Anti-p53 Antibodies Research Articles

Paradoxical Reductions in Serum Anti-p53 Autoantibody Levels by Chemotherapy in Unresectable Colorectal Cancer: An Observational Study

Objective: The anti-p53 autoantibody is an emerging tumor marker that is commonly produced in response to p53 mutations. The usefulness of this antibody has been suggested in screening for and the monitoring of recurrence in colorectal cancer; however, its significance as a marker during chemotherapy remains largely unknown. Methods: We measured serum anti-p53 antibody levels in patients with unresectable colorectal cancer who underwent first-line systemic chemotherapy. Tumor responses were evaluated by computed tomography. We determined whether temporal changes in this antibody during therapy are associated with radiological responses. Results: Of the 83 patients in our study, 29 (35%) had elevated serum anti-p53 antibody levels before chemotherapy. Among these, antibody levels decreased in all 14 responders. In contrast, among those patients with elevated pretherapeutic serum anti-p53 antibody levels, 89% showed a paradoxical decrease in antibody levels and exhibited disease progression after chemotherapy. Moreover, serum anti-p53 antibody levels before and after chemotherapy were not associated with survival. Conclusion: These results suggest that serum anti-p53 antibody levels are of limited value in the evaluation of responses to palliative chemotherapy in patients with colorectal cancer.

P53 codon 72 polymorphism and its overexpression in patients with laryngeal carcinoma: Prognostic implications.

Abnormalities in the p53 gene are the most common genetic alterations seen in laryngeal carcinoma. No data exist regarding the association between laryngeal carcinoma and a distinct codon 72 variant and its expression. We conducted a prospective study (1) to analyze the p53 codon 72 polymorphic variants in patients with laryngeal carcinoma, (2) to analyze the expression of p53 mRNA in tissues of patients with laryngeal carcinoma using the reverse transcriptase-polymerase chain reaction (RT-PCR) assay, and (3) to detect p53 antibodies in the plasma of patients with laryngeal carcinoma before and after treatment. Tissue and blood samples were taken from 40 patients with laryngeal carcinoma-36 men and 4 women, aged 40 to 65 years (mean: 56)-and 20 age-matched controls with laryngeal conditions other than carcinoma. RT-PCR was used to measure p53 mRNA expression, and PCR-restriction fragment length polymorphism was used to determine p53 polymorphism. In addition, p53 antibodies were detected in plasma by Western blot testing. The 40 patients were treated with either surgery (total laryngectomy or conservation surgery) or radiotherapy. Tissue and blood samples were analyzed before treatment and 4 weeks after treatment. The findings were compared with those of the 20 controls. The results revealed that (1) homozygosity of the Pro72 variant of p53 was present in 26 laryngeal carcinoma patients (65%), (2) heterozygosity for the Pro/Arg genotype was present in 13 patients (32.5%), and (3) the Arg72 variant of the p53 allele was present in 1 patient (2.5%) before treatment. Overexpression of p53 mRNA was found in all patients with laryngeal carcinoma and in none of the controls before treatment; the difference was approximately 3.3 folds higher in the carcinoma group. However, p53 expression was not related to the biologic aggressiveness of these tumors. It is interesting that 4 weeks after definitive therapy, the expression levels of p53 mRNA in the 40 patients were comparable to those of the controls. The p53 antibodies were detected in the plasma of all patients with laryngeal carcinoma prior to definitive therapy and in none of them afterward, indicating that these antibodies represent a prognostic marker in laryngeal carcinoma. Our findings suggest that there is a correlation between p53 overexpression and the development of laryngeal carcinoma. Anti-p53 antibodies can be used as a prognostic marker in laryngeal carcinoma, and they can be exploited in the future to control the response to therapy and to monitor for certain early recurrences before they become clinically detectable.

Tumour-Associated Autoantibodies as Diagnostic Biomarkers for Breast Cancer: A Systematic Review and Meta-Analysis.

