Rise and fall of an anti-MUC1 specific antibody.

BACKGROUND: So far, human antibodies with good affinity and specificity for MUC1, a transmembrane protein overexpressed on breast cancers and ovarian carcinomas, and thus a promising target for therapy, were very difficult to generate. RESULTS: A human scFv antibody was isolated from an immune library derived from breast cancer patients immunised with MUC1. The anti-MUC1 scFv reacted with tumour cells in more than 80% of 228 tissue sections of mamma carcinoma samples, while showing very low reactivity with a large panel of non-tumour tissues. By mutagenesis and phage display, affinity of scFvs was increased up to 500fold to 5,7×10(-10) M. Half-life in serum was improved from below 1 day to more than 4 weeks and was correlated with the dimerisation tendency of the individual scFvs. The scFv bound to T47D and MCF-7 mammalian cancer cell lines were recloned into the scFv-Fc and IgG format resulting in decrease of affinity of one binder. The IgG variants with the highest affinity were tested in mouse xenograft models using MCF-7 and OVCAR tumour cells. However, the experiments showed no significant decrease in tumour growth or increase in the survival rates. To study the reasons for the failure of the xenograft experiments, ADCC was analysed in vitro using MCF-7 and OVCAR3 target cells, revealing a low ADCC, possibly due to internalisation, as detected for MCF-7 cells. CONCLUSIONS: Antibody phage display starting with immune libraries and followed by affinity maturation is a powerful strategy to generate high affinity human antibodies to difficult targets, in this case shown by the creation of a highly specific antibody with subnanomolar affinity to a very small epitope consisting of four amino acids. Despite these "best in class" binding parameters, the therapeutic success of this antibody was prevented by the target biology.

Improvement of survival in sex cord stromal tumors - an observational study with more than 25 years follow-up.

OBJECTIVE: To highlight aspects of malignant ovarian sex cord stromal tumors, effects of treatment, and developments over the past 28 years. DESIGN: Population-based cohort study. SETTING: Gynecological departments within the catchment-area of the Munich Cancer Registry and associated with the project group 'Malignant Ovarian Tumors' of the Munich Cancer Center. SAMPLE: One hundred and forty-five women with an invasive single sex cord stromal tumor diagnosed between 1978 and 2005. METHODS: Overall survival was estimated with the Kaplan-Meier method, relative survival was computed by the ratio of observed to expected survival rate. The impact of age, International Federation of Gynecology and Obstetrics (FIGO)-stage, residual tumor, and chemotherapy was examined by multivariate analysis (Cox regression model). MAIN OUTCOME MEASURES: Overall and relative survival and multivariate adjusted overall survival. RESULTS: Survival data showed a five-/10-year overall survival of 55.8%/42.8% (relative survival 58.6%/49.2%) for women diagnosed before 1988 and 89.1%/78.3% (relative survival 92.7%/85.2%) for women diagnosed after 1988. After adjustment for age and FIGO-stage, the following hazard ratios and 95% confidence intervals (95% CI) for treatment methods resulted: 3.3 (95% CI 1.5-7.0) for women with compared to women without residual tumor and 2.2 (95% CI 1.2-4.2) for women with chemotherapy compared to women where no chemotherapy was given. CONCLUSIONS: Improvements in survival may be attributed to a stage-shift toward more favorable stages at diagnosis and to advances in treatment such as improved surgery without residual tumor. There is no evidence for any benefit of adjuvant chemotherapy. Surgery remains the cornerstone of treatment, yet the benefit of postoperative therapy is still under debate.

Loss of Drop1 expression already at early tumor stages in a wide range of human carcinomas.

