Molecular analysis of familial endometrial carcinoma: a manifestation of hereditary nonpolyposis colorectal cancer or a separate syndrome?

PURPOSE: Familial clustering of endometrial carcinoma (EC) may occur as part of hereditary nonpolyposis colorectal cancer (HNPCC), a multiorgan cancer syndrome with mismatch repair (MMR) deficiency. Clustering of EC alone, termed as familial site-specific EC, may constitute a separate entity. Because its genetic basis is unknown, our purpose was to characterize such families molecularly. MATERIALS AND METHODS: Twenty-three families with site-specific EC were identified among 519 consecutive patients diagnosed with EC during 1986 to 1997. Tumor tissues were examined for MMR protein expression by immunohistochemical (IHC) analysis, and MMR genes pinpointed by IHC changes were screened for germline mutations by exon-by-exon sequencing, multiplex ligation-dependent probe amplification, and direct tests for mutations common in the population. RESULTS: Among 33 ECs from 23 families, MLH1 protein was lost in seven tumors (21%), MSH2 together with MSH6 was lost in four tumors (12%), and MSH6 alone was lost in five tumors (15%). A truncating germline mutation in MSH6 (3261insC) was identified in one family and a likely pathogenic missense mutation in MSH2 (D603N) was identified in another family. Among the original 519 patients, nine (all with colon cancer in the family) were diagnosed with HNPCC at the outset-six with MLH1 and three with MSH2 mutations. CONCLUSION: Our study gives a minimum overall frequency of 2.1% (11 of 519) for germline MMR defects ascertained through EC in the index patients. The fact that only two of 23 families with site-specific EC (8.7%) had germline mutations in MMR genes suggests another as yet unknown etiology in most families with site-specific EC.     

Clinical features and impact of p53 status on sporadic mismatch repair deficiency and Lynch syndrome in uterine cancer

The clinical features of sporadic mismatch repair deficiency (MMRd) and Lynch syndrome (LS) in Japanese patients with endometrial cancer (EC) were examined by evaluating the prevalence and prognostic factors of LS and sporadic MMRd in patients with EC. Targeted sequencing of five LS susceptibility genes (MLH1, MSH2, MSH6, PMS2, and EPCAM) was carried out in 443 patients with EC who were pathologically diagnosed with EC at the National Cancer Center Hospital between 2011 and 2018. Pathogenic variants in these genes were detected in 16 patients (3.7%). Immunohistochemistry for MMR proteins was undertaken in 337 of the 433 (77.9%) EC patients, and 91 patients (27.0%) showed absent expression of at least one MMR protein. The 13 cases of LS with MMR protein loss (93.8%) showed a favorable prognosis with a 5-year overall survival (OS) rate of 100%, although there was no statistically significant difference between this group and the sporadic MMRd group (p = 0.27). In the MMRd without LS group, the 5-year OS rate was significantly worse in seven patients with an aberrant p53 expression pattern than in those with p53 WT (53.6% vs. 93.9%, log-rank test; p = 0.0016). These results suggest that p53 abnormalities and pathogenic germline variants in MMR genes could be potential biomarkers for the molecular classification of EC with MMRd.     

Human leukocyte antigen class I antigen expression is an independent prognostic factor in ovarian cancer.