Tumour-associated autoantibodies may be promising biomarkers that could facilitate breast cancer (BC) diagnosis and improve patient outcomes. This review aims to identify the tumour-associated autoantibodies with the greatest diagnostic potential. Systematic searches were conducted using PubMed and Web of Science. The most studied tumour-associated autoantibody was included in a meta-analysis, and its clinical value was determined using Fagan's nomogram. The analysis included 84 studies regarding tumour-associated autoantibodies with the diagnostic value. Anti-p53 antibody was the most frequently studied autoantibody, followed by autoantibodies against MUC1, HER2 and cyclin B1. Although individual tumour-associated autoantibodies showed low diagnostic sensitivity, combinations of autoantibodies offered relatively high sensitivity. Enzyme-linked immunosorbent assay (ELISA) was the most common detection method, and nucleic acid programmable protein microarrays appeared preferable to common protein microarrays. As the most commonly studied autoantibody, anti-p53 antibody was included in a meta-analysis. When it had been detected using ELISA and cut-off values were defined as the mean +2 or 3 standard deviations, the summary area under the receiver operating characteristic curve for the presence of BC was 0.78. Fagan's nomogram showed post-test probabilities of 32% and 6% for positive and negative results, respectively. Mammography might be supplemented by the use of tumour-associated autoantibodies as biomarkers for BC diagnosis in younger women with increased risks of BC. Even though several studies have investigated the diagnostic use of tumour-associated autoantibodies as biomarkers for BC detection, a high-quality prospective study is needed to validate their diagnostic value in practice.

Clinicopathological and prognostic significance of programmed cell death ligand-1 expression in lung adenocarcinoma and its relationship with p53 status

IntroductionPD-L1 expression is a predictive biomarker for response to anti-programmed cell death-1 (PD-1)/programmed cell death-ligand 1 (PD-L1) immune checkpoint inhibitors and can be evaluated by immunohistochemistry. Results of the clinicopathologic characteristics of PD-L1-positive lung adenocarcinoma have been inconsistent in previous studies, and there are no reports on the relationship between PD-L1 expression and p53 status in lung adenocarcinoma. MethodsWe examined PD-L1 and p53 expression in a total of 323 surgically resected lung adenocarcinoma cases using anti-PD-L1 (clone SP142) and anti-p53 (clone DO-7) antibodies, and analyzed the clinicopathologic characteristics of PD-L1-positive cases and their relationship with p53 status. ResultsPD-L1 expression in tumor cells was positive in 60 of 323 cases (18.6%). Higher PD-L1 expression (≥50%) was more prevalent in former or current smokers (p=0.026) and was associated with more pack-years (p=0.016). PD-L1-positive tumors were significantly associated with solid predominant type (p<0.001), p53 aberrant expression (p<0.001), and PD-L1 expression in tumor-infiltrating immune cells (p<0.001). Patients with stage I to III tumors harboring PD-L1-positive tumor cells showed poor recurrence-free survival (p<0.001) and overall survival (p<0.001) on univariate analysis. ConclusionsPD-L1 expression in tumor cells, solid predominant histology, p53 aberrant expression, and PD-L1 expression in tumor-infiltrating immune cells are closely related. These variables should be considered when analyzing the clinical outcomes of patients with lung adenocarcinomas treated with anti-PD1/PD-L1 immune checkpoint inhibitors.

NAT10 regulates p53 activation through acetylating p53 at K120 and ubiquitinating Mdm2.

