Association of human papillomavirus type 16 E7 and HLA class I antigen expression in cervical premalignant and malignant lesions

In the present experiment we studied the correlation between HPV16 infection and expression of HLA-I antigen in cervical premalignant and malignant lesions (cervicitis, CIN, cervical squamous carcinomas and adenocarcinoma samples). The HPV16 E7 DNA load and the expression of HLA-I antigen in the samples were measured by real-time fluorescence quantitative polymerase chain reaction (RFQ-PCR) and immunohistochemical S-P staining, respectively. Our data indicate that HPV16 E7 load was highly and positively associated with the development of cervical lesions (Spearman's correlation coefficient r=0.848, p<0.001), the negative rate of HLA-I antigen was significantly distinguished among groups (p<0.001), and HPV16 E7 infection and downregulation of HLA-I antigen were highly correlated in cervical lesions (Pearson's correlation coefficient r=-0.487, p<0.001). HPV16 E7 may play an important role in the downregulation of HLA-I antigen in cervical lesions, which results in the immune escape of the virus and the occurrence, development, invasion and metastasis of cancer. Furthermore, quantitative PCR for HPV16 E7 may play an important role in the early detection of cervical diseases and in guiding future therapy toward prevention.     

Analysis of the physical state of different human papillomavirus DNAs in intraepithelial and invasive cervical neoplasm.

The integration of human papillomavirus (HPV) DNA into the human genome has been generally accepted as a characteristic of malignant lesions. To gain a better understanding of this phenomenon, genomic DNA from 181 cervical biopsy specimens was isolated and analyzed for HPV type and physical state of the HPV genome. These specimens represented the full spectrum of cervical disease, from condyloma to invasive carcinoma. Discrimination between integrated and episomal HPV DNA was accomplished by the detection of HPV-human DNA junction fragments on Southern blots. In most cases in which ambiguous Southern blot results were obtained, the specimens were reanalyzed by two-dimensional gel electrophoresis. Of the 100 biopsy specimens of cervical intraepithelial neoplasia analyzed, only 3 showed integrated HPV DNA, in contrast to 56 (81%) of 69 cervical carcinomas (P less than 0.001) showing integrated HPV DNA. Of the 40 carcinomas containing HPV 16 DNA, 29 (72%) had integrated HPV DNA, of which 8 (20%) also had episomal HPV DNA. In 11 (27%) cancers, only episomal HPV 16 DNA was detected. All 23 HPV 18-containing carcinomas had integrated HPV DNA, and 1 also had episomal HPV 18 DNA. The difference between HPV types 16 and 18 with respect to frequency of integration was statistically significant (P less than 0.01). The results of this study indicate that detectable integration of HPV DNA, regardless of type, occurs infrequently in cervical intraepithelial neoplasia. The absence of HPV 16 DNA integration in some carcinomas implies that integration is not always required for malignant progression. In contrast, the consistent integration of HPV 18 DNA in all cervical cancers examined may be related to its greater transforming efficiency in vitro and its reported clinical association with more aggressive cervical cancers.     

Study of infection by human papillomavirus in severe dysplasias and carcinomas in situ of the uterine cervix using immunohistochemistry and in situ hybridization.

Diagnosis of human papillomavirus (HPV) infection in uterine cervical lesions is usually based on histopathological criteria and, in some cases, is confirmed by immunohistochemistry. The recent development of in situ hybridization techniques has facilitated the detection of HPV in these lesions. Consequently, we carried out a study on 18 uterine cervical biopsy specimens histopathologically diagnosed as severe dysplasias and carcinomas in situ, using an immunohistochemical method with a rabbit polyclonal antibody against the HPV common structural antigen and in situ hybridization techniques with three biotinylated DNA probes for HPV types 6/11, 16/18, and 31/35/51. By immunohistochemistry only one case (5.5%) proved to be positive, whereas by in situ hybridization 12 HPV-positive cases were obtained (66.6%), of which 7 were positive for HPV types 16/18 (38.8%) and 6 for HPV types 31/35/51 (33.3%). One case was positive with positive with both DNA probes. From our results it can be inferred that in situ hybridization is a more sensitive technique than immunohistochemistry for confirming the presence of HPV in severe dysplasias and carcinomas in situ of the uterine cervix. Furthermore, in situ hybridization provides much more information than immunohistochemistry since it permits the identification of the HPV types causing the lesion.     