In a study on gene deregulation in ovarian carcinoma we found a mRNA coding for a 350 kDa protein, Drop1, to be downregulated 20- to 180-fold in the majority of ovarian and mammary carcinomas. The mRNA is encoded by a set of exons in the 5' region of the SYNE1 gene. Immunohistochemical staining for Drop1 protein by a specific monoclonal antibody corresponds to the pattern seen for the mRNA. cDNA arrays of matched pairs of tumor and normal tissue and in situ hybridizations confirmed the drastic loss of Drop1 mRNA as a common feature in uterus, cervix, kidney, lung, thyroid and pancreas carcinomas, already at early tumor stages and in all metastases. Two-hybrid studies suggest a role of this deficiency in the malignant progression of epithelial tumors.

Neoadjuvant letrozole in postmenopausal estrogen and/or progesterone receptor positive breast cancer: a phase IIb/III trial to investigate optimal duration of preoperative endocrine therapy.

BACKGROUND: In recent years, preoperative volume reduction of locally advanced breast cancers, resulting in higher rates of breast-conserving surgery (BCS), has become increasingly important also in postmenopausal women. Clinical interest has come to center on the third-generation nonsteroidal aromatase inhibitors (AIs), including letrozole, for such neoadjuvant endocrine treatment. This usually lasts 3-4 months and has been extended to up to 12 months, but optimal treatment duration has not been fully established. METHODS: This study was designed as a multicenter, open-label, single-arm, exploratory phase IIb/III clinical trial of letrozole 2.5 mg, one tablet daily, for 4-8 months. The primary objective was to investigate the effect of neoadjuvant treatment duration on tumor regression and BCS eligibility to identify optimal treatment duration. Tumor regression (by clinical examination, mammography, and ultrasound), shift towards BCS eligibility, and safety assessments were the main outcome measures. Standard parametric and nonparametric descriptive statistics were performed. RESULTS: Letrozole treatment was received by 32 of the enrolled 33 postmenopausal women (median (range): 67.0 (56-85) years) with unilateral, initially BCS-ineligible primary breast cancer (clinical stage > or = T2, N0, M0). Letrozole treatment duration in the modified intent-to-treat (ITT; required 4 months' letrozole treatment) analysis population (29 patients) was 4 months in 14 patients and > 4 months in 15 patients. The respective per-protocol (PP) subgroup sizes were 14 and 11. The majority of partial or complete responses were observed at 4 months, though some beneficial responses occurred during prolonged letrozole treatment. Compared with baseline, median tumor size in the ITT population was reduced by 62.5% at Month 4 and by 70.0% at final study visit (Individual End). Similarly, in the PP population, respective reductions were 64.0% and 67.0%. Whereas initially all patients were mastectomy candidates, letrozole treatment enabled BCS (lumpectomy) in 22 ITT (75.9%) and 18 PP (72.0%) patients. CONCLUSION: Over half of patients become BCS-eligible within 4 months of preoperative letrozole treatment. While prolonged treatment for up to 8 months can result in further tumor volume reduction in some patients, there is no clear optimum for treatment duration. Letrozole has a favorable overall safety and tolerability profile. TRIAL REGISTRATION: ClinicalTrials.gov identifier NCT00535418.

Antigen loading of dendritic cells with apoptotic tumor cell-preparations is superior to that using necrotic cells or tumor lysates.

BACKGROUND: Loading of dendritic cells (DCs) with tumor cell (TC) preparations is an attractive method for vaccine preparation because the entire antigen repertoire of a tumor is processed and presented by the DCs, thus allowing the simultaneous stimulation of T-helper cells and cytotoxic T-lymphocytes. However, optimal loading conditions have still to be defined. MATERIALS AND METHODS: DCs were pulsed either with tumor lysates, apoptotic or necrotic preparations of a breast cancer cell line and subsequently used to stimulate autologous T-lymphocytes. Antigen loading was quantified using immunofluorescent-based methods. RESULTS: Four hours co-incubation of apoptotic TCs or tumor lysates with DCs undergoing maturation resulted in effective DC-loading. However, the DCs pulsed with apoptotic TCs were best in stimulating interferon-gamma (INF-gamma) secretion as the effector function of autologous T-cells. CONCLUSION: Tumor lysates are in common use for DC-based vaccine manufacturing. However, our data indicate an advantage of apoptotic TC-preparations in regard to antigen loading effectiveness as well as the loaded DC's capacity to activate T-cells.

