Application Of Peptide Microarray Based Serum Antibody Detection In The Diagnosis Of Prostate Cancer Patients

The experiment consists of the following two parts:Part?: Mapping of antibody spectrum against HPV-16 in serum of patients with prostate cancerObjective:Mapping of antibody spectrum against HPV-16 in serum of patients with prostate cancer.Methods:Seven functional proteins of HPV-16(HPV-L1,HPV-L2,HPV-E1,HPV-E2,HPV-E4,HPV-E6 and HPV-E7)were imbricated into peptide sequences(with the length of 20 aa and the repetition of 10aa)and synthesized 233 peptides.The synthesized peptides were immobilized on the i PDMS membrane to prepare peptide microarrays.Serological screening of 212 serum samples from patients with clinically and pathologically diagnosed prostate cancer and 229 serum samples from healthy persons was performed by using the peptide microarrays containing these HPV peptides.The antibody spectrum of HPV-16 protein in serum of patients with prostate cancer was drawn by serological screening results.Results:The 233 peptides of HPV-16 proteins can respond to both prostate cancer serum and healthy human serum,and the response to prostate cancer serum is significantly stronger.The results indicated that there are more specific antibodies against the HPV-16 peptides in prostate cancer.According to the response distribution of prostate cancer serum to the seven functional proteins,there was no strong response to HPV-E5 protein polypeptide in prostate cancer serum and healthy human serum.Among other proteins,the response coverage of multiple peptides to prostate cancer serum was more than 10%,while the response coverage of a few peptides to healthy human serum was also higher than 10%,such as HPV-206,suggests that the serum of healthy people may also have a strong response to HPV-16 peptides.Conclusions:The response of HPV-16 peptides in serum of prostate cancer and healthy people may be the specific response of antibody against its specific epitope,or the molecular mimicry,mimic epitope and nonspecific-interaction between antibody and peptide in serum.Therefore,healthy human serum can also respond to HPV-16 peptides,even response strongly to some peptides.At the same time,prostate cancer serum has stronger correlation and specificity to HPV-16 peptides than healthy samples,so its response to 233 HPV peptides is significantly stronger than that of healthy samples to these peptides.The presence of specific antibodies against seven functional proteins of HPV-16 in the serum of prostate cancer indicates that HPV-16 infection is associated with prostate cancer,and its correlation can be detected by serology.Part ?: Application of HPV-16 protein peptide in the diagnosis of prostate cancerObjective:To screen the specific antigen epitopes of prostate cancer in seven functional proteins of HPV-16,and to explore the application of antigen epitopes of HPV-16 in diagnosis of prostate cancer.Methods:The epitopes of seven functional proteins of HPV-16 were predicted by IEDB.After serological screening of HPV polypeptide in 212 serum samples of prostate cancer and 229 serum samples of healthy people,the highly responsive peptides with statistical difference in prostate cancer was obtained by SAM analysis,and its epitope sequence was obtained by sequence analysis.The combination analysis was used to identify the peptides combination for the diagnosis of prostate cancer,and ROC curve analysis was used to obtain the optimum positive judgment threshold for the diagnosis of prostate cancer,so that the combination of polypeptide combinations could obtain the optimum sensitivity and specificity for the diagnosis of prostate cancer.Results:By IEDB analysis,six proteins were predicted to have multiple potential epitopesequences.No epitope sequences were predicted on HPV-E5.SAM analysis revealed 57 highly expressed peptide in prostate cancer samples.The epitope sequences of 57 peptides were identified by sequence analysis.Sequence analysis showed that there were 11 epitope sequences in HPV-L1 protein,7 epitope sequences in HPV-L2 protein,10 epitope sequences in HPV-E1 protein,6 epitope sequences in HPV-E2 protein,2 epitope sequences in HPV-E4,HPV-E6 and HPV-E7 protein,and no epitope sequences in HPV-E5 protein.Comparing the predicted epitope sequence with the predicted epitope sequence,we found that the predicted epitope sequence was different from the predicted epitope sequence(for example,the identified epitope sequence of several proteins was inaccurate and different from the predicted epitope sequence,and many predicted epitopes could not be identified in the experiment),but most of them were consistent.A combination of thirteen peptides were identified by combination analysis.ROC analysis showed that the best threshold of counts of serum response to the peptides combination was 2.When the response of serum to any two of the 13 peptides was positive,the serum was judged to be prostate cancer.Under the positive judgment threshold of count ? 2,we got a combination of sensitivity(80.2%)and specificity(88.2%)for the diagnosis of prostate cancer.Conclusions:The antigenic epitope sequences of the seven functional proteins of HPV-16 obtained by the experiment were well consistent with those predicted by IEDB,which indicated that the corresponding epitope sequences might be real,and also showed the correctness and credibility of the experimental results.Specific antibodies against six functional proteins of HPV-16(except HPV-E5)in the serum of prostate cancer patients can be detected by HPV-16 peptide microarray.Specific antibodies in blood can be used as a promising biomarker for diagnosis of cancer.The results of this study also show that peptides from HPV-16 proteins has potential prospects and clinical value for diagnosis of prostate cancer.Its further clinical application and promotion need to be further explored.