Causal association between human papilloma virus infection and head and neck and esophageal squamous cell carcinoma

HPVs commonly cause proliferative lesions of squamous epithelium, and infection with certain HPV types carries a high risk of malignant transformation. We used molecular techniques to detect and type HPV in papillomas and carcinomas in the oral cavity and esophagus. DNA was extracted from 150 fresh or paraffin embedded biopsy specimens, and analyzed for HPV by PCR with 15 sets of consensus primers directed to conserved regions of L1 gene, three sets of HPV16E6 primers (specific for the HPV 16 prototype and L83V variant), and sets of primers specific for the E6 gene of other mucosa type HPVs including HPV 6, 11, 16, 18, 52, 58, 66 and 73. Overall, HPV sequences were detected in 61 of 150 specimens. HPV DNA sequences were detected in 16/32 specimens in the oropharyngeal region, in 13/36 specimens in larynx and 32/82 specimens in esophagus. Papillomas contained only the episomal form of HPV 16. In the esophagus, the most common type was HPV 73. In all specimens examined, HPV 6/11 (4/150), HPV 16 (23/150), HPV 35 (1/150), HPV 45 (1/150), HPV 54 (1/150), HPV 58 (1/150), HPV 61 (1/150), HPV 66 (1/150), HPV 68 (2/150), HPV 70 (3/150), HPV 72 (1/150), HPV 73 (16/150), double HPV infection (2/150), and unidentified HPV type (4/150) was detected. Interestingly, HPV was found in all verrucous carcinomas and in 18/22 basaloid squamous cell carcinomas. HPV16E6 T350G mutant were observed only in two of eight carcinomas. Using correspondence analysis, a segregation of specific virus types in specific clinico-pathologic lesions (verrucous carcinoma and basaloid squamous cell carcinoma) was proved. It was shown that the relative rates of the HPV positive tumors were significantly higher in women than in men. The synergic action of mucosal irritation and HPV infection may be necessary for the development of the papillomas and the specific types of carcinomas in the oral cavity and in the esophagus.     

Detection of human papillomavirus in non-small cell lung carcinoma by polymerase chain reaction

Human papillomavirus (HPV) infection is one of the risk factors contributing to the pathogenesis of lung cancer. The aim of the study was to determine the presence of HPV in non-small cell carcinomas of the lung. The study included 40 tumors: 22 squamous cell carcinomas, 13 adenocarcinomas and 5 large cell carcinomas. HPV was found in 4 cases (10%). High risk HPV was present in 3 tumors: in one squamous cell carcinoma, one large cell carcinoma and one adenocarcinoma, while low risk HPV was detected in one adenocarcinoma.     

Head and neck squamous cell carcinoma: role of the human papillomavirus in tumour progression

High risk human papilloma viruses (HPVs) have been shown to be independent risk factors for anogenital tract cancers, and have also been detected in head and neck squamous cell carcinomas (HNSCC). The aim of our study was to determine the prevalence of HPV DNA in a group of 47 squamous cell carcinomas of the oropharynx and the oral cavity, and to compare the clinical behaviour of HPV positive and negative tumours. We also assessed the proliferation index, as evaluated by Ki67 immunohistochemistry positivity, and the level of p53 reactivity. HPV DNA was found in 50% of carcinomas of the oropharynx and 36% in those of the oral cavity, the only genotype detected being HPV 16. Patients with HPV-positive carcinomas had a better overall survival than those with HPV-negative carcinomas. Our data suggest that HPV-positive oropharyngeal cancers comprise a distinct disease entity with an improved prognosis.     

利用重组HPV16L1抗原检测宫颈癌抗L1或VLP抗体的对比

为了评价重组大肠杆菌表达的HPV16L1蛋白和重组腺病毒表达的HPV16L1 VLP两种抗原在检测宫颈癌抗 16L1或VLP抗体及在宫颈癌血清学诊断意义上的差别 ,应用PCR技术从宫颈癌组织的DNA中扩增出全长15 35bp的HPV16L1基因片段 ,克隆至 pUC18 T载体中 ,进行DNA测序鉴定。然后 ,将HPV16L1基因克隆至pGEX 2T表达载体中 ,并诱导表达HPV16L1融合蛋白 ,分子量为 83kD ,能被HPV16L1单克隆抗体所识别。经GST柱层析法纯化后 ,与重组腺病毒表达的HPV16L1 VLP分别经酶联免疫吸附 (ELISA)法检测 12份宫颈癌患者和 35份献血员血清。 12例宫颈癌血清标本中 ,抗HPV16L1蛋白的抗体阳性率为 7例 (占 5 8.3% ) ;抗HPV16L1 VLP的抗体阳性率为 8例 (占 6 6 .7% )。经大肠杆菌表达的重组抗原HPV16L1检测为HPV16抗体IgG( )的 7份患者血清 ,利用HPV16L1 VLP试剂盒检测均阳性 ;经大肠杆菌表达的重组抗原检测为HPV16抗体IgG( )的 5份患者血清 ,利用HPV16L1 VLP试剂盒检测有 1份阳性。两者对HPV16抗体的阳性检出率并无显著差异 (P >0 .0 5 )。本实验结果说明HPV16与宫颈癌高度相关 ,利用大肠杆菌表达的重组抗原HPV16L1和HPV16L1 VLP重组抗原检测抗体的敏感性并不受影响。利用重组抗原HPV16L1对宫颈癌的抗体进行定性、定量分析有助于该疾病     