As a genome guardian, p53 maintains genome stability by arresting cells for damage repair or inducing cell apoptosis to eliminate the damaged cells in stress response. Several nucleolar proteins stabilize p53 by interfering Mdm2–p53 interaction upon cellular stress, while other mechanisms by which nucleolar proteins activate p53 remain to be determined. Here, we identify NAT10 as a novel regulator for p53 activation. NAT10 acetylates p53 at K120 and stabilizes p53 by counteracting Mdm2 action. In addition, NAT10 promotes Mdm2 degradation with its intrinsic E3 ligase activity. After DNA damage, NAT10 translocates to nucleoplasm and activates p53‐mediated cell cycle control and apoptosis. Finally, NAT10 inhibits cell proliferation and expression of NAT10 decreases in human colorectal carcinomas. Thus, our data demonstrate that NAT10 plays a critical role in p53 activation via acetylating p53 and counteracting Mdm2 action, providing a novel pathway by which nucleolar protein activates p53 as a cellular stress sensor.

Abstract A156: Anti-p53 auto-antibody serum profiling using high-density peptide arrays

Abstract Cancer is a result of a number of genetic alterations that disturb normal, controlled cell growth and differentiation. Mutational events leading to the activation of oncogenes or the inactivation of tumor-suppressor genes have been linked causally to the formation of tumors. p53 is one of the most important regulators of transcription, cellular cycle, DNA repair and apoptosis detected to date. Anti-p53 antibodies have been detected in the serum of cancer patients. This immune response is probably due to a self-immunization process linked to the strong immunogenicity of the p53 protein, and is associated predominantly with p53 missense mutation and p53 accumulation in the tumor. Auto-antibodies have also been proposed as potential diagnostic biomarkers for early stage diagnosis of cancers, since an increase in serum levels of certain auto-antibodies has been shown to precede the development of disease symptoms and correlate with cancer incidence for various cancers including breast and lung cancer. Here we systematically evaluate reactivity of antibodies in p53 positive serum samples and identify reactive epitopes to normal and mutant peptides using a high-density (2.9 Million) peptide microarray. We assess the effect of linker length, peptide length, and flanking serines on antibody detection. We propose peptide array design parameters that can be applied to a whole proteome level to enable biomarker discovery and validation of novel auto-antibody epitopes associated with cancer. Citation Format: Ken C. Lo, John C. Tan, Eric Sullivan, Ryan Bannen, Todd Richmond, Florian Grupp, Stefan Weiser, Dieter Heindl, Klaus-Peter Stengele, Albert Thomas. Anti-p53 auto-antibody serum profiling using high-density peptide arrays. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr A156.

The Correlation Between 1p/19q Codeletion, IDH1 Mutation, p53 Overexpression and Their Prognostic Roles in 41 Turkish Anaplastic Oligodendroglioma Patients.

To observe the correlation between 1p/19q codeletion, isocytrate dehydrogenase-1 (IDH1) mutation and p53 protein overexpression and their prognostic value in Turkish anaplastic oligodendroglioma patients who were treated with adjuvant radiotherapy and temozolomide chemotherapy. We retrospectively evaluated 41 patients who were diagnosed as anaplastic oligodendroglioma. Thirty-five patients received standard radiotherapy. Twenty-six patients received standard temozolomide chemoterapy concurrent to radiotherapy. Chromosome 1p/19q codeletion was observed in 19 of 41 patients (46%) via Fluorescent In Situ Hybridisation (FISH) technique. Twenty-six patients (63%) showed positive immunoreaction with anti-IDH1 antibody. Six patients (15%) showed positive immunoreaction with anti-p53 antibody. A statistically significant correlation was determined between chromosome 1p/19q codeletion and IDH1 mutation (p < 0.0001). The patients who had tumors with chromosome 1p/19q codeletion and p53 overexpression were mutually exclusive. The mean estimated Progression Free Survival (PFS) of the patients who had tumors with chromosome 1p/19q codeletion and/or IDH1 mutation was determined to be significantly longer than that of the patients without these genetic changes, regardless of the treatment modality (p=0.006, p=0.004). PFS of the patients who received adjuvant chemotherapy and whose tumors had chromosome 1p/19q codeletion or IDH1 mutation was significantly longer than that of the patients without these genetic changes (p=0.001, p < 0.0001). Chromosome 1p/19q codeletion and/or IDH1 mutation are favorable prognostic factors in anaplastic oligodendroglioma patients, in terms of PFS.

