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Stomach Cancer Early Diagnosis

#3909


2D Proteomic Expression Analysis of Human Stomach Cancer: Selection of Serum Markers for Diagnosis and Prognosis

Tech Area / Field

  • BIO-CGM/Cytology, Genetics and Molecular Biology/Biotechnology
  • MED-DID/Diagnostics & Devices/Medicine

Status
8 Project completed

Registration date
15.10.2008

Completion date
13.12.2011

Senior Project Manager
Melnikov V G

Leading Institute
Cancer Research Institute, Siberian Branch, Russian Academy of Medical Sciences, Russia, Tomsk reg., Tomsk

Supporting institutes

  • Institute of Highly Pure Biopreparations, Russia, St Petersburg

Collaborators

  • Karolinska Institute, Sweden, Stockholm

Project summary

Stomach cancer has been the leading cause of death for a large part of the twentieth century. Today, more than one million new cases of stomach cancer are registered annually all over the world and about 50,000 new cases are documented in Russia alone. The complexity of modern-day cancer diagnosis and treatment testifies to the need for further development, as well as application in clinical practice, of molecular tools for detection, prognosis and monitoring of the various types of common tumors. This study will focus on the application of these tools specifically to stomach, or gastric, cancer.

Project Goal

The main aim of this project is to identify, evaluate and validate tumor specific protein markers, which can be used for:

  • early serum diagnosis of gastric tumors, applicable to massive screens of patients at risk
  • prognosis of the disease outcome
  • determination of optimal treatment and assessment of its efficiency in a clinical setting

This proposal is based on the following recent developments in genomics:
  1. significant improvements in analysis sensitivity of tumor proteome using two-dimensional gel electrophoresis
  2. easy access to public databases of human cDNA sequences representing tens of thousands of genes with the data on their expression level in normal and tumor cells. It has been proven that extensive comparative analysis of tumor transcriptome and proteome provides an opportunity for the identification of markers that can be useful in diagnosis, prognosis, treatment, and post-operative management of the various types of tumors.

The successful treatment of cancer patients is predominantly defined by early diagnosis. However, the specificity and sensitivity of existing biomarkers of gastric cancer are insufficient for application not only in the early, but even in the very late stages of the disease.

Within this proposal, we plan to focus on the detailed comparative analysis of proteomes of gastric tumors and normal tissues, aiming to:

  1. identification of diagnostic and prognostic markers of the intestinal and diffuse-type tumors of the stomach
  2. implementation of sets of antibodies to the identified markers that can be useful in early serum diagnosis, prognosis and post operative management of gastric tumors in the clinical practice.

Staff participants of the current project have extensive expertise and long-term technical experience in the field of cancer research. In sum, they have tens of publications in international journals and have obtained several patents for innovations in early diagnosis of different tumors (see the reference list). Several project participants worked abroad for substantial periods of time in university medical schools, hospitals, and cancer centers (Memorial Sloan-Kettering Cancer Center, New York, USA; Karolinska Institute, Stockholm, Sweden; Cold Spring Harbor Laboratory, Cold Spring Harbor, USA; University of Pennsylvania, Philadelphia, USA). Project participants from the Institute of Highly Pure Biopreparations (Saint-Petersburg) add their profound knowledge of protein chemistry and immunochemistry to the group.

Experimental Objectives:

The experimental objectives of the project are:

  1. Identification of proteins which have a significantly elevated level of expression in tumors of the intestinal and diffuse types by comparative analysis of proteomes of normal tissues and gastric tumors using two-dimensional gel electrophoresis.
  2. Analysis of changes in expression of the corresponding genes (by RT-PCR in real time) in the vast collection of frozen gastric cancer biopsies of the Karolinsky Institute (Sweden; see letter of collaboration). Identification of prognostic markers based on statistically significant correlations between the elevated expression level of markers and long-term patient survival data. Identification of secreted proteins (by methods of bioinformatics), that are overexpressed most frequently and most significantly in the intestinal and diffuse-type gastric tumors and could be used for cancer diagnosis.
  3. Production of antibodies to prognostic and diagnostic markers and assessment of their efficiency by immunohistochemistry and immunoprecipitation, respectively. Design of tests for: diagnosis of gastric tumors, detection of metastasis and tumor recurrence after an operation, and prognosis of patients’ outcome.

In the event of project success, we plan to apply for patents in order to protect our intellectual property rights for the newly developed prognostic and diagnostic biomarkers of intestinal and diffuse-type gastric tumors for subsequent licensing of the tests in Russia, Europe and the USA for commercial distribution.

Our application will use the following methodologies:

  • methods of collection of samples – preparation and storage of biopsies, surgical materials samples; histological analysis, compilation of computer registry
  • proteomic methods – extraction of soluble proteins, 2-D analysis of proteome expression, protein identification by MALDI-TOF mass-spectrometry
  • protein chemistry – expression of epitope tagged His 6-proteins, purification of proteins by the FPLC
  • methods of DNA and RNA analysis – purification of DNA and RNA, reverse transcription, real time RT-PCR, DNA cloning, sequencing of PCR products
  • methods of immunochemistry – immunization, production of IgG antibodies, immunoaffinity chromatography, immunohistochemistry, immunoprecipitation, Western blotting
  • bioinformatics – comparative computer analysis of the existing databases (dbEST, Gene Cards etc.)

The realization of current project objectives will provide a research opportunity to the scientists of the State Research Institute of Highly Pure Biopreparations (IHPB), prior involved in medical and biological testing of chemical and biological weapons. These scientist will receive a chance to reorient their efforts and to apply their knowledge and experience to the development of novel approaches to Cancer Research in the Public Interest.


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