Computer-assisted analysis of p53 and PCNA expression in oral lesions infected with human papillomavirus

OBJECTIVE: To carry out a retrospective study to determine whether human papillomavirus (HPV) infection and immunohistochemical expression of p53 and proliferating cell nuclear antigen (PCNA) are related to the risk of oral cancer. STUDY DESIGN: Fifty-seven oral biopsies, consisting of 30 oral squamous papillomas (OSPs) and 27 oral squamous cell carcinomas (OSCCs) were tested for the presence of HPV 6/11 and 16/18 by in situ hybridization using catalyzed signal amplification and in situ hybridization. p53 And PCNA expression was analyzed by immunohistochemistry and evaluated quantitatively by image analysis. RESULTS: Nineteen of the 57 oral lesions (33.3%) were positive for HPV. HPV 6/11 was found in 6 of 30 (20%) OSPs and 1 of 27 (3.7%) OSCCs. HPV 16/18 was found in 10 of 27 (37%) OSCCs and 2 of 30 (6.7%) OSPs. Sixteen of the 19 HPV-positive cases (84.2%) were p53 negative; 5 (9%) were HPV 6/11 and 11 (19%) HPV 16/18, with an inverse correlation between the presence of HPV DNA and p53 expression (P = .017, P < .05). PCNA expression appeared in 18 (94.7%) of HPV positive cases, showing that HPV 16/18 was associated with intensity of PCNA expression and with OSCCs (P = .037, P < .05). CONCLUSION: Quantitative evaluation of p53 by image analysis showed an inverse correlation between p53 expression and HPV presence, suggesting protein degradation. Image analysis also demonstrated that PCNA expression was more intense in HPV DNA 16/18 OSCCs. These findings suggest involvement of high-risk HPV types in oral carcinogenesis.     

Ferritin immunohistochemical localization in normal and neoplastic colonic mucosa

High levels of ferritin have been detected in serum and tumoral extracts of gastrointestinal neoplasms. However, its histological localization is not well known. An immunoperoxidase technique (PAP) was used for detecting ferritin in 30 colorectal carcinomas, 20 polyps and 8 cases of non-neoplastic mucosae. Ferritin staining was detected in stromal cells (98%) much more than in epithelial cells (21%). Connective cells were positive in 5 cases of normal mucosae (62%), 19 polyps (95%) and all carcinomas (100%). The number of positive cells gradually rose from normal mucosa to carcinoma with an intermediate score in adenomas. However, no relation could be found between the stromal ferritin score and dysplasia in polyps. Likewise, no relation was found between the stromal ferritin score and the differentiation grade, invasion or metastases in carcinomas. The positive epithelial pattern seen in 12 cases (21%) suggests non-specific staining due to passive diffusion from the stroma. Thus, these immunohistochemical findings suggest that in colonic neoplasms, ferritin could be a tumor marker produced mainly by stromal cell reaction more than by the epithelial cells.     

Galectin-3 Overexpression in Invasive Laryngeal Carcinoma, Assessed by Computer-assisted Analysis

