CN105759062B - A kind of cancer risk assessment kit based on biomarker spectrum for China Rural Population people at highest risk - Google Patents
A kind of cancer risk assessment kit based on biomarker spectrum for China Rural Population people at highest risk Download PDFInfo
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Abstract
The present invention relates to a kind of cancer risk assessment kit based on biomarker spectrum for China Rural Population people at highest risk.Described kit includes the reagent of measure following four biomarker Serum protein expression quantity：Gastrin release propetide, carcinomebryonic antigen, the fragment of Cyfra21-1 and squamous cell carcinoma antigen.It is combined with other clinical indices ages, smoking history, malignant tumour history, sex, available for Accurate Prediction lung-cancer-risk.
Description
Technical field
The present invention relates to medical biotechnology field, specifically, is related to a kind of composed based on biomarker and is directed to China
The cancer risk assessment kit of people in the countryside people at highest risk.
Background technology
Lung cancer is common in world wide and is fatal rate highest cancer.In China, lung cancer morbidity rate and the death rate
It is in first of each malignant tumour.According to world institute of oncology of the World Health Organization (The International Agency
For Research on Cancer, IARC) statistics, China's lung cancer morbidity rate in 2008 is 35/,100,000 people, and the mechanism is estimated
For meter to 2025, China will increase lung cancer 1,000,000 newly every year.According to the Ministry of Public Health of China health statistics yearbook in 2010, cut-off
2009, national lung cancer mortality was 30.83/10 ten thousand people, and the liver cancer higher than the second high mortality exceedes 4/,100,000 people.Although lung cancer
5 years survival rates there was only 15%, but account for 16% early diagnosis and can reach 49% for the patient of lung cancer its 5 years survival rates, it is seen that
Early diagnosis is an important factor for improving patients with lung cancer survival rate.
The early diagnosis of lung cancer depends on the early diagnosis of iconography at present, and current research is found by high-risk people
Low-dose CT examination is carried out in group, it is possible to reduce 20% death rate caused by lung cancer.But in the wide people in the countryside in China
In, carry out extensive CT examinations at present and be difficult in a short time, therefore cause when making a definite diagnosis lung cancer, in medium and advanced lung cancer,
Lose the chance of surgical radical treatment.Therefore need badly one kind independent of CT images technology, suitable for the high-risk people of China Rural Population
The cancer risk assessment kit of group, but had not been reported at present on this kind of kit.
The content of the invention
The purpose of the present invention is to be directed to deficiency of the prior art, there is provided for predicting the composition of lung-cancer-risk.
Another purpose of the present invention is to provide the purposes of described composition.
Another purpose of the present invention is to provide a kind of kit for being used to predict lung-cancer-risk.
To realize above-mentioned first purpose, the present invention adopts the technical scheme that：
A kind of composition for being used to predict lung-cancer-risk, including the Serum protein expression of measure following four biomarker
The reagent of amount：Gastrin release propetide, carcinomebryonic antigen, the fragment of Cyfra21-1 and squamous cell carcinoma antigen.
Preferably, including following reagent：Four kinds of biomarkers and its enzyme labelled antibody, pH value are 9.6 carbonate buffer
Liquid, pH value be 7.4 phosphate buffer, haemocyanin dilution, terminate liquid, tetramethyl benzidine substrates solution.
It is highly preferred that also include following reagent：Normal human serum and positive control serum.
To realize above-mentioned second purpose, the present invention adopts the technical scheme that：
As above application of any described composition in the kit for preparing prediction lung-cancer-risk.
Described prediction lung-cancer-risk is directed to China Rural Population.
To realize above-mentioned 3rd purpose, the present invention adopts the technical scheme that：
A kind of kit for being used to predict lung-cancer-risk, including the Serum protein expression of measure following four biomarker
The reagent of amount：Gastrin release propetide, carcinomebryonic antigen, the fragment of Cyfra21-1 and squamous cell carcinoma antigen.
Preferably, described kit also includes recording the carrier such as drag and evaluation method：
Pernicious probability=ex/(1+ex)
X=-2.9169+ (0.03 × age)+(1.0721 × smoking history)+(0.306 × malignant tumour history)+(- 0.7012
× sex)+(- 0.000155 × ProGRP)+(0.0151 × SCC-Ag)+(0.1238 × CYFRA21-1)+(- 0.00043 ×
CEA)
It is low danger when pernicious risk probability≤0.21；When pernicious risk probability>It is middle danger when 0.21 and≤0.65；And
When pernicious probability>It is high-risk when 0.65；
Previously there are smoking history, smoking history=1, the past non-smoking history, smoking history=0；Previously there is malignant tumour history, it is pernicious swollen
Knurl history=1, previously without malignant tumour history, malignant tumour history=0；Sex is male, sex=1, and sex is women, sex=0.
