CN113862352B - Product for detecting liver cancer or prognosis of liver cancer based on PRSS35 pathway and liver cancer therapeutic agent based on PRSS35 pathway - Google Patents
Product for detecting liver cancer or prognosis of liver cancer based on PRSS35 pathway and liver cancer therapeutic agent based on PRSS35 pathway Download PDFInfo
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- CN113862352B CN113862352B CN202010630380.2A CN202010630380A CN113862352B CN 113862352 B CN113862352 B CN 113862352B CN 202010630380 A CN202010630380 A CN 202010630380A CN 113862352 B CN113862352 B CN 113862352B
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Abstract
The invention discloses a product for detecting liver cancer or prognosis of liver cancer based on a PRSS35 pathway and a liver cancer therapeutic agent based on the PRSS35 pathway. The product comprises a substance for detecting PRSS35 gene or PRSS35 protein. The detection substance is one of a primer pair for specifically amplifying the PRSS35 gene, a reagent for specifically recognizing the PRSS35 protein and a reagent for specifically binding the PRSS35 protein. The invention discovers that the expression of PRSS35 in liver cancer tumor in vivo is obviously down-regulated compared with normal liver tissue; the protein level of PRSS35 in serum of liver cancer patients is also obviously down-regulated compared with normal people. Exogenous overexpression of PRSS35 in the liver of mice significantly slowed the progression of liver cancer induced in mice. Based on the above, the PRSS35 pathway molecule can be a tumor auxiliary diagnosis marker, can be an effective target for tumor treatment, and can also be a survival prognosis index of a liver cancer patient.
Description
Technical Field
The invention relates to the field of liver cancer detection, in particular to a product for detecting liver cancer or prognosis of liver cancer based on a PRSS35 pathway and a liver cancer therapeutic agent based on the PRSS35 pathway.
Background
Liver cancer is one of the most common malignant tumors seriously harming human health, and according to the cancer report of China in 2019, primary liver cancer is the current cause of death of the 4 th common malignant tumor and the 2 nd tumor in China, and seriously threatens the life and health of people in China. Primary liver cancer mainly comprises 3 different pathological types including hepatocellular carcinoma (Hepatocellular carcinoma, HCC), intrahepatic bile duct carcinoma (Intrahepatic cholangiocarcinoma, ICC) and HCC-ICC, and 3 are greatly different in pathogenesis, biological behaviors, tissue morphology, treatment method, prognosis and the like, wherein HCC accounts for 85% -90% (therefore, the patent takes HCC as a main study object, and the later-mentioned liver cancers are all HCC). However, the current treatment mode of liver cancer is mainly surgery and TACE interventional therapy, and the medicine is usually only used as an auxiliary treatment means; diagnosis and treatment means are mainly various imaging examinations, and biomarker detection such as AFP and DFP detection is usually only used as an auxiliary means because a plurality of negative patients exist. Therefore, the key molecular mechanism in the liver cancer occurrence and development process is further clarified, and the method is particularly critical for developing new liver cancer therapeutic drugs and early diagnosis markers.
Numerous reports have been made today to demonstrate that many proteins and their mediated molecular pathways in liver cells are altered during the development of liver cancer to meet the demands of tumor cell survival and proliferation. In recent years, with the development and perfection of sequencing technology and mass spectrometry technology, research on molecular mechanism of liver cancer gradually shifts from small flux to large flux, for example, in 2019 He Fu beginner, by applying high-flux unlabeled proteomics technology, liver cancer tissues of 110 liver cancer patients and proteomes of paracancerous tissues are compared, intracellular protein molecules with significantly changed protein levels when liver cancer occurs are systematically analyzed, and 110 liver cancer patients are subjected to more careful molecular typing according to the changes, so that the method has important significance and value for treating and diagnosing liver cancer.
Meanwhile, the liver is one of the most main endocrine organs of the human body, and the secretory protein group of the liver is obviously changed in the development process of liver cancer. The secretory proteome is a generic term for proteins secreted by cells into the extracellular environment, and secreted proteins are involved in the important roles of intercellular signaling among multicellular organisms. Therefore, the secreted protein has important significance for diagnosis and treatment of liver cancer, and is more suitable for being used as a target for detection and targeted treatment because of the characteristic that the secreted protein of the liver exists in a large amount in blood. However, since the collection and analysis of secreted proteins have a series of technical difficulties, the research of intracellular molecules and signal pathways has been relatively delayed. However, with the development of mass spectrometry technology, researchers have made a series of progress by analyzing and researching cell line level secreted proteome and serum proteome, and 21 secreted proteins have been used as diagnosis and treatment targets for various cancers and have entered 26 clinical trials. However, there is a large number of gaps and unknowns in this field, compared to the extent of the study of intracellular molecules and from the total number of secreted proteins.
Disclosure of Invention
In order to overcome the defects or drawbacks of the prior art, the invention provides a product for detecting liver cancer or prognosis of liver cancer based on a PRSS35 pathway and a liver cancer therapeutic agent based on the PRSS35 pathway.
The object of the invention is achieved by at least one of the following technical solutions.
The invention provides application of molecules in the PRSS35 pathway as potential liver cancer occurrence and development inhibiting molecules in diagnosis, treatment and prognosis relations of liver cancer based on functions and mechanisms of the PRSS35 pathway in the liver cancer occurrence and development inhibiting process.
The invention provides a new theoretical basis and a brand new molecular path for treating liver cancer, and also provides a new product for auxiliary diagnosis and prognosis diagnosis of liver cancer.
The product for detecting liver cancer or liver cancer prognosis based on the PRSS35 pathway provided by the invention comprises a PRSS35 gene or a detection substance of PRSS35 protein. The molecular pathway which can obviously inhibit liver cancer is a molecular pathway mediated by PRSS35 gene/protein.
Further, the product for detecting liver cancer or liver cancer prognosis based on the PRSS35 pathway comprises a kit for detecting PRSS35 genes and a kit for detecting PRSS35 proteins. These two kits used the PRSS35 gene/protein and the pathway molecules mediated thereby, respectively, as detection targets.
Further, the detection substance is one of a primer pair for specifically amplifying the PRSS35 gene, a reagent for specifically recognizing the PRSS35 protein and a reagent for specifically binding to the PRSS35 protein. The reagent that specifically recognizes the PRSS35 protein and the reagent that specifically binds to the PRSS35 protein are preferably antibodies.
Further, the primer pair for specifically amplifying the PRSS35 gene comprises sequences shown as SEQ ID No. 0:1 and SEQ ID No. 0:2.
Further, the method for detecting the PRSS35 gene comprises fluorescence real-time quantitative PCR; the detection of PRSS35 protein includes one of Western blotting, immunohistochemistry, immunofluorescence and enzyme-linked immunosorbent assay (ELISA).
Further, when the expression level of the PRSS35 gene in the liver tissue in the detected sample is more than 2 times higher than that of the control hepatocellular carcinoma sample, the detected sample is positive.