PURPOSE: Despite improvements in cancer treatment, the prognosis of ovarian cancer remains low and imperfectly predicted by traditional pathologic criteria. Biomarkers that predict prognosis independently of such criteria shed light on important molecular variations, aiding in the development and targeting of novel therapies. Previous work has shown human leukocyte antigen (HLA) class I antigen expression to be independently predictive of prognosis in colorectal and breast cancer. We investigated the prognostic potential of HLA class I antigen expression by studying a large series of ovarian cancers. EXPERIMENTAL DESIGN: A tissue microarray of 339 ovarian cancer cases linked to prospectively recorded clinicopathologic and follow-up data was constructed. This was stained following a standard immunohistochemical protocol for HLA class I heavy chain (HC-10) and beta(2)-microglobulin (beta(2)-m). HLA class I antigen expression was compared with clinicopathologic factors and overall disease-specific survival using the Pearson chi(2) test, Kaplan-Meier curves, and the log-rank test. Cox regression was used to test for the independence and magnitude of effects. RESULTS: There were no univariate correlations between HLA class I antigen expression and clinicopathologic factors. Deviation from an HC-10(+)/beta(2)-m(+) phenotype correlated with reduced survival in univariate analysis (log-rank, 5.69; P = 0.017); a retained HC-10(+)/beta(2)-m(+) phenotype predicted improved prognosis independently of age, stage, level of cytoreduction, and chemotherapy usage on multivariate analysis (hazard ratio, 0.587; 95% confidence interval, 0.442-0.781; P < 0.001). CONCLUSIONS: HLA class I antigen expression is an independent prognostic marker in ovarian cancer, its loss correlating with a poor prognostic outcome.     

Alterations in classical and nonclassical HLA expression in recurrent and progressive HPV‐induced usual vulvar intraepithelial neoplasia and implications for immunotherapy

Immunotherapy of usual vulvar intraepithelial neoplasia (uVIN) is promising; however, many patients still fail to show clinical responses, which could be explained by an immune escape through alterations in human leukocyte antigen (HLA) expression. Therefore, we analyzed a cohort of patients with a primary (n = 43) and subsequent recurrent uVIN lesion (n = 20), vaccine‐treated uVIN patients (n = 12), patients with human papillomavirus (HPV)‐induced vulvar carcinoma (n = 21) and healthy controls (n = 26) for the expression of classical HLA‐class I/II and nonclassical HLA‐E/‐G and MHC class I chain‐related molecule A (MICA). HLA‐class I was downregulated in 70% of uVIN patients, including patients with a clinical response to immunotherapy. Downregulation of HLA‐class I is probably reversible, as only 15% of the uVIN cases displayed loss of heterozygosity (LOH) and HLA‐class I could be upregulated in uVIN keratinocyte cultures by interferon γ. HLA‐class I downregulation is more frequently associated with LOH in vulvar carcinomas (25–55.5%). HLA‐class II was found to be focally expressed in 65% of uVIN patients. Of the nonclassical molecules, MICA was downregulated in 80% of uVIN whereas HLA‐E and ‐G were expressed in a minority of cases. Their expression was more prominent in vulvar carcinoma. No differences were found between the alterations observed in paired primary and recurrent uVIN. Importantly, downregulation of HLA‐B/C in primary uVIN lesions was associated with the development of recurrences and progression to cancer. We conclude that downregulation of HLA is frequently observed in premalignant HPV‐induced lesions, including clinical responders to immunotherapy, and is associated with worse clinical outcome. However, in the majority of cases downregulation may still be reversible.     

Loss of MSH3 protein expression is frequent in MLH1-deficient colorectal cancer and is associated with disease progression

Mononucleotide repeat sequences are particularly prone to frameshift mutations in tumors with biallelic inactivation of the mismatch repair (MMR) genes MLH1 or MSH2. In these tumors, several genes harboring mononucleotide repeats in their coding region have been proposed as targets involved in tumor progression, among which are also the MMR genes MSH3 and MSH6. We have analyzed the expression of the MSH3 and MSH6 proteins by immunohistochemistry in 31 colorectal carcinomas in which MLH1 was inactivated. Loss of MSH3 expression was identified in 15 tumors (48.5%), whereas all tumors expressed MSH6. Frameshift mutations at coding microsatellites were more frequent in MSH3 (16 of 31) than in MSH6 (3 of 31; Fisher's exact test, P < 0.001). Frameshift mutations and allelic losses of MSH3 were more frequent in MSH3-negative tumors compared with those with normal expression (22 mutations in 30 alleles versus 8 mutations in 28 alleles; chi(2), P = 0.001). Biallelic inactivation was evident or inferred for 60% of MSH3-negative tumors but none of the tumors with normal MSH3 expression. In contrast, we did not identify frameshift mutations in the (A)8 tract of MSH3 in a control group of 18 colorectal carcinomas in which the MMR deficiency was based on the inactivation of MSH2. As it has been suggested that mutations of MSH3 might play a role in tumor progression, we studied the association between MSH3 expression and disease stage assessed by lymph node and distant metastases status. Dukes stages C and D were more frequent in primary tumors with loss of MSH3 expression (9 of 13), compared with tumors with retained expression (1 of 14; Fisher's exact test, P = 0.001), suggesting that MSH3 abrogation may be a predictor of metastatic disease or even favor tumor cell spread in MLH1-deficient colorectal cancers.     

