Combinatorial Chemistry

1) Combinatorial chemistry (or CombiChem) is an innovative method of synthesizing

many different substances quickly and at the same time.
2) Combinatorial chemistry contrasts with the time-consuming and labor intensive
methods of traditional chemistry where comounds are synthesized individually! one
at a time.
") #hile combinatorial chemistry is rimarily used by organic chemists who are
seeking new drugs! chemists are also now alying combinatorial chemistry to other
fields such as semiconductors! suerconductors! catalysts and olymers.
$) CombiChem is used to synthesize large number of chemical comounds by
combining sets of building blocks.
%) &ach newly synthesized comound's comosition is slightly different from the
revious one. ( traditional chemist can synthesize 1))-2)) comounds er year.
*) ( combinatorial robotic system can roduce in a year thousands or millions
comounds which can be tested for otential drug candidates in a high-throughut
screening rocess.
+) ,ver the last few years! the combinatorial chemistry has emerged as an e-citing
new aradigm for the drug discovery. .n a very short time the toic has become the
focus of considerable scientific interest and research efforts.
Development of Combinatorial Chemistry
1) /rom a historical ersective! the research efforts made in classical combinatorial
chemistry can be briefly outlined in three hases0
2) .n the early 111)s! the initial efforts in the combinatorial chemistry arena were
driven by the imrovements made in high-throughut screening (234) technologies.
3his led to a demand for access to a large set of comounds for biological screening.
") 3o kee u with this growing demand! chemists were under constant ressure to
roduce comounds in vast numbers for screening uroses. /or ractical reasons!
the molecules in the first hase were simle etides (or etide-like) and lacked
the structural comle-ity commonly found in modern organic synthesis literature.
$) 3he second hase started in the late 111)s! when chemists became aware that it is
not 5ust about numbers6 but something was missing in comounds roduced in a
combinatorial fashion. &mhasis was thus shifted towards quality rather than
quantity.
%) 7ike the first hase! the third hase had its origin in rogress made by the
biomedical community.
*) (s the scientific community moved into the ost-genomic chemical biology age!
there was a growing demand in understanding the role of newly discovered roteins
and their interactions with other bio-macromolecules (i.e. other roteins and 89( or
:9().
+) /or e-amle! the early goals of the biomedical research community were centered
on the identification of small-molecule ligands for biological targets! such as ;-
rotein-couled recetors (;<C:s) and enzymes.
=) 2owever! the current challenges are moving in the direction of understanding bio-
macromolecular (i.e. rotein-rotein! rotein-89(>:9() interactions and how small
molecules could be utilized as useful chemical robes in systematic dissection of
these interactions.
1) ?y no means will this be a trivial undertaking@ 3he develoment of biological assays
towards understanding biomacromolecular interactions is equally challenging as the
need for having access to useful small molecule chemical robes.
Methods of Combinatorial Chemistry
1) 3here are two ma5or aroaches in combinatorial chemistry. 3he Aool and slit
methodA involves attaching the starting comounds to olymer beads.
2) 3he beads are then slit into %) grous and reacted with the second set of reagents.
(fter this reaction! all the beads are ooled! mi-ed together! and slit into %) grous
again.
") .f mi-ing is efficient! each grou should contain aro-imately equal numbers of
beads reresenting each of the 1))) first-generation roducts.
$) 3he grous of beads are then reacted with the ne-t set of reagents. .f the beads are
tagged in some way after each reaction (for e-amle! with a different fluorescent
label to identify each reagent) the combination of tags will characterize each of the
%) ))) second-generation roducts e-actly.
%) 3he finished beads can be screened for their ability to bind a articular target
rotein in the body. 3he comound with the best erformance can be identified and
tested further.
*) (dditional rounds of ooling and slitting allow libraries with millions of comounds
to be generated and sub5ected to high-throughut screening (234).
+) 3he second method is called Aarallel synthesisA. (ll the different chemical structure
combinations are reared searately! in arallel! using thousands of reaction
vessels and a robot rogrammed to add the aroriate reagents to each one.
=) 3his method is unsuitable for the creation of very diverse libraries but is very useful
for the develoment of smaller and more secialized libraries based on a articular
skeleton (starting comounds).
