WO2008024009A1 - Niveau de transcription du gène timp3 utilisé en tant que marqueur servant à diagnostiquer le cancer du poumon non à petites cellules - Google Patents

Niveau de transcription du gène timp3 utilisé en tant que marqueur servant à diagnostiquer le cancer du poumon non à petites cellules Download PDF

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WO2008024009A1
WO2008024009A1 PCT/RU2006/000430 RU2006000430W WO2008024009A1 WO 2008024009 A1 WO2008024009 A1 WO 2008024009A1 RU 2006000430 W RU2006000430 W RU 2006000430W WO 2008024009 A1 WO2008024009 A1 WO 2008024009A1
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gene
lung cancer
transcription
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Russian (ru)
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Tamara Dmitrievna Mashkova
Nina Yurievna Oparina
Olga Leonidovna Zinovieva
Evgeny Pavlovich Kopantsev
Tatyana Viktorovna Vinogradova
Marina Valerievna Zinovieva
Irina Borisovna Zborovskaya
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Institut Molekulyarnoi Genetiki Rossiiskoi Akademii Nauk (Img Ran)
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Priority to PCT/RU2006/000430 priority Critical patent/WO2008024009A1/fr
Priority to EA200701894A priority patent/EA010571B1/ru
Publication of WO2008024009A1 publication Critical patent/WO2008024009A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
    • C12Q1/6886—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material for cancer
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/158—Expression markers

Definitions

  • the present invention relates to medicine, in particular oncology, and molecular biology and can be used for early diagnosis of non-small cell lung cancer, including squamous cell cancer.
  • Lung cancer remains one of the main causes of cancer death in the world. Mortality from lung cancer exceeds mortality from cancer of the breast, colon and prostate combined [Jamal A., Siegel R., Wagd E., Murrau T., Xu J., Smigl C, Thun MJ. 2006. Sapser statists, 2006. CA Sapser J Slip. 56, 6-30]. In Russia, mortality rates from lung cancer in men are 68.2 cases per 100 thousand of the population, in women - 6.8 cases [Zaridze D.G. 2004. Epidemiology and etiology of malignant neoplasms, pp. 29-85 - in the book. Carcinogenesis / Ed. D.G. Zaridze. -M .: Medicine].
  • NSCLC includes squamous cell lung cancer (PRL), lung adenocarcinoma (AKL), and large cell cancer.
  • PRL squamous cell lung cancer
  • ADL lung adenocarcinoma
  • large cell cancer adenocarcinoma
  • the main methods for diagnosing lung cancer are instrumental - x-ray and endoscopic. Sputum analysis for atypical cells, lung tissue biopsy, computed tomography, and fluorescence fibroscopy are also used. Immunological diagnostic methods have so far limited clinical use.
  • the definition of tumor markers is of practical importance: CEA - cancer embryonic antigen - tumor marker of colon cancer, but can be used in the evaluation of lung cancer; NSE - neuron-specific enolase - in some cases used in assessing the status of patients with lung cancer; tissue polypeptide antigen (TPA) - a fragment of cytokeratin, a more specific marker lung cancer. These markers are used to monitor treatment, to identify relapses, but not to diagnose PJI at an early stage.
  • SCC markers are widely used - squamous cell carcinoma antigen and a fragment of cytokeratin-19 (CYFRA 21.1) [Rastor A., Mepepes MJ, Rastor V., Lloris R., Azpar Jc 1997, Diagpost cclu CEA and CYFRA 21.1 ip hmg sapper: a Vauesiap apalusis. European Res. J. 10, 603-609; Wussheri G., Torshio R., Ferrigno D. 2003.
  • Diagnostic signs of malignant transformation of cells can also be changes in the genome in tumor cells — point mutations in the coding and regulatory regions of genes, microsatellite instability, allelic losses, changes in the level of transcription or translation of genes, methylation of promoter regions of the gene, etc. Unlike protein molecular genetic markers have significantly greater sensitivity. Lung cancer is accompanied by a change in the functional activity of many genes.
  • the TIMP gene [Ketupep E., Anttila S., Serrapep JK, Karjalaipep A., Edgren H., Lindstrom I, Salovara R., Nissep AM, SaIo J. are considered.
