Identification of Novel Mt-Guab2 Inhibitor Series Active against M. tuberculosis

Tuberculosis (TB) remains a leading cause of mortality worldwide. With the emergence of multidrug resistant TB, extensively drug resistant TB and HIV-associated TB it is imperative that new drug targets be identified. The potential of Mycobacterium tuberculosis inosine monophosphate dehydrogenase (IMPDH) as a novel drug target was explored in the present study. IMPDH exclusively catalyzes the conversion of inosine monophosphate (IMP) to xanthosine monophosphate (XMP) in the presence of the cofactor nicotinamide adenine dinucleotide (NAD+). Although the enzyme is a dehydrogenase, the enzyme does not catalyze the reverse reaction i.e. the conversion of XMP to IMP. Unlike other bacteria, M. tuberculosis harbors three IMPDH-like genes, designated as Mt-guaB1, Mt-guaB2 and Mt-guaB3 respectively. Of the three putative IMPDH's, we previously confirmed that Mt-GuaB2 was the only functional ortholog by characterizing the enzyme kinetically. Using an in silico approach based on designed scaffolds, a series of novel classes of inhibitors was identified. The inhibitors possess good activity against M. tuberculosis with MIC values in the range of 0.4 to 11.4 µg mL−1. Among the identified ligands, two inhibitors have nanomolar Kis against the Mt-GuaB2 enzyme

DNA building blocks: keeping control of manufacture

Ribonucleotide reductase (RNR) is the only source for de novo production of the four deoxyribonucleoside triphosphate (dNTP) building blocks needed for DNA synthesis and repair. It is crucial that these dNTP pools are carefully balanced, since mutation rates increase when dNTP levels are either unbalanced or elevated. RNR is the major player in this homeostasis, and with its four different substrates, four different allosteric effectors and two different effector binding sites, it has one of the most sophisticated allosteric regulations known today. In the past few years, the structures of RNRs from several bacteria, yeast and man have been determined in the presence of allosteric effectors and substrates, revealing new information about the mechanisms behind the allosteric regulation. A common theme for all studied RNRs is a flexible loop that mediates modulatory effects from the allosteric specificity site (s-site) to the catalytic site for discrimination between the four substrates. Much less is known about the allosteric activity site (a-site), which functions as an on-off switch for the enzyme's overall activity by binding ATP (activator) or dATP (inhibitor). The two nucleotides induce formation of different enzyme oligomers, and a recent structure of a dATP-inhibited α6β2 complex from yeast suggested how its subunits interacted non-productively. Interestingly, the oligomers formed and the details of their allosteric regulation differ between eukaryotes and Escherichia coli Nevertheless, these differences serve a common purpose in an essential enzyme whose allosteric regulation might date back to the era when the molecular mechanisms behind the central dogma evolved

Sequence analysis of beta-subunit genes of the 20S proteasome in patients with relapsed multiple myeloma treated with bortezomib or dexamethasone

Variations within proteasome beta (PSMB) genes, which encode the beta subunits of the 20S proteasome, may affect proteasome function, assembly, and/or binding of proteasome inhibitors. To investigate the potential association between PSMB gene variants and treatment-emergent resistance to bortezomib and/or long-term outcomes, in the present study, PSMB gene sequence variation was characterized in tumor DNA samples from patients who participated in the phase 3 Assessment of Proteasome Inhibition for Extending Remissions (APEX) study of bortezomib versus high-dose dexamethasone for treatment of relapsed multiple myeloma. Twelve new PSMB variants were identified. No associations were found between PSMB single nucleotide polymorphism genotype frequency and clinical response to bortezomib or dexamethasone treatment or between PSMB single nucleotide polymorphism allelic frequency and pooled overall survival or time to progression. Although specific PSMB5 variants have been identified previously in preclinical models of bortezomib resistance, these variants were not detected in patient tumor samples collected after clinical relapse from bortezomib, which suggests that alternative mechanisms underlie bortezomib insensitivity. This study is registered at www.clinicaltrials.gov as NCT00048230. (Blood. 2012;120(23):4513-4516