Practical Fragments has
written repeatedly about various assay artifacts (vide infra). Different
technologies are susceptible to different interference mechanisms, making
general rules difficult. Earlier this year we
wrote about the Metal Ion Interference
Set, or MIIS: a collection of a dozen salts that could be used to assess the
sensitivity of assays to metal contaminants. In a recent open-access
JACS Au
paper, Huabin Hu (Uppsala University), Jonathan Baell (Monash University), and
collaborators extend the concept to small molecules.
The researchers have compiled a Collection
Of useful Nuisance compounds, or CONS, perhaps with a nod to “Chemical con artists
foil drug discovery”
published a decade ago, which we highlighted
here. The 103
members of the CONS are divided into three categories.
The first set contains five aggregators:
molecules that have been shown to form colloidal clusters that non-specifically
interfere with biological assays, as discussed
here.
The largest set, at 67 members, consists
of PAINS, or pan-assay interference compounds, which we first
wrote about in
2010. These are themselves divided into various subcategories: non-specific
electrophiles such as
curcumin and an
isothiazolone, redox cyclers such as
quinones,
contaminants such as the decomposition products of certain fused
tetrahydroquinolines, miscellaneous,
metal chelators, and additional mechanisms including optical interference and
singlet oxygen quenchers, which are particularly problematic in AlphaScreen
assays.
The last set consists of 31
compounds that can cause problems in phenotypic assays. Some of these non-specifically
disrupt cell membranes. Others have well-defined but toxic effects, such as interfering
with tubulin or intercalating into DNA. Such bioactivity is not always a bad
thing: some of these molecules, such as topotecan and colchicine, are approved
drugs, but it’s useful to be aware of whether these types of activities will affect
your assay.
One criticism of the PAINS
concept is that it lumps together multiple mechanisms. (Pete Kenny
wrote about
this recently.) Another criticism is that, by focusing on chemical
substructures, true hits may be
unfairly deprioritized based on structure alone.
What’s nice about the CONS list is that the potentially interfering mechanisms
of each molecule are documented and categorized so they can be considered when establishing
an assay. For example, you may not care whether a compound interferes in a
phenotypic assay if you are performing a screen on an isolated enzyme.
The entire set of compounds is available
from
Enamine, and additional vendors are provided in a supplementary table. If
you’re doing a lot of assays, particularly on new targets and mechanisms, it
may be worth testing the CONS to understand what kinds of false positives might
occur.
