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Molecular detection of chromosome aberrations

a chromosome aberration and molecular detection technology, applied in the field of biotechnology, can solve the problems of inconvenient use, high labor intensity, and inconvenient use of cytogenetic analysis by conventional chromosome banding techniques, and achieve the effects of improving sensitivity, specificity and efficacy of analysis

Inactive Publication Date: 2006-01-19
DAKOAS
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

"The present invention provides nucleic acid probes that can be used for diagnostic testing of chromosome aberrations with high sensitivity and specificity. These probes can hybridize in situ with complementary nucleic acid molecules, such as mRNA or DNA, and can be labeled with different reporter molecules for detection. The probes are designed to be distinct and balanced, with one probe hybridizing to a different region of a non-aberrant chromosome, to avoid false-positive diagnosis. The probes can also be tested to ensure they do not hybridize with repetitive sequences. The invention also provides a method for mapping and checking the relative positions of the probes. Overall, the probes offer a more accurate and reliable tool for diagnostic testing of chromosome aberrations."

Problems solved by technology

Traditional techniques such as cytogenetic analyses by conventional chromosome banding techniques are, although highly precise, very labor intensive, require skilled personal and are expensive.
Automated karyotyping is useful for some diagnostic applications, such as prenatal diagnosis, but is ineffective in analyzing the complex chromosomal aberrations of many malignancies.
Furthermore, the above techniques require fresh (cultured) cells, which are not always available.
However, even with this modern technology, several disadvantages can be found that hamper the application of these diagnostic techniques in the rapid screening for chromosomal aberrations related to such malignancies can be found.
Southern blotting lasts 3 to 4 weeks, which is too slow for efficient diagnosis and choice of therapy in malignancies, and allows only 10-15 kb of nucleic acid sequences to be analyzed per probe analysis.
PCR, although, in essence, well-suited for rapid and massive diagnostic testing or even screening, allows only 0.1 to 2 kb of nucleic acid to be analyzed per PCR analysis, which greatly hampers rapid screening of vast stretches of chromosomes and breakpoint cluster regions within the chromosomes.
An additional disadvantage of PCR is its inherent sensibility to mismatched primers.
Small, normal, and physiological alterations which can always be present in the nucleic acid sequence of the gene fragment complementary to the primer hamper the reliable application of PCR and eventually give rise to false-negative results, which renders a PCR-based diagnostic test, albeit very specific, not sensitive enough for reliable diagnosis.
Using large probes renders the FISH technique very sensitive.
However, even the currently used FISH protocols have inherent disadvantages.
These disadvantages mainly relate to the selection of nucleic acid probes employed in the current FISH protocols, which can give false-positive results in the diagnosis of chromosomal aberrations.
For example, probes directed against different chromosomes with a juxtaposition of signals in the case of translocation create a rather large risk of false-positive results.
Hence, the diagnostic tests, although sensitive, are not specific enough to employ standard FISH techniques in massive or rapid diagnostic testing, let alone in automated testing or screening.
False-positive results are especially detrimental to rapid diagnosis if rapid or routine screening of patients is needed to detect malignancies or in evaluating treatment protocols.
A false-positive result then necessitates cumbersome retesting of patients, or even unsuspecting clients that have been submitted to routine screening protocols, and can greatly alarm these people.
However, in practice, 2 to 4% of normal interphase cells tested by FISH will show false-positive results due to the fact that the two probes colocalize by chance.
An additional disadvantage of the current FISH protocols is that it is, in practice, necessary to know both chromosomes that are involved in the translocation as well as the relevant breakpoint regions of both chromosomes to define the nucleic acid probes enabling the detection of the specified translocation, while as yet unknown or ill-defined translocations originating from a well-known gene and an unknown partner gene remain undetected.

Method used

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Embodiment Construction

[0019] Each year, worldwide, many cases of hematopoietic malignancies are being diagnosed. In the European Union (˜375 million inhabitants) this concerns ˜98,000 patients per year. The estimated number of patients in the USA (˜250 million inhabitants) is ˜65,500 per year. The majority of hematological malignancies are of lymphoid origin: acute lymphoblastic leukemias (“ALL”), chronic lymphocytic leukemias, most malignant lymphomas, and multiple myelomas. The nonHodgkin's lymphomas (“NHL”) form the largest group, representing approximately half of all hematopoietic malignancies. Furthermore, European epidemiological studies show that the incidence of NHL is gradually increasing (˜5% per year), which indicates that NHL poses a significant public health problem in Europe and most probably throughout the Western world. Although the annual number of patients diagnosed with ALL is smaller than for NHL, ALL has a high prevalence in children, representing the most frequent malignancy in chi...

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Abstract

The invention relates to the field of cytogenetics and the application of genetic diagnostic techniques in pathology and hematology. Specifically, the invention relates to nucleic acid probes that can be used in hybridization techniques for the detection of chromosomal aberrations and other gene rearrangements such as immunoglobulin and T-cell receptor gene rearrangements. The probes provided by the invention are a distinct and balanced set of probes of comparable size, each preferably being from 1 to 100 kb, or smaller, and flanking a potential breakpoint in a chromosome.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS [0001] This application is a continuation of co-pending application Ser. No. 09 / 439,040, filed Nov. 12, 1999, now U.S. Pat. No. ______ which is a continuation of pending International Application No. PCT / NL98 / 00270, filed on 13 May 1998, designating the United States of America, the contents of which are incorporated by this reference.TECHNICAL FIELD [0002] The invention relates generally to biotechnology, and more particularly to the field of cytogenetics and the application of genetic diagnostic techniques in pathology and hematology. Specifically, the invention relates to nucleic acid probes that can be used in hybridization techniques for the detection of chromosomal aberrations and other gene rearrangements such as immunoglobulin (Ig) and T-cell receptor (TCR) gene rearrangements. BACKGROUND [0003] Chromosomal aberrations are a leading cause of genetic disorders or diseases, including congenital disorders and acquired diseases such as mal...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): C12Q1/68C07H21/04C12N15/09C12Q1/6841C12Q1/6876C12Q1/6886
CPCC12Q1/6841C12Q2600/156C12Q1/6886C12Q1/6876
Inventor VAN DONGEN, JACOBUS JOHANNUS MARIALANGERAK, ANTHONIE WILLEM
Owner DAKOAS
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