Note : Les descriptions sont présentées dans la langue officielle dans laquelle elles ont été soumises.
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SYSTEMS AND METHODS TO FACILITATE MULTIPLE ORDER
COMBINATORIAL CHEMICAL PROCESSES
BACKGROUND OF THE INVENTION
This invention relates generally to the field of chemistry, and in particular
to techniques for synthesizing chemical entities and evaluating reactions
between the
chemical entities when subjected to certain reaction conditions. More
specifically, the
invention relates to the creation of diverse chemical libraries and to the
identification of
reactions that occur between members of the libraries.
Recent trends in the area of research for novel chemicals, including
pharmacological agents, have been concentrated on the preparation of so-called
"chemical libraries". Chemical libraries are intentionally created collections
of differing
molecules or chemical entities which can be prepared either synthetically or
biosynthetically. Following synthesis, the chemical entities may be used in
various
assays or combined with other chemicals and then screened for biological
activity or
chemical reactivity.
One way to produce chemical libraries is by synthesizing the various
chemicals to individual solid supports, which typically take the form of resin
beads. A
variety of techniques have been proposed for making chemical libraries which
utilize
individual solid supports to which the chemicals are tethered. One such method
is the
"discrete" method where solid supports are placed into multiple reaction
vessels. Various
chemicals are then synthesized onto the solid supports while the solid
supports remain
within the reaction vessels. After completing the synthesis process, the
chemical
compound on each solid support may be identified simply by identifying the
reaction
vessel from which the solid support was removed. Because of the need to
maintain the
solid supports within a given reaction vessel, the size of the resulting
chemical library is
limited by the number of reaction vessels used.
In an attempt to greatly increase the size of a chemical library, the mix and
split technique was developed. In the mix and split method, solid supports are
placed into
individual reaction vessels and a first building block is synthesized onto
each of the solid
supports. Solid supports are then mixed together and redistributed to the
reaction vessels
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where a second building block is synthesized onto the solid supports. The
solid supports
may once again be mixed and redistributed where another building block may be
synthesized onto the solid supports. This process may be repeated as many
times as
necessary. Examples of mix and split techniques are described generally in
U.S. Patent
No. 5,503,805, the complete disclosure of which is herein incorporated by
reference.
Once a mix and split chemical library has been produced, the compound
may be cleaved from the solid supports and tested to determine if the compound
produces
a desired result. If so, the particular compound needs to be identified.
However, since
the solid support was mixed and split one or more times during the synthesis
process,
identifying the compound on the solid support can be challenging. A variety of
techniques have been proposed for identifying the compounds, such as by the
use of
identifier tags as described in U.S. Patent No. 5,708,153, or by the use of
identification
codes as described generally in PCT International Application No.
PCT/LTS97/05701, and
in H. Mario Geysen, et al., Isotope or Mass Encoding of Combinatorial
Libraries, Chem.
& Biol. Vol. III, No. 8, pp. 679-688, August 1996, the complete disclosures of
which are
herein incorporated by reference.
Although a variety of techniques exist for creating diverse chemical
libraries and for identifying the resulting chemical entities of the
libraries, little has been
done in the way of improving the efficiency of processes that utilize the
resulting
chemical libraries. For example, it may be desirable to attempt to react the
chemical
entities of one library with the chemical entities of another library. For
instance, it may
be desirable to attempt to react multiple catalysts with a chemical library to
evaluate the
usefulness of various catalysts.
To perform such reactions using existing techniques, the chemicals are
typically cleaved from their solid supports and then combined in a well with
cleaved
chemicals from another library under certain reaction conditions. If
reactivity occurs, the
combined chemicals still need to be identified as previously mentioned.
Unfortunately,
such a process can be unpractical for even moderately sized libraries. For
example, if
each library had 1,000 members, then the total number of required reactions
would be
1,000,000. Individual cleavage and placement of chemicals into 1,000,000 wells
is
simply impractical.
Hence, the invention is related to techniques and chemical constructs
which enable multiple chemical libraries to be created, reacted with each
other and
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screened in an efficient manner. Once chemical reactivity has been identified,
the
invention also provides techniques for identifying the particular chemical
entities
involved in the reactions.
