The expression of P-glycoprotein does influence the distribution of novel fluorescent compounds in solid tumour models

Abstract

Solid tumours display a complex drug resistance phenotype that involves inherent and acquired mechanisms. Multicellular resistance is an inherent feature of solid tumours and is known to present significant barriers to drug permeation in tumours. Given this barrier, do acquired resistance mechanisms such as P-glycoprotein (P-gp) contribute significantly to resistance? To address this question, the multicellular tumour spheroid (MCTS) model was used to examine the influence of P-gp on drug distribution in solid tissue. Tumour spheroids (TS) were generated from either drug-sensitive MCF7 WT cells or a drug-resistant, P-gp-expressing derivative MCF7 Adr . Confocal microscopy was used to measure time courses and distribution patterns of three fluorescent compounds; calcein-AM, rhodamine123 and BODIPY-taxol. These compounds were chosen because they are all substrates for P-gp-mediated transport, exhibit high fluorescence and are chemically dissimilar. For example, BODIPY-taxol and rhodamine 123 showed high accumulation and distributed extensively throughout the TS WT , whereas calcein-AM accumulation was restricted to the outermost layers. The presence of P-gp in TS Adr resulted in negligible accumulation, regardless of the compound. Moreover, the inhibition of P-gp by nicardipine restored intracellular accumulation and distribution patterns to levels observed in TS WT . The results demonstrate the effectiveness of P-gp in modulating drug distribution in solid tumour models. However, the penetration of agents throughout the tissue is strongly determined by the physico-chemical properties of the individual compounds.

Multiple signaling pathways must be targeted to overcome drug resistance in cell lines derived from melanoma metastases

“…The fact that spheroids derived from the radial growth phase and vertical growth phase lesions, which are the same size as those from the metastatic lines, are exquisitely sensitive to these inhibitors suggests that there are no issues with either permeability or drug absorbance onto the matrix. In addition, previous studies using fluorescent drug conjugates, such as BOD-IPY-taxol, have shown the rapid (<360 minutes) transport of drugs to the core of the spheroid (22).…”

Effect of the cyclin-dependent kinases inhibitor p27 on resistance of ovarian cancer multicellular spheroids to anticancer chemotherapy

“…They were grown according to the ATCC's instruction. Multicellular spheroids were prepared by a liquid overlay method (Martin et al 2003). Briefly, the wells were coated with 0.5 ml of 1.5% agarose containing serum-free medium.…”

“…Development of P-gp -mediated drug resistance in tumors is another major barrier for the effectiveness of cancer therapy (Martin et al 2003). It is well demonstrated that after repetitive treatment of cancer cells with chemotherapeutics, increased expression of the P-gp is associated with a phenotype of multidrug resistance (MDR) (Dusek et al 2000).…”

“…A drug-resistant phenotype generated by sequential treatment of tumor-bearing mice with anticancer drugs was only reproducible in vitro when the same tumor cells were cultivated as multicellular spheroids, but not as monolayer culture (Wartenberg et al 2002;Walker et al 2004). Intrinsic drug resistance is expressed in the areas of quiescent cells in large prostate tumor spheroids, but not in small spheriods containing proliferating cells (Wartenberg et al 2000;Martin et al 2003).…”

Drug Penetration and Therapeutic Resistance

“…Diffusion-limited penetration of molecules into the cultures means that as well as producing oxygen and nutrient gradients they can also develop an acidic central region due to lactate accumulation (Walenta et al, 2000) and have been used to study relations between central pH and medium buffering capacity (Acker et al, 1987). They have been grown using a number of cell types including normal and drug resistant variants (Martin et al, 2003) and have been shown to create an extracellular matrix (Glimelius et al, 1988).…”