Further, a sample is detected positive when the serum PRSS35 gene expression level in the sample is greater than 20 ng/ml.
The medicine based on the PRSS35 channel for treating liver cancer or increasing the killing of liver cancer by immune cells comprises a substance for improving the expression level of PRSS35 protein or a human recombinant protein of PRSS35. The medicine adopts the means of improving the expression level of PRSS35 protein or regulating and controlling the mediated channel molecules.
Further, the substance for increasing the expression level of the PRSS35 protein is a homologous recombination expression protein of PRSS35 or an estrogen.
According to the invention, through proteomics experiments, molecular biology and cell biology experiments, PRSS35 is found to be significantly down-regulated in liver cancer. The invention screens secreted proteins with obvious difference in a liver cancer cell line and a normal liver cell line, and discovers that PRSS35 in the liver cancer cell line is obviously down-regulated; then detecting the expression level of PRSS35 in serum of a clinical patient with liver cancer and normal person, and finding that PRSS35 in serum of the patient with liver cancer is also obviously down-regulated; further, comparing the expression level of PRSS35 in liver cancer tissue of clinical liver cancer patient and corresponding paracancerous tissue, it is found that PRSS35 is still significantly down-regulated in liver cancer tissue; taken together, the present invention concludes that PRSS35 is significantly down-regulated in liver cancer, suggesting that the PRSS35 molecules of the present invention have the potential to be liver cancer-aiding diagnostic molecules. Therefore, liver cancer or prognosis of liver cancer can be detected based on the PRSS35 pathway, and liver cancer can be treated based on the PRSS35 pathway.
The invention discovers that the molecules in the PRSS35 pathway have potential to be targets for liver cancer treatment and diagnosis. Specific inhibition of liver cancer cell PRSS35 inhibiting molecule and PRSS35 downstream substrate level by preparing targeting medicine (such as neutralizing antibody); or the method and means for preparing medicines for improving the protein amount of the PRSS35 in liver cancer cells have clinical application value in treating liver cancer, and provide a new strategy and an access point for the effective treatment of liver cancer. This also suggests that molecules on the PRSS35 pathway have the potential to be prognostic indicators for survival in liver cancer patients.
Compared with the prior art, the invention has the following advantages and beneficial effects:
(1) According to the invention, PRSS35 is regulated and controlled by the transcription level of ERa channel through bioinformatics prediction, molecular biology and cell biology experiments; firstly, predicting potential transcription factor binding sites of a PRSS35 promoter region through bioinformatics, and finding that ERa can be combined with the PRSS35 promoter; then, through combined experiments in a liver cancer cell line, exogenous estrogen is added or not under the condition of having or not overexpressing ERa, and then the level of PRSS35 of liver cancer cells in each state is detected, the fact that the level of PRSS35 can be obviously up-regulated by the estrogen and the overexpressing ERa is found, and the ERa channel regulates the expression of PRSS35 at the transcriptional level is suggested, so that more cut-in targets and theoretical basis are provided for treating liver cancer through intervening in the PRSS35 channel in the future;
(2) According to the invention, in vivo and in vitro experiments show that the PRSS35 pathway is required to be dependent on an immune microenvironment for inhibiting liver cancer; the inventor firstly overexpresses the protein of the PRSS35 in a liver cancer cell line, and discovers that the PRSS35 has no influence on proliferation of liver cancer cells basically; then, experiments are carried out in an immunodeficient Balb/c-nude mouse, subcutaneous injection is carried out on two groups of cells of liver cancer cell HepG2 (marked as HepG2-PRSS 35) which over-expresses PRSS35 and liver cancer cell HepG2 (marked as HepG2-PRSS 35) which is not treated to establish a subcutaneous tumor transplantation model, and no obvious difference is found in proliferation of the two groups of tumors; furthermore, in the immune sound ICR mice, the in-situ liver cancer model of the mice is induced by injecting YAP5SA over-expression plasmid into the tail vein of high pressure, and under the condition of existence/non-existence of co-injection of PRSS35 over-expression plasmid, the occurrence and development conditions of the in-situ liver cancer of the mice are detected, and as a result, the PRSS35 can obviously inhibit the occurrence and development of the liver cancer in the mice. Meanwhile, protein cleavage substrates of the PRSS35 are obtained through screening by a proteomics combined transcriptomics experiment, and most of the cleavage substrates of the PRSS35 are known to be reported as molecules related to immunization; the experimental data prove that the PRSS35 can inhibit the occurrence and development of liver cancer, but the invention needs to rely on tumor microenvironment in vivo or immune microenvironment in vivo, so that the invention further understands the possible mechanism of the function of the PRSS35 pathway, and provides more cut-in targets and theoretical basis for treating the liver cancer through intervening the PRSS35 pathway in the future.
Drawings
FIG. 1 is a graph showing the results of screening a liver cancer cell line for secreted proteins that differ significantly from a normal liver cell line by SILAC marker secreted proteomics;
FIG. 2A is a graph showing the results of detecting the expression of PRSS35 in cancer tissue and paracancestor tissue of a clinical sample of liver cancer by Western blotting;
FIG. 2B is a graph showing the results of detecting the expression of PRSS35 in serum of liver cancer patients and normal persons by ELISA;
FIG. 3A is a graph showing the results of detecting proliferation of liver cancer cells in various liver cancer cell lines under the condition of overexpressing PRSS35;
FIG. 3B is a graph showing the results of subcutaneously injecting two groups of cells, hepG2-EV and HepG2-PRSS35, into an immunodeficient Balb/c-nude mouse to establish a subcutaneous tumor graft model, and detecting the proliferation of the cells;
FIG. 3C is a graph of experimental results of detecting whether PRSS35 can influence the occurrence and development of in-situ liver cancer in immunocompetent ICR mice by inducing a mouse in-situ liver cancer model by high-pressure injection of YAP5SA into tail vein;
FIG. 4A is a graph of the results of a study to verify that PRSS35 is regulated at the protein level by the transcriptional level of the ERa pathway;
FIG. 4B is a graph of the results of a study to verify that PRSS35 is regulated at the mRNA level by the transcriptional level of the ERa pathway;
FIG. 5A is a graph of the results of a combined proteomic and transcriptomic experiment demonstrating that the highest scoring substrate molecule is the substrate for PRSS35;
FIG. 5B is a graph showing the results of immunoblotting experiments to verify that IGFBP1 is the substrate molecule of PRSS35.
Detailed Description
The present invention will be further illustrated with reference to specific examples. The experimental procedures, which are not specified in the following examples, are generally carried out under conventional conditions, such as those described in the guidelines for molecular cloning experiments (third edition), or according to the manufacturer's recommendations.
