Antibody Competition Test
Tumors are often resistant to chemotherapy, as a result of a decreased,
subtoxic intracellular concentration of the chemotherapeutical agents. Frequently
the P-glycoprotein (Pgp, see Fig.1), an ABC transporter capable of extruding
a wide spectrum of cytostatic drugs from the cell, is responsible for the
failure of therapy due to the multidrug resistance connected to its activity.
The exact mechanism of substrate binding and export are subjects of intense
research.
Fig.1
We have implemented a conformation-sensitive, antibody-based method for
detection of Pgp-related multidrug resistance in hematological malignancies,
and for screening of potential reversing agents. A novel assay, the antibody
competition test (ACT), based on the combinative use of different anti-Pgp
antibodies exhibiting conformation sensitive competition has been elaborated.
The ACT assay is based
on a novel phenomenon observed with two competing monoclonal antibodies*:
The first anti-Pgp antibody (UIC2 mAb) only mildly affects the binding of
a second one (MM12.10), unless the Pgp-expressing cells are treated with
certain modulators/substrates when UIC2 preincubation completely abolishes
subsequent MM12.10 binding. Two basic modalities of reversing agents are
seen, those belonging to the first group elicit the conformational change
leading to UIC2 dominance over MM12.10, while the others do not (Fig.2a).
Fig 2a, 2b.
The feasibility of application of the method as a screening procedure
has been fully confirmed: About 50 different modulators have been compared
and found to be either completely verapamil-like (ACT-negative), or completely
cyclosporin-like (ACT-positive), suggesting that our method will be useful
in screening for “strong” reversing agents (Fig. 2b).
Patent pending: PCT/HU02/00025.
ACT is a cost effective, easy-to use, high throughput screening tool of
reversing compounds that enables highly accurate and precise distinction
between the "weak" verapamil-like (ACT-negative), or "strong", cyclosporin-like
(ACT-positive) reversing agents.
We welcome the opportunity to discuss your needs for testing of reversing
agents: Click here to contact us.
*H. Nagy et al.: P-glycoprotein conformational changes detected
by antibody competition. Eur. J. Biochem. 268(8): 2416-20. 2001