Biomedical Engineering Reference
In-Depth Information
just started to engage in apoptosis. These considerations need to be an integral
part of the experimental design for testing compounds to understand as a whole
the mechanisms of action taking place in the dismissal of cells by a compound
under study.
Here again, the cell line chosen to model a response needs to be considered
carefully to provide the best interpretation of screening or further mechanistic data
obtained from analytical results. This, together with the previous explanation for the
sequence of events described for the apoptosis, can introduce variables that need to be
considered carefully. These variables can introduce difficulties and pitfalls in analysis
of apoptosis, as pointed out by Darzynkiewicz et al. [25].
5.4 EXPERIMENTAL EXAMPLES
To address the cell cycle profiles as a multiparameter tool in oncology drug discovery,
specific models need to be established according to the mechanisms of action in
question. Here we present examples for two different scenarios:
1. Compounds inducing a high cell cycle phase-specific arrest (e.g., topoisome-
rase targeting agents and tubulin targeting agents)
2. Compounds inducing a mild arrest better defined as a slow traverse time across
the different phases (e.g., farnesyltransferase inhibitors).
Examples of each of these approaches are presented below.
5.4.1 Topoisomerase Targeting Agents
Topoisomerases are validated targets for cancer chemotherapy and agents capable of
interacting with either topoisomerase I or topoisomerase II have demonstrated the
potential for increased efficacy in the clinic. Camptothecin analogues that act upon
topoisomerase I and the two related analogues etoposide and teniposide that act
upon topoisomerase II are clinically active agents with their respective and
apparently mutually exclusive mechanisms of action [25-29]. Our earlier efforts
to bring a noncamptothecin compounds into the clinic as an anticancer agent lead to
the demonstration that topoisomerase I was the target responsible for the cytotoxi-
city of BMS-250749 [30-33]. We also found that Topo II was the specific target for
TAS-103, a proposed topoisomerase I and topoisomerase II dual interactive com-
pound [34, 35].
Both of these compounds were assayed by an experimental design that include the
“cell cycle components” and the “specific activities” previously described in this
chapter. In addition, each compound was tested on cell line pairs with specific
topoisomerase deficiencies. P388/Topo I is a mouse leukemia cell line that is
1000-
fold more resistant to camptothecin than parental P388, because it lacks any
detectable levels of topoisomerase I [36]. HCT116/Topo II is a human colon
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