Penetration of paclitaxel and 5-fluorouracil in multicellular layers of human colorectal cancer cells

Abstract

Abstract. Limited efficacy of chemotherapeutic drugs against solid tumors has been attributed to poor drug penetration into tumor tissues. Multicellular layer (MCL) cultures recapitulate barriers to drug penetration and distribution and have been used successfully in the production of clinically relevant data. In the present study, we evaluated the characteristics of paclitaxel (PTX) and 5-fluorouracil (5-FU) penetration and their effects on tissue penetration using MCLs of human colorectal cancer cells (DLD-1 and HT-29) grown in Transwell inserts. Drug concentration in conditioned media after MCL penetration was estimated using % survival of cells exposed to the conditioned media, and the penetration rate was calculated as % drug concentration relative to the expected concentration after penetration of cell-free MCLs. PTX showed limited penetration in both MCLs in contrast to the full penetration seen by 5-FU. The penetration rate measured after 24 h by cytotoxicity of the conditioned media was 40 and 38% in DLD-1 (20 μM) and HT-29 MCLs (1 μM), respectively, at which concentration the conditioned media produced 50% growth inhibition in monolayers. The penetration profile obtained using [14 C]-paclitaxel also showed slow and limited penetration with concentration-and cell line-dependency. In HT-29 MCL, full penetration of PTX was obtained at 10 μM after 48 h, whereas only 80% was obtained at 1 μM. In DLD-1 MCLs, penetration of PTX was minimal, especially at 1 μM, showing penetration rates as low as 10 and 20% after 24 and 96 h, respectively. When PTX and 5-FU were allowed to penetrate in sequential combination, no effect on the penetration rate was observed. Overall, our results demonstrated limited penetration of PTX in human colorectal cancer MCLs along with concentration-, time-, and cell line-dependency. Assessment of penetration using cytotoxicity of the conditioned media used in the present study may be useful in early stage screening of anticancer agents for their potential in tissue penetration.

NMR-Based Chemical Profiling, Isolation and Evaluation of the Cytotoxic Potential of the Diterpenoid Siderol from Cultivated Sideritis euboea Heldr.

“…For example, these IC 50 values are lower compared to those presented by the widely-studied terpene α-pinene, which shows a lower inhibitory effect in HepG2, A549, PC-12, and MCF-7 cell lines [31], or they are in the same range with the IC 50 values displayed by the prominent flavonoid quercetin [32]. On the other hand, the IC 50 values presented by siderol are higher compared to those of established chemotherapeutic drugs such as fluorouracil (2.95, 17.21, and 8.13 µM in DLD1 [33], HeLa [34], and A549 [34] cells, respectively) and the plant-derived paclitaxel (133, 2.6, and 4.1 nM in DLD1 [33], HeLa [35], and A549 [35] cells, respectively). Thus, siderol has great potential to be exploited as a potent anticancer agent, and its derivatization or formulation should be considered in order to enhance even more its cytotoxic efficacy.…”

In-depth phenotypic characterization of multicellular tumor spheroids: Effects of 5-Fluorouracil

“…The presence of these living cells may be attributed to (1) a decrease in drug penetration in the center of the spheroid and/or (2) the presence of cells showing an increase resistance to 5-FU treatment in a 3D environment. Numerous studies in various tumoral cell lines have described a good penetration of 5-FU into spheroids compared to other drugs [ 34 – 37 ]. Cell resistance to 5-FU treatment may thus rather be related to a difference in cell-cell and cell-matrix interactions within the spheroid, compared to cells in 2D cultures.…”

Tissue dynamics spectroscopy for phenotypic profiling of drug effects in three-dimensional culture

“…Three-dimensional tissue culture includes multicellular layers (MCL) [29,30], multicellular tumor spheroids (MCT) [31,32], and matrix-based tissue growth [33,34] in which cancer morphogenesis progression and metastasis are strongly dependent on a three-dimensional environment. Thin layers and small nodules can be imaged using conventional techniques, such as confocal fluorescence [35,36], two-photon [37,38], optical projection tomography (OPT) [39] and single-plane illumination projection (SPIM) [40].…”