Nonredundant split/pool synthesis of combinatorial libraries

• US 20010041344A1
• Filed: 02/02/2001
• Published: 11/15/2001
• Est. Priority Date: 02/03/2000
• Status: Active Grant

• Current Assignee: Nanoscale Combinatorial Synthesis, Inc.
• Original Assignee: Nanoscale Combinatorial Synthesis, Inc.
• Inventors: Baum, Stephen A., Ostrem, James A., Issakova, Olga L., Sepetov, Nikolai F.
• Granted Patent: US 6,799,120 B2
• Time in Patent Office: Days
• US Class Current: 435/7.1
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00306 Reactor vessels in a multiple arrangement
• CPC Class Codes: B01J 2219/00459 Beads
• CPC Class Codes: B01J 2219/00468 by manipulation of individual beads
• CPC Class Codes: B01J 2219/00497 Features relating to the solid phase supports
• CPC Class Codes: B01J 2219/005 Beads
• CPC Class Codes: B01J 2219/00581 Mass
• CPC Class Codes: B01J 2219/00585 Parallel processes
• CPC Class Codes: B01J 2219/0059 Sequential processes
• CPC Class Codes: B01J 2219/00592 Split-and-pool, mix-and-divide processes
• CPC Class Codes: B01J 2219/00596 Solid-phase processes
• CPC Class Codes: B01J 2219/00689 using computers
• CPC Class Codes: B01J 2219/00695 Synthesis control routines, e.g. using computer programs
• CPC Class Codes: B01J 2219/00698 Measurement and control of process parameters
• CPC Class Codes: B01J 2219/007 Simulation or vitual synthesis
• CPC Class Codes: B01J 2219/00702 Processes involving means for analysing and characterising the products
• CPC Class Codes: B01J 2219/00707 separated from the reactor apparatus
• CPC Class Codes: B01J 2219/0072 Organic compounds
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B01J 2219/00725 Peptides
• CPC Class Codes: B01J 2219/00745 : Inorganic compounds
• CPC Class Codes: B01J 2219/00752 : Alloys
• CPC Class Codes: C07B 2200/11 : Compounds covalently bound to a solid support
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 40/10 : Libraries containing peptides or polypeptides, or derivatives thereof
• CPC Class Codes: C40B 40/14 : Libraries containing macromolecular compounds and not covered by groups C40B40/06 - C40B40/12
• CPC Class Codes: C40B 40/18 : Libraries containing only inorganic compounds or inorganic materials
• CPC Class Codes: C40B 60/14 : for creating libraries
• CPC Class Codes: C40B 70/00 : Tags or labels specially adapted for combinatorial chemistry or libraries, e.g. fluorescent tags or bar codes

Abstract

The present invention includes methods for generating combinatorial libraries on solid phase supports in which increased productivity is achieved by pooling all common steps in synthesis without using a tracking or coding system to record the synthetic history of each compound. Methods for generating combinatorial libraries in which containers with random mixtures of solid phase particles are divided and combined together in non-random ways without the exchange of particles between containers are also provided. Various products are optionally produced in multi-stage syntheses according to the invention, such as oligomers and synthetic non-repetitive organic molecules. The methods additionally relate to the identification of each library component without adding extra synthetic, physical, optical, or electronic encoding steps during library synthesis. Combinatorial synthetic systems are also provided.

Claims

1. A method of identifying selected members of a synthesized library of materials, comprising:

(a) providing at least n*m*f solid phase synthesis units, wherein n is equal to a number of choices of different first components in a first stage of synthesis, m is equal to a number of choices of different second components in a second stage of the synthesis, and f is equal to a number of solid phase synthesis units to comprise identical materials upon completion of the synthesis;

(b) segregating the solid phase synthesis units into n separate first stage reaction vessels, wherein each separate first stage reaction vessel comprises at least m*f solid phase synthesis units;

(c) reacting the solid phase synthesis units in each of the separate first stage reaction vessels with a different first component in the first stage of the synthesis;

(d) segregating the solid phase synthesis units of (c) into m separate second stage reaction vessels by distributing at least one of the solid phase synthesis units from each of the separate first stage vessels into each separate second stage reaction vessel such that each of the separate second stage reaction vessels comprises at least n*f solid phase synthesis units;

(e) reacting the solid phase synthesis units in each of the separate second stage reaction vessels with a different second component in the second stage of the synthesis, thereby synthesizing the library of the materials;

(f) detecting one or more distinguishing physical properties of selected members of the library; and

,

(g) identifying the selected members based on the one or more detected distinguishing physical properties.

2. The method of

claim 1

, wherein the method is completely or partially computer implemented.

3. The method of

claim 1

, wherein each different first and second component independently comprises an organic or an inorganic component.

4. The method of

claim 1

, wherein the library of the materials comprises a combinatorial chemical library.