P16 protein is upregulated in a stepwise fashion in colorectal adenoma and colorectal carcinoma.

Background/Aims:p16 is tumor suppressor gene acting as a cell cycle regulator. The present study was conducted to compare p16 expression in normal, dysplastic, and malignant colonic mucosae, and to explore its relation to clinicopathological variables and follow-up data in colorectal carcinoma (CRC).Patients and Methods:Tissue microarrays were performed from 25 normal colonic mucosae, 41 colonic adenomas, and 191 CRC, with corresponding 50 nodal metastases. Immunohistochemistry was performed using anti-p16 antibody, sections were scored, and statistical analysis was performed. K-ras mutation detection was also performed.Results:Immunoexpression of p16 was significantly higher in CRC than in adenomas (P = 0.033) and normal colonic mucosa (P = 0.005). There was no statistically significant difference between p16 expression in CRC and nodal metastasis. There was no significant association between p16 immunoexpression in CRC and all clinicopathological data and survival probability. K-ras mutations were detected in 34% of CRC. However, there was no correlation between K-ras status and p16 expression (P = 0.325).Conclusion:Absence of p16 expression is correlated to a benign course of CRC adenomas. p16 has a key role in CRC progression and can be used as a marker for colorectal adenoma. On the other hand, it has no role as a predictive and/or prognostic factor in CRC. Further extended studies are required to explore the role of p16 as indicator of premalignant lesions in the colon and to test its relation with CRC histological grade, as well as to test its value as a new therapeutic target.

HPV genotyping and p16 expression in Xingu Indigenous Park, Brazil.

The association between high-risk human papillomavirus (HPV) genotypes and p16 expression in indigenous women from the Xingu Indigenous Park, Brazil, was unknown. This study evaluated p16 expression in women with a histological diagnosis of cervical intraepithelial neoplasia (CIN) 3 or higher and correlated this expression with HPV genotypes to determine possible discrepancies in the expression of this marker. We evaluated 37 previously collected samples with different HPV genotypes and high-grade lesions diagnosed based on cytology, histology, and colposcopy. Immunohistochemical analysis was performed using paraffin-embedded tissue sections and the CINtec® Histology Kit. p16 protein expression was investigated by immunostaining with an anti-p16 antibody. HPV genotyping was performed by reverse hybridization. The age of the study population ranged from 22-75 years (43.81 ± 15.89 years) and parity ranged from 1-11 (5.92 ± 2.58). Thirteen different HPV genotypes were found using the INNO-LiPA kit. Single and multiple infections by HPV were found with prevalence of single infections (P = 0.029). Comparison between HPV genotype and simple or multiple infections was highly significant; it was observed more HPV 52 followed by HPV 16 in single infections (P < 0.001). p16 expression was predominantly diffuse, which was observed in 91.7% of lesions, whereas 8.3% were focal (P < 0.001). HPV 52, HPV 16 and 31 were the most prevalent HPV types in high-grade CIN in these indigenous women. Diffuse p16 expression in high-grade CIN was not influenced by the viral genotype; however, more studies are necessary to further our understanding of this restricted group.

Evaluation of microvessel density and p53 expression in pancreatic adenocarcinoma

OBJECTIVE:To evaluate the prognostic significance of microvessel density and p53 expression in pancreatic cancer.METHODS:Between 2008 and 2012, 49 patients with pancreatic adenocarcinoma underwent resection with curative intention. The resected specimens were immunohistochemically stained with anti-p53 and anti-CD34 antibodies. Microvessel density was assessed by counting vessels within ten areas of each tumoral section a highpower microscope.RESULTS:The microvessel density ranged from 21.2 to 54.2 vessels/mm2. Positive nuclear staining for p53 was found in 20 patients (40.6%). The overall median survival rate after resection was 24.1 months and there were no differences in survival rates related to microvessel density or p53 positivity. Microvessel density was associated with tumor diameter greater than 3.0 cm and with R0 resection failure.CONCLUSIONS:Microvessel density was associated with R1 resection and with larger tumors. p53 expression was not correlated with intratumoral microvessel density in pancreatic adenocarcinoma.

