ABSTRACT
Arsenic trioxide (ATO) mediates PML-RARA (promyelocytic leukemia-retinoic acid receptor-α) oncoprotein degradation via the proteasome pathway and this degradation appears to be critical for achieving cure in acute promyeloytic leukemia (APL). We have previously demonstrated significant micro-environment-mediated drug resistance (EMDR) to ATO in APL. Here we demonstrate that this EMDR could be effectively overcome by combining a proteasome inhibitor (bortezomib) with ATO. A synergistic effect on combining these two agents in vitro was noted in both ATO-sensitive and ATO-resistant APL cell lines. The mechanism of this synergy involved downregulation of the nuclear factor-κB pathway, increase in unfolded protein response (UPR) and an increase in reactive oxygen species generation in the malignant cell. We also noted that PML-RARA oncoprotein is effectively cleared with this combination in spite of proteasome inhibition by bortezomib, and that this clearance is mediated through a p62-dependent autophagy pathway. We further demonstrated that proteasome inhibition along with ATO had an additive effect in inducing autophagy. The beneficial effect of this combination was further validated in an animal model and in an on-going clinical trial. This study raises the potential of a non-myelotoxic proteasome inhibitor replacing anthracyclines in the management of high-risk and relapsed APL.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Arsenicals/therapeutic use , Leukemia, Promyelocytic, Acute/drug therapy , Oxides/therapeutic use , Proteasome Inhibitors/therapeutic use , Animals , Arsenic Trioxide , Autophagy/drug effects , Bortezomib/therapeutic use , Cell Line, Tumor , Cell Transplantation , Drug Synergism , Humans , Leukemia, Promyelocytic, Acute/pathology , Mice , NF-kappa B/drug effects , Reactive Oxygen Species/agonists , Transplantation, Heterologous , Tumor Cells, Cultured , Unfolded Protein Response/drug effectsABSTRACT
Thirty-four Aspergillus flavus isolates were recovered from sorghum seeds sampled across five states in India. Our study included (1) species confirmation through PCR assay, (2) quantification of total aflatoxin concentrations by the indirect competitive-ELISA (ic-ELISA) method, and (3) analysis of molecular diversity among the A. flavus isolates using ß-tubulin, ITS, and ISSR markers. Among the isolates studied, 28 were found to be positive for the production of aflatoxins. ITS and ß-tubulin phylogenetic analysis segregated the A. flavus sample population into two major groups or clades with little to no subdivision based on geography. In contrast, ISSR analysis also separated the A. flavus isolates into two main clusters, showing a distance of 0.0-0.5, with one cluster exhibiting a high level of diversity though no geographic or chemotype subdivision could be observed. The majority of sampled A. flavus isolates were highly toxigenic, and also highly diversified in terms of toxin-producing potential in-vitro. Genetic diversity among the sorghum isolates of A. flavus further warrants the development of appropriate farming management practices as well as improved aflatoxin detection measures in India.
Subject(s)
Aflatoxins/analysis , Aspergillus flavus/chemistry , Aspergillus flavus/genetics , Sorghum/microbiology , Aflatoxins/chemistry , Aspergillus flavus/classification , Aspergillus flavus/isolation & purification , Enzyme-Linked Immunosorbent Assay , Genetic Variation , India , Phylogeny , Polymerase Chain Reaction , Seeds/microbiologyABSTRACT
Nuclear Overhauser effect spectroscopy (NOESY) gave full assignments of the 1H-NMR spectra of the picrotoxane neurotoxins tutin, hyenanchin, picrotoxinin and picrotin, as well as the solution conformations of these compounds, consistent with molecular modelling. Fully assigned 13C-NMR data are reported.