SUMMARY OF THE INVENTION
The invention provides various screening techniques along with novel
constructs that may be used when screening for reactive chemicals. In one
specific
embodiment, a method is provided to screen for reactive chemicals by
configuring a
plurality of constructs such that each construct of the set includes a
pairwise combination
of a chemical entity A~-A; of a chemical library A and a chemical entity B1-B~
of a
chemical library B. Further, the set of constructs include essentially every
possible
pairwise combination of the chemical entities A1-A; of the chemical library A
and the
chemical entities B~-B~ of the chemical library B. In this way, each chemical
entity from
library A and from library B are unambiguously associated, e.g. on a solid
support, so that
every possible combination of chemical entities from two or more libraries may
be tested
for reactivity. The constructs are exposed to a given set of conditions to
facilitate a
reaction or interaction between the chemical entity A1- A; and the chemical
entity B1- B~
of each construct. The constructs are then screened to identify where a
reaction or an
interaction occurred. If any reactions or interactions are identified, the
chemical entity
A~- A; and the chemical entity B1- B~ of the associated constructs are
determined.
In one aspect, each construct includes at least a pair of sites. Further, the
chemical entity AI- A; is synthesized to one of the sites of each construct
while the other
site is blocked. The other site of each construct is then unblocked, and the
chemical
entity BI- B~ is synthesized to the other site of each construct. In another
aspect, the
constructs are formed using a combinatorial processes to achieve the desired
pairwise
combinations. For example, the constructs may be mixed together after
synthesizing the
chemical entities A1- A; and then split into groups such that each group has
constructs
with essentially all other chemical entities A1- A;. The chemical entities B~-
B~ may then
be synthesized onto the constructs such that each group receives a different
chemical
entity B~- B~. Optionally, further combinatorial processes may be used when
synthesizing
library A and/or library B onto the constructs. For example, a combination of
chemicals
may be synthesized on each construct to create each A~- A; chemical entity and
each B1-
B~ chemical entity, e.g. using a mix and split technique. In this way, the
chemical entities
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of each library may be constructed of a single chemical building block or
multiple
chemical building blocks.
In another aspect, the constructs are screened by sensing for a change in
temperature to indicate that a reaction or an interaction has occurred.
Screening may also
be accomplished by measuring for the mass of any chemical products, e.g. using
a mass
spectrometer. Other screening techniques include the use of ultraviolet light
to test for a
color change or phosphorescence resulting from the creation of a chemical
product,
colored chromophotography, and the like.
In still another aspect, the specific chemical entities may be determined by
evaluating the masses of the unreacted chemical entities A~- A; and the
unreacted
chemical entities B~- B~ using mass spectrometry, and correlating each mass
with an
associated chemical entity of each library, e.g. by using a look-up table.
Alternatively,
each chemical entity A1- A; and each chemical entity B~- B~ may be encoded
with a code.
The codes may then be decoded to determine the specific chemical entities.
Conveniently, the codes may be decoded by evaluating the mass of the codes
using mass
spectrometry and correlating each mass with an associated chemical entity,
e.g. by using
look-up tables.
In one particular aspect, the chemical entities A~- A; or B1- B~ comprise
catalysts. In this way, multiple chemicals may be reacted with multiple
catalysts in an
efficient manner. In another particular aspect, multiple libraries of
constructs are
provided that each include the same pairwise combinations of chemical entities
A1- A;
chemical entities B~- B~. Further, each library of constructs is exposed to a
different set of
conditions. In this way, multiple libraries of chemicals may be exposed to
multiple
conditions in a high throughput manner. In an alternative aspect, each library
of
constructs may be exposed to a metal in one of its oxidation states as part of
a third or
higher order combinatorial process.
The invention further provides a method for making a library of constructs.
The constructs may be formed on a plurality of solid supports that each
include at least
two sites. A chemical entity from a chemical library A having A~- A; chemical
entities is
synthesized to one of the sites of each solid support while the other site is
blocked. The
blocked site for each solid support is then unblocked and a chemical entity
from a
chemical library B having BI- B~ chemical entities is synthesized to the
unblocked sites.
The chemical entities A~- A; and the chemical entities Bl- B~ are synthesized
to the sites
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to form a set of constructs that includes essentially every possible pairwise
combination
of the chemical entities A,-A; of the chemical library A and the chemical
entities B1-B~ of
the chemical library B.
In one aspect, the constructs are mixed after synthesizing the chemical
5 entities A1- A; and are split into groups such that each group has
constructs with
essentially all other chemical entities AI- A;. The chemical entities B1- B~
are then
synthesized onto the constructs such that each group receives a different
chemical entity
B1- B~. Optionally, the chemicals may be synthesized by synthesizing a
combination of
chemicals on each construct to create each A~- A; chemical entity and each B~-
B~
chemical entity. In this way, each chemical entity may be constructed of a
single
chemical building block or multiple building blocks. Conveniently, a mix and
split
technique may be employed to synthesize the chemicals. Optionally, each
chemical
entity A~- A; and each chemical entity B,- B~ may be encoded with an
identification code
to facilitate identification of the chemical entities following screening.