  • Tissue inhibitor of metalloproteinase 3 - TIMRZ participates in intercellular interactions and induces apoptosis, can inhibit the growth of many tumors, including HMPJI, angiogenesis, germination and metastasis [Dr. A., Quax PH, Neumann M., vap der Laap WH, Pap G, Dupda S ., Meipeske L, Kekow J., Neumapp W., Huizip TW, Naumpp M., Kopig W., Pare T. 2005.
  • the TIMPS gene is considered a potential tumor suppressor gene.
  • the loss of activity of such genes occurs as a result of point mutations, alleles of deletions, homozygous deletions, or hypermethylation of the CpG island of genes, especially in the promoter regions [Velipsku SA, Klipge DM, Dekker JD, Smith MW, Vklage TJ., Gilapre FD, DD, Stidle CA, Risshi M.A. 2005. Geothermother metulatiip ip rasma apd srutum ipreseses with lupg sappeg risk. CUn. Sapser Res. 11, 6505-6511].
  • This invention was made possible by the authors of a comparative analysis of the level of transcription of the TIMRZ gene in tumor tissues of the lungs of various types and at different stages of their malignant transformation and the unexpected discovery of the fact that even the early stages of the development of malignant transformation are accompanied by a significant decrease in the level of transcription of the TMPS gene.
  • the present invention in its first aspect, relates to a new marker for the diagnosis of non-small cell lung cancer, which is the level of transcription of the JJMPP gene.
  • the reduced level in the tissue of human tissue suspected of being cancer compared with its level in healthy tissue is a diagnostic sign of non-small cell lung cancer.
  • lung cancer the marker of which is the level of transcription of the TIMRI gene, is squamous cell lung cancer.
  • the present invention relates to the use of the transcription level of the ⁇ MP3 gene as a marker for the diagnosis of non-small cell lung cancer.
  • lung cancer for which the transcription level of the TIMPZ gene serves as a diagnostic marker, is squamous cell lung cancer.
  • the present invention relates to a method for the diagnosis of non-small cell lung cancer.
  • This method includes the following steps: a) obtaining an initial pair of tissue samples from a patient, where one of the samples was obtained from tissue suspected of being affected by cancer, and the second was obtained from adjacent histologically normal (conditionally normal) tissue; b) isolation and purification of RNA preparations from the initial pair of samples; c) synthesis of single-stranded or double-stranded cDNA on an RNA template using oligonucleotide primers; d) normalization of the concentration of TIMRZ cDNA according to the control gene, the transcription level of which is constant in normal conditions and in lung cancer; d) conducting a quantitative or semi-quantitative reaction of amplification of a fragment of the gene TJMP using cDNA obtained on stage c), as a matrix and a pair of gene-specific oligonucleotide primers; f) comparing the amount of the amplified TJMPS DNA fragment for
  • the oligonucleotide primers used to synthesize single or double stranded cDNA are selected from among oligo (dT) -containing primers, random hexamers, or a combination thereof, as well as gene-specific primers.
  • oligonucleotide primers selected in such a way that they specifically hybridize with cDNA even in the presence of an impurity of genomic DNA are used to amplify the cDNA in step d).
  • the primer sequence is represented by SEQ ID NO: 1 and 2.
  • the quantitative or semi-quantitative amplification reaction of a TIMPS gene fragment is real-time PCR or standard RT-PCR.
  • step d) the GAPDH gene encoding a glyceraldehyde-3-phosphate dehydrogenase protein is used as a control gene.
  • Another aspect of the present invention is a set of primers for the implementation of the polymerase chain reaction to determine the level of transcription of the TIMPS gene having the sequence of SEQ W NO: 1 and 2.
  • FIG. 1 shows the results of selecting conditions for determining the level of transcription of the TIMPS gene in lung tissue samples.
  • M is the molecular weight marker for DNA (plasmid DNA ⁇ BR222 / LM).
  • FIG. 2 shows the results of an RT-GShR analysis of the level of transcription of the TIMP 3 gene in PRL with centralized tumor location (after 30 cycles). Sample numbers are indicated above the tracks. T - tumor, N - conditional norm (tissue adjacent to the tumor). Nl - NZ - normal. The GAPDH gene (28 PCR cycles) was used as an internal control. Electrophoretic separation of PCR products was carried out on a 1.8% agarose gel.
