[Assessment of P glycoprotein expression by immunocytochemistry and flow cytometry coupled with functional efflux analysis: application to acute myeloid leukemia].
P glycoprotein (Pgp) expression is associated with failure of anticancer chemotherapy in acute myeloid leukemia (AML). However, a consensus has been difficult to reach, due to the variable results obtained by different methods. Samples of 27 patients with AML were studied here according to international recommendations (Beck, et al. , Cancer Research 1996; 56: 3010-20). Pgp expression was performed by immunocytochemistry (ICC) using the avidin-biotin peroxidase technique with JSB1 and UIC2 monoclonal antibodies. Flow cytometry (FCM) analysis of Pgp was investigated using UIC2 in an indirect immunofluorescent assay. UIC2 staining was measured by the Kolmogorov-Smirnov statistical test and fluorescence intensity ratio. Finally, the rhodamine 123 test (Rh 123) with or without verapamil was performed to detect functional activity.


RESULTS
by ICC, results of JSB1 and UIC2 were consistent in 94% of the cases. In 74% of the cases, concordant conclusions were observed by ICC and FCM. Overall, Pgp expression was detected in 67% of the cases (ICC/JSB1+ and ICC/UIC2+ or FCM/UIC2+). Functional activity of Pgp was shown in 59% of the patients. Rh 123 efflux was correlated with Pgp expression in 70% of the 27 studied cases but 3 cases were Pgp-/Rh 123+ and 5 Pgp+/Rh 123-. In conclusion, assessment of Pgp expression by ICC and FCM using two different monoclonal antibodies coupled with functional efflux test is required to identify discordant expression/function cases suggesting a non functional Pgp or another alteration of drug transport.
Poulain
S
S
Laboratoire d'hématologie A, Hôpital Calmette, CHU, 59037 Lille Cedex.
Lepelley
P
P
Cambier
N
N
Wattel
E
E
Fenaux
P
P
Cosson
A
A
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000032:analysis / Q000235:genetics*; D000208:Acute Disease; D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000911:Antibodies, Monoclonal; D002121:Calcium Channel Blockers; D002648:Child; D002675:Child, Preschool; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate; D019084:Fluorescent Antibody Technique, Indirect; D005456:Fluorescent Dyes; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007124:Immunoenzyme Techniques; D007150:Immunohistochemistry; D007951:Leukemia, Myeloid / Q000188:drug therapy*; D008875:Middle Aged; D020112:Rhodamine 123; D018709:Statistics, Nonparametric; D017211:Treatment Failure; D014700:Verapamil
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000911:Antibodies, Monoclonal; D002121:Calcium Channel Blockers; D005456:Fluorescent Dyes; D020112:Rhodamine 123; D014700:Verapamil; D016650:Fluorescein-5-isothiocyanate