Detection of p53 Mutations in Circulating DNA of Transplanted Hepatocellular Carcinoma Patients as a Biomarker of Tumor Recurrence.

p53 is the most commonly mutated gene in malignant human cancers. To detect p53 mutations in circulating DNA (cirDNA) of transplanted hepatocellular carcinoma (HCC) patients could be an interesting approach to know of any tumor recurrence. In this study, our objective was to determine the utility of this method in the diagnosis and the prognosis of HCC tumor recurrence.Twenty four liver transplanted HCC patients were included in the study together with a group of healthy controls. Detection of the specific p53 mutation in cirDNA was performed by high-resolution melting PCR (HRM-PCR) and COLD-PCR immediately before the transplantation. Serum anti-p53 was also determined using a p53-autoantibody ELISA kit.The results of the HRM-PCR and COLD-PCR showed two well-differentiated groups of transplanted patients after normalization by healthy controls. These data allow us to distinguish between patients with p53 mutated cirDNA and those with wild type cirDNA. Moreover, we have found that most of p53 mutated patients also presented elevated anti-p53 antibodies. The present results indicate that it is possible to detect mutated p53 genes with the cirDNA and that this could be used as a biomarker of tumor recurrence during the clinical evolution of the transplanted patients.

Enhanced antigen detection in immunohistochemical staining using a 'digitized' chimeric antibody.

The immunohistochemical (IHC) staining of mouse tissue sections using antibodies of mouse origin can result in high nonspecific background due to the staining of endogenous immunoglobulins (Igs) by enzyme-conjugated secondary antibodies. In order to obviate this issue, we developed a chimeric mouse-human anti-p53 monoclonal antibody (MH242) by grafting the variable regions of a known mouse antibody into a human Ig scaffold. This facilitated use of an anti-human secondary antibody, and resulted in near-zero background when compared with its parental mouse monoclonal antibody (PAb242). Furthermore, the chimeric antibody enabled reproducible detection of mutant p53 (homozygous R172H) expression in mouse tissue, an observation hitherto largely equivocal based on the use of existing antibodies. The approach we describe leads to the generation of tractable antibody reagents, whose integrity can be readily verified through DNA sequencing of expressor plasmids. The wide-spread adoption of such 'digitized' antibodies should reduce experimental disparities that can commonly arise through variations in antibody quality.

Does anti-p53 antibody status predict for clinical outcomes in metastatic colorectal cancer patients treated with fluoropyrimidine, oxaliplatin, plus bevacizumab as first-line chemotherapy?

BackgroundTP53 gene mutation is widely known as one of the determinants of impaired chemosensitivity. p53 is a tumor-suppressor protein in humans encoded by the TP53 gene. Some studies have shown that TP53 gene mutation and accumulation of the p53 protein are closely related with serum anti-p53 antibody positivity. This study aimed to evaluate the predictive significance of the serum p53 antibody status in metastatic colorectal cancer (mCRC) patients treated with fluoropyrimidine, oxaliplatin, plus bevacizumab as first-line chemotherapy.MethodsNinety patients treated with fluoropyrimidine, oxaliplatin plus bevacizumab as first-line chemotherapy were enrolled, including 70 whose KRAS genotype was revealed at the beginning of treatment. Before chemotherapy initiation, the serum p53 antibody level was quantified by enzyme-linked immunosorbent assay using MESACUP® anti-p53 test kits. The cutoff value for positivity was 1.3 U/mL, as calculated previously. The KRAS genotype of the tumor samples was analyzed using the Luminex® assay.ResultsOverall response rates of Response Evaluation Criteria in Solid Tumors criteria were 77.7 % (42/54) in anti-p53–negative patients and 69.4 % (25/36) in anti-p53–positive patients. The odds ratio was 1.07. Median overall survival was 36.1 months in the anti-p53–positive patients, and not available in the anti-p53–negative patients (hazard ratio, 0.81; 95 % confidence interval, 0.37–1.77; P = 0.61). The corresponding values for median progression-free survival were 13.3 months and 14.6 months (hazard ratio, 0.69; 95 % confidence interval, 0.41–1.17; P = 0.17), respectively.ConclusionsSerum anti-p53 antibody positivity did not predict chemoresistance in mCRC treated with fluoropyrimidine, oxaliplatin, plus bevacizumab as first-line chemotherapy.