MMR蛋白在515例子宫内膜样腺癌中表达及其与临床病理学特征的相关性分析

背景与目的：林奇综合征（Lynch syndrome,LS）相关的子宫内膜癌有着独特的临床病理学特征及治疗手段。对新发子宫内膜癌患者采用免疫组织化学（immunohistochemistry,IHC）染色的方法检测错配修复（mismatch repair,MMR）蛋白表达情况,可以有效地筛查LS相关的癌症患者。本研究探讨MMR蛋白（MLH1、MSH2、MSH6及PMS2）在子宫内膜样腺癌中的表达情况及其与患者临床病理学特征的关系。方法：收集中国医科大学盛京医院2018年1月—2020年8月共515例子宫内膜样腺癌连续性病例为研究对象,年龄范围为28 ~ 81（57.73±8.41）岁。采用IHC染色的方法检测癌组织中MLH1、MSH2、MSH6和PMS2蛋白表达情况,应用聚合酶链式反应（polymerase chain reaction,PCR）方法对MLH1蛋白表达缺失的标本进行基因的甲基化检测,并且分析MMR蛋白表达缺失情况与子宫内膜样腺癌临床病理学特征的关系。只要有一种MMR蛋白表达缺失即判定为MMR蛋白错配修复缺陷（deficient mismatch repair,dMMR）,蛋白全部阳性则判定为MMR表达完整（proficient mismatch repair,pMMR）。结果：515例子宫内膜样腺癌中有138例（26.8%）发生MMR蛋白表达缺失,MLH1、PMS2、MSH2及MSH6蛋白表达缺失率分别是16.3%（84/515）、19.0%（98/515）、5.4%（28/515）、8.0%（41/515）。MMR蛋白的缺失以MLH1和PMS2联合表达缺失（60.9%,84/138）为主;其次为MSH2和MSH6联合表达缺失（18.8%,26/138）;MSH2、MSH6和PMS2联合表达缺失有2例（1.4%,2/138）;PMS2、MSH2和MSH6蛋白单独表达缺失比例分别为8.0%（11/138）、1.4%（2/138）、10.1%（14/138）。对27例MLH1蛋白表达缺失标本进行甲基化检测,结果显示,阳性率为85.2%（23/27）。515例子宫内膜样腺癌组织中的MMR蛋白表达缺失与患者发病年龄、国际妇产科联合会（The International Federation of Gynecology and Obstetrics,FIGO）分期、组织学分化程度、浸润深度、脉管转移、神经侵犯、淋巴结转移、p53异常表达、肿瘤浸润淋巴细胞（tumor infiltrating lymphocyte,TIL）及肿瘤伴瘤周淋巴细胞有相关性,而与是否累及子宫下段无关。与pMMR的患者相比,dMMR的患者发病年龄更小,FIGO临床分期多为Ⅲ ~ Ⅳ期,组织学分化程度多为低分化,肿瘤多无肌层浸润,肿瘤多出现脉管神经侵犯及淋巴结转移,肿瘤浸润淋巴细胞增多,且肿瘤伴瘤周淋巴细胞更显著,MSH6蛋白缺失患者多无p53异常表达。结论：dMMR的子宫内膜样腺癌患者具有独特的临床病理学特征。应用免疫组织化学染色方法检测MMR蛋白表达情况,并对MLH1表达缺失的标本进行基因甲基化检测,能初步筛查LS患者,对肿瘤患者免疫治疗具有一定指导意义。     