The invention further provides an exemplary construct that comprises a
solid support having at least one arm and two or more sites branching from the
arm. A
chemical entity A is coupled to one of the sites, and a chemical entity B is
coupled to the
other site. Further, the pair of sites are configured such that the chemical
entity A is
spaced apart from the chemical entity B at a distance selected to facilitate a
reaction
between the chemical entity A and the chemical entity B. Optionally, an
identification
code may be coupled to the chemical entity A and the chemical entity B.
In another embodiment, the invention provides a library of chemical
constructs. The library includes a set of constructs that each comprise a
solid support
having at least one arm and at least a pair of sites branching from the arm. A
chemical
entity A~-A; of a chemical library A is coupled to one of the sites, and a
chemical entity
B1-B~ of a chemical library B is coupled to the other site. Further, the pair
of sites are
configured such that each chemical entity AI-A; is spaced apart from each
chemical entity
B1-B~ at a distance selected to facilitate a reaction or an interaction
between each chemical
entity A~-A; and each chemical entity B~-B~.
In one aspect, the set of constructs includes essentially every possible
pairwise combination of the chemical entities A~-A; of the chemical library A
and the
chemical entities B~-B~ of the chemical library B. In another aspect, the
chemical entities
A~-A; of the chemical library A and/or the chemical entities B~-B~ of the
chemical library
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B each comprise multiple chemical building blocks that have been synthesized
to the
sites. In still another aspect, the library A or the library B comprises
catalysts.
In an alternative embodiment, a library of chemical constructs comprises a
set of constructs that each comprise a solid support having at least a pair of
sites. A
chemical entity A1-A; of a chemical library A is coupled to one of the sites,
with the
chemical entity Al-A; comprising two or more chemical building blocks that
have been
synthesized to the site. A chemical entity B1-B~ of a chemical library B
coupled to the
other site. For example, the chemical library B may comprise catalysts.
Optionally, the
chemical entity B 1-B~ may also be constructed of two or more building blocks.
In one aspect, the set of constructs includes essentially every possible
pairwise combination of the chemical entities A~-A; of the chemical library A
and the
chemical entities B1-B~ of the chemical library B. In another aspect, an
identification
code may be coupled to each of the chemical entities.
The invention further provides techniques for evaluating combinations of
catalysts to determine which combinations are the most efficient in producing
end
products. For example, in one method, a set of constructs are configured such
that each
construct of the set includes a pairwise combination of a chemical entity A,-
A; of a
catalyst library A and a chemical entity B~-B~ of a catalyst library B. The
constructs are
exposed to a substrate in solution phase to facilitate potential reactions
involving the
chemical entity A1- A; and the chemical entity B~- B~ of each construct. The
constructs
may then be screened to identify any reactions or interactions, and the
chemical entity A~-
A; and the chemical entity B1- BJ of any constructs where reactions or
interactions
occurred may be identified. In one aspect, the set of constructs may include
essentially
every possible pairwise combination of the chemical entities A~-A; of the
catalyst library
A and the chemical entities BI-B~ of the catalyst library B. In this way, a
comprehensive
analysis of the reaction or interaction of two catalysts libraries with a
substrate may be
performed.
BRIEF DESCRIPTION OF THE DRAWINGS
Fig. 1 is a schematic view of a solid support having a plurality of tether
sites according to the invention.
Fig. 2 is a graph illustrating the distribution of distances between tether
sites for the solid support of Fig. 1.
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Fig. 3 is a schematic view of a solid support having an alternative
arrangement of tether sites according to the invention.
Fig. 4 is a graph illustrating the distance distribution between chemical
entities for the solid support of Fig. 3.
Fig. 5A is a flow chart illustrating a second order reaction process
according to the invention.
Fig. 5B is a flow chart illustrating a second order catalysis process
according to the invention.
Fig. 6 is a flow chart illustrating a third order combinatorial process
according to the invention.
Fig. 7 is a schematic view of an analytical construct according to the
invention.
Fig. 7A illustrates a process for making one specific analytical construct
according to the invention.
Fig. 7B illustrates one specific example of an alternative analytical
construct according to the invention.
Fig. 8 is a flow chart illustrating one possible method for synthesizing a
chemical library A onto solid supports according to the invention.
Fig. 9 is a schematic view of a library of constructs created from two
chemical libraries, A and B.
Fig. 10 illustrates a method for producing the library of constructs of Fig.
9.
Fig. 11 illustrates a method where the chemical entities on a library of
constructs are tested for reactions or interactions by subj ecting the
constructs to various
conditions.
Fig. 12 illustrates a resulting product C when two chemical entities A and
B react or interact.
Fig. 13 illustrates a method for producing a library of constructs and
identifying reacting or interacting chemicals.
