Synergistic effects of methyl 2-cyano-3,11-dioxo-18beta-olean-1,-12-dien-30-oate and erlotinib on erlotinib-resistant non-small cell lung cancer cells

Non-small cell lung cancer(NSCLC)is often characterized by an underlying mutation in the epidermal growth factor receptor(EGFR),contributing to aggressive metastatic disease.Methyl 2-cyano-3,11-dioxo-18 beta-olean-1,12-dien-30-oate(CDODA-Me),a glycyrrhetinic acid derivative,reportedly improves the therapeutic response to erlotinib(ERL),an EGFR tyrosine kinase inhibitor.In the present study,we performed a series of studies to demonstrate the efficacy of CDODA-Me(2μM)in sensitizing HCC827 R(ERL-resistant)cells to ERL.Herein,we first established the selectivity of ERL-induced drug resistance in the HCC827 R cells,which was sensitized when ERL was combined with CDODA-Me(2μM),shifting the IC50 from 23.48μM to 5.46μM.Subsequently,whole transcriptomic microarray expression data demonstrated that the combination of ERL+CDODA-Me elicited 210 downregulated genes(0.44%of the whole transcriptome(WT))and 174 upregulated genes(0.36%of the WT),of which approximately 80%were unique to the ERL+CDODA-Me group.Synergistic effects centered on losses to cell cycle progression transcripts,a reduction of minichromosome maintenance complex components(MCM2-7),all key components of the Cdc45·MCM2-7 GINS(CMG)complex,and replicative helicases;these effects were tantamount to the upregulation of processes associated with the nuclear factor erythroid 2 like 2 translational response to oxidative stress,including sulfiredoxin 1,heme oxygenase 1,and stress-induced growth inhibitor 1.Collectively,these findings indicate that the synergistic therapeutic effects of ERL+CDODA-Me on resistant NSCLC cells are mediated via the inhibition of mitosis and induction of oxidative stress.

Upscaling human mesenchymal stromal cell production in a novel vertical-wheel bioreactor enhances extracellular vesicle secretion and cargo profile

Human mesenchymal stromal cells(hMSCs)are mechanically sensitive undergoing phenotypic alterations when subjected to shear stress,cell aggregation,and substrate changes encountered in 3D dynamic bioreactor cultures.However,little is known about how bioreactor microenvironment affects the secretion and cargo profiles of hMSC-derived extracellular vesicles(EVs)including the subset,“exosomes”,which contain therapeutic proteins,nucleic acids,and lipids from the parent cells.In this study,bone marrow-derived hMSCs were expanded on 3D Synthemax II microcarriers in the PBS mini 0.1L Vertical-Wheel bioreactor system under variable shear stress levels at 25,40,and 64 RPM(0.1-0.3 dyn/cm^(2)).The bioreactor system promotes EV secretion from hMSCs by 2.5-fold and upregulates the expression of EV biogenesis markers and glycolysis genes compared to the static 2D culture.The microRNA cargo was also altered in the EVs from bioreactor culture including the upregulation of miR-10,19a,19b,21,132,and 377.EV protein cargo was characterized by proteomics analysis,showing upregulation of metabolic,autophagy and ROS-related proteins comparing with 2D cultured EVs.In addition,the scalability of the Vertical-Wheel bioreactor system was demonstrated in a 0.5L bioreactor,showing similar or better hMSC-EV secretion and cargo content compared to the 0.1L bioreactor.This study advances our understanding of bio-manufacturing of stem cell-derived EVs for applications in cell-free therapy towards treating neurological disorders such as ischemic stroke,Alzheimer’s disease,and multiple sclerosis.