In an attempt to exploit the hydrolytic mechanism by which β-lactamases degrade cephalosporins, we designed and synthesized a series of novel cephalosporin prodrugs aimed at delivering thiol-based... Show moreIn an attempt to exploit the hydrolytic mechanism by which β-lactamases degrade cephalosporins, we designed and synthesized a series of novel cephalosporin prodrugs aimed at delivering thiol-based inhibitors of metallo-β-lactamases (MBLs) in a spatiotemporally controlled fashion. While enzymatic hydrolysis of the β-lactam ring was observed, it was not accompanied by inhibitor release. Nonetheless, the cephalosporin prodrugs, especially thiomandelic acid conjugate (8), demonstrated potent inhibition of IMP-type MBLs. In addition, conjugate 8 was also found to greatly reduce the minimum inhibitory concentration of meropenem against IMP-producing bacteria. The results of kinetic experiments indicate that these prodrugs inhibit IMP-type MBLs by acting as slowly turned-over substrates. Structure–activity relationship studies revealed that both phenyl and carboxyl moieties of 8 are crucial for its potency. Furthermore, modeling studies indicate that productive interactions of the thiomandelic acid moiety of 8 with Trp28 within the IMP active site may contribute to its potency and selectivity. Show less
Targeted photodynamic therapy (PDT) has the potential to selectively damage tumor tissue and to increase tumor vessel permeability. Here we characterize the tissue biodistribution of two EGFR... Show moreTargeted photodynamic therapy (PDT) has the potential to selectively damage tumor tissue and to increase tumor vessel permeability. Here we characterize the tissue biodistribution of two EGFR-targeted nanobody-photosensitizer conjugates (NB-PS), the monovalent 7D12-PS and the biparatopic 7D12-9G8-PS. In addition, we report on the local and acute phototoxic effects triggered by illumination of these NB-PS which have previously shown to lead to extensive tumor damage.Methods: Intravital microscopy and the skin-fold chamber model, containing OSC-19-luc2-cGFP tumors, were used to investigate: a) the fluorescence kinetics and distribution, b) the vascular response and c) the induction of necrosis after illumination at 1 or 24 h post administration of 7D12-PS and 7D12-9G8-PS. In addition, dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) of a solid tumor model was used to investigate the microvascular status 2 h after 7D12-PS mediated PDT.Results: Image analysis showed significant tumor colocalization for both NB-PS which was higher for 7D12-9G8-PS. Intravital imaging showed clear tumor cell membrane localization 1 and 2 h after administration of 7D12-9G8-PS, and fluorescence in or close to endothelial cells in normal tissue for both NB-PS. PDT lead to vasoconstriction and leakage of tumor and normal tissue vessels in the skin-fold chamber model. DCE-MRI confirmed the reduction of tumor perfusion after 7D12-PS mediated PDT. PDT induced extensive tumor necrosis and moderate normal tissue damage, which was similar for both NB-PS conjugates. This was significantly reduced when illumination was performed at 24 h compared to 1 h after administration.Discussion: Although differences were observed in distribution of the two NB-PS conjugates, both led to similar necrosis. Clearly, the response to PDT using NB-PS conjugates is the result of a complex mixture of tumor cell responses and vascular effects, which is likely to be necessary for a maximally effective treatment. Show less
Food for human consumption is screened widely for the presence of antibiotic-resistant bacteria to assess the potential for transfer of resistant bacteria to the general population. Here, we... Show moreFood for human consumption is screened widely for the presence of antibiotic-resistant bacteria to assess the potential for transfer of resistant bacteria to the general population. Here, we describe an Enterobacter cloacae complex isolated from imported seafood that encodes two carbapenemases on two distinct plasmids. Both enzymes belong to Ambler class A β-lactamases, the previously described IMI-2 and a novel family designated FLC-1. The hydrolytic activity of the novel enzyme against aminopenicillins, cephalosporins, and carbapenems was determined. Show less