Reduced incidence of distant metastases and lower mortality in 1072 patients with breast cancer with a history of hormone replacement therapy.

OBJECTIVE: Substitution of estrogens (hormone replacement therapy [HRT]) is the most common therapy and prophylaxis of postmenopausal complaints. However, in most studies, long-term HRT has been associated with an increased risk for breast cancer, but the influence on a prognosis of breast cancer has been examined rarely. STUDY DESIGN: For further investigation, we analyzed 1072 patients aged 45-70 years at the time of first diagnosis of breast cancer with and without preoperative HRT with regard to the incidence of distant metastases and overall survival. Of these, 279 women were premenopausal (mean, 47.8 +/- 3.2 years); 793 women were postmenopausal (mean, 54.5 +/- 3.5 years); 320 women had received HRT over a minimum of 1 year (mean, 5.5 +/- 4.0 years; group HRT+); and 473 women had not received HRT (group HRT-). The median follow-up time was 73.2 months. RESULTS: Although body mass index, tumor size, and grading of group HRT- were significantly higher than in group HRT+, nodal status, S-phase fraction, hormone-receptor status, and local recurrence showed no significant differences. In regard to the incidence of distant metastases, women without HRT have significantly (P < .001) more metastases to bone (68 vs 20 women), lung (47:13 women), and liver (47:13 women). Overall survival was significantly lower in the HRT- group. CONCLUSION: We were able to show that the use of HRT before the diagnosis of breast cancer results in more favorable primary tumors, with a lower incidence of recurrences and a better overall survival rate. This might be due to normalized bone metabolism by the use of HRT, which may lower the conditions of tumor cell seeding.

Preeclampsia: increased expression of soluble ADAM 12.

Preeclampsia is a multisystemic pregnancy-associated disease affecting about 3-7% of pregnancies worldwide and is still a principal cause of fetal and maternal morbidity and mortality. To identify potential markers, we have compared gene expression profiles from control and preeclamptic placental tissues taken at various age-matched gestational stages using complementary DNA microarray analysis. Besides previously identified preeclampsia-associated genes, novel differentially expressed transcripts were found. The soluble form of the disintegrin metalloprotease ADAM 12 (a disintegrin and metalloproteinase 12; meltrin-alpha) represented the most upregulated transcript. This was confirmed by in situ hybridization of sections of preeclamptic placentas and by serum protein analysis of preeclamptic pregnant women. Thus, ADAM 12 could serve as an early biomarker for preeclampsia that may be of predictive and/or functional significance.

Long-term remission of excessive liver metastases in a breast cancer patient with chronic alcohol abuse using a monotherapy with trastuzumab.

We report on the successful treatment of a 43-year-old breast cancer patient with excessive liver metastases and chronic alcohol abuse. After first occurrence of hepatic metastases, systemic and interventional therapies were performed, and resulted in short-term partial remission. Finally, an excessive progression of the hepatic metastases was diagnosed. A systemic therapy with weekly trastuzumab (Herceptin) infusions was induced and a complete remission was achieved that is ongoing now for over 45 months.

Specifically activated memory T cell subsets from cancer patients recognize and reject xenotransplanted autologous tumors.

Bone marrow of breast cancer patients was found to contain CD8(+) T cells specific for peptides derived from breast cancer-associated proteins MUC1 and Her-2/neu. Most of these cells had a central or effector memory phenotype (CD45RA(-)CD62L(+) or CD45RA(-)CD62L(-), respectively). To test their in vivo function, we separated bone marrow-derived CD45RA(+) naive or CD45RA(-)CD45RO(+) memory T cells, stimulated them with autologous dendritic cells pulsed with tumor lysate, and transferred them into NOD/SCID mice bearing autologous breast tumors and normal skin transplants. CD45RA(-) memory but not CD45RA(+) naive T cells infiltrated autologous tumor but not skin tissues after the transfer. These tumor-infiltrating cells had a central or effector memory phenotype and produced perforin. Many of them expressed the P-selectin glycoprotein ligand 1 and were found around P-selectin(+) tumor endothelium. Tumor infiltration included cluster formation in tumor tissue by memory T cells with cotransferred dendritic cells. It was associated with the induction of tumor cell apoptosis and significant tumor reduction. We thus demonstrate selective homing of memory T cells to human tumors and suggest that tumor rejection is based on the recognition of tumor-associated antigens on tumor cells and dendritic cells by autologous specifically activated central and effector memory T cells.

