Cell-based assays:Layout 1 14/1/10 19:58 Page 72
Assays
before, we don’t in particular. The key criteria for you’re interested in antagonists, it really doesn’t
me is reproducibility. If different labs can’t get the matter if it’s a heterodimer or a dimer, as long as
same results given a similar target, similar you’re interfering with one of the ligand interac-
approaches and similar cell types, then you’ve got tions with the receptor.
to wonder, do the stars have to align and do the
waves have to be just right to see what someone Kevin Pfleger: It also depends how you define
else says they saw with a dimer? It’s a matter of antagonists.
how much time and effort am I going to put into
something like that. Martin Valler: There’s an increasing tendency
toward inverse agonists.
Guido Zaman: Yes I think I agree with what Kevin
said, that the proof is in the physiological rele- Arthur Christopoulos: Depending on the same
vances and there you need very sensitive technolo- receptor constant, whether it’s a mono or a dimer,
gy to make use of labelled compounds, labelled the same receptor, the same ligand it could be an
antibodies to look at native tissues. We know that inverse agonist on pathway A, a positive agonist on
in overexpression systems and depending on the pathway B, neutral for pathway C, in the same cell
technology that you can use here, you can see var- type. And then, a ligand is a ligand, maybe we
ious combinations, combinations that are some- should come up with a new term called ‘affinist’ –
times less specific, but then the next question is, if anything that has affinity. I’m not really proposing
indeed, this changes this heterodimer, does this we do that.
change the pharmacology of the natural ligands, What does this mean? I have no idea what this
and if it does not change the pharmacology, then means other than a headache for the screening, it
it’s not very interesting. means you’ve got to have multiple ligands for the
one target, for the one library unless you can pre-
Martin Valler: In the natural situation, that may dict where... I mean the interesting thing is that
be true, but for intervention, for drug finding, you may find the compounds cluster. If you do a
that could yield new information, new binding large scale screen you may find compounds cluster.
sites, new modalities for those compounds. So for This cluster will give us inverse agonism here, pos-
drug discovery that kind of interaction is itive agonism there. If this cluster gives us only
extremely relevant. inverse agonism, then you might take a small sub-
set of those, and they’re the ones you take forward
Guido Zaman: That’s true. for a more detailed approach.
Martin Valler: I agree fundamentally this under- David Marks: Yes, that’s another issue that you
pins this whole concept that we need morphologi- brought up, that is, even for antagonists there
cal systems to do our testing, and if we say the real could be inverse agonism. In the past, that really
answer is, we go from our compound that we find hasn’t been investigated very much. So I think that
to an animal model, well, that’s not my definition moving forward, that should be more interesting,
of high throughput screening. We need our high looking at how to do that.
throughput systems to be as relevant as possible,
and informative and not only to just represent Arthur Christopoulos: There’s an element of aes-
potential dimerisation, but they also need to repre- theticness, you know what I mean? A mechanis-
sent the signalling pathways. In all those areas, tic... you don’t like not understanding how these
we’ve got a long way to go. I see the dimerisation things tend to work but there is an element of
is only one aspect of that. The question is, do we empiricism when you’re screening a large number
have the relevant signalling pathways going on, do of compounds, and you can see... you can finger-
we have the relevant expression levels, do we have print compounds in these sorts of profiles without
the assay systems to detect relevant drug effects in necessarily knowing all the mechanisms and that
a system where maybe the receptor is extremely may be sufficient to take things forward. In other
low expression. words, compounds that make this go up, this go
down, and this do nothing, may work in vivo,
David Marks: Even for GPCRs, there are really whereas compounds that go in that direction don’t
two different types of drugs that you’re looking for. work in vivo. I don’t know why, but I already
The issues that we’ve been discussing is really only know they work in vivo, then I’ll spend time trying
relevant for if you’re interested in agonists. If to work out why.
72 Drug Discovery World Winter 2009/10
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