Muir–Torre syndrome or phenocopy? The value of the immunohistochemical expression of mismatch repair proteins in sebaceous tumors of immunocompromised patients

Primary and secondary immunodepressive conditions are associated with an increased incidence of sebaceous tumors. Microsatellite instability (MSI) and lack of expression of mismatch repair (MMR) proteins, typical markers of Muir–Torre/Lynch heredo-familial settings, can be recognized also in immunocompromised patients. We aimed to carry on a systematic examination of clinical, immunohistochemical, biomolecular features of sebaceous tumors arising in immunocompromised and immunocompetent patients between 1986 and 2012. Microsatellite screening, immunohistochemical analysis and genetic testing were performed for hMLH1, hMSH2 and hMSH6. Methylation status of MMR genes was checked in cases with immunohistochemistry (IHC) loss of MMR proteins expression and no germline mutations. Fifteen patients had a personal history of visceral carcinomas fulfilling diagnostic criteria for Muir–Torre syndrome. In this cohort, IHC analysis, MSI status and genetic testing were in agreement, showing eight MSH2 and two MLH1 germline mutations. Five patients were immunosuppressed and their sebaceous tumors showed a lack of MSH2/MSH6 expression, although just one case with positive family history for visceral cancer harbored a germline mutation. In immunosuppressed patients, loss of IHC for MMR proteins is not necessarily secondary to MMR germline mutations. IHC false positives are probably due to epigenetic alterations. MSI and lack of expression of MMR proteins can be recognized also in immunocompromised patients without MMR germline mutations.     

MLH1 promoter hypermethylation predicts poorer prognosis in mismatch repair deficiency endometrial carcinomas

ObjectiveThe antitumor effects of anti-PD-1 antibody against mismatch repair deficiency (MMR-D)-associated cancers have been reported. MMR-D is found in approximately 20%–30% of endometrial carcinomas (ECs) and frequently occurs due to MLH1 promoter hypermethylation (MLH1-PHM). ECs with MLH1-PHM are classified according to the molecular screening of Lynch syndrome (LS), but few detailed reports are available. The purpose of this study was to clarify the clinical features of EC with MLH1-PHM.MethodsImmunohistochemistry of MMR proteins (MLH1, MSH2, MSH6, and PMS2) was performed on specimens from 527 ECs treated at our university hospital from 2003 to 2018. MLH1 methylation analysis was added to cases with MLH1/PMS2 loss. ECs were classified as follows: cases that retained MMR proteins as “MMR-proficient;” cases with MLH1/PMS2 loss and MLH1-PHM as “met-EC;” and cases with other MMR protein loss and MLH1/PMS2 loss without MLH1-PHM as “suspected-LS.” The clinical features, including long-term prognosis, of each group, were analyzed.ResultsAccordingly, 419 (79.5%), 65 (12.3%), and 43 (8.2%) cases were categorized as “MMR-proficient,” “suspected-LS,” and “met-EC,” respectively. Significantly, “met-EC” had a lower proportion of grade 1 tumors (37.5%) and a higher proportion of stage III/IV tumors (37.2%) than the other groups. The overall and progression-free survival of “met-EC” were significantly worse than those of “suspected-LS” in all cases.ConclusionIn ECs with MMR-D, “met-ECs” were a subgroup with a poorer prognosis than “suspected-LS.” “Met-ECs” would be the main target for anti-PD-1 antibody treatment, and its clinical susceptibility should be verified individually.     

