NĒRx BioSciences (United States)

Bacterial antibiotic resistance has emerged as a significant global threat, making it increasingly challenging to effectively treat infections in patients. Nanomedicine technologies can be implemented for the targeted delivery of medications and drugs to patients. This work investigates the use of silica nanoparticles (SiNPs) loaded with the clindamycin and tetracycline antibiotics to treat Staphylococcus epidermidis infection in a 3D-bioprinted dermal model. SiNPs are stable, biocompatible, and can be loaded with small molecules like antibiotics. The SiNPs were synthesized and loaded with antibiotics. The loading efficiency of the SiNPs was determined by UV-Vis spectroscopy and high-performance liquid chromatography. Dome-shaped constructs containing fibroblasts were 3D printed using a fibrin-based bioink to mimic the dermis of skin. These constructs were then inoculated with bacterial cultures labeled with green fluorescent protein (GFP) 4 days post printing and then treated with antibiotic-loaded SiNPs to determine their effect on bacterial growth. After the incubation phase, the bacteria were cultured in broth to determine the colony-forming unit (CFU) count on toxin superantigen (TSA) plates containing 10 mg/mL chloramphenicol. The CFU count of the 3D-bioprinted human constructs samples treated with antibiotics was significantly lower than both the SiNP-treated and untreated samples. The results suggest that antibioticreleasing SiNPs can serve as a more efficient treatment for skin bacterial infections.

ABSTRACTEcopipam is a first‐in‐class dopamine D1 receptor antagonist under investigation for the treatment of Tourette syndrome. This open‐label, fixed‐sequence, three‐cohort study evaluated whether ecopipam induces or inhibits cytochrome P450 (CYP) 1A2, CYP2B6, CYP2C19, CYP2D6, CYP3A4, organic anion‐transporting polypeptide (OATP) 1B1, and p‐glycoprotein (P‐gp). Probe substrates were administered alone and after single or steady‐state dosing of ecopipam (1.8 mg/kg/day). Cohort 1 received standard doses of omeprazole (CYP2C19), caffeine (CYP1A2), and dextromethorphan (CYP2D6) orally, and a microdose of midazolam (CYP3A4) intravenously. Cohort 2 received a standard dose of bupropion (CYP2B6) orally. Cohort 3 received an oral solution containing microdoses of dabigatran (P‐gp), pitavastatin (OATP1B1), rosuvastatin (BCRP, OATP, and P‐gp), atorvastatin (BCRP, OATP, P‐gp, and CYP3A4), and midazolam. A total of 56 healthy individuals (median [range] age, 36.5 [19–54] years; 85.7% male) were enrolled, and 48 completed the study. Ecopipam was well tolerated in the presence and absence of probe substrates. Steady‐state ecopipam administration increased dextromethorphan exposure (> 100‐fold); decreased the AUCinf of midazolam, omeprazole, and dabigatran by 44.2%, 44.1%, and 37.9%, respectively; and decreased unconjugated bilirubin (UGT1A1) by 19.5%. Single‐dose ecopipam increased atorvastatin Cmax by 95% and rosuvastatin Cmax by 11%, whereas steady‐state ecopipam decreased the AUCinf of atorvastatin by 26.8% and rosuvastatin by 16.4%. There were no changes in caffeine, bupropion, or pitavastatin exposure. Ecopipam is a strong inhibitor of CYP2D6 and weak inducer of CYP3A4, CYP2C19, P‐gp, and UGT1A1. Ecopipam did not inhibit CYP3A4, CYP2C19, CYP2B6, CYP1A2, UGT1A1, P‐gp, or OATP1B1 and did not induce OATP1B1 or CYP2B6.

EDITORIAL article Front. Oncol., 27 September 2022Sec. Cancer Molecular Targets and Therapeutics Volume 12 - 2022 | https://doi.org/10.3389/fonc.2022.1023500