  • This invention provides a new genetic marker for the diagnosis of non-small cell lung cancer and based on determining the level of transcription of this marker, a simple, reliable method for the diagnosis of HMPJI at various stages of the development of malignant transformation, including the initial ones.
  • a reliably detectable difference in transcription of the TIMRZ gene in normal and tumor tissues can be used to detect lung cancer.
  • biopsy samples punctate, including material obtained by bronchoscopy with direct biopsy (central cancer of the lungs) and transthoracic (percutaneous) puncture of the tumor (peripheral cancer), can be used.
  • RNA isolation and purification include the following stages: grinding of tumor and normal tissue samples in liquid nitrogen, cell lysis, RNA isolation and purification, RNA quality control by electrophoresis in 1% agarose gel in the presence of ethidium bromide dye or in a denaturing polyacrylamide gel, as well as spectrophotometric determination of the amount of RNA.
  • Homogenization of tissue pieces can be carried out manually, grinding with a pestle in a ceramic mortar, or with the help of mechanical homogenizers, for example, Ompi Mikher or Micro-Dismotr U of Sartorius (Germany).
  • mechanical homogenizers for example, Ompi Mikher or Micro-Dismotr U of Sartorius (Germany).
  • RNA isolation methods In classical RNA isolation methods, strong chaotropic agents are used, such as guanidinochloride and guanidine isothiocyanate, which dissolve the proteins, and sequential extraction with phenol and chloroform to denature and remove proteins [Sambrook J., Fritskh EF, Mapiatis T., Molisilgr. A laborotoru Mapial. 2nd Editiop ed. 1989, CoId Sprippg Narbour: CSHL Regss]. The method using the Trizol reagent [GSCO / Life Testmologie] is also widely used.
  • Trizol reagent GSCO / Life Testmologie
  • inhibitors can be used, such as an RNAsip inhibitor of placental or recombinant origin, or, for example, a vanadyl riboside complex, a non-specific broad-spectrum RNAse inhibitor.
  • Rapid and high-quality RNA isolation can be carried out using a number of commercially available kits (Klonogen, St. Russia; RN Kochsu kits (Qiagep); SV Total RNA Isolatiop Sustem, Protega (USA), etc.).
  • RNA To exclude work with aggressive agents, to speed up and simplify the isolation of RNA, various devices are used, for example, QuickGene-810 (Life Science, Japan). For the extraction of RNA, this device uses an 80 ⁇ m porous membrane, which is 12.5 times thinner than the glass filter commonly used in such devices (1000 ⁇ m). This allows you to reduce the degradation of RNA and increase its output.
  • Reverse transcription reaction synthesis of cDNA on an RNA template isolated from lung tissue samples and its conversion into double-stranded form.
  • RNA matrix if necessary, with the completion of the second chain, allows us to switch from unstable RNA molecules to more stable DNA molecules and amplify using the polymerase chain reaction to the amounts necessary for detection.
  • RT-PCR amplification allows the use of very small amounts of the original RNA (at the level of 1 nanogram), and, consequently, the amount of the studied lung tissue from which RNA is isolated.
  • the reverse transcription reaction can be carried out using a number of commercially available reverse transcriptase preparations, such as Moloney mouse leukemia virus (M-MLV), avian myeloblastosis virus (AMV) reverse transcriptase, PoWerSpirt (point mutation M-MLV-RT), C.
  • M-MLV Moloney mouse leukemia virus
  • AMV avian myeloblastosis virus
  • PoWerSpirt point mutation M-MLV-RT
  • C C.
  • Thether Polr and others, with which you can get amplification products up to several thousand and even several tens of thousands of nucleotide pairs (kbp).
  • the thermo-stable DNA polymerase of Thermus thermorhilus (Th) can be used, which has reverse transcriptase activity in the presence of Mn 2+ ions .
  • primers for reverse transcription, various primers can be used, for example:
  • Oligo (dT) n -containing primers that bind to the endogenous polyA tail at the 3'-end of the mRNA (the number n is usually 12-18, but can reach a larger value). These primers are most often used to obtain full-size cDNA.
  • nucleotides A, C or G are often added at the 3'-end to "lock" the primer at the border of the transcript and the poly-A tract;
  • Random hexanucleotide primers that hybridize with RNA in numerous sites. When reverse transcription with these primers receive short cDNA. Random hexamers are used to overcome the difficulties associated with the strong secondary structure of RNA, they are more effective in reverse transcription of 5'-region of mRNA;
  • Hexamers or other short oligonucleotides (10-12 nucleotides) of random composition can also be used in combination with oligot-containing primers;
  • Gene transcription analysis can be performed using single-stranded or double-stranded and amplified cDNA.