Microsatellite instability and DNA mismatch repair protein deficiency in Lynch syndrome colorectal polyps

Colorectal cancers associated with Lynch syndrome are characterized by deficient DNA mismatch repair (MMR) function. Our aim was to evaluate the prevalence of microsatellite instability (MSI) and loss of MMR protein expression in Lynch syndrome-associated polyps. Sixty-two colorectal polyps--37 adenomatous polyps, 23 hyperplastic polyps, and 2 sessile serrated polyps (SSP)--from 34 subjects with germline MMR gene mutations were tested for MSI using a single pentaplex PCR for five mononucleotide repeat microsatellite markers, and also for expression of MLH1, MSH2, MSH6, and PMS2 proteins by immunohistochemistry. High-level MSI (MSI-H) was seen in 15 of 37 (41%) adenomatous polyps, one of 23 (4%) hyperplastic polyps, and one of two (50%) SSPs. Loss of MMR protein expression was seen in 18 of 36 (50%) adenomatous polyps, zero of 21 hyperplastic polyps, and zero of two SSPs. Adenomatous polyps 8 mm or larger in size were significantly more likely to show MSI-H [OR, 9.98; 95% confidence interval (CI), 1.52-65.65; P = 0.02] and deficient MMR protein expression (OR, 3.17; 95% CI, 1.20-8.37; P = 0.02) compared with those less than 8 mm in size. All (six of six) adenomatous polyps 10 mm or larger in size showed both MSI-H and loss of MMR protein expression by immunohistochemistry. Our finding that the prevalence of MMR deficiency increases with the size of adenomatous polyps suggests that loss of MMR function is a late event in Lynch syndrome-associated colorectal neoplasia. Although testing large adenomatous polyps may be of value in the diagnostic evaluation of patients with suspected Lynch syndrome, the absence of an MMR-deficient phenotype in an adenoma cannot be considered as a strong evidence against Lynch syndrome, as it is with colorectal carcinomas.     

Clinical Impact of Mismatch Repair Protein Testing on Outcome of Early Staged Colorectal Carcinomas

Introduction

Colorectal cancer is the third most common cancer in men and second most common in women globally. In the present study, we aimed to analyse the proportion of patients with loss of immunostaining for mismatch repair (MMR) proteins in all newly diagnosed stage II cases of colorectal cancer for the purpose of prognostication, for determination of further chemotherapeutic strategy and for familial screening.

Method

From January 2014 to December 2015, 62 consecutive newly diagnosed cases of stage II colorectal cancer were included in the study. Details of each patient related to their demographic profile and tumour profile were recorded. All the cases were grossed and staged according to College of American Pathologist (CAP) guidelines. The expression of MMR proteins (which was earlier validated on normal as well as tumour tissue) in FFPE tumour tissue using IHC for mut L homologue 1 (MLH1), mut S homologue 2 (MSH2), mut S homologue 6 (MSH6) and post-meiotic segregation increased 2 (PMS2) was studied. Information regarding stage, treatment, clinical outcome and overall survival was retrieved when available.

Results

Out of a total of 371 cases, 62 (16.7%) cases were of stage II CRC, out of which 43 (12%) were treatment naive. Among the selected 62 cases, 26 (41.9%) demonstrated loss of MMR proteins and 36 (58.0%) cases had intact nuclear expression. Out of the cases with MMR loss, 38.4% showed loss of MLH1 and PMS2, 30.7% showed loss of MSH2 and MSH6, 26.9% showed isolated loss of PMS2 and 3.8% showed isolated loss of MSH6. Right-sided location (57.6%) was more common than left-sided (19.2%) and transverse colon (23.0%). Majority of the cases were moderately differentiated (65.3%) in morphology. There was no intratumoural infiltrate in most of the cases (53.8%), and only 3.8% cases showed marked intratumoural infiltrate. Also, peritumoural lymphocytic infiltrate was mild to moderate in most of the cases (26.9%) and marked Crohn’s-like infiltrate was seen in only 7.6% cases.