Nontoxic virus nanofibers improve the detection sensitivity for the anti-p53 antibody, a biomarker in cancer patients.

The presence of anti-p53 antibody in serum is a biomarker for cancer. However, its high sensitivity detection is still an issue in cancer diagnosis. To tackle this challenge, we used fd phage, a human-safe bacteria-specific virus nanofiber that can be mass-produced by infecting host bacteria in an error-free manner, and genetically engineered it to display a peptide capable of recognizing and capturing anti-p53 antibody on its side wall. We employed the resultant phage nanofibers as a capture probe to develop a modified version of the enzyme-linked immunosorbent assay (ELISA) method, termed phage-ELISA. We compared it to the traditional ELISA method for the detection of anti-p53 antibody, p53-ELISA, which uses recombinant wild-type p53 protein to capture anti-p53 antibody. We applied phage-ELISA to detect anti-p53 antibody in an experimental group of 316 patients with various types of malignant tumors. We found that a detection rate of 17.7% (56 positive cases) was achieved by phage-ELISA, which was comparable to the detection rate of 20.6% for p53-ELISA (65 positive cases). However, when both phage and p53 were combined to form antibody-capturing probes for phage/p53-ELISA, a detection rate of 30.4% (96 positive cases) was achieved. Our work showed that owing to the combined capture of the anti-p53 antibody by both phage nanofibers and p53, the phage/p53-ELISA achieved the highest diagnostic accuracy and detection efficiency for the anti-p53 antibody in patients with various types of cancers. Our work suggests that a combination of nanofibers and antigens, both of which capture antibody, could lead to increased detection sensitivity, which is useful for applications in the life sciences, clinical medicine, and environmental sciences.

Abstract 544: Lung cancer classification using new immunohistochemical assay with anti-p40 (BC28) mouse monoclonal antibody: Comparison with the p40 Echelon assay and anti-p63 (4A4) antibody