  • Specific primers are most often used for the synthesis of the second chain and its amplification.
  • kits for cDNA synthesis based on the use of various reverse transcriptases and various primers for seed.
  • the SMART method has also been developed (swithchip mesapdism. At the 5 'end of RNA thermoplates of the transcripts), which is based on the property of reverse transcriptases to add several nucleotide residues to the 3' end of the first cDNA synthesized, mainly dC.
  • This oligo (dC) sequence serves as the annealing site for the oligonucleotide adapter having a complementary oligo (dG) sequence at the 3'-end.
  • Reverse transcriptase perceives the primer as an extension of the RNA template and continues the synthesis of the first chain [Schmidt WM, Mueller MW 1999. SarSelst: and high sepsitiv metod fog 5 ' ⁇ -depept de eshishhmept of fiill-lepgth with DNS IP PC-mediated ap of sr mRN ⁇ s. Nisleis Acids Res. 27, eZ l].
  • the first cDNA chain is flanked on one side by a 3'-primer sequence with oligo (dT) at the 3 'end, and on the other hand, by a sequence complementary to the adapter.
  • These primers have the same external sequence, differing only at the 3 'end.
  • the first chain is amplified in TTTTP with a primer corresponding to the outer part of the 3'-primer and adapter.
  • the nucleotide sequence of the common part of these primers is selected depending on further goals, for example, the production of clonotes, the use of subtractive hybridization, etc. The result is double-stranded DNA enriched with full-sized sequences.
  • RNA RNA
  • Kits for producing cDNA in this way are produced by various companies, for example, Evrogen, Russia (MINT kit), ⁇ loptesh, USA, etc.
  • thermostable DNA polymerase for example, Taq, Pfu, TfI, Tth, Tma, etc.
  • specific primers sites for which are located in the transcript of interest
  • thermostable DNA polymerase for example, Taq, Pfu, TfI, Tth, Tma, etc.
  • specific primers sites for which are located in the transcript of interest
  • conduct standard, semi-quantitative PCR in which the amplified transcript fragment is detected by simple gel electrophoresis, or real-time quantitative PCR.
  • the selection of specific primers is carried out by a method well known to specialists in this field. To select primers and annealing temperatures, it is advisable to use commercially available programs or programs that are freely available on the Internet.
  • the length of the primers can be selected in the range from 18 to 25 bp.
  • this invention does not use rhSase-free DNase.
  • primers for PCR are selected so that they specifically hybridize with cDNA even in the presence of an impurity of genomic DNA in the preparation. This can be achieved, for example, by selecting primers for different exons of the TIMRZ gene.
  • the length of the PCR product that is amplified with impurity genomic DNA will be significantly longer than the expected length of the PCR product amplified with cDNA. If at least one of the selected primers overlaps the boundary between exons, impurities of genomic DNA will not affect the results of the amplification reaction.
  • primers are used: TJRZJF (SEQ ID NO: 1) S'-ASSATSAAGSAGATGAAGATGTASS-3 'and TIMP3_R (SEQ W NO: 2) 5'-ACTTGATCTTGCAGTTACAACCCA-3' It will be understood by a person skilled in the art that others may pairs of primers that differ, for example, in their length, in their localization with respect to the transcript sequence of the ⁇ MP3 gene, which will provide specific and efficient amplification of the TJMPS gene transcript fragment even in the presence of an impurity of genomic DNA.
  • HMPJI shows the smallest spread in transcription levels in normal and tumor tissues.
  • PCR-RV real-time PCR
  • PCR-RV real-time PCR
  • the probe complementary to the middle part of the amplified fragment, contains a fluorophore and a quencher at the ends. When the fluorophore and quencher are associated with an oligonucleotide probe, only slight fluorescence emission is observed.
  • the fluorescent label passes into the solution, being released from the vicinity of the quencher, and generates a fluorescent signal that amplifies in real time in proportion to the accumulation of the amplification.