5. The method of

claim 1

, wherein the at least n*m*f solid phase synthesis units are subjected to one or more split/pool synthesis steps prior to (a).

6. The method of

claim 1

, wherein the solid phase synthesis units each comprise a single particle independently selected from one or more of:

a bead, a crown, a piece of paper, a piece of cotton, or a piece of polymer.

7. The method of

claim 1

, wherein one or more of the solid phase synthesis units comprise single functionalized particles.

8. The method of

claim 1

, wherein one or more of the solid phase synthesis units comprise single non-functionalized particles.

9. The method of

claim 1

, wherein at least two of the solid phase synthesis units comprise single particles having different functionalities attached thereto.

10. The method of

claim 1

, wherein at least one of the separate first stage reaction vessels comprises at least two solid phase synthesis units comprising different functionalities.

11. The method of

claim 1

, wherein each separate first stage reaction vessel in (b) comprises m*f solid phase synthesis units.

12. The method of

claim 1

, wherein (b) comprises providing one or more of the at least m*f solid phase synthesis units in one or more two-dimensional arrays in the separate first stage reaction vessels.

13. The method of

claim 1

wherein one or more of the at least m*f solid phase synthesis units in one or more of the separate first stage reaction vessels are non-arrayed.

14. The method of

claim 1

, wherein each of the separate second stage reaction vessels in (d) comprises n*f solid phase synthesis units.

15. The method of

claim 1

, wherein (d) comprises providing one or more of the at least n*f solid phase synthesis units in one or more two-dimensional arrays in the separate second stage reaction vessels.

16. The method of

claim 1

wherein one or more of the at least n*f solid phase synthesis units in one or more of the separate second stage reaction vessels are non-arrayed.

17. The method of

claim 1

, wherein the solid phase synthesis units of (d) are randomly arranged in at least one of the second stage reaction vessels.

18. The method of

claim 1

, wherein the solid phase synthesis units of (d) are non-randomly arranged in at least one of the second stage reaction vessels.

19. The method of

claim 1

, wherein each of the separate second stage reaction vessels designates which different second component reacted with the solid phase synthesis units therein to thereby permit structural identification of selected library members upon detecting the one or more distinguishing physical properties of the selected members in (f).

20. The method of

claim 1

, wherein the solid phase synthesis units each comprise multiple particles combined together.

21. The method of

claim 20

, wherein an array or a container comprises the multiple particles combined together.

22. The method of

claim 20

, wherein at least one of the multiple particles comprises a non-functionalized solid support.

23. The method of

claim 20

, wherein at least one of the multiple particles comprises a solid support having one or more functionalities attached thereto.

24. The method of

claim 20

, wherein at least two of the multiple particles comprise solid supports having one or more identical functionalities attached thereto.

25. The method of

claim 20

, wherein at least two of the multiple particles comprise solid supports having one or more different functionalities attached thereto.

26. The method of

claim 1

, wherein (a) comprises:

(i) segregating the at least n*m*f solid phase synthesis units into p separate third stage reaction vessels, wherein p is equal to a number of choices of different third components in a third stage of the synthesis, and wherein each separate third stage reaction vessel comprises at least n*m*f/p solid phase synthesis units;

(ii) reacting the solid phase synthesis units in each of the separate third stage reaction vessels with a different third component in the third stage of the synthesis; and

,

(iii) combining and mixing the solid phase synthesis units of (ii) in a single pool, thereby providing the at least n*m*f solid phase synthesis units.

27. The method of

claim 26

, wherein the at least n*m*f solid phase synthesis units comprise n*m*f*p solid phase synthesis units.

28. The method of

claim 26

, further comprising:

(iv) separating the at least n*m*f solid phase synthesis units of (iii) into n*m separate containers, wherein the n*m separate containers are segregated into the n separate first stage reaction vessels as the solid phase synthesis units of (b).

29. The method of

claim 26

, further comprising separating the at least n*m*f solid phase synthesis units of (c) into n*m separate containers, wherein the n*m separate containers are segregated into the m separate second stage reaction vessels as the solid phase synthesis units of (d).

30. The method of claims 28 or 29, wherein each of the n*m separate containers comprises multiple particles combined together.

31. The method of

claim 1

, wherein (f) further comprises cleaving the materials from the solid phase synthesis units prior to detecting the one or more distinguishing physical properties.

32. The method of

claim 1

, wherein the solid phase synthesis units of (e) each comprise multiple particles combined together, and wherein (f) further comprises separating selected particles from other particles and cleaving synthesized materials from the selected particles prior to detecting the one or more distinguishing physical properties.

33. The method of claims 1, 31, or 32, wherein the one or more distinguishing physical properties comprise different molecular masses.

34. The method of

claim 33

, wherein the different molecular masses are detected by mass spectrometry.

35. The method of

claim 33

, wherein structural identification of the selected members comprises determining a fingerprint of library members in one or more of the separate second stage reaction vessels.