DNA aneuploidy and integration of human papillomavirus type 16 e6/e7 oncogenes in intraepithelial neoplasia and invasive squamous cell carcinoma of the cervix uteri.

PURPOSE: Increasingly deregulated expression of the E6-E7 oncogenes of high-risk human papillomaviruses (HR-HPVs) has been identified as the major transforming factor in the pathogenesis of cervical dysplasia and derived cancers. The expression of these genes in epithelial stem cells first results in chromosomal instability and induces chromosomal aneuploidy. It is speculated that this subsequently favors integration of HR-HPV genomes into cellular chromosomes. This in turn leads to expression of viral cellular fusion transcripts and further enhanced expression of the E6-E7 oncoproteins. Chromosomal instability and aneuploidization thus seems to precede and favor integration of HR-HPV genomes. EXPERIMENTAL DESIGN: To prove this sequential concept, we analyzed here the sequence of events of DNA aneuploidization and integration in a series of HPV-16-positive cervical dysplastic lesions and carcinomas. Eighty-five punch biopsies of HPV-16-positive cervical lesions (20 CIN1/2, 50 CIN3, and 15 CxCa) were analyzed for DNA ploidy by DNA flow cytometry and for integration of HPV E6/E7 oncogenes using the amplification of papillomavirus oncogene transcripts assay, a reverse transcription-PCR method to detect integrate-derived human papillomavirus oncogene transcripts. RESULTS: DNA aneuploidy and viral genome integration were both associated with increasing dysplasia (P < 0.001, chi(2) test for trend). In addition, DNA aneuploidy was associated with increased viral integration (P < 0.01, Fisher's exact test). Nineteen of 20 (95%) lesions with integrated viral genomes had aneuploid cell lines; however, only 19 of 32 (59%) lesions with aneuploid cell lines had integrated viral genomes. CONCLUSIONS: These data support the hypothesis that aneuploidization precedes integration of HR-HPV genomes in the progression of cervical dysplasia. Accordingly, deregulated viral oncogene expression appears to result first in chromosomal instability and aneuploidization and is subsequently followed by integration of HR-HPV genomes in the affected cell clones.

Only grading has independent impact on breast cancer survival after adjustment for pathological response to preoperative chemotherapy.

Our objective was to determine pretreatment factors with an independent impact on survival after adjusting for response to preoperative chemotherapy and to describe parameters predictive for achieving a pathological complete remission (pCR) after preoperative chemotherapy containing an anthracycline. We performed univariate and multivariate analyses to describe the impact of the following pretreatment characteristics of 240 primary breast cancer patients who received preoperative chemotherapy containing an anthracycline at our institution on disease-free survival (DFS), distant disease-free survival (DDFS) and overall survival (OS): age, stage, clinical tumor size, clinical nodal status, grading, and expression of estrogen receptor, progesterone receptor, Her2/neu, Ki67, Bcl-2 and p53. Afterwards, the response to preoperative chemotherapy was added to the multivariate model in order to evaluate which pretreatment parameters retained their prognostic impact. In addition, univariate analysis was performed to describe pretreatment variables predictive for achieving a pCR. With a median follow-up of 6.4 years (range 0-10.4), only grading retained its independent impact on DFS, DDFS and OS [hazard ratio (HR) 1.5, 1.7 and 2.9, respectively; p<0.05] after adjusting for the strongest independent prognostic factors pathological T category at surgery (HR 1.6, 1.8 and 1.7, respectively; p<0.001) and pathological N category at surgery (HR 2.3, 2.4 and 2.1, respectively; p<0.001). Predictive factors for the achievement of pCR (p<0.05) were age under 35 years, lower stage or smaller clinical tumor size and higher expression of Bcl-2 at diagnosis. We conclude that only grading retained its independent prognostic impact on DFS, DDFS and OS after adjusting for pathological response of breast tumor and axillary lymph node metastases to preoperative chemotherapy. According to our data, it could be hypothesized that young patients with early tumor stage and small primary tumors might profit most from preoperative chemotherapy.