Example 1: down-regulation of PRSS35 expression in clinical liver cancer samples
1. SILAC-labeled proteomics to detect secretory protein expression level of liver cancer cell line and normal liver cell line and analyze differential protein
The method comprises the following steps:
(1) Isotopically labeled cell culture and protein sample collection:
respectively establishing PLC cell lines for stably transfecting PCDH-EV and PCDH-PRSS35 plasmids, adding light-standard lysine and arginine into the EV group, adding heavy-standard lysine and arginine into the PRSS35 group, and culturing for 24 hours, wherein 1:2 passage 1 dish, adding puromycin (puromycin, 1.0 mg/ml), culturing for 24h, recording this as isotope labeled incorporation day 1; repeating the steps until the isotope labeling incorporation is cultivated to the 8 th day; the EV and PRSS35 group cells on day 8 are washed 3 times with PBS respectively, 5ml of serum-free corresponding culture medium is added for 24 hours; after 24h of treatment, the medium was filtered with a 0.22 micron filter and the medium of both groups of cells was collected separately into a centrifuge tube; adding protease inhibitor solution into the filtered culture medium, ultrafiltering the filtered culture medium with 3kd-5ml ultrafilter tube, centrifuging 4000g for 30min to minimum volume, and transferring the sucked protein solution into 2ml protein low adsorption EP tube.
(2) Reduction, alkylation, enzymolysis and concentration of protein samples:
respectively taking 50 mu g of protein from EV group and PRSS35 group, respectively adding 100mM HEPES (hydroxyethyl piperazine ethylsulfuric acid) and 8M GuHCL solution (guanidine hydrochloride) (pH 7.0-8.0) with the same volume, uniformly mixing to make GuHCL concentration 4M, and standing at room temperature for 10min; then, mixing the two groups of proteins uniformly, adding the mixture into a 3kd-0.5ml ultrafiltration tube, centrifuging for 15min at 14000g, and discarding filtrate; adding 400 μl of 50mM HEPES and 4M GuHCL solution (pH 7.0-8.0), centrifuging for 15min at 14000g, discarding the filtrate, repeating for 5 times, and centrifuging until the minimum volume is 20-30 microlitres; 425 μl of 50mM HEPES and 4M GuHCL solution (pH 7.0-8.0), 1000g, centrifuging for 2min in an inverted manner, collecting into a new collecting tube, adding 25 μl of 0.1M DTT solution, and making the final concentration of DTT (DL-dithiothreitol) about 5mM in a water bath at 56 deg.C for 30min; cooling the sample to room temperature, adding 25 μl of 0.3M IAM (iodoacetamide) solution, and reacting in dark at room temperature for 30min; thereafter, 100. Mu.l of a 0.1M DTT solution was added to a final concentration of about 20mM, and the IAM reaction was terminated; adding the mixed sample into a 3kd-0.5ml ultrafiltration tube, centrifuging for 15min and discarding the filtrate; 400 microliters of 50mM HEPES solution (pH 7-8), 14000g, centrifuged for 15min, and the filtrate discarded and repeated 3 times; finally, 100. Mu.l of 50mM HEPES solution (pH 7-8), 1000g, were added, centrifuged upside down for 2min into a new collection tube at a ratio of 1:50, 2. Mu.g of pancreatin was added, digested for 18h, and after a further addition of 2. Mu.g of pancreatin, digested for 4h. Then, an ultrafiltration tube of 0.5ml-10kd of the enzymatically hydrolyzed sample was centrifuged for 15min at 14000g, and then 400. Mu.l of ultrapure water was added, and 14000g was centrifuged for 15min, and the filtrate was collected and repeated 3 times, and the filtrate was transferred to an EP tube, and 1.3. Mu.l of TFA (trifluoroacetic acid) solution was added to a final concentration of approximately 0.1% by volume, and then the sample was separated into 8 fractions by a fractionation kit, and the sample was evaporated by vacuum spin drying, and then 15. Mu.l of 0.1% by volume of formic acid was added to dissolve the sample, and 12000g was centrifuged for 5min, and the whole sample was transferred to an upper sample tube, and the sample was prepared.
(3) Upper machine
Carrying out sample analysis on the prepared sample by using a QE-plus mass spectrometer of the Siemens femto company;
(4) Data analysis
The data obtained in (3) were subjected to data analysis using the proteomic analysis software PD2.2 of sameira.
Results: as shown in fig. 1, PRSS35 was found to vary most significantly in the liver cancer cell line PLC (drop) compared to the normal liver cell line THLE 3.
Analysis of results: PRSS35 is significantly expressed in the secretory proteome of liver cancer cell lines. Therefore, the protein level of PRSS35 in the clinical sample is detected by mass spectrometry, which can provide evidence for clinical diagnosis of liver cancer.
2. Immunoblotting (Western blot) test of expression level of gene PRSS35 in clinical liver cancer tissue sample
The method comprises the following steps: crushing clinical liver cancer tumor tissues and corresponding normal liver tissues of each component into tissue homogenate by using a tissue crusher, crushing cells of each component by using a cell lysate, dissolving cell contents (including proteins) into the cell lysate, quantitatively adjusting the proteins of the cell lysate under the same conditions, and finally analyzing each component sample with uniform protein content by Western bolt detection, wherein the method comprises the following specific steps of: after separating proteins by SDS-PAGE electrophoresis, transferring the proteins to a nitrocellulose membrane, standing a spring red staining chart, and sealing the proteins by skimmed milk; the method comprises the steps of fully binding an antibody specifically binding to two proteins, namely PRSS35 and ACTIN (the PRSS35 designs 3 antibodies aiming at 3 functional domains, the design of the antibody is completed by the inventor, the preparation is completed by the Abcloanl company of Wuhan), then binding a secondary antibody specifically binding to the primary antibody, and then carrying out fluorescence development and fixation; the pictures were scanned and analyzed.
Results: as shown in FIG. 2A, western blot analysis showed that the protein expression level of PRSS35 of cancer tissue in clinical tissue samples of 7 liver cancer patients was significantly lower than that of their corresponding paracancestor tissue samples. Fig. 2A is a tissue sample test, which shows that PRSS35 was significantly down-regulated in liver cancer tissue samples.
Analysis of results: PRSS35 is low expressed in clinical liver cancer samples. Therefore, abnormal low expression of PRSS35 in clinical slice samples can be detected by using an auxiliary diagnostic kit containing an IHC antibody that specifically recognizes PRSS35, and it can be further confirmed that the tissue sample is a tumor sample. Analysis of clinical samples further suggests that PRSS35 may be a target for aiding diagnosis.
ELISA detection of the expression level of the Gene PRSS35 in clinical liver cancer serum samples
The method comprises the following steps: and taking serum with the same protein amount from 50 normal people and 50 clinical liver cancer patients, quantitatively adjusting under the same condition, and finally detecting and analyzing serum samples of the normal people and the clinical liver cancer patients with uniform protein content by ELISA. The specific steps include incubation of serum and a reagent plate, cleaning, addition of a pairing antibody, incubation, cleaning, addition of a developing reagent, cleaning, and detection of absorbance of a specific OD value.
Results: as shown in fig. 2B, ELISA analysis showed that the protein expression level of PRSS35 in the clinical serum samples of 50 liver cancer patients was significantly lower than that of the normal human serum samples. Fig. 2B is a serum sample test, which shows that PRSS35 was also significantly down-regulated in the serum of liver cancer patients.
