Immunohistochemical detection and quantitation of P-glycoprotein in multiple drug-resistant human myeloma cells: association with level of drug resistance and drug accumulation

Abstract

Using several multiple drug-resistant human myeloma cell lines as standards, we developed an immunohistochemical staining technique and means of quantitating P-glycoprotein in individual myeloma cells. The level of staining intensity for P-glycoprotein in individual myeloma cells was quantitated by measuring the average optical density of each cell with a microscopic computerized cell analysis system. Using this system, we observed that the level of P-glycoprotein for individual cells within a cell population of known drug sensitivity was very homogeneous (coefficient of variation less than or equal to 13%). Analysis of cell lines with gradually increasing levels of multidrug resistance (8226/S, 8226/Dox6 and 8226/Dox40) demonstrated a close association between the level of resistance to doxorubicin, defined by the mean lethal dose (D0) and the amount of P-glycoprotein on individual cells determined by the optical density (r = 0.82, P less than 0.0005). Intracellular doxorubicin (DOX) accumulation in the individual cell lines was inversely related to the level of drug resistance expressed as D0. P-glycoprotein was also detected in the marrow-derived myeloma cells of patients with drug refractory disease using immunohistochemical staining. The amount of P-glycoprotein in the cells of one patient was directly compared to the amount found in the simultaneously stained standard cell lines (8226/Dox6 and 8226/Dox40) by comparing the optical densities for individual cells. Using this immunohistochemical technique to detect and quantitate P-glycoprotein in patient myeloma cells and comparing it to standard multidrug resistant myeloma cell lines may be of value in determining the contribution of P-glycoprotein to clinical drug resistance in patients with multiple myeloma.

Phase I Study of Intravenous PSC‐833 and Doxorubicin: Reversal of Multidrug Resistance

“…The bioassay was performed using 8226/Dox 6 , a doxorubicin-resistant cell line of human myeloma RPMI 8226. 29,30) 8226/Dox 6 and RPMI 8226 were kind gifts from Dr. William S. Dalton (University of Arizona Cancer Center, Tucson, AZ). The method for the bioassay has been described in detail elsewhere (Uchiyama-Kokubu, N. et al, in preparation).…”

A lack of detectable modification of topoisomerase II activity in a series of human tumor cell lines expressing only low levels of etoposide resistance

“…Indeed, our objective was to establish sublines expressing marginal orders of resistance, such as those likely to be encountered clinically. For example, cancer cells obtained freshly from human malignancies have been shown to have a low level of resistance and P-glycoprotein mRNA expression compared with in-vitro-derived resistant cell lines (Dalton et al, 1989;Goldstein et al, 1989); and in a series of human ovarian carcinoma cell lines established from tumours from patients who were clinically refractory to platinum-based chemotherapy, orders of resistance expressed were generally less than 10-fold, compared with lines derived from tumors from untreated patients (Langdon et al, 1988;Hamilton et al, 1989). Indeed, Kuiper et al (1990) have stressed the importance of studying drug resistance in low-level resistant cell lines to allow meaningful comparison with clinically-derived tumor cells.…”

Correlation between the uptake of Tc-99m-sestaMIBI and prognostic factors in patients with multiple myeloma

“…It has been reported that Tc-99m-MIBI is an agent recognized by the human MDR-1 P-glycoprotein, by which transported out of malignant cells . It has been confirmed by an immunohistochemical method that P-glycoprotein expression is associated with MDR in myeloma cells (Dalton et al, 1989).…”