Fig. 13A illustrates a screening method to screen for reactive or
interactive chemicals.
Fig. 14 is a graph produced when analyzing one of the constructs of Fig.
10 after a reaction or interaction has occurred using mass spectroscopy.
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Fig. 15 illustrates a look-up table associating atomic mass units with
chemical entities.
Fig. 16 is a schematic view of an alternative construct having codes
associated with the chemical entities according to the invention.
Fig. 17 illustrates a method for producing a mass encoded library of
constructs and using mass codes to identify reacting or interacting chemicals.
Fig. 18 is a graph produced when analyzing one of the constructs of Fig.
16 after a reaction or interaction has occurred using mass spectroscopy.
Fig. 19 illustrates a look-up table associating atomic mass units with
codes.
Fig. 20 illustrates a method for producing a chemical library involving
coordination complexes having metals placed in their centers.
Fig. 21 illustrates a method for producing and evaluating constructs having
catalysts when using mass codes.
Fig. 22 illustrates a method for producing a chemical library and then
reacting or interacting the chemicals on the constructs using a variety of
conditions.
Fig. 23 is a schematic view of an analytical construct having chemical
entities from two separate libraries and a reagent.
Fig. 24 illustrates a method for producing and evaluating a chemical
library involving multiple catalysts.
DESCRIPTION OF THE SPECIFIC EMBODIMENTS
The invention provides for the creation of libraries of chemical constructs
using chemical entities (i.e., single chemical building blocks or two or more
synthesized
chemical building blocks) from two or more chemical libraries in order to
determine
reactivity or interactivity between the chemical entities of one library with
the chemical
entities of another library. For example, in one embodiment, the invention
provides a set
of constructs that each have a member of a chemical library A and a member of
a
chemical library B. The set of constructs are configured such that each member
of library
A is unambiguously associated with one of the members of library B. In this
way,
reactions may be attempted between each member of library A with each member
of
library B.
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The constructs of the invention may comprise any chemical arrangement
that allows for the attachment of one or more chemical entities. For example,
the
constructs may comprise solid supports which include one or more tether sites.
The
constructs may further comprise chemical entities that are linked or tethered
to the
various sites. The solid supports of the invention may be constructed from one
or more
materials upon which combinatorial chemistry synthesis can be performed.
Example of
solids supports that may be used include beads, solid surfaces, solid
substrates, particles,
pellets, discs, capillaries, hollow fibers, needles, solid fibers, cellulose
beads, pore glass
beads, silica gels, polystyrene beads optionally crosslinked with
divinylbenzene, grafted
copoly beads, polyacrylamide beads, latex beads, dimethylacrylamide beads,
optionally
cross-linked N, N'-bis-acryloyl ethylene diamine, glass particles coated with
a
hydrophobic polymer, fullerenes and soluble supports, such as iow molecular
weight,
noncrosslinked polystyrene, and the like.
One convenient way for producing the constructs is by linking the
chemical entities from library A to one or more sites of the solid supports
while other
sites are blocked, e.g. with a protecting group. The remaining sites are
unblocked and the
chemical entities from library B are linked to the remaining sites. The
members of the
libraries may synthesized to the sites in relatively close proximity to
facilitate reactions
between the members of the different libraries.
One way of arranging sites 2 on a solid support 4 to facilitate reactions is
shown in Fig. 1. Sites 2 are randomly assigned, and a sufficient number of
sites are
provided so that a reasonable number of the members of libraries A and B are
within
reacting distance as shown in the graph of Fig. 2. In this way, if the members
on a given
solid support are reactive, enough of the members will react to permit
adequate screening
and identification. As shown in the example of Fig. 2 (which includes
arbitrary units), the
reacting distance may be in the range from about 15 distance units to about 30
distance
units. Fig. 2 also illustrates the fraction of sites 2 that fall within the
range.
Fig. 3 illustrates another arrangement of tether sites on a solid support 6 to
facilitate reactions between the members of the different libraries. Solid
support 6
includes multiple arms 7 that each have a pair of sites 8 and 9 branching from
arms 7.
Sites 8 and 9 may be constructed to maximize the potential for reactions or
interactions
occurring between the chemical entities. For example, the sites may be
constructed to
maximize the time averaged fraction of chemical entities that will be within
the reacting
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or interacting range as shown in Fig. 4. The manner of constructing the sites
so as to
optimize the time averaged fraction is described in greater detail
hereinafter.