Induction of remission in a patient with metastatic breast cancer refractory to trastuzumab and chemotherapy following treatment with gefitinib ('Iressa', ZD1839).

Despite new therapies and several treatment options, metastatic breast cancer (MBC) remains incurable. One reason for the low median survival rate may be intense cross-talk between growth factor receptors such as the epidermal growth factor receptor (EGFR/HER1) and the HER2 growth factor receptor. This report describes the case history of a patient with MBC whose disease had progressed despite surgery, radiotherapy and four different chemotherapy regimens, including trastuzumab (a monoclonal antibody that specifically blocks HER2) combined with docetaxel. However, treatment with 500 mg/day gefitinib ('Iressa', ZD1839), an EGFR tyrosine kinase inhibitor, and trastuzumab (2 mg/kg/week) caused a rapid and sustained regression of breast cancer metastases in skin and lymph nodes. Thus, for patients with MBC whose tumors co-express EGFR and HER2, gefitinib in combination with trastuzumab may prevent receptor cross-talk, improving the outcome of MBC.

Her-2/neu gene amplification and protein expression in primary male breast cancer.

The role of HER-2/ neu in male breast cancer is not well defined. The purpose of the current study was to measure the frequency of HER-2/ neu expression in primary male breast cancer, to demonstrate HER-2/ neu gene amplification in cases found to be positive for protein overexpression, and to correlate HER-2/ neu positivity with clinicopathological variables. Formalin-fixed, paraffin-embedded archival material from 99 primary male breast carcinomas was evaluated by immunohistochemistry (IHC) using the HercepTest (DAKO Corp., Hamburg, Germany). Scoring was performed according to established guidelines. All cases demonstrating HER-2/ neu staining by IHC (1+/2+ and 3+) were analyzed for HER-2/ neu gene amplification by fluorescence in situ hybridization (FISH) utilizing the PathVysion assay (Vysis Corp., Downers Grove, Illinois) to assess HER-2/ neu amplification status. The immunohistochemical staining of the HER-2/ neu protein revealed HER-2/ neu positivity in 15/99 (15.1%) cases, eight tumors showed 2+ and 7 tumors 3+ staining. HER-2/ neu gene amplification was observed in 11/99 cases (11,1%), and all of the 3+ and 4/8 from the 2+ cases were amplified. HER-2/ neu gene amplification/protein overexpression did not correlate with tumor state, histological grade or estrogen/progesterone receptor status nor the axillary lymph node status. This is the first comprehensive study of HER-2/ neu gene amplification by FISH analysis in primary male breast cancer. Compared to female primary breast cancer the percentage of HER-2/ neu positivity in our study was lower. Our data provide first evidence for HER-2/ neu gene amplification in male breast cancer. Further studies should be addressed on the potential application of the monoclonal rhuMAB HER-2/ neu antibody for treatment of HER-2/ neu positive male breast cancer.

Neoadjuvant therapy with gemcitabine in breast cancer.