The larynx is the most common site of malignancy in the upper aerodigestive tract. In Brazil, malignant laryngeal lesions represent 2% of all cancers, with ∼3000 annual deaths. The association between human papillomavirus (HPV) and laryngeal cancer is still controversial. The aim of the present retrospective study was to determine the expression of galectin-3 immunoperoxidase in laryngeal carcinoma by examining paraffin-embedded larynx biopsies from 65 patients, 10 in situ laryngeal carcinomas, 27 laryngeal carcinomas without metastases, and 28 with metastases. Twenty-eight cervical lymph nodes from patients with metastatic lesions were also evaluated. Nested PCR was performed to detect and type HPV DNA. Galectin-3 expression was assessed by immunohistochemistry using a computer-assisted system. Among 65 patients, 55 (84.6%) were positive to beta-globin (internal control); 10 (15.4%) patients were beta-globin negative and were excluded from the HPV evaluation. Thus, 7 (12.7%) out of 55 patients were HPV positive and 48 (87.3%) out of 55 patients were HPV negative. High expression of galectin-3 was observed in invasive laryngeal tumors, suggesting that galectin-3 could be associated with the invasiveness and aggressiveness of laryngeal carcinomas. (J Histochem Cytochem 57:665–673, 2009)     

Up-regulation of Na(+),K(+)-ATPase alpha 3-isoform and down-regulation of the alpha1-isoform in human colorectal cancer

We investigated expression levels of Na(+),K(+)-ATPase alpha-isoforms and their ATPase activities in human colorectal cancer tissue and the accompanying normal mucosa. A decrease in expression of the alpha1-isoform protein was observed in all sampled cancer tissues compared with the normal mucosae. The level of ouabain (5 microM)-sensitive Na(+),K(+)-ATPase activity in carcinomas was 81+/-5% that of in the normal mucosae. The mRNA expression of alpha2- and alpha 4-isoforms was decreased in almost all the carcinoma samples. Interestingly, the expression level of the alpha 3-isoform protein in the cancer tissue was higher than that of the normal mucosa. These results indicate that a decrease in the alpha1-isoform expression and an increase in the alpha 3-isoform expression may be associated with colorectal cancer.     

Production of human papillomavirus type 33 L1 major capsid protein and virus-like particles from Bacillus subtilis to develop a prophylactic vaccine against cervical cancer

We developed a bacterial expression system to produce human papillomavirus (HPV) type 33 L1 major capsid protein and virus-like particles from a recombinant Bacillus subtilis strain. For the first time, we have isolated self-assembled virus-like particles (VLPs) of HPV type 33 from B. subtilis, a strain generally recognized as safe (GRAS). The gene encoding the major capsid protein L1 of HPV type 33 was amplified from viral DNA isolated from a Korean patient and expressed in B. subtilis; a xylose-induction system was used to control gene activity. HPV33 L1 protein was partially purified by 40% (w/v) sucrose cushion centrifugation and strong cation exchange column chromatography. Eluted samples exhibited immunosignaling in fractions of 0.5-1.0 M NaCl. The HPV33 L1 protein was shown to be approximately 56 kDa in size by SDS-PAGE and Western blotting; recovery and purity were quantified by indirect immuno-ELISA assay. The final yield and purity were approximately 20.4% and 10.3%, respectively. Transmission electron microscopic analysis of fractions immunoactive by ELISA revealed that the L1 protein formed self-assembled VLPs with a diameter of approximately 20-40 nm. Humoral and cellular immune responses provoked by the B. subtilis/HPV33 L1 strain were approximately 100- and 3-fold higher than those of the empty B. subtilis strain as a negative control, respectively. Development of a VLP production and delivery system using B. subtilis will be helpful, in that the vaccine may be convenient production as an antigen delivery system. VLPs thus produced will be safer for human use than those purified from Gram-negative strains such as Escherichia coli. Also, use of B. subtilis as a host may aid in the development of either live or whole cell vaccines administered by antigen delivery system.     

Detection of Merkel cell polyomavirus in cervical squamous cell carcinomas and adenocarcinomas from Japanese patients