The members of each chemical library may comprise a single chemical
building block or may be a synthesized chemical entity formed from two or more
monomers or chemical building blocks. Conveniently, mix and split techniques
may be
employed when synthesizing multiple building blocks onto the tether sites of
the solid
supports. In this way, two or more relatively large chemical libraries may be
reacted with
each other in a rapid and convenient manner. For example, as shown in Fig. 5A,
a
chemical library A may be formed such that each member A1_; is constructed
from three
10 building blocks (X,Y,Z). If each building block comprises ten chemicals,
then three mix
and split steps will result in chemical library A having 103 members. If a
similar process
were followed for a chemical library B, it would also have 103 members. By
associating
every member of library A with every member of library B, 106 different
combinations
are provided. Hence, 106 potential reactions or interactions may be evaluated.
As shown
in Fig. 5B, a similar process may be used with a chemical library A and a
catalysis
library.
Third or higher order combinatorial processes are also possible. For
example, as shown in Fig. 6, a set of constructs that includes a library of
catalysts may be
subjected to another combinatorial process where the constructs may be
loaded/reacted
with metals in one or more of their possible oxidation states in order to
insert metal ions
into the catalysts. Hence, using the example of Fig. 5, if 103 metals were
used, then the
number of potential reactions becomes 109.
After forming the constructs, the chemical entities from each of the
libraries may subjected to certain conditions to determine if any of the
chemical entities
are reactive or interactive with each other. Hence, reactions between two or
more
different chemical libraries may be attempted simply by synthesizing the
chemical
entities from each of the libraries onto constructs such that each chemical
entity from
each library is associated with all other entities from all other libraries
The constructs are
then subjected to appropriate conditions to provide an environment where the
chemical
entities on the constructs may potentially react or interact with each other.
The constructs may be screened to determine the constructs where
chemical reactions or interactions occurred. A variety of techniques may be
employed to
screen for chemical reactivity or interactivity, including the use of
thermography as
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described generally in Steven J. Taylor, et al., "Thermographic Selection of
Effective
Catalysts from an Encoded Polymer-Bound Library", Science, Volume 280, pp. 267-
270,
April 10, 1998, the complete disclosure of which is herein incorporated by
reference.
Another screening technique is to clip the link with the solid support and use
mass
spectroscopy to evaluate whether any chemical products were produced. Use of
mass
spectrometry as a screening tool may also be used to identify any starting
materials as
described below. Another screening technique is the use of ultraviolet light
to test for a
color change or phosphorescence of any products. Such a process may be rapidly
accomplished using, for example, a FACS sorter. Still another screening
technique
utilizes colored chromophotography as described generally in Matthew T.
Burger, et al.,
"Enzymatic, Polymer-Supported Formation of an Analog of the Trypsin Inhibitor
A90720A: A Screening Strategy for Macrocyclic Peptidase Inhibitors", J. Am.
Chem.
Soc. 1997, 119, 12697-12698.
The constructs experiencing chemical reactions or interactions may be
separated out for analysis to identify the chemical entities that were
reactive. A variety of
techniques may be employed, alone or in combination, to identify the chemical
entities.
Such techniques include, for example, the measurement of any unreacted
chemicals using
mass spectroscopy, the use of mass based codes, performing one or more
synthesizing
steps as discrete steps, and the like.
The invention may be employed to attempt to react or interact a variety of
chemical libraries. The chemical libraries included on the constructs may be
those
creating using any type of synthesis, including combinatorial synthesis
processes, as
known in the art. As one specific example, one of the libraries may comprise a
group of
catalysts that are reacted with a group of chemical entities. As another
example, the
constructs may also include various reagents that may be involved in the
reactions.
Hence, with the techniques of the invention, multiple combinatorial chemical
libraries
may be reacted or interacted, screened and evaluated in a rapid and efficient
manner.
Referring now to Fig. 7, one embodiment of an analytical construct 10 will
be described. Construct 10 comprises a solid support 12 that has been
engineered to
include an arm 13 that has two branching tether sites 14 and 16. Although only
one arm
is shown, it will be appreciated that multiple arms and associated tether
sites may be
provided on solid support 12.
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Coupled to site 14 is a chemical entity 18, and coupled to site 16 is a
chemical entity 20. Chemical entity 18 may comprise any chemical entity from a
library
A of chemical entities A~-A;, and chemical entity 20 may comprises an chemical
entity
from a library B of chemical entities B~-B~ (where i and j may or may not be
equal).
Sites 14 and 16 are configured to maximize the probability that chemical
entities 18 and 20 will be within reacting distance for a sufficient time to
permit reactions
or interactions to occur, i.e., sites 18 and 20 may be configured to optimize
the time
averaged distance between the sites to increase the probability that an
observable reaction
or interaction will occur. Under appropriate conditions, at least some of the
chemical
entities will react or interact with each other to form a product, or, if one
of the libraries
comprises catalysts, one entity will react or interact with the catalyst to
produce a product,
while the catalyst remains unchanged.
