Methods for treating cancer using anti-Wnt2 monoclonal antibodies and siRNA

Inactive Publication Date: 2006-02-23
RGT UNIV OF CALIFORNIA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0014] This invention provides a method of inhibiting the proliferation of a cell that overexpresses a Wnt2. The method compri

Problems solved by technology

Surgery, chemotherapy, and radiation have been used with generally unsatisfactory results in advanced disease.
Since MPM usually presents at an advanced stage, a curative resection is rarely possible.
Radiotherapy has failed to show clinical benefit as a single treatment modality, and the administration of chemotherapy is mostly restricted to the advanced stage with limited efficiency (Kindler, Lung Cancer 45 Suppl 1:S125-7 (2004)).
Alternative strategies based on pleural injections of recombinant cytokines have similarly proven unsatisfactory (Bard et al.
Since current interventions offer only limited benefit, and overall survival is low, there is an urgent need to develop new therapeutic agents based on a greater understanding of MPM's underlying molecular mechanisms.
However, the use of polyclonal and monoclonal antibodies in humans is severely restricted when the polyclonal monoclonal antibodies are produced in a non-human animal.
Repeated injections in humans of a “foreign” antibod

Method used

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  • Methods for treating cancer using anti-Wnt2 monoclonal antibodies and siRNA
  • Methods for treating cancer using anti-Wnt2 monoclonal antibodies and siRNA
  • Methods for treating cancer using anti-Wnt2 monoclonal antibodies and siRNA

Examples

Experimental program
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example 1

Materials and Methods

[0221] 1. Cell Lines and Tissue Samples

[0222] Human non-small-cell lung cancer (NSCLC) cell lines (A549, NCI-H1703, H460, and H1299), mesothelioma cancer cell lines (NCI-H2052, H28 and H513), melanoma cell lines (LOX, FEM, FEMX, and SK-Mel-2), breast cancer cell lines (MCF-7 and HuL100, and colon cancer cell lines (HCT116 and SW480) were from American Type Culture Collections (ATCC, Manassas, Va.). Mesothelioma cancer cell lines were obtained from the following sources: LRK1A and REN through a generous gift from Dr. Steven Albelda (University of Pennsylvania, Philadelphia, Pa.), NCI-H2052, H28 and H513 from American Type Culture Collections (ATCC, Manassas, Va.), MS-1 and NCI-H290 from NIH (Frederick, Md.) and LP9 were from the Cell Culture Core Facility at Harvard University (Boston, Mass.). All cell lines except LP9 were cultured in RPMI 1640 supplemented with 10% fetal bovine serum, penicillin (100 IU / ml) and streptomycin (100 μg / ml). LP9 was cultured in M1...

example 2

Identification of Antigenic Wnt2 Peptides

[0243] Antigenic peptides of human Wnt2 protein were determined using various methods. For example, the EMBOSS (Parker et al., Biochemistry 25:5425-5432 (1986)) finds antigenic sites in proteins. Antigenic peptides were also determined using the method of Kolaskar and Tangaonkar (K&T; FEBS Lett. (1990) 276(1-2):172-4). Both methods led to the identification of similar antigenic peptides of human Wnt2 (Table 1). While most of the antigenic peptide sequences identified can be used to generate antibodies that specifically bind to human Wnt2, some antibodies may also bind to other Wnt proteins due to amino acid sequence homology among various Wnt proteins with human Wnt2. For example, the amino acid sequences of SEQ ID NO:42 and SEQ ID NO:44 have homology to human Wnt2B (Wnt13); the amino acid sequence of SEQ ID NO:43 has homology to human Wnt2B (Wnt13), Wnt3, Wnt3A, Wnt5B, and Wnt10A; the amino acid sequence of SEQ ID NO:45 has homology to huma...

example 3

Generation of Anti-Wnt2 Monoclonal Antibodies and Antibody Incubation with Cells

[0244] The antigen used to raise monoclonal anti-Wnt2 antibodies was a synthetic peptide corresponding to amino acid residues 49-63 of the human Wnt2 (Ac-SSQRQLCHRHPDVMR-amide, SEQ ID NO:2). This antigen was chosen bioinformatically based on its hydrophilicity (Parker et al., (1986) Biochemistry 25(19):5425-32), antigenicity (Welling et al., (1985) FEBS Lett. 188(2):215-8), accessibility (Janin, Nature 277:491-2 (1979)), sequence homology (BLAST search), and N-terminal vicinity.

[0245] The anti-Wnt2 mouse monoclonal antibody (IgG1) was custom-made at Rockland Inc. (Gilbertsville, Pa.). Several hybridoma cell lines were generated of which five were characterized in detail: (1) 17F7.G7 (Subclone A; FIG. 14); (2) 17F7.G7 (Subclone B; FIG. 18); (3) 17F7.E5 (FIG. 16); (4) 8B11.D2 (FIG. 15); and (5) 8B11.H6 (FIG. 17). Two different light chains were found in 8B11.H6, termed 8B11.H6 (Chain1) and 8B11.H6 (Chain...

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Abstract

This invention relates to methods of inhibiting the growth of cells, in particular cancer cells, that overexpress Wnt2. The methods comprise contacting the cell with an agent that binds to Wnt2 mRNA or Wnt2 protein, interferes with Wnt2 signaling or inhibits binding of the Wnt2 protein to another protein, such as a Frizzled receptor.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application claims benefit of provisional application Ser. No. 60 / 571,323, filed May 14, 2004, the disclosure of which is incorporated in its entirety herein by reference.FIELD OF THE INVENTION [0002] This invention relates to methods of inhibiting the growth of cancer cells that overexpress Wnt2 protein. The methods comprise contacting the cell with an agent that binds to Wnt2 mRNA or Wnt2 protein, interferes with Wnt2 signaling, or inhibits binding of the Wnt2 protein to other proteins, such as the Frizzled receptor. BACKGROUND OF THE INVENTION [0003] Lung Cancer is the leading cause of cancer death in the United States and worldwide, with >170,000 newly diagnosed cases each year in the US and nearly a million cases worldwide (Minna et al. Cancer Cell. 1(1):49-52 (2002)). Despite aggressive approaches made in the therapy of lung cancer in the past decades, the 5-year survival rate for lung cancer remains under 15% (Minna et al...

Claims

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Application Information

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IPC IPC(8): A61K48/00A61K39/395C07K16/18C12N15/113C12P21/06
CPCA61K39/3955A61K45/06A61K2039/505C07K16/18C07K2316/96C07K2317/565C12N2310/14C07K2317/73C12N15/113A61K2300/00C07K2317/75A61P1/04A61P1/16A61P1/18A61P11/00A61P13/08A61P13/12A61P15/00A61P17/00A61P25/00A61P35/00A61P35/02A61P43/00A61P5/14
Inventor YOU, LIANGHE, BIAOXU, ZHIDONGJABLONS, DAVID
Owner RGT UNIV OF CALIFORNIA
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