P-Glycoprotein: Clinical Significance and Methods of Analysis

Abstract

Multidrug resistance (MDR) is responsible for a decrease in sensitivity of tumor cells tumor cells to unrelated, naturally occurring anticancer drugs. This resistance is correlated with expression and activity of a membrane protein, P-gp 170, functioning as a drug-extruding pump. It has been well described in in vitro situations; however, the clinical detection and implications are not yet clear. Multiple detection assays have been developed based on the discovery of the MDR gene family and the corresponding protein. Southern, Northern, or Western blot analysis, S1 nuclease protection or PCR-based assays, immunohistochemical detection or functionality tests by flow cytometry have been used extensively. However, by use of these techniques on clinical material, both normal and malignant, contradictory results have emerged. The sensitivity and specificity of a certain technique are always limited by unavoidable parameters, for example, skill of the technician. Moreover, the complexity of the development of resistance against anticancer agents (external determinants), such as the diversity of tumor tissues, the simultaneous presence of other resistance mechanisms, and the low expression level, make MDR detection equivocal and can lead to contradictory results. Previous treatment influencing the MDR profile and inappropriate timing of the test make a possible correlation between MDR expression and chemotherapeutic resistance difficult to establish and can lead to discordant results. In this review, the need for proper criteria is stressed. No single detection technique provides the ideal test to detect MDR. Tandem testing could give more certainty, although small sample size limit this application. Formulation of a standard assay with better definition of a positivity is essential before clinical trials are started.

99mTc tetrofosmin scintigraphy in acute leukaemia: the relationship between marrow uptake of tetrofosmin and P-glycoprotein and chemotherapy response

“…Similar discordant results also exist for ALL [5,45]. Discordant results about the prognostic value of Pgp in leukaemia may be explained by several factors, including the variety of methods used for detecting Pgp, the simultaneous existence of multiple resistance mechanisms, the existence of Pgp in normal tissue or cells, the heterogenous expression of Pgp, and the expression Pgp without functional capacity [6,10,15].…”

“…Several methods have been established for the determination of MDR1 with the hope of optimizing therapy protocols [6][7][8][9][10]. While some methods (e.g.…”

“…reverse transcriptase polymerase chain reaction, slot blot, northern blot) have aimed to detect MDR1 messenger RNA, others (e.g. flow cytometry, immunohistochemistry) have aimed to show the increase of the protein expression of Pgp [11][12][13][14], but these methods may reveal discordant results, with each method having its own disadvantages [6,10,[15][16][17].…”

P-glycoprotein and alloimmune T-cell activation

“…Multiple assays have been developed and employed for the detection and functional analysis of the MDR1 gene and its corresponding gene product P-gp [23]. Gene-specific methods directed at detection of MDR1 messenger ribonucleic acid (mRNA), including the polymerase chain reaction (PCR), Northern analysis and RNAse protection assays have been used to study P-gp expression; however, P-gp expression and function has most commonly been examined at the protein level using Western analysis, immunohistochemistry, fluorescent dye transport studies, most commonly involving rhodamine-123 (Rh-123) [15,24] or immunostaining-based flow cytometry.…”

“…These assays differ in their specificity for MDR1 P-gp, and it is now well-recognized that certain fluorescent dye probes are transport substrates for related ABC transporters in addition to MDR1 P-gp. Furthermore, among the panel of well-characterized anti-P-gp mAbs that recognize either internal (C-219, C494, JSB-1) or external (Hyb-612, Hyb-241, MRK16, MRK17, 265/F4, 4E3.16, UIC2) epitopes of the transporter [23,[25][26][27][28][29], only a subset is thought to be highly specific for MDR1 P-gp, including Hyb-241 and MRK-16, whereas others, such as C219, are known to bind epitopes conserved among all known P-gp molecules and additional ABC transporters. These limitations of the currently available methods for the specific study of the MDR1 gene and its gene product have to be recognized when critically examining the evidence for P-gp expression and function in immune cells, and account in part for the considerable controversy surrounding these issues.…”