Analysis of results: PRSS35 is low expressed in clinical liver cancer samples. Therefore, the ELISA auxiliary diagnostic kit containing the antibody pair specifically recognizing PRSS35 can detect the abnormal low expression of PRSS35 in the serum of clinical patients, and can further confirm that the patients are liver cancer patients. Analysis of clinical samples further suggests that PRSS35 may be a target for aiding diagnosis.
Example 2: a cell line stably and highly expressing PRSS35 was constructed.
1. Construction of stable PRSS35 highly expressed cell lines
(1) Construction of expression plasmid for PRSS35
The cDNA sequence of the CDs region of PRSS35 was amplified by PCR, and the purified CDs sequence of PRSS35 was constructed into a pCDH-lenti-puro expression vector (available from Addgene) to obtain PRSS35 overexpressing plasmid pCDH-lenti-puro-PRSS35.
(2) Transfecting human liver cancer cell
The obtained pCDH-lenti-puro-PRSS35 and pCDH-lenti-puro empty vector as a control were transferred into HEK293T cells using Lipofectamine2000 (Invitrogen, # 11668), respectively. And after 6 hours the DMEM medium (DMEM; gibco) was replaced with 10% foetal calf serum (Gibco). Lentiviral supernatants were collected 48 hours later and infected with human hepatoma cells HepG2 cells. A stable PRSS35 high-expression cell strain HepG2-PRSS35 can be obtained by screening with puromycin (1 mug/mL; sigma-aldrich) for about 10 days, and a control cell strain is HepG2-EV control.
(3) Culturing stable cell lines highly expressing PRSS35
Liver cancer cells HepG2 were cultured using DMEM medium (DMEM; gibco) supplemented with 10% fetal bovine serum (Gibco) and a semi-inhibitory concentration of puromycin, and cultured in a sterile cell incubator containing 5% carbon dioxide at 37 ℃.
Example 3: functional study of PRSS35 in liver cancer cells
1. Detection of tumor cell proliferation
(1) In vitro experiments: cell growth experiments
Cell number statistics is carried out on cells with different components by a bromophenol blue staining method, and the change of the growth rate of the exogenous liver cancer cells with high expression of PRSS35 is detected. The following is a brief description: the experimental group cells (HepG 2-PRSS 35) or the Control group cells (HepG 2-EV Control) were cultured at a ratio of 2X 10 4 The cells in the corresponding wells are digested and blown into single cells every 24 hours, the cell dilution suspension is dripped into a blood cell counting plate after the bromophenol blue is diluted to a certain multiple, the cell number is counted under a microscope, and finally the cell number of each well is obtained by multiplying the dilution multiple, and the experiment is repeated for 3 times in parallel.
Results: as shown in FIG. 3A, compared with the Control cell HepG2-EV Control (EV curve in the figure), the proliferation of the liver cancer cell highly expressing the exogenous PRSS35 has no significant change (PRSS 35 curve in the figure).
Analysis of results: the cell count experiment result shows that at the cell level, the over-expression of PRSS35 has no obvious effect on the proliferation of liver cancer cells, and the effect of PRSS35 on liver cancer is likely to depend on the microenvironment in vivo.
(2) In vivo experiments: mouse heterogeneous subcutaneous transplantation tumor experiment
The method comprises the following steps: preparing sufficient quantities of HepG2-EV and HepG2-PRSS35 cells; preparing an immunodeficient mouse Balb/c-nude mouse, weighing and recording the day before injection, and randomly separating cages; directly sucking the cells into a 1ml syringe by using a needle, and injecting the cells into the back of the mouse subcutaneously; the size of the subcutaneous transplantation tumor of the mice was measured after 9 days of injection, until day 30, and the mice were killed and the subsequent experiments were performed.
Results: as shown in fig. 3B, PRSS35 did not affect proliferation of subcutaneous engraftment tumors in immunodeficient mice.
Analysis of results: PRSS35 did not affect proliferation of immunodeficient mice subcutaneous transplants, suggesting that the effect of PRSS35 on liver cancer according to the present invention is likely to depend on the immune microenvironment in vivo.
(3) In vivo experiments: spontaneous liver cancer induction experiment of mice
The method comprises the following steps: pBL-CMV-lenti-puro-EV (shown as SEQ ID N0: 3), pBL-lenti-puro-RFP (shown as SEQ ID N0: 4) and pBL-CMV-lenti-puro-YAP5SA (shown as SEQ ID N0: 5) plasmids (the tail vein high pressure injection of pBL-CMV-lenti-puro-YAP5SA plasmids can induce spontaneous liver cancer of mice); amplifying cDNA sequence of PRSS35 CDs region by PCR, constructing purified CDs sequence of PRSS35 into pBL-CMV-lenti-puro-EV expression vector to obtain PRSS35 over-expression plasmid pBL-CMV-lenti-puro-PRSS35; pBL-CMV-lenti-puro-RFP was used as a group, and pBL-CMV-lenti-puro-RFP and pBL-CMV-lenti-puro-YAP5SA were co-injected as a group; pBL-CMV-lenti-puro-PRSS35 and pBL-CMV-lenti-puro-YAP5SA were co-injected as a group, and an equal amount of plasmid was injected into 5 ICR mice of 4 weeks of age by tail vein high pressure injection, respectively. Mice were dissected 3 months after injection and observed for intrahepatic tumor formation and tumor size.
Results: as shown in fig. 3C, the RFP group did not form tumors, whereas the yap5sa+prss35 group had significantly less tumors than the yap5sa+rfp group.
Analysis of results: experimental results show that spontaneous liver cancer caused by YAP5SA can be remarkably inhibited by high expression of PRSS35 in liver. From fig. 3A, 3B and 3C, it was found that PRSS35 did not affect proliferation of liver cancer cells at the cell line level, did not affect tumor proliferation in immunodeficient mice, and significantly inhibited development of liver cancer in immunocompetent mice.
2. Investigation of the upstream regulatory mechanism of PRSS35
The method comprises the following steps: in liver cancer cell lines, mRNA and protein level changes of PRSS35 were detected by WB and qPCR with/without overexpression of ERa, exogenous presence/absence of added estrogen. The primer pair for qPCR detection comprises sequences shown as SEQ ID No. 0:1 and SEQ ID No. 0:2.
Results: as shown in fig. 4A and 4B, fig. 4A is a graph of results at protein level, and fig. 4B is a graph of results at mRNA level, where both estrogen and ERa can up-regulate mRNA and protein levels of PRSS35.
Analysis of results: the demonstration that PRSS35 is subject to transcriptional regulation by the ERa pathway suggests that intervention of the present invention in PRSS35 to treat liver cancer may be carried out by the ERa pathway.