Primary systemic therapy (ie, preoperative or neoadjuvant) increases the possibility for breast-conserving surgery in patients with primary breast cancer. Patients with pathologic complete response to primary systemic therapy have improved survival compared with those with persistent tumors. Several phase II trials have evaluated gemcitabine-containing doublet or triplet regimens as primary systemic therapy for breast cancer, results of which have shown promising clinical and pathologic response rates with manageable toxicity. Results of a phase I/II study of gemcitabine (Gemzar)/epirubicin (Ellence)/docetaxel (Taxotere), or GEDoc, with prophylactic filgrastim (Neupogen), as primary systemic therapy in 77 evaluable patients with primary breast cancer are reported herein. Dose-limiting toxicities were grade 3 febrile neutropenia (n = 1) and grade 3 diarrhea (n = 2) at the fourth dose level of GEDoc tested (gemcitabine at 800 mg/m2 days 1 and 8, epirubicin at 90 mg/m2 day 1, and docetaxel at 75 mg/m2 day 1). As assessed by ultrasound, 92% of patients responded overall (22% complete response), and 79% of patients could undergo breast-conserving surgery. The pathologic complete response rate in resected breast tissue was 26%.

Serum levels of soluble E-selectin are associated with the clinical course of metastatic disease in patients with liver metastases from breast cancer.

Increasing evidence indicates that the expression of the endothelial adhesion molecule E-selectin is associated with progression and metastasis of breast cancer. Patients with liver metastases also show increased serum levels of the soluble form of E-selectin. It was our aim to compare serum levels of soluble E-selectin (sES) in such patients with the biology of the primary tumor and the course of the metastatic disease under therapy. We examined 69 patients with liver metastases from breast cancer who were selected to receive systemic tumor therapy because of progressive disease (n = 44) or newly detected liver metastases (n = 25). Serum concentrations of sES were measured before each therapy cycle using a specific ELISA. Serum concentrations of sES before the start of therapy were compared to clinical parameters and histopathological findings referring to the primary tumor. Secondly, serum levels of sES were compared to serum concentrations of the corresponding tumor markers. We observed a possible trend for certain unfavorable prognostic parameters (e.g., young women, low-graded tumors, human epidermal growth factor receptor 2 overexpression) to be related to higher serum levels of sES. Serum levels of sES were correlated with tumor marker levels in a logarithmical relation (r = 0.44, P < 0.0005). In some cases it could be demonstrated that serum levels of sES changed similarly to the course of tumor marker levels. We conclude that serum levels of sES are associated with the clinical course of liver metastases from breast cancer. Further investigations are needed to clarify if serum levels of sES may serve as tumor marker in certain clinical situations. E-selectin should be evaluated as a possible target for antimetastatic therapy studies.

Raw and cooked vegetables, fruits, selected micronutrients, and breast cancer risk: a case-control study in Germany.

In 1998-2000, a case-control study of breast cancer was conducted in Heidelberg, Germany. Three hundred ten consecutively recruited cases with primary breast cancer were matched according to 10-yr age groups to 353 controls with conditions unrelated to diet or endocrine disorders. Intake of raw vegetables, total vegetables, and whole-grain products was inversely associated with breast cancer risk (highest vs. lowest quartile adjusted odds ratio [OR] 0.51, 95% confidence interval [CI] 0.31-0.84; OR = 0.62, 95% CI = 0.38-1.02; and OR = 0.57; 95% CI = 0.34-0.95, respectively). Also, high intake of some selected vitamins and minerals possessing putative DNA-stabilizing properties displayed significant inverse risk associations. Adjusted ORs were as follows: vitamin C (OR = 0.49, 95% CI = 0.2-0.88), folate equivalents (OR = 0.47, 95% CI = 0.25-0.88), b-carotene (OR = 0.46, 95% CI = 0.27-0.80), zinc (OR = 0.35, 95% CI = 0.15-0.78), and copper (OR = 0.51, 95% CI = 0.31-1.03). In contrast, no significant association with risk was seen for an increased intake of fruits, cooked vegetables, fiber, calcium, manganese, or iron. In this population of German women, components of raw vegetables and some micronutrients appear to decrease breast cancer risk.

Breast cancer in young women (< or = 35 years): Genomic aberrations detected by comparative genomic hybridization.

Sporadic breast cancer in young women is different from the one in older patients regarding pathological features and aggressiveness of the tumors, but the spectrum of genetic alterations are largely unknown. We used comparative genomic hybridization (CGH) to analyze DNA copy number changes in 88 tumor samples from women

Centrosomal aberrations in primary invasive breast cancer are associated with nodal status and hormone receptor expression.