  • probes for PCR-RV can be carried out in accordance with the standard recommendations of the manufacturer of devices for PCR-RV. If there is no need for multiplex analysis of several genes simultaneously, an economical alternative can be a system using SYBR Green dye specific for double-stranded DNA, the fluorescence intensity of which increases in real time in proportion to the increase in the number of amplicons. In this embodiment, primers without a probe can be used.
  • PCR-RV PCR-RV
  • the use of PCR-RV also reduces the risk of contamination and automates the diagnostic process.
  • the process lasts two to three hours (50 or 60 PCR cycles, respectively) and includes simultaneous PCR, detection of a fluorescent signal, data processing and their presentation in graphical form (full kinetic curve) using special software.
  • a comparative method (relative measurement method, RQ method) is usually chosen, based on the relative measurement of the number of studied polynucleotide sequences, which allows a double comparison of the results for control and target genes, as well as for normal and tumor samples cDNA
  • Example 1 Samples of lung tissue.
  • TNM tumor samples were characterized according to the international system of clinical and morphological classification of TNM tumors, where T (tumo) - T0-T4 - categories reflecting the increase in the size and / or local distribution of the primary tumor, N (on) - NO-NZ - categories reflecting different the degree of defeat by metastases of the regional lymph nodes; M (metastasis) - M0-M1 - characterizes distant metastases. All possible combinations of TNM are combined into larger groups - stages that reflect the course of the tumor process. The first stage of the disease was found in six patients, the P-th - in 18, Sh-th - in five. Signs of distant metastases were not observed in any of the patients. None of the patients were exposed to radiation therapy and chemotherapy before surgery.
  • Example 2 The selection of RNA from tissue samples
  • RNA preparations were purified using the Rpaseu Ms kit kit (Qiagep, USA) according to the protocol attached by the manufacturer. Such a three-stage RNA purification procedure allowed us to effectively get rid of not only soluble glycoprotein precipitates, but also low molecular weight RNA.
  • the quality of the RNA preparation was checked by electrophoresis on a 1% agarose gel in the presence of ethidium bromide. The amount of RNA was determined spectrophotometrically [Sambrook et al, 1989, supra].
  • CDS (5'-AGCAGTGGTATCAACGCAGAGTAC (T) zON- l N-Z l ), and the reverse transcriptase of the Powerscrypt (Clophesh, USA).
  • RNA (1 ⁇ g / ⁇ l) 1.0 ⁇ l SMART primer (6 ⁇ M) 2.0 ⁇ l CDS primer (10 ⁇ M) 1.0 ⁇ l sterile deionized water 1.0 ⁇ l
  • the sample was heated at 72 ° C for 3 min and placed on ice.
  • Amplification conditions 95 0 C, 1.5 min 1 cycle
  • the number of cycles was selected, allowing to obtain the same amount of amplified material.
  • Example 5 Selection of conditions for determining the level of transcription of the TIMPZ gene in lung tissue samples.
  • double-stranded cDNA was amplified using gene-specific primers TIMP ⁇ _F (SEQ ⁇ NO: 1) and TIMP ⁇ _R (SEQ ⁇ NO: 2), which were matched to different exons of the TIMPZ gene, one of the primers overlapping the border between exons. Under these conditions, genomic DNA impurities do not affect the amplification results.
  • the size of the PCR fragment was 279 bp
  • the selection of the conditions for the NDP was carried out on several samples of cDNA. All primers were selected using the Rgimer Dosigper program developed at the IMB RAS.
  • Amplification was carried out in 25 ⁇ l of a mixture containing: 67 mM Tris-Hcl, pH 8.8, 16.6 mM (NI ⁇ ) 2 SO 4 , 0.1% Twep-20, 2.5 mM MgCl 2 , 0.2 mM each of dNTP, 0.1 ⁇ g cDNA, 0.2 ⁇ M each of the primers, 2 units the activity of Bio-Taq DNA polymerase (Dialat Ltd., Moscow).
  • Conditions for conducting PTTR The composition of the reaction mixture (25 ⁇ l):
  • the cDNA samples were normalized by the GAPDH control gene encoding glyceraldehyde-3-phosphate dehydrogenase protein. Used primers selected for different exons of the GAPDH gene, and one of the primers overlaps the border between exons:
  • GAPDHJ S'-GGAGTSAASGGATTTGGTS-3 1 and GAPDHJL: 5'-TGGGTGGAATCAT ATTGGAACAT-3 ',
  • the size of the PCR fragment is 139 bp
  • the selection of the conditions for TTfTP was carried out on several cDNA samples. Amplification conditions: preheating at 94 0 C 2 min; 94 0 C, 30 s, 56 0 C, 30 s and 72 0 C, 30 s - 28 and 30 cycles; 72 0 C, 5 min - 1 cycle.