Preferably, described kit includes following reagent：Four kinds of biomarkers and its enzyme labelled antibody, pH value 9.6
Carbonate buffer solution, pH value be 7.4 phosphate buffer, haemocyanin dilution, terminate liquid, tetramethyl benzidine substrates
Solution.
It is highly preferred that described kit also includes following reagent：Normal human serum and positive control serum.
Herein, according to《Primary bronchogenic carcinoma of lung early diagnoses Chinese Consensus of experts》In definition, people from Chinese countryside
Mouthful people at highest risk, which is defined as living in Chinese countryside, has the crowd of following High Risk of Lung Cancer factor：55~80 years old age；Smoking refers to
>=400 years branch (or 20 bag years) of number；High-risk Exposed history；There are malignant tumour medical history or lung cancer family history；There is chronic obstructive pulmonary disease, diffuse
Property pulmonary fibrosis and pulmonary tuberculosis medical history.
The invention has the advantages that：
1st, present invention firstly provides the fragment and squamous that propetide, carcinomebryonic antigen, Cyfra21-1 are discharged including gastrin
4 kinds of biomarkers and the combination of other clinical indices including cell carcinoma antigen can turn into the leading indicator of screening lung cancer.
2nd, including gastrin release propetide, carcinomebryonic antigen, the fragment of Cyfra21-1 and squamous cell carcinoma antigen
4 kinds of biomarkers and other clinical indices combine as prediction index compared with the low dosage chest CT currently used for examination lung cancer
Index and other clinical indices are more objective, and it is simple, easy, reliable to determine such biomarker expression level.
3rd, disclosure is particularly well suited to the cancer risk assessment of China Rural Population people at highest risk.
Brief description of the drawings
Fig. 1 is bent for the cancer risk assessment model ROC of China Rural Population people at highest risk based on biomarker spectrum
Line.
Embodiment
Embodiment provided by the invention is elaborated below in conjunction with the accompanying drawings.
Embodiment 1
The composition of kit of the present invention：
By biomarker and its enzyme labelled antibody, pH value is 9.6 carbonate buffer solution, and the phosphate that pH value is 7.4 delays
Fliud flushing, haemocyanin dilution, terminate liquid, tetramethyl benzidine substrates solution, normal human serum and positive control serum composition.
Described biomarker includes gastrin release propetide, carcinomebryonic antigen, the fragment and squamous of Cyfra21-1
Cell carcinoma antigen；The carbonate buffer solution that the pH value is 9.6 is that 1.59 grams of sodium carbonate and 2.93 grams of sodium acid carbonates are added into 1L
Obtained in distilled water；The phosphate buffer that the pH value is 7.4 is by 0.2 gram of potassium dihydrogen phosphate, 2.9 gram of 12 hypophosphite monohydrate
Disodium hydrogen, 8.0 grams of sodium chloride, 0.2 gram of potassium chloride, 0.5mL Tween-20s are added in 1L distilled water and obtained；Described haemocyanin
Dilution is that 0.1 gram of cow's serum, sheep blood serum or rabbit serum proteins add to 100mL pH value and obtained for 7.4 phosphate buffer；
Described terminate liquid is 2M sulfuric acid solutions；Described tetramethyl benzidine substrates solution is the tetramethyl that 0.5mL concentration is 2mg/mL
The ethanol solution of base benzidine, the peroxide that the substrate buffer solution and 32 μ L concentration that 10mL pH value is 5.0 are 0.75vol%
Change aqueous solution of hydrogen to be obtained by mixing, the substrate buffer solution that the pH value is 5.0 is disodium hydrogen phosphate containing 0.2M and 0.1M citric acids
Distilled water solution.
The application method of kit of the present invention：
1st, it is coated with：Biomarker is diluted into protein content with the carbonate buffer solution that 0.05M pH value is 9.6 is
1-10μg/mL.Add 0.1mL in the reacting hole of each polystyrene, in 4 DEG C overnight.Next day, solution in hole is discarded, uses pH value
Washed 3 times, every time 3 minutes for 7.4 phosphate buffer.
2nd, it is loaded：Respectively plus measuring samples, normal human serum and positive control after certain haemocyanin diluted
Serum 0.1mL into above-mentioned coated reacting hole, put 37 DEG C be incubated 1 hour after with pH value be 7.4 phosphate buffer wash 3
It is secondary, 3 minutes every time.
3rd, horseradish peroxidase labeling antibody is added：The horseradish peroxidase mark that diluted fresh is added in each reacting hole resists
Body 0.1mL, is washed 3 times, every time 3 minutes after 37 DEG C of incubation 0.5-1 hours with the phosphate buffer that pH value is 7.4.