3. Screening for substrate molecules of PRSS35
The method comprises the following steps: preparation of 5X 10 each of the cells of the experimental group (PLC-PRSS 35) and the cells of the Control group (PLC-EV Control) 6 Collecting cells, and extracting mRNA in half for transcriptome sequencing;
the other half is extracted protein for proteomics detection. And selecting a molecular listing list with significantly reduced protein level and no significantly changed mRNA level under the condition of over-expressing PRSS35, taking the molecular listing list as a candidate substrate molecule of the PRSS35, and selecting a molecule with the highest mRNA level/protein level value for verification by an immunoblotting experiment (Western blot).
Results: the results of the combined transcriptomic and proteomic analysis are shown in FIG. 5A, and the molecular IGFBP1 with the highest mRNA/protein level was selected for immunoblotting, and was indeed down-regulated by PRSS35, as shown in FIG. 5B.
Analysis of results: confirming that IGFBP1 is a downstream substrate of PRSS35, suggesting that intervention of PRSS35 by the present invention to treat liver cancer may be practiced by directly modulating IGFBP1 and downstream thereof; it is also suggested that IGFBP1 and its downstream molecules of the invention have potential as targets for liver cancer diagnosis. Meanwhile, the invention discovers from a screened substrate list that the substrate contains a plurality of molecules related to immunity, which suggests that the PRSS35 inhibits the molecular mechanism of liver cancer possibly related to immunity.
The above examples are only preferred embodiments of the present invention, and are merely for illustrating the present invention, not for limiting the present invention, and those skilled in the art should not be able to make any changes, substitutions, modifications and the like without departing from the spirit of the present invention.
Sequence listing
<110> university of North China
<120> products for detecting liver cancer or prognosis of liver cancer based on PRSS35 pathway and therapeutic agent for liver cancer based on PRSS35 pathway
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tcatttaaaa catcagaatg agtatttggt ttagagtttg gcaacatatg cccatatgct 480
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tgtccattcc ttattccata gaaaagcctt gacttgaggt tagatttttt ttatattttg 600
ttttgtgtta tttttttctt taacatccct aaaattttcc ttacatgttt tactagccag 660
atttttcctc ctctcctgac tactcccagt catagctgtc cctcttctct tatggagatc 720
cctcgacctg cagcccaagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt 780
gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg 840
gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt 900
cgggaaacct gtcgtgccag cggatcgatc tgacaatgtt cagtgcagag actcggctac 960
gcctcgtgga ctttgaagtt gaccaacaat gtttattctt acctctaata gtcctctgtg 1020
gcaaggtcaa gattctgtta gaagccaatg aagaacctgg ttgttcaata acattttgtt 1080
cgtctaatat ttcactaccg cttgacgttg gctgcacttc atgtacctca tctataaacg 1140
cttcttctgt atcgctctgg acgtcatctt cacttacgtg atctgatatt tcactgtcag 1200
aatcctcacc aacaagctcg tcatcgcttt gcagaagagc agagaggata tgctcatcgt 1260
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tcaagaatgc atgcgtcaat tttacgcaga ctatctttct agggttaaaa aagatttgcg 1620
ctttactcga cctaaacttt aaacacgtca tagaatcttc gtttgacaaa aaccacattg 1680
tggccaagct gtgtgacgcg acgcgcgcta aagaatggca aaccaagtcg cgcgagcgtc 1740
gacctcgagg gggggcccgg tacccagctt ttgttccctt tagtgagggt taattgcgcg 1800
cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc 1860
acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta 1920
actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca 1980
gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc 2040
cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc 2100
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aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc 2340
tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt 2400
ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa 2460
gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta 2520
tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa 2580
caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa 2640
ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt 2700
cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt 2760
ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat 2820
cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat 2880
gagattatca aaaaggatct tcacctagat ccttttaatt aaaaatgaag ttttaaatca 2940
atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca 3000
cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag 3060
ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac 3120
ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc 3180
agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct 3240
agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc 3300
gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg 3360
cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc 3420
gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat 3480
tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag 3540
tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat 3600
aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg 3660
cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca 3720
cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga 3780
aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc 3840
ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata 3900
tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg 3960
ccacctgacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc 4020
gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt 4080
ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc 4140
cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt 4200
agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt 4260
aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt 4320
gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa 4380
aaatttaacg cgaattttaa caaaatatta acgcttacaa tttccattcg ccattcaggc 4440
tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga 4500
aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac 4560
gttgtaaaac gacggccagt gagcgcgcgt aatacgactc actatagggc gaattggagc 4620
tccaccgcgg ccgcccggtt tatcgttaat atggatcaat ttgaacagtt gattaacgtg 4680
tctctgctca agtctttgat caaaacgcaa atcgacgaaa atgtgtcgga caatatcaag 4740
tcgatgagcg aaaaactaaa aaggctagaa tacgacaatc tcacagacag cgttgagata 4800
tacggtattc acgacagcag gctgaataat aaaaaaatta gaaactatta tttaacccta 4860
gaaagataat catattgtga cgtacgttaa agataatcat gcgtaaaatt gacgcatgtg 4920
ttttatcggt ctgtatatcg aggtttattt attaatttga atagatatta agttttatta 4980
tatttacact tacatactaa taataaattc aacaaacaat ttatttatgt ttatttattt 5040
attaaaaaaa aacaaaaact caaaatttct tctataaagt aacaaaactt ttaaacattc 5100
tctcttttac aaaaataaac ttattttgta ctttaaaaac agtcatgttg tattataaaa 5160
taagtaatta gcttaactta tacataatag aaacaaatta tacttattag tcagtcagaa 5220
