Background: 5-Fluorouracil (5-FU) has been commonly prescribed for patients with colorectal cancer (CRC), but resistance to 5-FU is one of the main reasons for failure in CRC. Recently, microRNAs (miRNAs) have been established as a means of reversing the dilemma by regulating signaling pathways involved in initiation and progression of CRC. However, how to safely and effectively deliver miRNA to target cells becomes a main challenge. Results: In this study, Engineered exosomes were exploited to simultaneously deliver an anticancer drug 5-FU and miR-21 inhibitor oligonucleotide (miR-21i) to Her2 expressing cancer cells. Purified engineered exosomes from the donor cells loaded with 5-FU and miR-21i via electroporation to introduce into 5-FU-resistant colorectal cancer cell line HCT-116 5FR. Furthermore, systematic administration of 5-FU and miR-21i loaded exosomes in tumor bearing mice indicated a significantly anti-tumor effect. The results showed that the engineered exosome-based 5-FU and miR-21i co-delivery system could efficiently facilitate cellular uptake and significantly down-regulate miR-21 expression in 5-FU resistant HCT-116 5FR cell lines. Consequently, the down-regulation of miR-21 induced cell cycle arrest, reduced tumor proliferation, increased apoptosis and rescued PTEN and hMSH2 expressions, regulatory targets of miR-21. Of particular importance was the significant reduction in tumor growth in a mouse model of colon cancer with systematic administration of the targeting miR-21i. More excitedly, the combinational delivery of miR-21i and 5-FU with the engineered exosomes effectively reverse drug resistance and significantly enhanced the cytotoxicity in 5-FU-resistant colon cancer cells, compared with the single treatment with either miR-21i or 5-FU. Conclusion: The strategy for co-delivering the functional small RNA and anticancer drug by exosomes foreshadows a potential approach to reverse the drug resistance in CRC and thus to enhance the efficacy of the cancer treatment.
Safe, efficient and cancer cell targeted delivery of CRISPR/Cas9 is important to increase the effectiveness of available cancer treatments. Although cancer derived exosomes offer significant advantages, the fact that it carries cancer related/inducing signaling molecules impedes them from being used as a reliable drug delivery vehicle. In this study, we report that normal epithelial cell-derived exosomes engineered to have HN3 (HN3LC9-293exo), target tumor cells as efficiently as that of the cancer cell-derived exosomes (C9HuH-7exo). HN3LC9-293exo were quickly absorbed by the recipient cancer cell in vitro. Anchoring HN3 to the membrane of the exosomes using LAMP2, made HN3LC9-293exo to specifically enter the GPC3+ HuH-7 cancer cells than the GPC3− LO2 cells in a co-culture model. Further, sgIQ 1.1 plasmids were loaded to exosomes and surprisingly, in combination with sorafenib, synergistic anti-proliferative and apoptotic effect of loaded HN3LC9-293exo was more than the loaded C9HuH-7exo. While cancer-derived exosomes might induce the drug resistance and tumor progression, normal HEK-293 cells-derived exosomes with modifications for precise cancer cell targeting like HN3LC9-293exo can act as better, safe and natural delivery systems to improve the efficacy of the cancer treatments.
Background
Sorafenib resistance poses therapeutic challenges in HCC treatment, in which cancer stem cells (CSCs) plays a crucial role. CRISPR/Cas9 can be utilized as a potential technique to overcome the drug resistance. However, a safe, efficient and target specific delivery of this platform remains challenging. Extracellular vesicles (EVs), the active components of cell to cell communication, hold promising benefits as delivery platform.
Results
Herein we report the normal epithelial cell –derived EVs engineered with HN3(HLC9-EVs) show competing tumor targeting ability. Anchoring HN3 to the membrane of the EVs through LAMP2, drastically increased the specific homing of HLC9-EVs to GPC3+Huh-7 cancer cells rather than co-cultured GPC3−LO2 cells. Combination therapy of HCC with sorafenib and HLC9-EVs containing sgIF to silence IQGAP1 (protein responsible for reactivation of Akt/PI3K signaling in sorafenib resistance) and FOXM1 (self-renewal transcription factor in CSCs attributed to sorafenib resistance), exhibited effective synergistic anti-cancer effect both in vitro and in vivo. Our results also showed that disruption of IQGAP1/FOXM1 resulted in the reduction of CD133+ population that contribute to the stemness of liver cancer cells.
Conclusion
By reversing sorafenib resistance using combination therapeutic approach with engineered EVs encapsulated CRISPR/Cas9 and sorafenib, our study foreshadows a path for a better, accurate, reliable and successful anti-cancer therapy in the future.
Graphical Abstract
Aim:
To assess the cytotoxicity level of newly introduced poly vinyl ether silicone (PVES) compared to poly vinyl siloxane (PVS) and polyether (PE) elastomeric impression materials.
Settings and Design:
Comparative -Invitro study design.
Materials and Methods:
Mouse cell line NIH/3T3 was grown in Dulbecco's modified Eagle's medium. Samples of three elastomers were dissolved in dimethyl sulfoxide and were tested at various concentrations. Twenty-four well plates with NIH/3T3 cells with different concentrations of elastomeric solutions were incubated at 37°C. 3-(4,5-dimethylthiazol-2-yl)-2-5-diphenyltetrazolium bromide assay was performed on day 1, 3, and 7, with a time interval of 15 min, 30 min, 60 min, and 24
th
h to estimate the cytotoxicity for all three elastomers.
Statistical Analysis Used:
Kruskal–Wallis ANOVA test and the period effect within the subjects, repeated-measure ANOVA was done using the Greenhouse–Geisser correction method.
Results:
The mean cell viability (survival rate) of NIH 3T3 cells at the concentrations tested was measured. A repeated-measure Kruskal–Wallis ANOVA determined the mean survival concentration on day 1, 3, and 7. PVES showed significant decrease in the survival rate on day 1 than PVS and PE, while PVS and PE had significant decrease in the survival rates of cells on day 3 and 7 which were statistically significant (
P
< 0.001).
Conclusion:
PVES shows early cytotoxic signs as compared to PVS and PE, and cell viability for PVS was the highest among all. When making impression with PVES and PE, it is always better to evaluate the impression and gingival sulcus carefully with magnification to prevent adverse reaction, if any material is left inadvertently for longer period of time.
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