ABSTRACT: BACKGROUND: Merkel cell polyomavirus (MCPyV) was identified originally in Merkel cell carcinoma (MCC), a rare form of human skin neuroendocrine carcinoma. Evidence of MCPyV existence in other forms of malignancy such as cutaneous squamous cell carcinomas (SCCs) is growing. Cervical cancers became the focus of our interest in searching for potentially MCPyV-related tumors because: (i) the major histological type of cervical cancer is the SCC; (ii) the uterine cervix is a common site of neuroendocrine carcinomas histologically similar to MCCs; and (iii) MCPyV might be transmitted during sexual interaction as demonstrated for human papillomavirus (HPV). In this study, we aimed to clarify the possible presence of MCPyV in cervical SCCs from Japanese patients. Cervical adenocarcinomas (ACs) were also studied. RESULTS: Formalin-fixed paraffin-embedded tissue samples from 48 cervical SCCs and 16 cervical ACs were examined for the presence of the MCPyV genome by polymerase chain reaction (PCR) and sequencing analyses. PCR analysis revealed that 9/48 cervical SCCs (19 %) and 4/16 cervical ACs (25 %) were positive for MCPyV DNA. MCPyV-specific PCR products were sequenced to compare them with reference sequences. The nucleotide sequences in the MCPyV large-T (LT)-sequenced region were the same among MCPyV-positive cervical SCCs and AC. Conversely, in the MCPyV viral protein 1 (VP1)-sequenced, two cervical SCCs and three cervical ACs showed several nucleotide substitutions, of which three caused amino acid substitutions. These sequencing results suggested that three MCPyV variants of the VP1 were identified in our cases. Immunohistochemistry showed that the LT antigen was expressed in tumor cells in MCPyV-positive samples. Genotyping of human HPV in the MCPyV-positive samples revealed that infected HPVs were HPV types 16, 31 and 58 for SCCs and HPV types 16 and 18 for ACs. CONCLUSIONS: This study provides the first observation that MCPyV coexists in a subset of HPV-associated cervical cancers from Japanese patients. The prevalence of MCPyV in these lesions was close to that observed in the cutaneous SCCs. Further worldwide epidemiological surveys are warranted to determine the possible association of MCPyV with pathogenesis of cervical cancers.     

Germline mutation in RNASEL predicts increased risk of head and neck, uterine cervix and breast cancer

THE BACKGROUND: Ribonuclease L (RNASEL), encoding the 2'-5'-oligoadenylate (2-5A)-dependent RNase L, is a key enzyme in the interferon induced antiviral and anti-proliferate pathway. Mutations in RNASEL segregate with the disease in prostate cancer families and specific genotypes are associated with an increased risk of prostate cancer. Infection by human papillomavirus (HPV) is the major risk factor for uterine cervix cancer and for a subset of head and neck squamous cell carcinomas (HNSCC). HPV, Epstein Barr virus (EBV) and sequences from mouse mammary tumor virus (MMTV) have been detected in breast tumors, and the presence of integrated SV40 T/t antigen in breast carcinomas correlates with an aggressive phenotype and poor prognosis. A genetic predisposition could explain why some viral infections persist and induce cancer, while others disappear spontaneously. This points at RNASEL as a strong susceptibility gene. METHODOLOGY/PRINCIPAL FINDINGS: To evaluate the implication of an abnormal activity of RNase L in the onset and development of viral induced cancers, the study was initiated by searching for germline mutations in patients diagnosed with uterine cervix cancer. The rationale behind is that close to 100% of the cervix cancer patients have a persistent HPV infection, and if a defective RNase L were responsible for the lack of ability to clear the HPV infection, we would expect to find a wide spectrum of mutations in these patients, leading to a decreased RNase L activity. The HPV genotype was established in tumor DNA from 42 patients diagnosed with carcinoma of the uterine cervix and somatic tissue from these patients was analyzed for mutations by direct sequencing of all coding and regulatory regions of RNASEL. Fifteen mutations, including still uncharacterized, were identified. The genotype frequencies of selected single nucleotide polymorphisms (SNPs) established in the cervix cancer patients were compared between 382 patients with head and neck squamous cell carcinomas (HNSCC), 199 patients with primary unilateral breast cancer and 502 healthy Danish control individuals. We found that the genotype frequencies of only one of the 15 mutations, the yet uncharacterized 5'UTR mutation rs3738579 differed significantly between cancer patients and control individuals (P-value: 4.43x10(-5)). CONCLUSION/SIGNIFICANCE: In conclusion, we have discovered an increased risk, a heterozygous advantage and thereby a protective effect linked to the RNASEL SNP rs3738579. This effect is found for patients diagnosed with carcinoma of the uterine cervix, HNSCC, and breast cancer thus pointing at RNASEL as a general marker for cancer risk and not restricted to familial prostate cancer.     

The biology and significance of human papillomavirus infections in the genital tract.

A variety of human papillomavirus (HPV) types infect the anogenital mucosa, giving rise to lesions that differ in clinical appearance, histology, and risk of malignant progression. Certain high-risk types (HPVs 16, 18, 31, 33, 35 and 39) have a strong association with high-grade epithelial neoplasia and invasive carcinomas of the anogenital tract. Cancer appears to have a multifactorial etiology, and HPV infection alone is probably insufficient for malignant transformation. The consistent association between HPV infection and anogenital cancers emphasizes, however, that the sexually transmitted papillomaviruses may have a necessary role in carcinogenesis. Hence, there is a prospect that vaccination programs may one day allow public health control of HPV infection, thereby eliminating an important risk factor.     

