REVIEW | doi:10.20944/preprints202103.0050.v1
Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: antisense; antibiotic resistance; RNase P; RNase H; nucleotide analogs
Online: 2 March 2021 (09:17:47 CET)
Antisense technologies consist of the utilization of oligonucleotides or oligonucleotide analogs to interfere with undesirable biological processes, commonly through inhibition of expression of selected genes. This field holds a lot of promise for the treatment of a very diverse group of diseases including viral and bacterial infections, genetic disorders, and cancer. To date, drugs approved for utilization in clinics or in clinical trials target diseases other than bacterial infections. Although several groups and companies are working on different strategies, the application of antisense technologies to prokaryotes still lags with respect to those that target other human diseases. In those cases where the focus is on bacterial pathogens, a subset of the research is dedicated to produce antisense compounds that silence or reduce expression of antibiotic resistance genes. Therefore, these compounds will be adjuvants administered with the antibiotic to which they reduce resistance levels. A varied group of oligonucleotide analogs like phosphorothioate or phosphorodiamidate morpholino residues, as well as peptide nucleic acids, locked nucleic acids and bridge nucleic acids, the latter two in gapmer configuration, have been utilized to reduce resistance levels. The major mechanisms of inhibition include eliciting cleavage of the target mRNA by the host’s RNase H or RNase P, and steric hindrance. The different approaches targeted resistance to β-lactams including carbapenems, aminoglycosides, chloramphenicol, macrolides, and fluoroquinolones.
REVIEW | doi:10.20944/preprints202101.0178.v1
Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: metalloproteins; zinc transporters; metal chelators; antibiotic resistance; antimicrobials
Online: 11 January 2021 (10:20:01 CET)
Zinc is a redox-inert trace element that is second only to iron in abundance in biological systems. In cells, zinc is typically buffered and bound to metalloproteins, but may also exist as a labile or chelatable (free ion) form. Zinc plays a critical role in prokaryotes and eukaryotes ranging from structural to catalytic to replication to demise. This review discusses the influential properties of zinc on various mechanisms of bacterial proliferation and synergistic action as anti-microbial element. We also touch upon the significance of zinc among eukaryotic cells and how it may modulate their survival and death through its inhibitory or modulatory effect on certain receptors, enzymes, and signaling proteins. A brief discussion on zinc chelators is also presented and chelating agents may be used with or against zinc to affect therapeutics against human diseases. Overall, the multidimensional effects of zinc in cells attest to the growing numbers of scientific research that reveal the consequential prominence of this remarkable transition metal in human health and disease.
ARTICLE | doi:10.20944/preprints202103.0552.v1
Subject: Biology And Life Sciences, Anatomy And Physiology Keywords: Huma Serum Albumin; Acinetobacter baumannii; quorum sensing; iron; human fluids.
Online: 22 March 2021 (15:49:16 CET)
Acinetobacter baumannii is a nosocomial pathogen capable of causing serious infections associated with high rates of morbidity and mortality. Due to its antimicrobial drug resistance profile, A. baumannii is categorized as an urgent priority pathogen by the Centers for Disease Control and Prevention in the United States and priority group 1 critical microorganism by the World Health Organization. Understanding how A. baumannii adapts to different host environments may provide critical insights into strategically targeting this pathogen with novel antimicrobial and biological therapeutics. Exposure to human fluids was previously shown to alter the gene expression profile of a highly drug susceptible A. baumannii strain A118 leading to persistence and survival of this pathogen. Herein, we explore the impact of human pleural fluid (HPF) and human serum albumin (HSA) on the gene expression profile of a highly multi-drug resistant strain of A. baumannii AB5075. Differential expression was observed for ~30 genes, whose products are involved in quorum sensing, quorum quenching, iron acquisition, fatty acid metabolism, biofilm formation, secretion systems and type IV pilus formation. Phenotypic and further transcriptomic analysis using quantitative RT-PCR confirmed RNA-seq data and pointed out a distinctive role of HSA as the molecule involved in A. baumannii response.