Abstract The DNA-dependent protein kinase (DNA-PK) plays a critical role in the non-homologous end joining (NHEJ) double-strand break (DSB) repair pathway and the DNA damage response (DDR). Consequently, blocking DNA-PK kinase activity is a novel anti-cancer therapeutic strategy in combination with ionizing radiation (IR). Towards developing a new class of DNA-PK inhibitors, our laboratory has exploited the mechanism of DNA-PK activation which requires binding to DNA termini via the Ku 70/80 heterodimer. We have previously reported the development of Ku-DNA binding inhibitors (Ku-DBi’s) that act via this novel mechanism of action to inhibit DNA-PK catalytic kinase activity. Ku-DBi’s display nanomolar activity in vitro, possess cellular DNA-PK and NHEJ inhibitory activity and sensitize non-small cell lung cancer (NSCLC) cells to DSB generating chemotherapeutics bleomycin and etoposide. In this study, we demonstrate that pre-incubation of our novel Ku-DBi’s can potentiate the cellular effects of bleomycin and IR and also induce p53 phosphorylation through the activation of the ATM pathway, which is concomitant with a decrease in DNA-PKcs autophosphorylation events at the S2056 (pS2056) cluster. Using a combination of Western blot and immunofluorescence assays in the NSCLC NCI-H460 and A549 cell lines, Ku-DBi’s treatment in combination with DNA DSBs-inducing agents such as bleomycin and IR, showed a significant reduction of autophosphorylation events of DNA-PKcs at the S2056 cluster compared to DNA DSBs-inducing agent alone. In addition, analysis of phospho-ATM and phospho-p53 protein levels in these NSCLC cell lines, suggested activation of the ATM pathway as a function of Ku-DBi’s treatment followed by bleomycin, evidenced by an increase of phosphorylation of ATM at Ser1981, and a modest increase of p53 phosphorylation at Ser15. Our findings demonstrate that Ku-DBi’s block DNA-DSB dependent DNA-PKcs autophosphorylation, resulting in potentiating cellular sensitivity to bleomycin and IR, and a likely effect on the ATM-dependent signaling pathway. These data are consistent with Ku-DBi’s possessing a novel mechanism of action that abrogates autophosphorylation of DNA-PKcs to impact DSB repair and potentially DDR signaling, becoming in a promising approach as part of an anticancer therapeutic strategy in combination with DNA DSBs-inducing agents. Citation Format: Pamela L. Mendoza-Munoz, Navnath S. Gavande, Pamela S. VanderVere-Carozza, Katherine S. Pawelczak, Joseph R. Dynlacht, Joy E. Garrett, John J. Turchi. Impact of novel Ku-DNA binding inhibitors on the DNA DSBs-induced DNA damage response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2596.

NMR studies can provide unique information about protein conformations in solution. In CASP14, three reference structures provided by solution NMR methods were available (T1027, T1029, and T1055), as well as a fourth data set of NMR-derived contacts for an integral membrane protein (T1088). For the three targets with NMR-based structures, the best prediction results ranged from very good (GDT_TS = 0.90, for T1055) to poor (GDT_TS = 0.47, for T1029). We explored the basis of these results by comparing all CASP14 prediction models against experimental NMR data.

Simple SummaryGenome replication, repair, recombination, and the DNA damage response (DDR) rely on a cadre of DNA binding proteins to execute and regulate these critical pathways. There are many structural motifs that support DNA-binding and included in these is the oligonucleotide/oligosaccharide binding fold (OB-fold). OB-folds are found across the tree of life and play critical roles in chromosome maintenance and stability. This paper reviews the current state of knowledge regarding structure, function, and involvement of OB-fold proteins in the DNA damage response, replication, and repair. We highlight a group of OB-fold DNA binding proteins and discuss how disruption of these critical protein–DNA interactions can be exploited in cancer therapy.Genome stability and maintenance pathways along with their requisite proteins are critical for the accurate duplication of genetic material, mutation avoidance, and suppression of human diseases including cancer. Many of these proteins participate in these pathways by binding directly to DNA, and a subset employ oligonucleotide/oligosaccharide binding folds (OB-fold) to facilitate the protein–DNA interactions. OB-fold motifs allow for sequence independent binding to single-stranded DNA (ssDNA) and can serve to position specific proteins at specific DNA structures and then, via protein–protein interaction motifs, assemble the machinery to catalyze the replication, repair, or recombination of DNA. This review provides an overview of the OB-fold structural organization of some of the most relevant OB-fold containing proteins for oncology and drug discovery. We discuss their individual roles in DNA metabolism, progress toward drugging these motifs and their utility as potential cancer therapeutics. While protein–DNA interactions were initially thought to be undruggable, recent reports of success with molecules targeting OB-fold containing proteins suggest otherwise. The potential for the development of agents targeting OB-folds is in its infancy, but if successful, would expand the opportunities to impinge on genome stability and maintenance pathways for more effective cancer treatment.

Replication protein A (RPA) is the major human single stranded DNA (ssDNA)-binding protein, playing essential roles in DNA replication, repair, recombination, and DNA-damage response (DDR). Inhibition of RPA-DNA interactions represents a therapeutic strategy for cancer drug discovery and has great potential to provide single agent anticancer activity and to synergize with both common DNA damaging chemotherapeutics and newer targeted anticancer agents. In this letter, a new series of analogues based on our previously reported TDRL-551 (4) compound were designed to improve potency and physicochemical properties. Molecular docking studies guided molecular insights, and further SAR exploration led to the identification of a series of novel compounds with low micromolar RPA inhibitory activity, increased solubility, and excellent cellular up-take. Among a series of analogues, compounds 43, 44, 45, and 46 hold promise for further development of novel anticancer agents.