One way to construct sites 14 and 16 to optimize the time averaged
distance between the sites is by using techniques associated with cyclic
molecules as
described generally in Ernest L. Eliel, Stereochemistry of Organic Compounds,
John
Wiley & Sons, Inc. pp. 675- 685, the complete disclosure of which is herein
incorporated
by reference. When constructing sites 14 and 16, an appropriate number of
bonds may be
provided to increase the probability that the chemical entities of libraries A
and B will be
placed in close enough contact to permit reactions to occur. One particular,
non-limiting
example of how to product a construct having a pair of sites branching from an
arm is
illustrated in Fig. 7A. Fig. 7B illustrates one alternative construct having a
pair of sites
linking members of two different chemical libraries.
Conveniently, chemical libraries A and B may each be created using a
combinatorial process where each chemical entity is formed from two or more
chemical
building blocks. For example, as shown in Fig. 8, each chemical entity 18 of
library A
may be formed from three sets of chemical building blocks X, Y and Z. In such
as case,
chemical building blocks X1-X " are initially synthesized to site 14. Solid
supports 12 are
then mixed and split into groups where chemical building blocks Y,-Yn are
synthesized to
site 14. This process is repeated for building blocks Z~-Z~. Once library A
has been
synthesized, a similar process may be used for the B library if it is to be
constructed from
multiple building blocks.
Fig. 9 illustrates one simplified example of a library 22 of constructs 24-34
formed from two chemical libraries A and B, with library A having chemical
entities A,
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and A2, and library B having chemical entities B 1, BZ and B3. In so doing, it
will be
appreciated that, in practice, both libraries may be significantly larger.
Library 22 is
constructed such that each construct includes a different pairwise combination
of
chemical entities from libraries A and B. In other words, a given chemical
entity from
library A will be associated with every one of the chemical entities from
library B (on
different constructs), and vice versa. As such, the number of constructs is
determined by
2 X 3 = 6. Hence, library 22 may be constructed so that it is a combination of
two or
more separate combinatorial libraries.
Fig. 10 illustrates one method for forming library 22 of Fig. 9. Initially, a
large number of solid supports are provided, with only one solid support 36
being shown
for convenience of illustration. Solid support 36 includes a pair of sites 38
and 40. The
solid supports are configured such that sites 40 are provided with a
protecting group 42.
Using a synthesis process, site 38 of each solid support receives a chemical
entity A1_; of a
library A. Optionally, a combinatorial synthesis process may be employed if
the
chemical entities contain more than one building block. For example, three mix
and split
processes may be used so that each chemical entity has three building blocks.
Protecting
group 42 is then removed from each site 40, and a synthesis process is
employed to
synthesize chemical entities B 1 ~ of a library B onto sites 40. This may be
accomplished,
for example, by forming groups of constructs that each include a complete set
of
members from library A. Each of these groups then receives a different member
of
library B. If library B is to be constructed of more than one building block,
one or more
combinatorial processes may be employed in a manner similar to that previously
described.
As shown in Fig. 11, once the library of constructs has been created, the
constructs are placed under a certain set of conditions to facilitate
reactions or
interactions. In one application, all of the constructs are placed under the
same set of
conditions. Such conditions may include, for example, a certain temperature
and a certain
reagent. Under such conditions, at least some of the constructs will have one
of their
chemical entities react or interact with the other chemical entity to form a
product C
(assuming one of the libraries does not contain catalysts) as shown in Fig.
12. A
summary of this process involving the construct of Fig. 10 is illustrated in
Fig. 13.
To determine which constructs experienced reactions or interactions, a
screening process may be performed. For example, one screening process is a
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14
thermography process to detect changes in temperature of the constructs to
indicate that a
reaction has occurred. Other techniques include mass measurement of any
chemical
products, luminescence or phosphorescence resulting from the creation of a
product,
colored chromophotography, and the like. The constructs where a reaction or
interaction
was detected may then be separated from the remainder of the constructs for
further
evaluation. A variety of separating techniques may be used, including the use
of a bead
picker.
One specific example of a screening technique is illustrated in Fig. 13A.
In Fig. 13A, solid support 36 is shown with sites 38 and 40. A mass
identification code is
linked between the chemical entities of libraries A and B as described in
greater detail
with reference to Fig. 16. Also linked to site 38 is a label 39 of some
description. If no
reaction occurs, cleavage of chemical entity A removes label 39 from site 38
as shown.
When construct 36 is scanned, label 39 will not be detected, thus indicating
that no
reaction or interaction occurred. On the other hand, if a reaction or
interaction does
occur, cleavage at site 38 will not release label 39 from construct 36 as
shown. Hence,
when construct 36 is scanned, label 39 will be detected to indicate that a
reaction or
interaction has occurred. A variety of labels may be used to label the
constructs,
including, for example, immunological labels, radio isotope labels,
chromophore labels,
and the like.
