Intracellular concentration of reactive oxygen species (e.g., H2O2) in cancer cells is elevated over 10-fold as compared to normal cells. This feature has been used by us and several other research groups to design cancer specific prodrugs, for example, N-alkylaminoferrocene (NAAF)-based prodrugs. Further improvement of the efficacy of these prodrugs can be achieved by their targeting to intracellular organelles containing elevated reactive oxygen species (ROS) amounts. For example, we have previously demonstrated that lysosome-targeted NAAF-prodrugs exhibit higher anticancer activity in cell cultures, in primary cells and in vivo (Angew. Chem. Int. Ed. 2017, 56, 15545). Mitochondrion is an organelle, where electrons can leak from the respiratory chain. These electrons can combine with O2, generating O2−• that is followed by dismutation with the formation of H2O2. Thus, ROS can be generated in excess in mitochondria and targeting of ROS-sensitive prodrugs to these organelles could be a sensible possibility for enhancing their efficacy. We have previously reported on NAAF-prodrugs, which after their activation in cells, are accumulated in mitochondria (Angew. Chem. Int. Ed. 2018, 57, 11943). Now we prepared two hybrid NAAF-prodrugs directly accumulated in mitochondria and activated in these organelles. We studied their anticancer activity and mode of action. Based on these data, we concluded that ROS produced by mitochondria is not available in sufficient quantities for activation of the ROS-responsive prodrugs. The reason for this can be efficient scavenging of ROS by antioxidants. Our data are important for the understanding of the mechanism of action of ROS-activatable prodrugs and will facilitate their further development.
Human cytomegalovirus (HCMV) infection causes severe clinical conditions in the face of immature or suppressed immune defense functions. The virus is, however, readily controlled by an immunocompetent host. Central to this process is the stimulation and maturation of dendritic cells (DC) (1, 2). Monocytederived, immature DC (iDC) are fully permissive to HCMV replication (3), but infection of these cells is restricted to strains expressing the viral genes UL128 to UL131A (4). Infection of iDC by HCMV and its impact on their maturation and function have been studied by several laboratories (5-10). There is consensus that T cell costimulatory molecules, such as CD40, CD80, and CD86 and MHC classes I and II, are downregulated on the surfaces of iDC following HCMV infection (2). As a result, iDC are impaired in their capacity to stimulate T cell responses (5-9), although this effect may depend on time after infection (4). With iDC function impaired, direct HCMV infection of iDC may not be sufficient to prime and sustain the vigorous T cell response against the virus that is seen during natural infection. Uptake and cross-presentation of HCMV antigen by uninfected iDC was identified as one mechanism to bypass HCMV-mediated immunosuppression of iDC function (11-13). Here we address whether exposure of iDC to subviral, noninfectious dense bodies (DB) of HCMV in the absence of viral infection is suitable for inducing activation and maturation, thereby rendering these cells competent for antigen presentation.(Part of this research was conducted by C. Sauer in partial fulfillment of the requirements for a doctoral degree from Johannes Gutenberg University, Mainz, Germany.)To investigate the impact of DB on iDC, we generated iDC from peripheral blood mononuclear cells (PBMC) of HCMV-seronegative individuals (14). Over 99% of the cells were CD3 negative, excluding lymphocyte contaminations (data not shown). Human foreskin fibroblasts (HFF; 3,6 ϫ 10 7 cells for each virus) were infected with either strain RV-HB5 (a derivative of the laboratory strain AD169) or the endotheliotropic strain RV-TB40/E. DB and virions were purified from 400 ml of culture supernatants at 7 days postinfection using glycerol-tartrate gradient ultracentrifugation (15). After high-speed sedimentation of the material collected from the bands, DB (RV-HB5) fractions were resuspended in 300 l of phosphate-buffered saline (PBS; 1 mg), virions from RV-HB5 were resuspended in 100 l of PBS (175 g), and virions from TB40/E were resuspended in 100 l of PBS (130 g). DB fractions were either used directly or exposed to UV light (DB UV ; 254 nm of UV light for 2 min) to inactivate residual infectious virus. HFF and iDC were incubated with 10 g of each of these fractions in order to determine infectivity (by staining for HCMV immediate early protein 1 [IE1] expression) or antigen uptake (by staining for the tegument protein pp65) ( Table 1). The results showed that DB contained very little virus (DB fraction) or no virus (DB UV fraction) that would infect iDC. By co...
The human cytomegalovirus (HCMV) replicates to high titers in primary human fibroblast cell cultures. A variety of primary human cells and some tumor-derived cell lines do also support permissive HCMV replication, yet at low levels. Cell lines established by transfection of the transforming functions of adenoviruses have been notoriously resistant to HCMV replication and progeny production. Here, we provide first-time evidence that a permanent cell line immortalized by adenovirus type 5 E1A and E1B (CAP) is supporting the full HCMV replication cycle and is releasing infectious progeny. The CAP cell line had previously been established from amniotic fluid cells which were likely derived from membranes of the developing fetus. These cells can be grown under serum-free conditions. HCMV efficiently penetrated CAP cells, expressed its immediate-early proteins and dispersed restrictive PML-bodies. Viral DNA replication was initiated and viral progeny became detectable by electron microscopy in CAP cells. Furthermore, infectious virus was released from CAP cells, yet to lower levels compared to fibroblasts. Subviral dense bodies were also secreted from CAP cells. The results show that E1A/E1B expression in transformed cells is not generally repressive to HCMV replication and that CAP cells may be a good substrate for dense body based vaccine production.
The oxidation of alkyl thiols to disulfides has been achieved under mild conditions using a chemiluminescent 1,2-dioxetane as a stoichiometric oxidant. Besides the mild and biocompatible reaction conditions, this approach offers the possibility to monitor the presence of thiols through oxidation and chemiluminescence of the remaining dioxetane.
Optical storage and photon quantification systems based on sensitive photoreactions have numerous applications. Herein, we report a highly efficient photocatalytic reaction, in which ruthenium photoredox catalysis is combined with a 1,2-dioxetane from which chemiluminescence can be triggered. In this system, blue light irradiation as optical input enables a defined inverse correlation with base-triggered, blue light emission as optical output. Comparison of readout by 1 H NMR and chemiluminescence, relative to previous optical input, underlines the reliability and usefulness of the ruthenium-dioxetane system for optical storage, sensing and ruthenium detection.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.