Conclusion

Our study shows that the routine evaluation of MMR proteins is achievable and essential for the purpose of prognostication, planning of treatment strategies and ascertaining a hereditary basis of CRC. The incidence of MMR protein loss was quite high in our study compared to other studies probably due to a difference in ethnicity. Though a right-sided predominance was supported, none of the typical morphological features of microsatellite instability (MSI) tumours were substantiated by our study, highlighting the lack of importance of histology for predicting MSI, and emphasising the point that MSI testing should be done as a routine procedure in all stage II CRC. A short follow-up was done for all our cases and comparison between the survival of the chemotherapy treated MSI cases versus those which were treatment naïve was performed and revealed that chemotherapy (CT) did not provide additional benefit to survival; MSI tumours in general are a better prognostic category and do not require additional chemotherapy.

     

Mismatch Repair Protein Expression and Microsatellite Instability in Cutaneous Squamous Cell Carcinoma

There exist relatively sparse and conflicting data on high-level microsatellite instability (MSI-H) and deficient mismatch repair (dMMR) in cutaneous malignancies. We aimed to determine the expression profiles of MMR proteins (MSH2, MSH6, MLH1, and PMS2) in different progression stages of cutaneous squamous cell carcinoma (cSCC, 102 patients in total) by immunohistochemistry, and search for MSI-H in patients with low-level MMR or dMMR using multiplex-PCR. Low-level MMR protein expression was observed in five patients: One patient with primary cSCC < 2 mm thickness and low-level MLH1, three patients with primary cSCC > 6 mm (including one with low-level MSH2, as well as MSH6 expression, and two with low-level PMS2), and one patient with a cSCC metastasis showing low-level MSH2 as well as MSH6. Intergroup protein expression analysis revealed that MLH1 and MSH2 expression in actinic keratosis was significantly decreased when compared to Bowen’s disease, cSCC < 2 mm, cSCC > 6 mm, and cSCC metastasis. In cases with low-level MMR, we performed MSI-H tests revealing three cases with MSI-H and one with low-level MSI-L. We found low-level MMR expression in a small subset of patients with invasive or metastatic cSCC. Hence, loss of MMR expression may be associated with tumour progression in a small subgroup of patients with non-melanoma skin cancer.     

Immunohistochemical expression pattern of MMR protein can specifically identify patients with colorectal cancer microsatellite instability

The microsatellite instability (MSI) pathway is found in most cases of hereditary nonpolyposis colorectal cancer (HNPCC) and in 12 % of sporadic colorectal cancer (CRC). It involves inactivation of deoxyribonucleic acid mismatch repair (MMR) genes MLH1, MSH2, PMS2, and MSH6. MMR germline mutation detections are an important supplement to HNPCC clinical diagnosis. It enables at-risk and mutation-positive relatives to be informed about their cancer risks and to benefit from intensive surveillance programs that have been proven to reduce the incidence of CRC. In this study, we analyzed for the first time in Tunisia the potential value of immunohistochemical assessment of MMR protein to identify microsatellite instability in CRC. We evaluate by immunohistochemistry MMR protein expression loss in tumoral tissue compared to positive expression in normal mucosa. Immunohistochemistry revealed loss of expression for MLH1, MSH2, MSH6, and PMS2 in 15, 21, 13, and 15 % of cases, respectively. Here, we report a more elevated frequency of MSI compared to data of the literature. In fact, by immunohistochemistry, 70 % of cases were shown to be MSS phenotype, whereas 30 % of cases, in our set, were instable. Moreover, according to molecular investigation, 71 % of cases were instable (MSI-H) and remaining cases were stable (29 %). Thus, we found a perfect association between MMR immunohistochemical analyses and MSI molecular investigation. Immunohistochemical analysis of MMR gene product expression may allow one to specifically identify MSI phenotype of patients with colorectal carcinomas.     

Loss of colonic HLA antigens in familial adenomatous polyposis.