Abstract Background: The precise differentiation of lung adenocarcinoma (ADC) and lung squamous cell carcinoma (SCC) is essential for the determination of appropriate cancer therapy. Currently, the anti-p63 (4A4) antibody is the most frequently used marker for the lung SCC classification. A major limitation of this antibody is low specificity due to its reactivity in a substantial proportion of lung ADCs. The p63 antibody recognizes two p63 isoforms - 1) long TAp63 isoform and 2) a truncated variant designated as p40. In contrast to the anti-p63 antibody, the anti-p40 (BC28) antibody recognizes only the p40 isoform. The goal of this study was to compare a newly optimized IHC assay using the anti-p40 (BC28) antibody and OptiView IHC DAB Detection Kit with 1) the commercially available p40 Echelon assay, and with 2) an IHC assay using the anti-p63 (4A4) antibody in various lung tumors. Methods: Lung tumors from 271 patients were evaluated for p40 and p63 expression using IHC on VENTANA BenchMark XT platform with OptiView detection. The p40 expression was also evaluated by the commercially available anti-p40 (BC28) Echelon assay (Biocare Medical Inc.) using the manufacturer's recommended protocol. After the initial classification of these tumors by H&amp;E, anti-Napsin A (MRQ-60), anti-Cytokeratin 5/6 (D5/16B4) or anti-TTF-1 (SP141) antibodies were used for the evaluation of the discordant cases when H&amp;E and p40/p63 status did not correlate. Results: The anti-p40 and anti-p63 antibodies showed nuclear staining with no or minimal background. After resolving discordant cases, the new optimized Ventana assay using anti-p40 (BC28) antibody demonstrated superior sensitivity to the p40 Echelon assay in SCC classification. 92.9% of cases (52/56) showed p40 positive staining with the new p40 assay compared to 73.2% SCC cases (41/56) stained with p40 Echelon assay. In addition, the new p40 IHC assay demonstrated increased specificity compared to the anti-p63 antibody for ADC classification. 7.7% lung ADCs (13/169) were positive with anti-p63 antibody but only 3.0% (5/169) ADCs were positive with anti-p40 antibody. The new p40 assay correctly classified 96.0% SCC and ADC cases while 91.2% and 93.8% SCC and ADC case were correctly classified with p40 Echelon assay and anti-p63 antibody, respectively. All of the other lung tumors were negative for p40 and p63 except one bronchioloalveolar carcinoma case (1/14) and 6 adenosquamous carcinoma cases (6/13). Conclusion: In summary, the optimized p40 assay shows an improved sensitivity over the p40 Echelon assay, while both these p40 assays demonstrate a higher specificity compared to IHC assay using anti-p63 (4A4) antibody. Citation Format: Katerina Dvorak, Christian Roessler, John Palting, Akira Moh. Lung cancer classification using new immunohistochemical assay with anti-p40 (BC28) mouse monoclonal antibody: Comparison with the p40 Echelon assay and anti-p63 (4A4) antibody. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 544. doi:10.1158/1538-7445.AM2015-544

Characteristics of keratinocytes in facial solar lentigo with flattened rete ridges: comparison with melasma.

Flattened rete ridges occurring on the face are not uncommon in solar lentigo (SL). To investigate the morphological changes of keratinocytes in facial SL with flattened rete ridges and melasma. In total, 25 patients with facial SL showing flattened rete ridges and 20 patients with melasma, in which rete ridge flattening is also a common feature, were included in the study. Skin biopsies were performed on lesional skin and perilesional normal skin. The histological findings showed that the epidermis was significantly thicker in SL skin with flattened rete ridges compared with perilesional normal skin. Comparative quantitative analysis of epidermal morphology revealed that the individual keratinocytes were larger in size in lesional skin, without a significant change in the number of cells. Anti-p16 antibody staining was more intense in lesional epidermis, suggesting senescence of keratinocytes in the thickened epidermis. By contrast, melasma samples showed no significant difference in epidermal thickness or keratinocyte morphology in terms of number or size compared with normal skin. These findings suggest that senescent changes in keratinocytes are important in the development of SL, even in the absence of rete ridge elongation, and the removal of keratinocytes harbouring melanin could be a possible strategy for SL treatment.

Diagnostic relevance of autoantibody detection against inhibitors of apoptosis proteins in colon cancer and colon adenoma.