acaactttgg cacatatcaa tattatgctc tcgacaaata acttttttgc attttttgca 5280
cgatgcattt gcctttcgcc ttattttaga ggggcagtaa gtacagtaag tacgtttttt 5340
cattactggc tcttcagtac tgtcatctga tgtaccaggc acttcatttg gcaaaatatt 5400
agagatatta tcgcgcaaat atctcttcaa agtaggagct tctaaacgct tacgcataaa 5460
cgatgacgtc aggctcatgt aaaggtttct cataaatttt ttgcgacttt gaaccttttc 5520
tcccttgcta ctgacattat ggctgtatat aataaaagaa tttatgcagg caatgtttat 5580
cattccgtac aataatgcca taggccacct attcgtcttc ctactgcagg tcatcacaga 5640
acacatttgg tctagcgtgt ccactccgcc tttagtttga ttataataca taaccatttg 5700
cggtttaccg gtactttcgt tgatagaagc atcctcatca caagatgata ataagtatac 5760
catcttagct ggcttcggtt tatatgagac gagagtaagg ggtccgtcaa aacaaaacat 5820
cgatgttccc actggcctgg agcgactgtt tttcagtact tccggtatct cgcgtttgtt 5880
tgatcgcacg gttcccacaa tggttaattc gagctcgccc aaaccgggcg cgccgggcat 5940
ttaaatggaa ttcctttatc cagccctcac tccttctcta ggcgccggaa ttgaagatct 6000
gggggatcga tcctctagag tccgttacat aacttacggt aaatggcccg cctggctgac 6060
cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa 6120
tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag 6180
tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc 6240
ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct 6300
acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg 6360
gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt 6420
tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga 6480
cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct cgtttagtga 6540
accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga agacaccggg 6600
accgatccag cctccgcggc cgggaacggt gcattggaac ggacctgcag gaattgatcc 6660
gcggccgcac catg 6674
<210> 4
<211> 7227
<212> DNA
<213> Synthesis by hand (Artificial sequence)
<400> 4
ttcgaagcta attcactcct caggtgcagg ctgcctatca gaaggtggtg gctggtgtgg 60
ccaatgccct ggctcacaaa taccactgag atctttttcc ctctgccaaa aattatgggg 120
acatcatgaa gccccttgag catctgactt ctggctaata aaggaaattt attttcattg 180
caatagtgtg ttggaatttt ttgtgtctct cactcggaag gacatatggg agggcaaatc 240
atttaaaaca tcagaatgag tatttggttt agagtttggc aacatatgcc catatgctgg 300
ctgccatgaa caaaggttgg ctataaagag gtcatcagta tatgaaacag ccccctgctg 360
tccattcctt attccataga aaagccttga cttgaggtta gatttttttt atattttgtt 420
ttgtgttatt tttttcttta acatccctaa aattttcctt acatgtttta ctagccagat 480
ttttcctcct ctcctgacta ctcccagtca tagctgtccc tcttctctta tggagatccc 540
tcgacctgca gcccaagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt 600
tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt 660
gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg 720
ggaaacctgt cgtgccagcg gatcgatctg acaatgttca gtgcagagac tcggctacgc 780
ctcgtggact ttgaagttga ccaacaatgt ttattcttac ctctaatagt cctctgtggc 840
aaggtcaaga ttctgttaga agccaatgaa gaacctggtt gttcaataac attttgttcg 900
tctaatattt cactaccgct tgacgttggc tgcacttcat gtacctcatc tataaacgct 960
tcttctgtat cgctctggac gtcatcttca cttacgtgat ctgatatttc actgtcagaa 1020
tcctcaccaa caagctcgtc atcgctttgc agaagagcag agaggatatg ctcatcgtct 1080
aaagaactac ccattttatt atatattagt cacgatatct ataacaagaa aatatatata 1140
taataagtta tcacgtaagt agaacatgaa ataacaatat aattatcgta tgagttaaat 1200
cttaaaagtc acgtaaaaga taatcatgcg tcattttgac tcacgcggtc gttatagttc 1260
aaaatcagtg acacttaccg cattgacaag cacgcctcac gggagctcca agcggcgact 1320
gagatgtcct aaatgcacag cgacggattc gcgctattta gaaagagaga gcaatatttc 1380
aagaatgcat gcgtcaattt tacgcagact atctttctag ggttaaaaaa gatttgcgct 1440
ttactcgacc taaactttaa acacgtcata gaatcttcgt ttgacaaaaa ccacattgtg 1500
gccaagctgt gtgacgcgac gcgcgctaaa gaatggcaaa ccaagtcgcg cgagcgtcga 1560
cctcgagggg gggcccggta cccagctttt gttcccttta gtgagggtta attgcgcgct 1620
tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac 1680
acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac 1740
tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc 1800
tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg 1860
cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc 1920
actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt 1980
gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc 2040
ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa 2100
acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc 2160
ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg 2220
cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc 2280
tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc 2340
gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca 2400
ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact 2460
acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg 2520
gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt 2580
ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct 2640
tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga 2700
gattatcaaa aaggatcttc acctagatcc ttttaattaa aaatgaagtt ttaaatcaat 2760
ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc 2820
tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat 2880
aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc 2940
acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag 3000
aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag 3060
agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt 3120
ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg 3180
agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt 3240
tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc 3300
tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc 3360
attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa 3420
taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg 3480
aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc 3540
caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag 3600
gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt 3660
cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt 3720
tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc 3780
acctgacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt 3840
gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct 3900
cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg 3960
atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag 4020
tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa 4080
tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga 4140
tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa 4200
atttaacgcg aattttaaca aaatattaac gcttacaatt tccattcgcc attcaggctg 4260
cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa 4320
gggggatgtg ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt 4380
tgtaaaacga cggccagtga gcgcgcgtaa tacgactcac tatagggcga attggagctc 4440
caccgcggcc gcccggttta tcgttaatat ggatcaattt gaacagttga ttaacgtgtc 4500
tctgctcaag tctttgatca aaacgcaaat cgacgaaaat gtgtcggaca atatcaagtc 4560
gatgagcgaa aaactaaaaa ggctagaata cgacaatctc acagacagcg ttgagatata 4620
cggtattcac gacagcaggc tgaataataa aaaaattaga aactattatt taaccctaga 4680
aagataatca tattgtgacg tacgttaaag ataatcatgc gtaaaattga cgcatgtgtt 4740
ttatcggtct gtatatcgag gtttatttat taatttgaat agatattaag ttttattata 4800
