectability and overall image quality within these studies. Secondly, we present a method designed to mitigate the influence of rapid rat movements, which resulted in a medium loss of events of around 30%, primarily observed during the initial phase of the data acquisition. We validated our study with PET acquisitions from an awake rat within the acceptable conditions of activity and motion compensation parameters. This acquisition revealed an 8% reduction in resolution compared to a sedated animal, along with a 6% decrease in signal-to-noise ratio (SNR). These outcomes affirm the viability of our method for conducting awake preclinical brain studies.

Innovative cross-over study designs were explored in non-human primate (NHP) studies to determine the value of this approach for the evaluation of drug efficacy against tuberculosis (TB). Firstly, the pharmacokinetics (PK) of each of the drugs Isoniazid (H), Rifampicin (R), Pyrazinamide (Z) and Ethambutol (E), that are standardly used for the treatment of tuberculosis, was established in the blood of macaques after oral dosing as a monotherapy or in combination. Two studies were conducted to evaluate the pharmacokinetics and pharmacodynamics of different drug combinations using cross-over designs. The first employed a balanced, three-period Pigeon design with an extra period; this ensured that treatment by period interactions and carry-over could be detected comparing the treatments HR, HZ and HRZ using H37Rv as the challenge strain of Mycobacterium tuberculosis (M. tb). Although the design accounted for considerable variability between animals, the three regimens evaluated could not be distinguished using any of the alternative endpoints assessed. However, the degree of pathology achieved using H37Rv in the model during this study was less than expected. Based on these findings, a second experiment using a classical AB/BA design comparing HE with HRZ was conducted using the M. tb Erdman strain. More extensive pathology was observed, and differences in computerized tomography (CT) scores and bacteriology counts in the lungs were detected, although due to the small group sizes, clearer differences were not distinguished. Type 1 T helper (Th1) cell response profiles were characterized using the IFN-γ ELISPOT assay and revealed differences between drug treatments that corresponded to decreases in disease burden. Therefore, the studies performed support the utility of the NHP model for the determination of PK/PD of TB drugs, although further work is required to optimize the use of cross-over study designs.

Developing more efficient methods for antibiotic susceptibility testing is a pressing issue in novel drug development as bacterial resistance to antibiotics becomes increasingly common. Microfluidic devices have been demonstrated to be powerful platforms that allow researchers to perform multiplexed antibiotic testing. However, the level of multiplexing within microdevices is limited, evidencing the need of creating simple, low-cost and high-resolution imaging systems that can be integrated in antibiotic development pipelines. This paper describes the design and development of an epifluorescence inverted microscope that enables long-term monitoring of bacteria inside multiplexed microfluidic devices. The goal of this work is to provide a simple microscope powerful enough to allow single-cell analysis of bacteria at a reduced cost. This facilitates increasing the number of microscopes that are simultaneously used for antibiotic testing. We prove that the designed system is able to accurately detect fluorescent beads of 100 nm, demonstrating comparable features to high-end commercial microscopes and effectively achieving the resolution required for single-cell analysis of bacteria. The proposed microscope could thus increase the efficiency in antibiotic testing while reducing cost, size, weight, and power requirements, contributing to the successful development of new antibiotic drugs.

Real-time positron emission tomography (PET) may provide information from first-shot images, enable PET-guided biopsies, and allow awake animal studies. Fully-3D iterative reconstructions yield the best images in PET, but they are too slow for real-time imaging. Analytical methods such as Fourier back projection (FBP) are very fast, but yield images of poor quality with artifacts due to noise or data incompleteness. In this work, an image reconstruction based on the pseudoinverse of the system response matrix (SRM) is presented. w. To implement the pseudoinverse method, the reconstruction problem is separated into two stages. First, the axial part of the SRM is pseudo-inverted (PINV) to rebin the 3D data into 2D datasets. Then, the resulting 2D slices can be reconstructed with analytical methods or by applying the pseudoinverse algorithm again. The proposed two-step PINV reconstruction yielded good-quality images at a rate of several frames per second, compatible with real time applications. Furthermore, extremely fast direct PINV reconstruction of projections of the 3D image collapsed along specific directions can be implemented.

The synthesis of 9,9'-[1,2-ethanediylbis(oxymethylene)]bis-2-amino-1,9-dihydro-6H-purin-6-one, a minor impurity of acyclovir, is described. Starting with commercial N-(9-acetyl-6-oxo-1H-purin-2-yl)acetamide, the process uses an acid catalysed phase transfer catalysis (PTC) process to produce the selective alkylation at the 9 position of the guanine ring.