eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 12

Some currently approved drugs reduce the activity
of PARP1 and PARP2 by competing with NAD+
for binding to the catalytic site. Without NAD+,
PARP fails to PARylate and remains bound to
the damaged DNA, shielding it from other DDR
proteins. This prevents DNA repair and increases
cellular toxicity, potentiating the effect of these
drugs. Thus, the cytotoxic effects of this class of
drugs depend primarily on how efficiently they
trap PARP1 on damaged DNA.9
Scientists recently found that trapping PARP1, but
not PARP2, to DNA with PARPi resulted in increased
cytotoxicity towards cancer cells. Therefore, PARPi
drug screens should include assays that quantify
PARP-trapping ability and distinguish an inhibitor's
selectivity to PARP1 or PARP2.
Most commercially available PARP activity assays
quantify PARylation of target proteins, such as
histones, and test only one PARP enzyme at a time.
In contrast, the PARPtrap™ Combo Assay Kit for
PARP1 and PARP2 compares a molecule's ability
to trap PARP1 versus PARP2 in the same assay. The
assay uses fluorescently labeled DNA probes that
emit polarized light depending on PARP1 or PARP2
binding. These probes have high fluorescence
polarization (FP) when PARP1 or PARP2 is bound.
When scientists add NAD+ to the assay, PARylated
enzymes detach from the probe, reducing FP
levels. If, instead, they add NAD+ and a PARPi, the
inhibitor's trapping ability increases FP in a dosedependent
manner. This homogeneous, simple
assay can be incorporated into high-throughput
drug discovery screens for molecules that enhance
PARP1 or PARP2 trapping on DNA. This PARPtrap™
assay allows researchers to efficiently screen their
libraries for the most specific and effective PARPis.
Figure 6: Assay principle (left panel) and representative results (right panels) showing drug-induced trapping of PARP1 and PARP2
(BPS Bioscience #80584 and #78296). This innovative assay was designed to screen for small molecule PARP inhibitors that can trap
the enzyme on DNA, a feature unique to PARP1 and PARP2.
12

eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development

Table of Contents for the Digital Edition of eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development

eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 1
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 2
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 3
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 4
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 5
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 6
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 7
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 8
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 9
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 10
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 11
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 12
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 13
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 14
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 15
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 16
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 17
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 18
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 19
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 20
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 21
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 22
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 23
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 24
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 25
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 26
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 27
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 28
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 29
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 30
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 31
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 32
eBook: Biochemical and Cell-Based Assays for Targeted Cancer Drug Discovery and Development - 33
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