Once the constructs have been separated, the chemical entities of each
construct are then identified. As shown in Fig. 13, product C is cleaved from
solid
support 36 to facilitate evaluation. One convenient way to then identify the
chemical
entities is by a mass deconvolution process using mass spectroscopy (MS) where
it is
assumed that at least some of the chemical entities have not reacted and
remain attached
to the solid support. Further, with such a process, it is assumed that none of
the chemical
entities is isobaric. The products and remaining chemical entities cleaved
from the solid
supports are placed in a mass spectrometer where the atomic mass of each
chemical entity
and product is measured. The mass spectrometer may further be configured to
produce a
graph illustrating the outcome. One example of such a graph is illustrated in
Fig. 14. A
look-up table, such as the table of Fig. 15, may then be employed to determine
the mass
for each chemical entity of the A library. A similar process occurs for the B
library.
In this way, two relatively large combinatorial chemical libraries may be
reacted with each other, and any reactions or interactions identified in a
rapid and
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efficient manner. Merely by way of example, two combinatorial libraries of
1,000
members each may be reacted with each other to produce a 1,000,000 member
library.
This library may be rapidly screened for chemical activity, and then, using
mass
deconvolution, may have the reactive or interactive chemical entities rapidly
identified.
An alternative way to identify the chemical entities on the solid supports
following screening is by the use of mass codes. Fig. 16 illustrates one
example of a
construct 44 that includes such mass codes and may be used to create a library
formed
from multiple combinatorial chemical libraries. Construct 44 comprises a solid
support
46 having a pair of sites 48 and 50 similar to the other constructs described
herein.
10 Coupled to site 48 is a mass code 52 which in turn is coupled to a chemical
entity 54 from
a chemical library A. Coupled to site 50 is a mass code 56 which in turn is
coupled to a
chemical entity 58 from a chemical library B. Each mass code is assigned to a
specific
chemical entity and is stored in a look-up table as described hereinafter.
Fig. 17 is a summary of the process used to produce construct 44 (when
15 library B comprises catalysts). To form construct 44, mass code 52 is
linked to site 48
and chemical entity A is synthesized to mass code 52 while site 50 is blocked
with a
protecting group. Site 50 is then unblocked and mass code 56 is linked and
chemical
entity 58 is synthesized. The manner in which the mass codes may be assigned
and
linked, as well as techniques for combinatorially synthesizing the chemical
entities are
described in PCT International Application No.PCT/US97/05701, and in H. Mario
Geysen, et al., Isotope or Mass Encoding of Combinatorial Libraries, Chem. &
Biol. Vol.
III, No. 8, pp. 679-688, August 1996, previously incorporated by reference.
After synthesizing the chemicals onto constructs 44, the constructs are
subjected to certain reaction conditions and the constructs are screened for
any chemical
activity in a manner similar to that previously described. For constructs
where chemical
activity is found, the codes, catalyst and any products are cleaved and placed
into a mass
spectrometer to measure the atomic mass of the codes. Conveniently, the mass
spectrometer may be configured to graphically display the results as
illustrated in Fig. 18.
Look-up tables, such as those illustrated in Fig. 19, may then be employed to
relate the
atomic mass of each measured code to a specific code. In turn, the identified
code may
be correlated with the chemical entity as described in PCT International
Application
No. PCT/LTS97/05701, and in H. Mario Geysen, et al., Isotope or Mass Encoding
of
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16
Combinatorial Libraries, Chem. & Biol. Vol. III, No. 8, pp. 679-688, August
1996,
previously incorporated by reference.
Fig. 20 illustrates an example of a third order combinatorial process. This
specific process utilizes a construct 60 that comprises a solid support 62
having a pair of
sites 64 and 66. Synthesized to site 64 is a substrate 68 that is part of a
combinatorial
library of substrates A. Synthesized to site 66 is a coordination complex 70
that is part of
a library of coordination complexes. In this way, a library of constructs may
be created
with every possible pairwise combination of substrates and coordination
complexes in a
manner similar to that previously described. As shown in Fig. 10, k such
libraries are
created, with each library placed into a discrete vessel 72. Each vessel then
receives a
metal that is in one of its oxidation states and that is to be placed into the
center of the
coordination complexes. Hence, by providing i substrates, j coordination
complexes, and
k metals, a third order combinatorial library may be created having i X j X k
members.