The loss of HLA antigens by neoplastic cells is considered important for tumor growth and metastasis, since it may allow tumors to escape immune surveillance. We studied the expression of HLA class I and II antigens in the colons of 10 patients with familial adenomatous polyposis (FAP), a condition which leads inevitably to colorectal cancer. Expression of HLA class antigens was studied by immunohistochemistry in (a) adenomas from patients with FAP, (b) histologically normal mucosa distant from the adenomas, and (c) histologically normal colonic mucosa from normal subjects. The expression of HLA class I and II antigens was decreased in histologically normal mucosa from FAP patients compared to normal controls. Adenomas showed a similar but quantitatively more pronounced reduction (or loss) of HLA antigen expression. The reduction of HLA expression in adenomas was comparable to that observed in sporadic colon carcinomas. This generalized suppression of HLA gene expression in the colon of FAP patients, which precedes the onset of overt histological manifestations of neoplasia, may be an important early event in colon carcinogenesis.     

Mismatch repair markers in preoperative and operative endometrial cancer samples; expression concordance and prognostic value

Background The endometrial cancer mismatch repair (MMR) deficient subgroup is defined by loss of MSH6, MSH2, PMS2 or MLH1. We compare MMR status in paired preoperative and operative samples and investigate the prognostic impact of differential MMR protein expression levels.Methods Tumour lesions from 1058 endometrial cancer patients were immunohistochemically stained for MSH6, MSH2, PMS2 and MLH1. MMR protein expression was evaluated as loss or intact to determine MMR status, or by staining index to evaluate the prognostic potential of differential expression. Gene expression data from a local (n = 235) and the TCGA (n = 524) endometrial cancer cohorts was used for validation.Results We identified a substantial agreement in MMR status between paired curettage and hysterectomy samples. Individual high expression of all four MMR markers associated with non-endometrioid subtype, and high MSH6 or MSH2 strongly associated with several aggressive disease characteristics including high tumour grade and FIGO stage, and for MSH6, with lymph node metastasis. In multivariate Cox analysis, MSH6 remained an independent prognostic marker, also within the endometrioid low-grade subgroup (P < 0.001).Conclusion We demonstrate that in addition to determine MMR status, MMR protein expression levels, particularly MSH6, may add prognostic information in endometrial cancer.Subject terms: Endometrial cancer, DNA mismatch repair, Endometrial cancer, Prognostic markers, Gynaecological cancer     

E5 protein of human papillomavirus type 16 selectively downregulates surface HLA class I

Papillomaviruses have evolved mechanisms that result in escape from host immune surveillance. The E5 protein is expressed early in papillomavirus infection in the deep layers of the infected epithelium. It is localized to the Golgi apparatus (GA) and endoplasmic reticulum. The E5 protein of bovine papillomavirus (BPV) impairs the synthesis and stability of major histocompatibility (MHC) class I complexes and prevents their transport to the cell surface due to retention in the GA. Here we show that human papillomavirus type 16 (HPV-16) E5 also causes the retention of MHC (HLA) class I complexes in the GA and impedes their transport to the cell surface, which is rescued by treatment with interferon. Unlike BPV E5, HPV-16 E5 does not affect the synthesis of HLA class I heavy chains or the expression of the transporter associated with antigen processing TAP. These results show that downregulation of surface MHC class I molecules is common to both BPV and HPV E5 proteins. Moreover, we determined that HPV-16 E5 downregulates surface expression of HLA-A and HLA-B, which present viral peptides to MHC class I-restricted cytotoxic T lymphocytes (CTLs), but not the natural killer (NK) cell inhibitory ligands HLA-C and HLA-E. Selective downregulation of cell surface HLA class I molecules may allow the virus to establish infection by avoiding immune clearance of virus-infected cells by both CTLs and NK cells.     

NK cell activating ligands on human malignant cells: molecular and functional defects and potential clinical relevance

Malignant transformation of cells is frequently associated with HLA class I antigen downregulation or loss. This abnormality provides malignant cells with a mechanism to escape control by HLA class I antigen-restricted, tumor antigen-specific cytotoxic T lymphocytes. Surprisingly, HLA class I antigen downregulation or loss by tumor cells is not associated with control of tumor growth by natural killer (NK) cells, as it would be predicted by the "missing-self" hypothesis. Here, we discuss the role of NK cell activating ligand abnormalities as well as HLA class I molecule and ICAM-1 shedding in the lack of control of tumor growth by NK cells with emphasis on their molecular mechanisms. In addition, we discuss the impact of these abnormalities on cancer immune surveillance.     