Solid-Phase Synthesis
.n 11*" Berrifield reorted the first e-amles of 4olid-hase synthesis of etides
using chloromethylated-olystyrene containing immobilized 9-rotected amino acids
building blocks. 3his chemistry develoed over the ensuing decade and became the
basis for much of the rogress in etide chemistry.
3he ability to synthesize comounds on an inert olymeric resin bead! to force a
reaction to comletion by the addition of e-cess of reagents and monomers! and then
remove all the unwanted material by simle filtration and wash it is at the heart of most
library synthesis. 3he use of solid suorts for chemical (no etides and etide
molecules) as well as biological synthesis (roteins! etides! olynucleotides) relies on
three interconnected requirements0
1) ( cross-linked! insoluble! olymeric material that is inert to the condition of
synthesis.
2) ( chemical rotection strategy to allow selective orthogonal rotection and
derotection of reactive grous in the monomers.
") 4ome means of linking the substrate to this solid hase that ermits the cleavage of
some or the entire roduct from the solid suort during synthesis for analysis of the
e-tent of reaction! and ultimately to give the final roduct of interest.
Solution Phase Synthesis
8esite the focus on the use of solid-hase techniques for the synthesis of
combinatorial libraries! there have been a few e-amles where libraries have
successfully been made and screened in solution. .ndeed some grous have e-ressed
a reference for solution libraries because there is no rior requirement to develo
workable solid-hase couling and linking techniques. <anlabs have recently disclosed
an interest in making large numbers of comounds as individual comonents using
arallel! reliable solution chemistry.
3he idea behind the method of generating libraries of small organic molecules is to
combine a rigid core molecule suorting multile reactive sites with a mi-ture of
building blocks to roduce a random mi-ture of olyfunctionalized structures! e.g. a
molecule such as cubane tetra acid chloride could be combined with four molar
equivalents of amines to roduce tetra-substituted cubane comounds. 3his method of
library generation has several advantages. /irst it is a owerful method of generating
molecular diversity6 in a single combinatorial ste! the cubane core and 2* amines
would roduce theoretically "= +)1 different tetra amides. :eactions are ushed to
comletion by the use of e-cess quantities of the reactive reagent! and are isolated by
the solvent-solvent e-traction. 3here is no further urification! and thus they refer to
describe these samles as Areaction roductsA.
3he grous from ;la-o and <irrung have synthesized dimeric comounds using amide!
ester or carbamate bond-forming reactions. &very library comound was reared twice
in mi-tures of differing comosition. 3esting all of these mi-tures allows identification of
likely active comounds without the need to resynthesise every comound in an active
mi-ture. 3he drawback for solution hase chemistry is the time-consuming urification
of intermediates that is often required in between synthetic stes.
High Throughput Screening
3he sets of comounds roduced by combinatorial chemistry are generally referred to
as libraries! which deending on how the solid-hase is handled! may be either
mi-tures or individual comounds. 3here are a range of otions for testing the libraries
in a biological assay0
Test mixture in solution-
(ll the comounds are cleaved from the beads and tested in solution. .f the resin beads
were intimately mi-ed! it is not ossible to test the roducts searately! but rather as a
mi-ture. .f activity in a harmacological screen is observed it is not ossible to say
which comound or comounds are active. .n order to identify the most active
comonent! it is necessary to resynthesize the comounds individually and thereby find
the most otent. 3his iterative rocess of resynthesis and screening is one of the most
simle and successful methods for identifying active comounds from libraries.
Test individual compounds in solution-
( second method is to searate the beads manually into individual wells and cleave the
comounds from the solid-hase. 3hese comounds can now be tested as individual
entities.
Test compounds on the beads
( third method for screening is testing on the beads! using a colorimetric or fluorescent
assay technique. .f there are active comounds! the aroriate beads can be selected
by color or fluorescence! CickedD out by micromaniulation and the roduct structure! if
a etide! determined by sequencing on the bead. 9on-etide structures would need
to be identified by one of the tagging methods. 4creening on the bead may be an
inaroriate method for drug discovery! as the bead and linker resent conformational
restrictions that may revent binding to the recetor. /urthermore for harmaceutical
alications comounds will be invariably need to act! and thus ideally need to be
tested in solution.