Investigation of P-Glycoprotein-Mediated Multidrug Resistance in Ovarian Cancer Through Biodynamic Imaging
Multidrug resistance is one of the main causes of ineffective treatment of ovarian cancer, which is the deadliest gynecological cancer today. Almost 80% of advanced cases have recurring malignancies after treatment, even after initial responsiveness to the usual platinum-based chemotherapeutics. One mechanism for resistance is the transmembrane drug efflux pump P-glycoprotein (Pgp), encoded by the gene MDR1. Overexpression of this protein in ovarian cancer is associated with poor chemotherapeutic response and multidrug resistance due to low intracellular accumulation of the drug. Biodynamic imaging (BDI) is a low-coherence form of digital holography that provides 3D functional information on intratumor motility. Also, it provides spectral signatures related to drug mode of action and for mitosis, apoptosis, and necrosis. It is a more accurate predictor of in vivo drug response than commonly used 2D cell-based assays. BDI was used to investigate if modulating Pgp affects spheroid drug response. MDR1 was silenced in ovarian cancer spheroids using inducible lentiviral shRNA. Holistic drug response was assessed by BDI and compared to drug response of spheroids expressing Pgp. For BDI, background spectral data was collected for 4 and then 9 h after drug treatment. Pgp expression was confirmed using Western blot and immunofluorescence. Sensitivity to drugs were measured in parallel experiments using XTT assays for 2D cultures and ATP viability assays for 3D cultures. MDR1 silencing shifts the drug response of spheroids treated with 10 μM paclitaxel towards a previously determined apoptotic spectral signature. Pgp inhibitors verapamil and zosuquidar changed spheroid drug response to 10 μM doxorubicin. Verapamil exhibited a strong increase in low- to mid-range frequency suppression. The increase in drug toxicity was seen in both 2D and 3D experiments as well. Results indicate that BDI could detect changes in drug response due to modulation of a single protein, Pgp. The impact of this in vitro study is its potential to predict how different Pgp levels in in vivo tumors affect drug response as assessed by BDI.
Turek, Purdue University.
Molecular biology|Cellular biology|Optics
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