Downregulation of HLA Class I molecules in the tumour is associated with a poor prognosis in patients with oesophageal squamous cell carcinoma

As antigenic peptides in the context of human leukocyte antigen (HLA) class I molecules are recognised by cytotoxic T lymphocytes (CTL), the downregulation of HLA class I molecules is one of the reasons why tumour cells can evade CTL-mediated anti-tumour immunity. In this study, we investigated HLA class I expression in oesophageal squamous cell carcinoma (ESCC) (n=70) and in their metastatic lesions (lymph nodes (n=40) and liver (n=3)), by immunohistochemistry with anti-HLA class I monoclonal antibody (EMR8-5). As a result, the downregulation of HLA class I expression in primary lesions of ESCC was observed in 43%, and that in metastatic lymph nodes was noted in 90%. Furthermore, patients with preserved HLA class I expression in primary tumours showed a better survival in comparison to those with downregulated HLA class I molecules (P<0.01). Furthermore, multivariate analysis using Cox's proportional hazards model revealed that the downregulated expression of HLA class I in primary lesions was an independent, unfavourable prognostic factor (P<0.01). In conclusion, the downregulation of HLA class I expression frequently occurred in primary tumour and, to a greater extent, in metastatic lesions of patients with ESCC and was associated with patient survival.     

Lack of MSH2 and MSH6 characterizes endometrial but not colon carcinomas in hereditary nonpolyposis colorectal cancer

Hereditary nonpolyposis colorectal cancer syndrome is associated with an inherited predisposition to primarily colorectal cancer (CRC) and endometrial cancer (EC); however, the biological basis of the organ involvement remains unknown. As an attempt to explore whether the expression levels of MLH1, MSH2, and MSH6 may play a role, we used immunohistochemistry to study 42 ECs and 35 CRCs from patients carrying the same predisposing mutations. Among MSH2 mutation carriers, MLH1 was expressed in both tumor types, whereas MSH2 and, in many cases, also MSH6, were absent. Remarkably, among MLH1 mutation carriers, 54% of ECs (21 of 39), but none of the CRCs (0 of 32), lacked the MSH2 and/or MSH6 protein in addition to lacking MLH1 protein expression. These results demonstrate a marked difference between hereditary nonpolyposis colorectal cancer-related CRCs and ECs and suggest that the development of the latter tumors is selectively associated with the MSH2/MSH6 protein complex deficiency.     

Haplotype loss of HLA class I antigen as an escape mechanism from immune attack in lung cancer

One of tumor escape mechanisms from the host's immunosurveillance system (i.e., a haplotype loss of HLA class I antigens) has been detected in various tumor cells. We hypothesize that the majority of tumor cells with normal HLA class I expression were attacked and eradicated by CTLs, and only a minority with an abnormal expression of HLA class I antigens could escape the host's immunosurveillance system. Using HLA class I-transfected tumor variants as stimulators in A904L lung cancer cell line, which has a haplotype loss of HLA class I antigens, both the transfected HLA-A26 and HLA-B39-restricted CTL lines were induced from autologous lymphocytes. However, only one HLA-B39-restricted CTL clone (CTL G3b) was established, and it was then used to identify the antigen. SGT1B [suppressor of G2 allele of SKP1 (SGT1), suppressor of kinetochore protein (SKP1)] was identified as the antigen recognized by CTL G3b. Further experiments using 13 subclones from a primary culture of A904L were found to confirm our above-mentioned hypothesis. Tumor cells with a normal HLA class I expression may thus be killed by CTL at an early stage of carcinogenesis, and only tumor cells with a haplotype loss of HLA class I antigens can escape an immune attack and develop into clinical cancer.