  • PCR was performed on a MasterSusler, Errepdorf (Germany) thermocycler with a heating cap or Tertsik thermocycler, DNA technology ( Russia). Amplification products were analyzed on a 1.8% agarose gel with 0.5 ⁇ g / ml ethidium bromide (see Fig. 1). As a result, the optimal conditions for OT- PCR (27-30-33 cycles), in which a linear relationship was obtained between the number of cycles and the number of PCR products. All amplification reactions were repeated three times.
  • the fluorescence intensity of the bands after electrophoretic separation of PCR products was quantified using the GeperProfiler photo densitometry program (https://www/scanalytics.com) and expressed as normal / tumor relative intensity values (N / T).
  • Amplified fragments of the TIMPZ gene were cloned in the pGEM®-T ⁇ facility redesign torque vector ( ⁇ romega) and sequenced. Their nucleotide sequences completely coincided with the sequences of the corresponding cDNA fragment. Sequencing was performed using the ABI PRISM® BigDue TM TERMIPATOR v reagent kit. 3.1 followed by analysis of the reaction products on an ABI PRISM 3100-Avant automated DNA sequencer.
  • Example 6 Determination of the level of transcription of the TIMRZ gene in samples of normal and tumor tissues.
  • Master-mix by mixing all PCR components except the matrix. Master-mix should be prepared at the rate of 1 reaction with a volume of 25 ⁇ l for each sample + 1 additional reaction with a volume of 25 ⁇ l.
  • the present invention also provides that determining the level of transcription of the T ⁇ MRZ gene will be useful in monitoring the effectiveness of anti-cancer therapy, and the analysis by the method of the present invention should be carried out before and after the course of treatment, and if necessary during the course of treatment.
  • An increase in the level of transcription of the TTMPS gene during or at the end of the course of treatment will indicate the restoration of the activity of the tumor suppressor gene TIMRZ, which may indicate positive changes in treatment.
  • a decrease in the level of transcription of the TIMRZ gene will indicate a further suppression of gene activity, which may indicate a lack of treatment effectiveness.
  • T0 - T4 - categories reflecting the local distribution of the primary tumor; N0 - N3 - categories reflecting a different degree of metastatic lesion of regional lymph nodes; M0-M1 - characterize the presence of distant metastases.

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Abstract

La présente invention concerne le domaine de la médecine et notamment l'oncologie et la biologie moléculaire. Selon l'invention, on utilise en tant que marqueur servant à diagnostiquer le cancer du poumon non à petites cellules un niveau de transcription du gène TIMP3. Un niveau de transcription réduit dans un tissu humain probablement affecté par le cancer, en comparaison à son niveau dans un tissu sain, sert d'indicateur diagnostique d'un cancer du poumon non à petites cellules. L'invention permet de diagnostiquer, avec un degré de véridicité élevé, le cancer du poumon non à petites cellules, y compris le carcinome de cellules squameuses, y compris au stade le plus précoce de la progression de la transformation tumorale.
PCT/RU2006/000430 2006-08-15 2006-08-15 Niveau de transcription du gène timp3 utilisé en tant que marqueur servant à diagnostiquer le cancer du poumon non à petites cellules WO2008024009A1 (fr)

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EA200701894A EA010571B1 (ru) 2006-08-15 2006-08-15 Способ диагностики немелкоклеточного рака легких и набор для его осуществления

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US10240209B2 (en) 2015-02-10 2019-03-26 The Chinese University Of Hong Kong Detecting mutations for cancer screening
US10633713B2 (en) 2017-01-25 2020-04-28 The Chinese University Of Hong Kong Diagnostic applications using nucleic acid fragments
US11261494B2 (en) 2012-06-21 2022-03-01 The Chinese University Of Hong Kong Method of measuring a fractional concentration of tumor DNA
US11581063B2 (en) 2015-07-23 2023-02-14 The Chinese University Of Hong Kong Analysis of fragmentation patterns of cell-free DNA

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