Method for screening combinatorial bead library, capturing cells from body fluids, and ligands for cancer cells

• US 6,670,142 B2
• Filed: 10/26/2001
• Issued: 12/30/2003
• Est. Priority Date: 10/26/2001
• Status: Expired due to Fees

• Current Assignee: Regents of the University of California (University of California)
• Original Assignee: Regents of the University of California (University of California)
• Inventors: Lam, Kit S., Lau, Derick H.
• Primary Examiner(s): WESSENDORF, TERESA D
• Application Number:
  US10/032,678
• Publication Number:
  US 20030170735A1
• Time in Patent Office: 795 Days
• Field of Search: 435/7.23, 435/7.1, 435/7.2, 435/7.21, 435/4, 435/6, 530/317, 530/328, 530/300
• US Class Current: 506/10
• CPC Class Codes: C07K 7/06 having 5 to 11 amino acids
• CPC Class Codes: G01N 33/5011 for testing antineoplastic activity
• CPC Class Codes: G01N 33/54313 the carrier being characterised by its particulate form

Abstract

The invention includes a cell-growth-on-bead assay for screening a one-bead-one-compound combinatorial bead library to identify synthetic ligands for cell attachment and growth or proliferation of epithelial cells. Cells are incubated with a compound bead library for 24 to 72-hours, allowing them to attach and grow on the beads. Those beads with cells growing are removed, and the ligand on the bead is identified. Also provided are ligands specific for epithelial cancer cells. The invention also includes a method of capturing epithelial cells from body fluids. In this method, beads are prepared with a known ligand specific for a particular type of cell and incubated for 24 to 72 hours with a sample of the body fluid being tested. Those cells attached to the beads are removed and identified.

Claims

1. A method of identifying a peptide ligand that promotes cell attachment and proliferation, comprising:

introducing a suspension of live mammalian cells to a one-bead-one-compound combinatorial peptide library, wherein said library comprises multiple beads with synthetic compounds attached thereto;

incubating said cells with said beads of said library for a period of about 24 to about 72 hours;

identifying a bead having cells growing thereon;

isolating said identified bead; and

determining the chemical structure of the compound attached to said identified bead.

2. A method for isolating and capturing epithelial mammalian cells of a particular type from body fluid, comprising:

introducing a sample of body fluid to a multiplicity of beads having a known ligand specific for a particular type of cell;

incubating said body fluid with said beads for a period of about 24 to about 72 hours;

identifying a bead having cells growing thereon;

isolating said bead; and

recovering the cells, growing on said isolated bead.

3. The method of claim 2, wherein said incubating is at a temperature of about 37°

C.

4. The method of claim 2, wherein said ligand is a peptide specific for a cancer cell.

5. The method of claim 2, wherein said ligand is specific for human epithelial lung cancer cells and has the chemical structure of cXGXGXXc, in which “

c”

is D-cysteine;

“

X”

is an amino acid selected from the group consisting of L-amino acids, D-amino acids, and unnatural amino acids; and

“

G”

is glycine.

6. The method of claim 5, wherein said ligand has the chemical structure cNGRGEQc, in which “

c”

is D-cysteine.

7. The method of claim 6, wherein said body fluid is human pleural fluid.

8. A ligand specific for human epithelial lung cancer cells, wherein said ligand has the chemical structure of cXGXGXXc, in which “

c”

is D-cysteine;

“

X”

is an amino acid selected from the group consisting of L-amino acids, D-amino acids, and unnatural amino acids; and

“

G”

is glycine.