Status of the human DNA papillomavirus in cervical tumors]

Cervical carcinoma is etiologically associated with the human papilloma virus (HPV), HPV 16 and HPV 18 being the most common. Viral DNA is thought to persist mostly in the episomal form in early tumor development, and in the integrated form in carcinomas. This assumption was checked with a new method that discriminated between RNAs transcribed from episomal and integrated HPV DNAs. Both forms were detected in carcinomas of Russian patients regardless of the disease stage. The data were verified by two other methods. RNA with sequences of the HPV transforming gene E7 proved to be transcribed from either DNA form. The results suggest that HPV integration is not crucial for carcinoma progression.     

The c-myc activation in cervical carcinomas and HPV 16 infections

Despite the prominent role for Human Papillomavirus (HPV) infection in the development of genital cancer, other genetic or environmental co-factors have also been involved. Studies of c-myc activation in cervical carcinomas have reported that gene over-expression (mainly gene amplification) are common in cervical squamous cell carcinomas and may correlate with the biologic behavior of the neoplasm. Using PCR based technology, DNAs from 79 normal cervical samples and 225 abnormal cervical tissue scrapes were analyzed for HPV detection and typing and for c-myc gene amplification. Significant differences were found between the different cyto/histology groups (P<0.0001) and also with HPV high-risk infected samples (P<0.0002). In this sense, we showed that the average c-myc copy number increased according to the histological grade of the lesion (OR=6.3, CI=2.1-18.8). Also, the results showed that the infection with HPV 16 was tightly associated with c-myc amplification (OR=10.6, CI=3.1-36). These results could indicate that oncogene amplification take place in pre-invasive stages of cervical disease and could cooperate not only in tumor progression but also in cell transformation. Moreover, the results strongly associate the c-myc gene amplification to the infection with the oncogenic HPV 16, showing that the pattern of virus infection and oncogene activation could be specific for different viral genotypes.     

Presence of papillomavirus genomes and amplification of the c-myc and C-Ha-ras oncogenes in invasive cancers of the uterine cervix

Invasive squamous cell carcinomas of the uterine cervix from 12 untreated patients were examined for the presence of human papillomavirus (HPV) genomes and for the state of the oncogenes c-myc and c-Ha-ras. Blot hybridization experiments have demonstrated the presence of the genome of HPV type 16 (HPV 16) in six tumors and that of the genomes of HPV types weakly related to HPV 16 or HPV 18 in five others. In the nine tumors corresponding to advanced stages of the disease (stages 3 and 4) there was a 3-30 fold amplification of c-myc and/or c-Ha-ras. A concomitant amplification of both oncogenes was found in eight cancers. In only one of the three tumors confined to the cervix (stage 1), the oncogene c-Ha-ras was weakly amplified. Neither HPV DNA sequences, nor oncogene amplification were detected in the leukocytes of five patients. Thus, it seems likely that specific HPV types play a role in the development of carcinomas of the uterine cervix, and that cellular oncogenes, activated through an amplification process, are involved in at least some steps of tumor progression.     

Expression of A and H blood-group and of CD44 antigens during chemical rat colonic carcinogenesis

Using an experimental model of rat colon adenocarcinoma, we have recently shown that the presence of H blood-group antigen on variants of the CD44 adhesion molecule carrying amino acids encoded by exon v6 (CD44v6), increased the cells' tumorigenicity. In the present study, colon adenocarcinomas were induced by 1,2-dimethylhydrazine treatment in rats. Using immunohistochemistry, biopsies of normal, precancerous and carcinomatous colon mucosa were evaluated for expression A and H blood group antigens and CD44s and CD44v6 antigens. Normal rat colon showed strong and homogeneous expression of blood-group antigen A, but weak expression of H antigen. Several weeks before the appearance of tumours, dysplastic glands were strongly stained with anti-H reagents, while their A antigen was lost. Expression of CD44v6 was weak and restricted to some cells at the bottom of normal crypts. No obvious change was observed before appearance of severe dysplasia. In carcinomas, a strong but irregular expression of A, H and CD44v6 antigens was observed. In moderately differentiated carcinomas, A and H antigens were present at the apical surface of cells, whereas CD44v6 was found at the basolateral side. Only carcinomatous cells with loss of polarity, found in poorly differentiated cancers or infiltrated in the muscularis mucosae, were found to coexpress blood-group H or A and CD44v6 antigens at their surface. This revised version was published online in November 2006 with corrections to the Cover Date.