36. The method of

claim 33

, wherein structural identification of the selected members comprises subtracting a mass of the different second component reacted in a particular separate second reaction vessel from the different detected masses of the selected members to thereby determine masses of different first components included in each of the selected members.

37. The method of

claim 36

, wherein the structural identification accounts for mass defects of reaction.

38. The method of

claim 33

, wherein structural identification of the selected members comprises correlating the different detected masses of the selected members to a physical or logical matrix comprising masses for each individual library member.

39. The method of

claim 38

, wherein at least one entry in the matrix comprises a summation of masses of different combinations of first and second components.

40. The method of

claim 38

, wherein correlations of the different detected masses to entries in the matrix account for mass defects of reaction.

41. The method of

claim 38

, wherein the correlation is computer implemented.

42. A combinatorial library synthesis system, comprising:

(a) a plurality of reaction vessels;

(b) a handling system configured to translocate solid phase synthesis units and reagents to and from the plurality of reaction vessels;

(c) a detection system to detect one or more distinguishing physical properties of selected members of the combinatorial library; and

,

(d) a computer operably connected to the handling and detection systems, the computer comprising system software which directs the handling or detection systems to;

(i) segregate the solid phase synthesis units into n separate first stage reaction vessels to provide m*f solid phase synthesis units in each of the n vessels, wherein n is equal to a number of choices of different first components in a first stage of a library synthesis, m is equal to a number of choices of different second components in a second stage of the library synthesis, and f is equal to a number of solid phase synthesis units which comprise identical materials on completion of the library synthesis;

(ii) deliver one or more of the different first components to each of the n separate first stage reaction vessels, thereby providing for reaction of the different first components with the solid phase synthesis units to provide first stage reacted solid phase members;

(iii) segregate the first stage reacted solid phase members from the n separate first stage reaction vessels into m separate second stage reaction vessels by distributing at least one of the first stage reacted solid phase members from each of the separate first stage reaction vessels into each second stage reaction vessels such that each second stage reaction vessel comprises at least n*f solid phase synthesis units;

(iv) deliver one or more different second components to the second stage reaction vessels, thereby providing for reaction of the different second components with the first stage reacted solid phase members to provide the combinatorial library; and

,

(v) detect one or more distinguishing physical properties of the selected members of the combinatorial library.

43. The combinatorial library synthesis system of

claim 42

, wherein the handling system comprises a bead handler.

44. The combinatorial library synthesis system of

claim 42

, wherein prior to (i) the system software directs the handling system to:

(1) segregate the at least n*m*f solid phase synthesis units into p separate third stage reaction vessels, wherein p is equal to a number of choices of different third components in a third stage of the library synthesis, and wherein each separate third stage reaction vessel comprises at least n*m*f/p solid phase synthesis units;

(2) deliver one or more of the different third components to each of the separate third stage reaction vessels, thereby providing for reaction of the different third components with the solid phase synthesis units to provide third stage reacted solid phase members; and

,

(3) combine and mix the third stage reacted solid phase members in a single pool, thereby providing the solid phase synthesis units for (i).

45. The combinatorial library synthesis system of

claim 44

, wherein the at least n*m*f solid phase synthesis units comprise n*m*f*p solid phase synthesis units.

46. The combinatorial library synthesis system of

claim 44

, wherein the system software further directs the handling system to:

(4) separate the solid phase synthesis units of (3) into n*m separate containers, wherein the n*m separate containers are segregated into the n separate first stage reaction vessels as the solid phase synthesis units of (i).

47. The combinatorial library synthesis system of

claim 44

, wherein the system software further directs the handling system to separate the solid phase synthesis units of (3) into n*m separate containers, wherein the n*m separate containers are segregated into the m separate second stage reaction vessels as the solid phase synthesis units of (iii).

48. The combinatorial library synthesis system of claims 46 and 47, wherein each of the n*m separate containers comprises multiple particles combined together.

49. The combinatorial library synthesis system of

claim 42

, wherein the system software directs the handling system in (iv) to effect cleavage of combinatorial library members from the solid phase synthesis units.

50. The combinatorial library synthesis system of claims 42 or 49, wherein the one or more distinguishing physical properties comprise different masses.

51. The combinatorial library synthesis system of

claim 50

, wherein the detection system comprises a mass spectrometer.

52. The combinatorial library synthesis system of

claim 51

, wherein the computer further comprises at least one database having a logical matrix corresponding to masses of members of a virtual library that are correlated with the detected masses of the combinatorial library members produced by the system to thereby identify chemical structures of the combinatorial library members.

53. The combinatorial library synthesis system of

claim 52

, wherein correlations account for mass defects of reaction of the detected masses.

54. The combinatorial library synthesis system of

claim 52

, wherein at least one entry in the logical matrix comprises a summation of masses of different combinations of first and second components.