Drug resistance cell models

Cell monolayers grown on permeable culture inserts form confluent mono-layers with barrier properties and can be used for drug absorption experiments. The most well-known cell line for the in vitro determination of intestinal drug permeability is the human colon adenocarcinoma Caco-2 [20, 21], The utility of the Caco-2 cell line is due to its spontaneous differentiation to enterocytes under conventional cell culture conditions upon reaching confluency on a porous membrane to resemble the intestinal epithelium. This cell model displays small intestinal carriers, brush borders, villous cell model, tight junctions, and high resistance [22], Caco-2 cells express active transport systems, brush border enzymes, and phase I and II enzymes [22-24], Permeability models... 

Other cell lines used in permeability studies include the T84 human colonic adenocarcinoma colonic crypt cell model. This line has a reduced carrier expression, secrets mucus, and has very high resistance [31, 32], The IEC cell line is a rat fetal intestinal epithelium cell with higher permeabilities than Caco-2 cells [33], LLC PKi is a pig kidney epithelial cell line with low expression of efflux systems, but expression systems for transport proteins [32], 2/4/A1 cells are a conditionally immortalized rat fetal intestinal epithelium line with crypt cell-like morphology and temperature-sensitive differentiation [34], They form differentiated monolayers with tight junctions, increased brush border enzymes when grown on extracellular matrices with laminin. Transport of drugs with LP in 2/4/A1 monolayers was comparable to that in the human jejunum and up to 300 times faster than that in Caco-2 monolayers. In contrast, the permeability of HP drugs was comparable in both cell lines [34],... 

The in vitro system has some limitations and it can be used only as a model to address acquired drug resistance, because cell lines may not accurately reflect the in vivo situation of patients treated with anticancer drugs. In vivo analysis using patient samples to study anti-cancer drug resistance are described in the published literature. Chang and coworkers (64) showed that the gene-expression patterns from mRNA derived from needle biopsies of breast cancer patients treated with the anticancer drug, docetaxel, could be predictive of response to the therapy. 

Fig. 11.12. The use of gel microdroplets and flow cytometry to assay drug sensitivity of bacterial cells. The figure shows side scatter and green fluorescence contour plots of gel microdroplets (GMDs) containing E. coli cells that have been stained with fluorescein isothiocyanate for total protein. The microdroplets have been analyzed in the flow cytometer either at time 0 or 2 h after incubation in control medium (left plots) or medium containing penicillin (right plots). A model system was created by mixing two strains of bacteria (susceptible or resistant to penicillin). The data show that a small subpopulation of resistant cells could be detected within 2 h because of its rapid growth in comparison to susceptible cells. From Weaver et al. (1991).

The choice of a cell line to study MDR modulator potency was very important for future potential application in human cancer treatment. PhM (12) that were quite effective in resistant mouse lymphoma cells were only slightly active in drug-resistant human sarcoma cell line MES-SA/Dx5 [198]. The drug-sensitive human sarcoma cell line MES-SA and its multidrug-resistant counterpart MES-SA/Dx5 were applied as a model system for evaluation of MDR modulator activities. Examination performed by the flow cytometric Rhl23 accumulation test demonstrated that the well-known P-gp modulators verapamil (79) and TFP (5) reduced MDR in MES-SA/Dx5 cells. In resistant MES-SA/Dx5 cells, verapamil (79) and TFP (5) restored the drug accumulation pattern which was typical for sensitive cells. However, the effectiveness of PhM (12) was very low. The most active compounds were derivatives with an H atom at position 2 of the phenothiazine ring, followed by Cl-substituted and CF3-substituted compounds. 

This type of tubulin activity has so far been exclusively found in the four above-mentioned natural products and some derivatives, although far more then 140000 synthetic compounds and extracts have been tested. Of these four compounds, epothilones appear to be the best candidates. They are equally or even more active, e.g. up to 35 000 times better then Taxol in resistant cell lines [2]. They also have better cytotoxic potential connected to the tubulin activity, as not all microtubule stabilizers lead to sufficient cell death, and they allow extensive derivatization much faster then Taxol or discodermolide [3, 4]. Also, improvements in the applicability to patients compared to the sparingly soluble Taxol arc expected, eliminating some of the severe side effects connected to the latter drug. Since the binding sites of Taxol and epothilones overlap, epitope comparisons and models of binding... 

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