4th, plus substrate solution develops the color：Tetramethyl benzidine substrates solution 0.1mL is added in each reacting hole, is incubated at 37 DEG C
10-30 minutes.
5th, terminating reaction：2M sulfuric acid 0.05mL is added in each reacting hole.
6th, result judgement：On ELISA detectors, at 450nm, each hole OD values are surveyed after being returned to zero with blank control wells.If
It is the positive more than defined 2.1 times of negative control OD value.
Using normal control product protein expression level as criterion, Serum of Patients with Lung Cancer proteinogram expression quantity divided by this mark
Standard, so that it is determined that expression ratio of patient's proteinogram relative to normal person, ratio is substituted into following formula respectively, calculate pernicious
Risk probability.
Pernicious probability=ex/(1+ex)
X=-2.9169+ (0.03 × age)+(1.0721 × smoking history)+(0.306 × malignant tumour history)+(- 0.7012
× sex)+(- 0.000155 × ProGRP)+(0.0151 × SCC-Ag)+(0.1238 × CYFRA21-1)+(- 0.00043 ×
CEA)
It is low danger when pernicious risk probability≤0.21；When pernicious risk probability>It is middle danger when 0.21 and≤0.65；And
When pernicious probability>It is high-risk when 0.65.
The model sensitiveness is 94.2%, and specificity is 94.5%.
Previously there are smoking history, smoking history=1, the past non-smoking history, smoking history=0；Previously there is malignant tumour history, it is pernicious swollen
Knurl history=1, previously without malignant tumour history, malignant tumour history=0；Sex is male, sex=1, and sex is women, sex=0.
Embodiment 2
The present invention passes through clinical 715 patients of people in the countryside for determining to have High Risk of Lung Cancer factor by collecting, eventually through
Definitive pathological diagnosis lung cancer 434, sampled 154 Serum of Patients with Lung Cancer by method of randomization；Simultaneously 281 non-lung cancer persons'
In Serum Bank, sampled 235 normal human serums by method of randomization.The sub- immunodetection detection of applied chemistry luminous particle
4 kinds of biomarkers in serum, while gather the clinical information such as patient age, smoking history, sex.Obtain model sensitiveness and spy
It is different in nature as shown in table 1.It is bent for the cancer risk assessment model ROC of China Rural Population people at highest risk based on biomarker spectrum
Line is as shown in Figure 1.
Cancer risk assessment model validation of the table 1 based on biomarker spectrum for China Rural Population people at highest risk
Described above is only the preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
Member, on the premise of the inventive method is not departed from, can also make some improvement and supplement, and these are improved and supplement also should be regarded as
Protection scope of the present invention.
Claims (3)
1. a kind of application of composition in the kit for preparing forecast China people in the countryside people at highest risk's lung-cancer-risk, described group
Compound includes the reagent of measure following four biomarker Serum protein expression quantity：Gastrin release propetide, carcinomebryonic antigen,
The fragment and squamous cell carcinoma antigen of Cyfra21-1, scoring model is and evaluation method is：
Pernicious probability=ex/(1+ex)
X=-2.9169+ (0.03 × age)+(1.0721 × smoking history)+(0.306 × malignant tumour history)+(- 0.7012 × property
Not)+(- 0.000155 × ProGRP)+(0.0151 × SCC-Ag)+(0.1238 × CYFRA21-1)+(- 0.00043 × CEA)
It is low danger when pernicious risk probability≤0.21；When pernicious risk probability>It is middle danger when 0.21 and≤0.65；And when evil
Property probability>It is high-risk when 0.65；
Previously there are smoking history, smoking history=1, the past non-smoking history, smoking history=0；Previously there are malignant tumour history, malignant tumour history
=1, previously without malignant tumour history, malignant tumour history=0；Sex is male, sex=1, and sex is women, sex=0；
Described China Rural Population people at highest risk, which is defined as living in Chinese countryside, has the crowd of following High Risk of Lung Cancer factor：
55~80 years old age；Cigarette smoking index >=400 year branch or 20 bag years；High-risk Exposed history；There are malignant tumour medical history or lung cancer man
Race's history；There are chronic obstructive pulmonary disease, diffusivity pulmonary fibrosis and pulmonary tuberculosis medical history.
2. application according to claim 1, it is characterised in that the composition includes following reagent：Four kinds of biological markers
Thing and its enzyme labelled antibody, pH value are 9.6 carbonate buffer solution, and pH value is 7.4 phosphate buffer, and haemocyanin dilutes
Liquid, terminate liquid, tetramethyl benzidine substrates solution.
3. application according to claim 2, it is characterised in that the composition also includes following reagent：Normal human serum
And positive control serum.
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