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Use of P2Y purinergic receptor expression for identifying preneoplastic and neoplastic states

A receptor and tumor technology, applied in the direction of anti-receptor/cell surface antigen/cell surface determinant immunoglobulin, antineoplastic drugs, microbial determination/examination, etc. Issues such as randomized and controlled trials of surgical radiation therapy outcomes

Inactive Publication Date: 2009-01-07
BIOSCEPTRE PTY LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Unfortunately, prostate cancer tissue is notoriously heterogeneous, and an important diagnostic feature may be easily overlooked on biopsy
Compounding this situation is the absence of randomized and controlled trials examining the outcomes of surgery or radiation therapy [2]

Method used

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  • Use of P2Y purinergic receptor expression for identifying preneoplastic and neoplastic states
  • Use of P2Y purinergic receptor expression for identifying preneoplastic and neoplastic states
  • Use of P2Y purinergic receptor expression for identifying preneoplastic and neoplastic states

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0057] Example 1. Production of Antibodies

[0058] Following the method adopted by Hansen et al. [15], human P2Y 2 [12], People P2Y 4 [13] Heren P2Y 6 [14] Clone the appropriate antigenic epitope of the receptor consensus sequence. used with P2Y 2 Non-homologous epitopes corresponding to the Leu226-Lys242 segment and equivalent P2Y 4 (Arg226-Arg242) and P2Y 6 (Cys220-Lys236) sequence. Various passes in P2Y 2 and P2Y 4 The epitope added N-terminal Cys was linked to diphtheria toxoid via a maleimide octanoate-N-hydroxysuccinimide cross-linker. All syntheses were performed in an ABI synthesizer using standard t-BOC chemistry [16]. The peptide-antigen cross-linked product was made into a 5 mg / ml suspension with water, and an aliquot was mixed with complete Freund's adjuvant to emulsify. This was followed by intramuscular injection of 1 ml emulsion containing 2 mg peptide and incomplete Freund's adjuvant, and the second, third, fourth, and fifth immunizations were perfor...

Embodiment 2

[0060] Example 2. Immunohistochemical procedure

[0061] Sections with a thickness of 8 μm were cut from unfixed frozen tissues or from paraffin-embedded tissues with a Reichert Jung 2800 cryogenic knife. After allowing the sections to air dry at room temperature for 1 hour, they were fixed in minus 20 acetone for 12 hours, and air dried at room temperature for 1 hour before antibody labeling. Combine it with monoclonal mouse, rabbit or sheep anti-P2Y 2 Incubate with primary antibody at room temperature. After washing, sections were incubated for 30 minutes with secondary antibodies: HRP-conjugated goat anti-mouse secondary antibody (Dako) diluted 1:30 for mouse primary antibody, HRP for rabbit primary antibody - Conjugated anti-rabbit secondary antibody or HRP-conjugated goat anti-sheep secondary antibody. After further washing the sections were immersed in 15% diaminobenzidine tetrahydrochloride (DAB-Sigma) for 10 minutes. Sections were rinsed, air dried and mounted in D...

Embodiment 3

[0063] Example 3. P2Y receptors in human (prostate) cancer tissue

[0064] In a study of 40 normal individuals and 40 human prostate cancer cases, P2Y in human prostate cancer tissues 2 Subtypes increased significantly. There are no visible markers in normal tissue or benign prostatic hyperplasia. P2Y in cancer tissue 2The labeling pattern was distinct, with a greater proportion of labeled acinar epithelial cells at all stages of prostate cancer, suggesting that oncogenic transformation is associated with P2Y 2 There is a direct correlation between the degree of acinar labeling, starting from the nucleus (stage 1; figure 1 ), developing into punctate markers outside the nucleus or in the cytoplasm (stage 2, figure 2 ), followed by deposition of apical and lateral epithelial cells (stage 3; image 3 ), accompanied by morphological changes in advanced stage 3 cases.

[0065] Combination of P2Y and P2X receptor antibodies

[0066] P2Xs are fast-acting ligand (ATP)-gated ...

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Abstract

The present invention relates to methods for identifying pre-neoplastic and neoplastic states in mammals and in particular to a method for identifying pre-neoplasia and neoplasia in cells and tissues based on differential expression of P2Y purineric receptors.

Description

technical field [0001] The present invention relates to methods for identifying preneoplastic and neoplastic states in mammals, particularly methods for identifying preneoplastic and neoplasia in cells and tissues based on differential expression of P2Y purinergic receptors. Background technique [0002] Cellular characteristics such as the degree of variability in cancer cell volume and shape, the proportion of actively dividing cells, and invasion of adjacent tissues in a biopsy sample must be considered when making a cancer diagnosis. Commonly used histological stains are hematoxylin (primary stain) and eosin (counterstain), which differentially label subcellular components. Other diagnostic methods employ antibodies against specific diagnostic molecules in cells (via intracellular epitopes) or on the cell / tissue surface (via extracellular epitopes), which, such as carcinoembryonic antigen (CEA) As seen in the analysis, some specific examples are described below. [000...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K39/395A61P35/00G01N33/48C07K16/28C07K16/30C12N15/09C12P21/08C12Q1/02C12Q1/68G01N33/52G01N33/53G01N33/574
CPCC07K16/28G01N2333/705G01N33/574A61P35/00
Inventor J·巴登M·斯莱特
Owner BIOSCEPTRE PTY LTD
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