After introducing the metals, the constructs may be screened for chemical
activity in a manner similar to that previously described. For constructs
where chemical
activity occurred, the associated metal may be easily be determined since the
last step was
performed as a discrete step, i.e., simply identify the vessel where the
construct was
obtained. The substrate and the coordination complex may be identified using a
mass
spectrometer and by the use of mass codes in a manner similar to that
previously
described. Fig. 21 illustrates a similar process with the use of mass codes 71
and 73 to
assist in identifying the members of library A and the catalysts in a manner
similar to that
previously described.
Fig. 22 illustrates another example of a third order combinatorial process.
The process of Fig. 22 is similar to that of Fig. 20 in that the last step is
performed as a
discrete step. The process of Fig. 22 utilizes a construct 74 that comprises a
solid support
76 and a chemical entity 78 from a combinatorial library A and a chemical
entity 80 from
a combinatorial library B. Construct 74 may be constructed in a manner similar
to that
previously described and may optionally include one or more mass codes in a
manner
similar to that previously described. Similar to the process of Fig. 20, k
libraries of
constructs are produced that each include constructs with every possible
pairwise
combination of chemical entities from libraries A and B. Each comprehensive
library is
then subjected to a different set of conditions as a discrete step as shown.
In this way, i
chemical entities from a library A may be reacted with j chemical entities
from a library
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17
B, each under k conditions. The constructs are then screened and chemical
entities of
interest (and associated reaction conditions) identified in a manner similar
to that
previously described.
The invention further provides constructs that include more than two sites.
In this way, nt" order combinatorial processes may be performed. An example of
such a
construct 82 is illustrated in Fig. 23. Construct 82 comprises a solid support
84 having
three sites 86, 88 and 90. A chemical entity 92 from a library A (which may
optionally be
produced combinatorially from multiple building blocks) is linked to site 86,
and a
chemical entity 94 from a library B (which may optionally be produced
combinatorially
from multiple building blocks) is linked to site 88 in a manner similar to
that described
with previously embodiments. A reagent 96 from a library of reagents is linked
to site 90.
With the use of constructs 82, i chemical entities from a library A may be
reacted with j chemical entities from a library B using k reagents. Screening
and
deconvolution, including the use of mass codes, may be performed in a manner
similar to
that previously described.
In another aspect of the invention, constructs may be formed that have
catalysts from two or more catalyst libraries to determine which combinations
of catalysts
are the most efficient in producing end products. For example, a set of
constructs may be
configured such that each construct of the set includes a pairwise combination
of a
chemical entity AI-A; of a catalyst library A and a chemical entity B1-B~ of a
catalyst
library B. Conveniently, the set of constructs may include essentially every
possible
pairwise combination of the chemical entities A~-A; of the catalyst library A
and the
chemical entities B1-B~ of the catalyst library B.
The constructs are exposed to a substrate in solution phase to facilitate
potential reactions or interactions involving the chemical entity A1- A; and
the chemical
entity B1- B~ of each construct. The constructs may then be screened to
identify any
reactions or interactions, and the chemical entity A1- A; and the chemical
entity BI- B~ of
any constructs where reactions or interactions occurred may be identified. In
this way, a
comprehensive analysis of the interaction of two catalyst libraries with a
substrate may be
performed.
One example of such a process is illustrated in Fig. 24. The process
employs a plurality of solid supports 100 (only one being illustrated for
convenience of
discussion). Solid support 100 has a pair of tether sites 102 and 104 that may
be
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18
constructed in a manner similar to the other embodiments described herein.
Optionally,
mass codes 106 and 108 may be linked to each site 102 and 104, respectively,
in a
manner similar to other embodiments and used to identify a particular catalyst
after a
reaction has been identified. Initially, site 104 includes a protecting group
(Pg) and a
member A1 of a catalyst library A is synthesized to site 102. As with other
embodiments
described herein, a mufti-step synthesis process may be used to produce member
Al.
Although not shown, each solid support may receive a different member of the
catalyst
library A in a manner similar to other embodiments.
Protecting group Pg is then removed and the above process is repeated to
synthesize a member B 1 of a catalyst library B to site 104. In this way, a
set of constructs
may be produced with every pairwise combination of catalysts from libraries A
and B.
The set of constructs is then exposed to a substrate 110 to potentially
produce a product
112 if a reaction occurs. The set of constructs may then be screened for
potential
reactions or interactions using any of the screening techniques described
herein. For
constructs where reactions or interactions are detected, codes 106 and 108 and
catalyst
members A, and B 1 may be cleaved from solid support 100. Codes 106 and 108
may
then be decoded using mass spectroscopy in a manner similar to that previously
described
to identify the particular catalysts involved in the reaction.
The invention has now been described in detail for purposes of clarity and
understanding. However, it will be appreciated that certain changes and
modifications
may be practiced within the scope of the appended claims.