tttacactta catactaata ataaattcaa caaacaattt atttatgttt atttatttat 4860
taaaaaaaaa caaaaactca aaatttcttc tataaagtaa caaaactttt aaacattctc 4920
tcttttacaa aaataaactt attttgtact ttaaaaacag tcatgttgta ttataaaata 4980
agtaattagc ttaacttata cataatagaa acaaattata cttattagtc agtcagaaac 5040
aactttggca catatcaata ttatgctctc gacaaataac ttttttgcat tttttgcacg 5100
atgcatttgc ctttcgcctt attttagagg ggcagtaagt acagtaagta cgttttttca 5160
ttactggctc ttcagtactg tcatctgatg taccaggcac ttcatttggc aaaatattag 5220
agatattatc gcgcaaatat ctcttcaaag taggagcttc taaacgctta cgcataaacg 5280
atgacgtcag gctcatgtaa aggtttctca taaatttttt gcgactttga accttttctc 5340
ccttgctact gacattatgg ctgtatataa taaaagaatt tatgcaggca atgtttatca 5400
ttccgtacaa taatgccata ggccacctat tcgtcttcct actgcaggtc atcacagaac 5460
acatttggtc tagcgtgtcc actccgcctt tagtttgatt ataatacata accatttgcg 5520
gtttaccggt actttcgttg atagaagcat cctcatcaca agatgataat aagtatacca 5580
tcttagctgg cttcggttta tatgagacga gagtaagggg tccgtcaaaa caaaacatcg 5640
atgttcccac tggcctggag cgactgtttt tcagtacttc cggtatctcg cgtttgtttg 5700
atcgcacggt tcccacaatg gttaattcga gctcgcccaa accgggcgcg ccgggcattt 5760
aaatggaatt cctttatcca gccctcactc cttctctagg cgccggaatt gaagatctgg 5820
gggatcgatc ctctagagtc cgttacataa cttacggtaa atggcccgcc tggctgaccg 5880
cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata 5940
gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta 6000
catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc 6060
gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac 6120
gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga 6180
tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg 6240
ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg 6300
caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctcg tttagtgaac 6360
cgtcagatcg cctggagacg ccatccacgc tgttttgacc tccatagaag acaccgggac 6420
cgatccagcc tccgcggccg ggaacggtgc attggaacgg acctgcagga attgatccgc 6480
ggccgcacca tggaacaaaa actcatctca gaagaggatc tggcgggcaa gatgagcgag 6540
ctgatcaagg agaacatgca catgaagctg tacatggagg gcaccgtgaa caaccaccac 6600
ttcaagtgca catccgaggg cgaaggcaag ccctacgagg gcacccagac catgaagatc 6660
aaggtggtcg agggcggccc tctccccttc gccttcgaca tcctggctac cagcttcatg 6720
tacggcagca aagccttcat caaccacacc cagggcatcc ccgacttctt taagcagtcc 6780
ttccctgagg gcttcacatg ggagagaatc accacatacg aagacggggg cgtgctgacc 6840
gctacccagg acaccagctt ccagaacggc tgcatcatct acaacgtcaa gatcaacggg 6900
gtgaacttcc catccaacgg ccctgtgatg cagaagaaaa cacgcggctg ggaggccaac 6960
accgagatgc tgtaccccgc tgacggcggc ctgagaggcc acagccagat ggccctgaag 7020
ctcgtgggcg ggggctacct gcactgctcc ttcaagacca catacagatc caagaaaccc 7080
gctaagaacc tcaagatgcc cggcttccac ttcgtggacc acagactgga aagaatcaag 7140
gaggccgaca aagagaccta cgtcgagcag cacgagatgg ctgtggccaa gtactgcgac 7200
ctccctagca aactggggca cagataa 7227
<210> 5
<211> 8097
<212> DNA
<213> Synthesis by hand (Artificial sequence)
<400> 5
aagccattcc tgggacaaat gtggaccttg gaacactgga aggagatgga atgaacatag 60
aaggagagga gctgatgcca agtctgcagg aagctttgag ttctgacatc cttaatgaca 120
tggagtctgt tttggctgcc accaagctag ataaagaaag ctttcttaca tggttatagg 180
aattcgaagc taattcactc ctcaggtgca ggctgcctat cagaaggtgg tggctggtgt 240
ggccaatgcc ctggctcaca aataccactg agatcttttt ccctctgcca aaaattatgg 300
ggacatcatg aagccccttg agcatctgac ttctggctaa taaaggaaat ttattttcat 360
tgcaatagtg tgttggaatt ttttgtgtct ctcactcgga aggacatatg ggagggcaaa 420
tcatttaaaa catcagaatg agtatttggt ttagagtttg gcaacatatg cccatatgct 480
ggctgccatg aacaaaggtt ggctataaag aggtcatcag tatatgaaac agccccctgc 540
tgtccattcc ttattccata gaaaagcctt gacttgaggt tagatttttt ttatattttg 600
ttttgtgtta tttttttctt taacatccct aaaattttcc ttacatgttt tactagccag 660
atttttcctc ctctcctgac tactcccagt catagctgtc cctcttctct tatggagatc 720
cctcgacctg cagcccaagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt 780
gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg 840
gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt 900
cgggaaacct gtcgtgccag cggatcgatc tgacaatgtt cagtgcagag actcggctac 960
gcctcgtgga ctttgaagtt gaccaacaat gtttattctt acctctaata gtcctctgtg 1020
gcaaggtcaa gattctgtta gaagccaatg aagaacctgg ttgttcaata acattttgtt 1080
cgtctaatat ttcactaccg cttgacgttg gctgcacttc atgtacctca tctataaacg 1140
cttcttctgt atcgctctgg acgtcatctt cacttacgtg atctgatatt tcactgtcag 1200
aatcctcacc aacaagctcg tcatcgcttt gcagaagagc agagaggata tgctcatcgt 1260
ctaaagaact acccatttta ttatatatta gtcacgatat ctataacaag aaaatatata 1320
tataataagt tatcacgtaa gtagaacatg aaataacaat ataattatcg tatgagttaa 1380
atcttaaaag tcacgtaaaa gataatcatg cgtcattttg actcacgcgg tcgttatagt 1440
tcaaaatcag tgacacttac cgcattgaca agcacgcctc acgggagctc caagcggcga 1500
ctgagatgtc ctaaatgcac agcgacggat tcgcgctatt tagaaagaga gagcaatatt 1560
tcaagaatgc atgcgtcaat tttacgcaga ctatctttct agggttaaaa aagatttgcg 1620
ctttactcga cctaaacttt aaacacgtca tagaatcttc gtttgacaaa aaccacattg 1680
tggccaagct gtgtgacgcg acgcgcgcta aagaatggca aaccaagtcg cgcgagcgtc 1740
gacctcgagg gggggcccgg tacccagctt ttgttccctt tagtgagggt taattgcgcg 1800
cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc 1860
acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta 1920
actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca 1980
gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc 2040
cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc 2100
tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat 2160
gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt 2220
ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg 2280
aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc 2340
tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt 2400
ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa 2460
gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta 2520
tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa 2580
caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa 2640
ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt 2700
cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt 2760
ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat 2820
cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat 2880
gagattatca aaaaggatct tcacctagat ccttttaatt aaaaatgaag ttttaaatca 2940
atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca 3000
cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag 3060
ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac 3120
ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc 3180
agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct 3240
agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc 3300
gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg 3360
cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc 3420
gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat 3480
tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag 3540
tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat 3600
aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg 3660
cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca 3720
cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga 3780
aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc 3840
ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata 3900
tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg 3960
ccacctgacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc 4020
gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt 4080
ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc 4140
cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt 4200
agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt 4260
aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt 4320
gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa 4380
aaatttaacg cgaattttaa caaaatatta acgcttacaa tttccattcg ccattcaggc 4440
tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga 4500
aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac 4560
gttgtaaaac gacggccagt gagcgcgcgt aatacgactc actatagggc gaattggagc 4620
tccaccgcgg ccgcccggtt tatcgttaat atggatcaat ttgaacagtt gattaacgtg 4680
tctctgctca agtctttgat caaaacgcaa atcgacgaaa atgtgtcgga caatatcaag 4740
tcgatgagcg aaaaactaaa aaggctagaa tacgacaatc tcacagacag cgttgagata 4800
tacggtattc acgacagcag gctgaataat aaaaaaatta gaaactatta tttaacccta 4860
gaaagataat catattgtga cgtacgttaa agataatcat gcgtaaaatt gacgcatgtg 4920
ttttatcggt ctgtatatcg aggtttattt attaatttga atagatatta agttttatta 4980
tatttacact tacatactaa taataaattc aacaaacaat ttatttatgt ttatttattt 5040
attaaaaaaa aacaaaaact caaaatttct tctataaagt aacaaaactt ttaaacattc 5100
tctcttttac aaaaataaac ttattttgta ctttaaaaac agtcatgttg tattataaaa 5160
taagtaatta gcttaactta tacataatag aaacaaatta tacttattag tcagtcagaa 5220
acaactttgg cacatatcaa tattatgctc tcgacaaata acttttttgc attttttgca 5280
cgatgcattt gcctttcgcc ttattttaga ggggcagtaa gtacagtaag tacgtttttt 5340
cattactggc tcttcagtac tgtcatctga tgtaccaggc acttcatttg gcaaaatatt 5400
agagatatta tcgcgcaaat atctcttcaa agtaggagct tctaaacgct tacgcataaa 5460
cgatgacgtc aggctcatgt aaaggtttct cataaatttt ttgcgacttt gaaccttttc 5520
tcccttgcta ctgacattat ggctgtatat aataaaagaa tttatgcagg caatgtttat 5580
cattccgtac aataatgcca taggccacct attcgtcttc ctactgcagg tcatcacaga 5640
acacatttgg tctagcgtgt ccactccgcc tttagtttga ttataataca taaccatttg 5700
cggtttaccg gtactttcgt tgatagaagc atcctcatca caagatgata ataagtatac 5760
catcttagct ggcttcggtt tatatgagac gagagtaagg ggtccgtcaa aacaaaacat 5820
cgatgttccc actggcctgg agcgactgtt tttcagtact tccggtatct cgcgtttgtt 5880
tgatcgcacg gttcccacaa tggttaattc gagctcgccc aaaccgggcg cgccgggcat 5940
ttaaatggaa ttcctttatc cagccctcac tccttctcta ggcgccggaa ttgaagatct 6000
gggggatcga tcctctagag tccgttacat aacttacggt aaatggcccg cctggctgac 6060
cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa 6120
tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag 6180
tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc 6240
ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct 6300
acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg 6360
gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt 6420
tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga 6480
cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct cgtttagtga 6540
accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga agacaccggg 6600
accgatccag cctccgcggc cgggaacggt gcattggaac ggacctgcag gaattgatcc 6660
gcggccgcac catggaacaa aaactcatct cagaagagga tctggcgggc aaggatcccg 6720
ggcagcagcc gccgcctcaa ccggcccccc agggccaagg gcagccgcct tcgcagcccc 6780
cgcaggggca gggcccgccg tccggacccg ggcaaccggc acccgcggcg acccaggcgg 6840
cgccgcaggc accccccgcc gggcatcaga tcgtgcacgt ccgcggggac gcggagaccg 6900
acctggaggc gctcttcaac gccgtcatga accccaagac ggccaacgtg ccccagaccg 6960
tgcccatgag gctccggaag ctgcccgact ccttcttcaa gccgccggag cccaaatccc 7020
actcccgaca ggccgctact gatgcaggca ctgcaggagc cctgactcca cagcatgttc 7080
gagctcatgc cgctccagct gctctgcagt tgggagctgt ttctcctggg acactgaccc 7140
ccactggagt agtctctggc ccagcagcta cacccacagc tcagcatctt cgacaggctg 7200
cttttgagat acctgatgat gtacctctgc cagcaggttg ggagatggca aagacatctt 7260
ctggtcagag atacttctta aatcacatcg atcagacaac aacatggcag gaccccagga 7320
aggccatgct gtcccagatg aacgtcacag cccccaccag tccaccagtg cagcagaata 7380
tgatgaactc ggcttcaggt cctcttcctg atggatggga acaagccatg actcaggatg 7440
gagaaattta ctatataaac cataagaaca agaccacctc ttggctagac ccaaggcttg 7500
accctcgttt tgccatgaac cagagaatca gtcagagtgc tccagtgaaa cagccaccac 7560
ccctggctcc ccagagccca cagggaggcg tcatgggtgg cagcaactcc aaccagcagc 7620
aacagatgcg actgcagcaa ctgcagatgg agaaggagag gctgcggctg aaacagcaag 7680
aactgcttcg gcaggagtta gccctgcgta gccagttacc aacactggag caggatggtg 7740
ggactcaaaa tccagtgtct tctcccggga tgtctcagga attgagaaca atgacgacca 7800
atagctcaga tcctttcctt aacagtggca cctatcactc tcgagatgag gctacagaca 7860
gtggactaag catgagcagc tacagtgtcc ctcgaacccc agatgacttc ctgaacagtg 7920
tggatgagat ggatacaggt gatactatca accaaagcac cctgccctca cagcagaacc 7980
gtttcccaga ctaccttgaa gccattcctg ggacaaatgt ggaccttgga acactggaag 8040
gagatggaat gaacatagaa ggagaggagc tgatgccaag tctgcaggaa gctttga 8097
Claims (1)
- Application of PRSS35 human recombinant protein in preparing medicine for treating liver cancer.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103097552A (en) * | 2010-07-14 | 2013-05-08 | 加利福尼亚大学董事会 | Biomarkers for diagnosis of transient ischemic attacks |
| CN104906110A (en) * | 2014-08-08 | 2015-09-16 | 遵义医学院附属医院 | Application of estrogen and its receptor stimulant in preparation of anti-hepatocellular carcinoma drugs |
| CN109628571A (en) * | 2018-12-10 | 2019-04-16 | 浙江省肿瘤医院 | Radiation pneumonitis incubation period sensitive gene quantitative detecting method |
| CN111154869A (en) * | 2018-11-08 | 2020-05-15 | 北京大学 | Biomarker for liver cancer diagnosis and kit thereof |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20110301095A1 (en) * | 2010-06-04 | 2011-12-08 | Sanford-Burnham Medical Research Institute California | Methods and compositions for assessing and treating metastasis, metastatic cancer, and potential for metastasis |
| US20150355181A1 (en) * | 2013-01-25 | 2015-12-10 | Beth Israel Deaconess Medical Center, Inc. | Methods and compositions for human epididymis protein-4 (he4) |
-
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- 2020-06-30 CN CN202010630380.2A patent/CN113862352B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103097552A (en) * | 2010-07-14 | 2013-05-08 | 加利福尼亚大学董事会 | Biomarkers for diagnosis of transient ischemic attacks |
| CN104906110A (en) * | 2014-08-08 | 2015-09-16 | 遵义医学院附属医院 | Application of estrogen and its receptor stimulant in preparation of anti-hepatocellular carcinoma drugs |
| CN111154869A (en) * | 2018-11-08 | 2020-05-15 | 北京大学 | Biomarker for liver cancer diagnosis and kit thereof |
| CN109628571A (en) * | 2018-12-10 | 2019-04-16 | 浙江省肿瘤医院 | Radiation pneumonitis incubation period sensitive gene quantitative detecting method |
Non-Patent Citations (2)
| Title |
|---|
| 必应.必应 PRSS35.《必应搜索》.2013,1. * |
| 毕锡麟等.牛卵泡颗粒细胞PRSS35表达模式及功能研究.《畜牧兽医学报》.2018,第49卷(第11期),摘要，第1.2节. * |
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