Autoantibodies against cancer-related antigens may be detected in the sera of patients with various types of cancer, although their clinical utility has not yet been established. In this study, we aimed to demonstrate the diagnostic relevance of autoantibody detection against inhibitor of apoptosis protein (IAP) family members in colon cancer, as compared to anti-p53 antibody, carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9. We established an ELISA system using original recombinant proteins of IAP family members (survivin, livin and X-linked IAP) and measured the expression levels in the sera of 62 healthy donors, 250 patients with colon polyps (adenoma) and 176 patients with colon cancer. When the cutoff value was set as the mean value + 2 standard deviations in healthy donors, anti-survivin exhibited the highest positivity rate (24.4%) among IAP autoantibodies in cancer patients. Furthermore, the anti-survivin antibody exhibited a high positivity rate in early-stage carcinoma and adenoma. In the combination assay, reflecting the significantly high positivity rate of CEA in stage IV tumors, the positivity rate was highest when combining the detection of anti-survivin antibody and CEA in cancer patients (50.0%), indicating that this combination may not be useful for the diagnosis of early-stage cancers. By contrast, reflecting the complete independencE of anti-survivin and anti-p53 antibodies, the combination of detecting these two antibodies resulted in the highest positivity rate (35.6%) in early-stage disease (stage 0-I). These results suggest that the combined measurement of anti-survivin and anti-p53 antibodies may be useful for the detection of early-stage colon cancer.

Circulating antibodies to p16 protein-derived peptides in breast cancer.

Overexpression of the p16 protein has been reported in breast cancer and may trigger the secretion of antibodies against itself. Circulating anti-p16 antibodies that were detected with a recombinant protein have been reported in breast cancer. The present study was designed to determine whether the levels of circulating IgG antibody to p16 protein-derived linear antigens are altered in breast cancer. An enzyme-linked immunosorbent assay (ELISA) was developed in-house to determine circulating IgG against peptide antigens derived from the p16 protein in 152 female breast cancer patients and 160 healthy female subjects. The Student's T-test revealed that breast cancer patients exhibited significantly higher levels of anti-p16 IgG antibody compared to control subjects (T=2.02, P=0.045). In addition, ductal cancer appeared to be the main type contributing to the increased levels of circulating anti-p16 antibodies (T=2.08, P=0.038). Of all four stages of breast cancer, stage I was associated with the highest levels of IgG antibody (T=2.02, P=0.045) and receiver operating characteristic (ROC) analysis demonstrated that the area under the ROC curve was 0.74 (95% confidence interval: 0.65-083) and that the sensitivity against a specificity of 90% was 30.3%. Therefore, the levels of circulating IgG antibody to the p16 protein may be a potential biomarker for early diagnosis of breast cancer.

Identification of the p53-Responsive Element in the Promoter Region of the Human Decorin Gene

We have obtained some evidence that shows that the decorin gene is the target of p53 transactivation. Luciferase reporter plasmid, which contained the promoter region between positions -252 and -205, was activated by p53 dose- dependently up to 170-fold. The promoter region involved a sequence, 5’-AGGCAAGTAG-3’, similar to p53-binding consensus sequence, 5’-PuPuPuC(A/T)(A/T)GPyPyPy-3’. Chromatin immunoprecipitation assay using p53 antibodies revealed that the region between -413 and -232 of the promoter of the decorin gene was co-precipitated with p53. p53- binding to this region was further demonstrated by electrophoretic mobility shift assay, in which the complex between decorin promoter DNA and proteins decreased by pretreatment with anti-p53 antibodies. The mRNA expression levels of decorin increased after treatment with p53-activating nutlin-3 greatly and with genotoxic reagent, adriamycin, to some extent. Consequently, decorin promoter is useful to evaluate the p53 transactivation ability.

A case of odontogenic myxoma in a cow

Tumours of odontogenic origin are rare in domestic animals. A case of odontogenic myxoma was recorded in 12 years old cow involving the periodontal region of the left mandible. Tumorous growth was slow growing but progressively increased, locally invasive and recurred after surgical removal. Grossly, the growth was soft and slimy. Histopathologically, it revealed myxoma characterized by large, stellate or triangular fibroblast like cells with cytoplasmic processes, vesicular nuclei and small nucleoli. Mitotic figures were rarely observed. Expression of anti-pancytokeratin and anti-p53 antibodies by immunohistochemistry was not detected. Based on anatomical location, radiographic, gross and histopathological examination, this tumour was diagnosed as an odontogenic myxoma.