Subject: Biology And Life Sciences, Biochemistry And Molecular Biology Keywords: ESKAPE; Acinetobacter; aminoglycosides; amikacin; acetyltransferase; silver; adjuvant
Online: 9 December 2020 (11:05:29 CET)
Clinical resistance to amikacin and other aminoglycosides is usually due to enzymatic acetylation of the antimicrobial molecule. A ubiquitous resistance enzyme among Gram-negatives is the aminoglycoside 6'-N-acetyltransferase type Ib [AAC(6')-Ib], which catalyzes acetylation using acetyl-CoA as donor substrate. Therapies that combine the antibiotic and an inhibitor of the inactivation reaction could be an alternative to treat infections caused by resistant bacteria. We had previously observed that metal ions such as Zn2+ or Cu2+ in complex with ionophores interfere with the AAC(6')-Ib-mediated inactivation of aminoglycosides and reduced resistance to susceptibility levels. Ag1+ recently attracted attention as a potentiator of aminoglycosides' action by mechanisms still in discussion. We found that silver acetate is also a robust inhibitor of the enzymatic acetylation mediated by AAC(6')-Ib in vitro. This action seems to be independent of other mechanisms, like increased production of reactive oxygen species and enhanced membrane permeability, proposed to explain the potentiation of the antibiotic effect by silver ions. The addition of this compound to aac(6')-Ib harboring Acinetobacter baumannii and Escherichia coli cultures resulted in a dramatic reduction of the resistance levels. Time-kill assays showed that the combination of silver acetate and amikacin was bactericidal and exhibited low cytotoxicity to HEK293 cells.
ARTICLE | doi:10.20944/preprints202108.0306.v1
Subject: Biology And Life Sciences, Immunology And Microbiology Keywords: aminoglycoside resistance; structure-activity relationship; aminoglycoside modifying enzymes, acetyltransferase, Acinetobacter
Online: 14 August 2021 (14:12:06 CEST)
The aminoglycoside 6′-N-acetyltransferase type Ib [AAC(6′)-Ib] is a common cause of resistance to amikacin and other aminoglycosides in Gram-negatives. Utilization of mixture-based combinatorial libraries and application of the positional scanning strategy identified an inhibitor of AAC(6′)-Ib. This inhibitor’s chemical structure consists of a pyrrolidine pentamine scaffold substituted at four locations (R1, R3, R4, and R5). The substituents are two S-phenyl (R1 and R4), an S-hydroxymethyl (R3), and a 3-phenylbutyl (R5) groups. Another location, R2, does not have a substitution, but it is named because its stereochemistry was modified in some compounds utilized in this study. Structure-activity relationship (SAR) analysis using derivatives with different functionalities, modified stereochemistry, and truncations were carried out by assessing the effect of the addition of each compound at 8 µM to 16 µg/ml amikacin-containing media and performing checkerboard assays varying the concentrations of the inhibitor analogs and the antibiotic. The results showed that: 1) the aromatic functionalities at R1 and R4 are essential, but the stereochemistry is essential only at R4, 2) the stereochemical conformation at R2 is critical, 3) the hydroxyl moiety at R3 as well as stereoconformation are required for full inhibitory activity, 4) the phenyl functionality at R5 is not essential and can be replaced by aliphatic groups, 5) the location of the phenyl group on the butyl carbon chain at R5 is not essential, 6) the length of the aliphatic chain at R5 is not critical, 7) all truncations of the scaffold resulted in inactive compounds. Molecular docking revealed that all compounds preferentially bind to the kanamycin C binding cavity, and binding affinity correlates with the experimental data for most of the compounds evaluated. The SAR results in this study will serve as the basis for the design of new analogs in an effort to improve their ability to induce phenotypic conversion to susceptibility in amikacin-resistant pathogens.