Liquid purification or separation – With means to add treating material – Chromatography
Reexamination Certificate
1999-03-25
2002-03-12
Therkorn, Ernest G. (Department: 1723)
Liquid purification or separation
With means to add treating material
Chromatography
C210S659000, C210S656000, C210S143000, C422S070000
Reexamination Certificate
active
06355163
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
This invention relates to apparatus for screening compound libraries, such as compound libraries generated using combinatorial chemistry techniques. The apparatus of this invention employ frontal chromatography in combination with mass spectrometry to screen a library of compounds to identify and rank those members of the library that bind to a target receptor. The apparatus of this invention also permit a compound library to be rapidly screened to determine if one or more members of the library have an affinity for a target receptor as measured by a pre-selected indicator compound.
2. References
The following publications, patents and patent applications are cited in this application as superscript numbers:
1
K. S. Lam,
Anti
-
Cancer Drug Des.
1997, 12, 145-167.
2
P. M. Sweetnam et al.,
In Burger's Medicinal Chemistry and Drug Discovery;
M. E. Wolff, Ed.; John Wiley & Sons: New York, 1995; pp 697-731.
3
R. H. Griffey et al., In
Proceedings of the
45
th
ASMS Conference on Mass Spectrometry and Allied Topics,
Palm Springs, Calif., Jun. 1-5, 1997; p. 400.
4
L. Fang et al., In
Proceedings of the
45
th
ASMS Conference on Mass Spectrometry and Allied Topics,
Palm Springs, Calif., Jun. 1-5, 1997; p. 401.
5
Y.-H. Chu et al.,
J. Am. Chem. Soc.
1996, 118, 7827-7835.
6
Y.-Z. Zhao et al.,
J. Med. Chem.
1997, 40, 4006-4012.
7
Y. F. Hsieh et al.,
J. Mol. Div.
1996, 2, 189-196.
8
R. W. Nelson et al.,
Anal. Chem.
1995, 67, 1153-1158.
9
D. C. Schriemer and L. Li,
Anal. Chem.
1996, 68, 3382-3387.
10
PCT/US97/07964 (International Publication No. WO 97/43641), published Nov. 20, 1997, entitled “Molecular Diversity Screening Device and Method.”
11
R. Wieboldt et al.,
Anal. Chem.
1997, 69, 1683-1691.
12
R. B. van Breemen et al.,
Anal. Chem.
1997, 69, 2159-2164.
13
M. L. Nedved et al.,
Anal. Chem.
1996, 68, 4228-4236.
14
PCT/US95/03355 (International Publication No. WO 95/25737), published Sep. 28, 1995, entitled “Method for Identifying Members of Combinatorial Libraries.”
15
PCT/EP97/02215 (International Publication No. WO 97/43301), published Nov. 20, 1997, entitled “Identification of Members of Combinatorial Libraries By Mass Spectrometry.”
All of the above publications, patents and patent applications are herein incorporated by reference in their entirety to the same extent as if each individual publication, patent or patent application was specifically and individually indicated to be incorporated by reference in its entirety.
State of the Art
In recent years, a large number of combinatorial chemistry techniques have been developed which permit vast libraries of diverse chemical compounds to be rapidly synthesized.
1
In combinatorial chemistry, a series of chemical reactions is typically conducted employing a plurality of reagents at each step to generate a library of compounds. Such techniques have the potential to greatly accelerate the discovery of new compounds having biologically useful properties by providing large collections of diverse chemical compounds for biological screening.
This ability to rapidly generate large collections of compounds using combinatorial chemistry techniques has created a need for new methods of screening compound libraries. The traditional approach of screening each compound individually in an assay to identify those compounds having the desired biological activity is no longer practical due to time and resource constraints. Thus, a need exists for new methods and apparatus which permit the rapid screening of compound libraries.
In this regard, various methods for screening compound libraries have been reported. Typically, these screening methods involve the use of target receptors which have been labeled with fluorescent or other reporter groups.
2
In these methods, the compound library, typically bound to a resin bead, is exposed to the labeled target receptor and those members binding to the labeled target receptor are identified and physically separated. The particular ligand binding to the target receptor is then identified. In many of these techniques, elaborate procedures are required to keep track of individual members of the library. For example, coded tags are often added during the synthesis of the combinatorial library to allow the structure of the individual members to be subsequently determined. Alternatively, combinatorial libraries can be prepared in an array and the individual members of the library subsequently identified by their location in the array. While such methods can be effective, the need to keep track of individual members of the library during their synthesis and screening is quite cumbersome and often limits the type of synthetic procedures that can be employed. Additionally, many of these techniques require that the synthetic procedures be conducted on a solid phase, thus further limiting the synthetic procedures and reagents that can be used.
As an alternative, mass spectrometry is emerging as an important tool for the interrogation of combinatorial libraries. To date, mass spectrometry has been used to assess library quality
3,4
and, when coupled with molecular recognition technologies, has allowed for some success in the isolation and characterization of active library compounds.
5-15
Typically, when screening compound libraries for biologically active members, mass spectrometry is used in combination with a “capture and release” methodology. In this methodology, compound mixtures are presented to the target receptor, which is often immobilized on a solid support, and the resulting ligand-receptor complexes are separated from the unbound members of the library. After separation, the ligand-receptor complexes are typically denatured, for example, with a solvent and the solvent mixture containing the previously bound ligands is presented to the mass spectrometer to permit identification of the high affinity ligands.
For example, ultrafiltration has been used in combination with electrospray mass spectrometry to screen combinatorial libraries.
10-12
In this method, ligands present in a compound library are allowed to bind to a receptor and the resulting ligand-receptor complexes are purified by ultrafiltration. The ligand-receptor complexes are then dissociated using a solvent, such as methanol, and the previously bound ligands are detected by an electrospray mass spectrometer.
Affinity capillary electrophoresis (ACE) has also been coupled with mass spectrometry to screen combinatorial libraries.
5
In this procedure, ACE is used to separate ligand-receptor complexes from unbound ligands and mass spectrometry is used to identify the high affinity ligands.
Similarly, compound libraries have been screened using affinity chromatography in combination with mass spectrometry. For example, WO 97/43301 describes a method for characterizing the members of a combinatorial library, which method utilizes affinity selection in combination with mass spectrometry. Specifically, the members of the library are brought into contact with a domain of interest to allow for binding, i.e., the formation of a complex. After binding, the complex is separated from the unbound members of the library, typically by washing the unbound members from the column containing the complexes. The complexes are then treated to elute the bound library components and the eluted components are analyzed by mass spectrometry. The elution methods described include the use of displacers, chaotrope agents, pH elution, salt gradients, temperature gradients, organic solvents, selective denaturants and detergents. Using such methods, the weakly bound members of the library are purportedly eluted first and analyzed by mass spectrometry, followed by the elution of the more strongly bound members.
There are several disadvantages associated with the “capture and release” methods for screening compound libraries that have been previously reported. First, the procedure used to “release” the bound ligands from the ligand-receptor complexes may alter the binding profile for the various bound
Hindsgaul Ole
Schriemer David C.
Burns Doane , Swecker, Mathis LLP
Hindsgaul Ole
Therkorn Ernest G.
LandOfFree
Apparatus for screening compound libraries does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Apparatus for screening compound libraries, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Apparatus for screening compound libraries will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2834620