Our purpose was to assess the presence of centrosomal aberrations as measured by immunohistochemistry in primary invasive breast cancer and their association with established and proposed prognostic factors. Tissue sections of 103 primary invasive breast cancers were examined using centrosome-specific antibodies to pericentrin and gamma-tubulin. At least 3 different tumor regions per case were examined to determine maximum centrosomal aberration levels, which represent the proportion of cells with abnormal centrosomes in the region with the highest percentage of cells with centrosomal aberrations. The chi(2) test was performed to evaluate the association of maximum centrosomal aberration levels with patient age; tumor size; nodal status; nuclear grade; hormone receptor and Her2/neu expression; proportion of Ki67-, p53- and Bcl-2-positive tumor cells; DNA index; S-phase fraction; and proliferation index. With pericentrin immunohistochemistry, maximum centrosomal aberration levels >35% were detectable in 92 of the 103 breast carcinomas (89%). We found a highly significant correlation of maximum centrosomal aberration levels above 35% with axillary nodal tumor involvement (p < 0.0001) and the absence of hormone receptors (p < 0.0001). In addition, there was a borderline significant relationship with age <50 years (p = 0.050) and Her2/neu overexpression (p = 0.050). Among node-negative patients, maximum centrosomal aberration levels >35% were also associated with an increased DNA index (p = 0.006). In a subset of patients, additional staining of centrosomes with a monoclonal anti-gamma-tubulin antibody essentially confirmed these results. In primary invasive breast cancer, centrosomal aberrations are associated with those factors predicting a more aggressive course of disease. This might indicate a fundamental role of centrosomal dysfunction in disease evolution, possibly as a result of chromosome missegregation during mitosis.

Similar contributions of BRCA1 and BRCA2 germline mutations to early-onset breast cancer in Germany.

This study was undertaken to investigate the prevalence of BRCA1 and BRCA2 germline mutations in 91 German patients unselected for family history, who were diagnosed with breast cancer before the age of 41 years. Clinical information and blood samples were obtained from all patients. A comprehensive BRCA1 and BRCA2 mutational analysis was performed using the protein truncation assay and single-strand conformational polymorphism analysis followed by DNA sequencing of variant signals detected by these assays. Five different deleterious germline mutations including four frameshift mutations and one missense mutation were identified, three in BRCA1 (3.3%) and two mutations (2.2%) in BRCA2. Both BRCA2 mutations are novel and might be specific for the German population. An additional BRCA1 missense mutation previously described and classified as an unknown variant was found. This mutation was also detected in two breast cancer patients of family P 328 and not in 140 healthy controls suggesting that it is disease associated. In addition, one common polymorphism and five novel intronic sequence variants with unknown significance were found. Our findings show that mutations in BRCA1 and BRCA2 may contribute similarly to early-onset breast cancer in Germany. Given current constraints on health-care resources, these results support the notion that BRCA1 and BRCA2 mutation screening may have the strongest impact on health-care when targeted to high-risk populations.

Immunocytochemical detection of HPV high-risk type L1 capsid proteins in LSIL and HSIL as compared with detection of HPV L1 DNA.

OBJECTIVE: To investigate the prevalence of HPV L1 capsid proteins in HPV-infected HSIL and LSIL. STUDY DESIGN: Cervical smears from 74 women with cytologically and histologically confirmed LSIL (n = 32) and HSIL (n = 42) were collected prospectively to detect HPV high-risk (hr) types 16, 18, 33, 35, 39, 45, 56 and 58 L1-DNA by standardized L1-consensus primer PCR (MY 09/11) and L1 capsid proteins by immunocytochemistry using monoclonal antibodies T31 (HPV16) and T16 (HPV hr) in a standardized protocol. RESULTS: In HSIL and LSIL, L1 DNA was found for HPV hr in 93% and 59% and for HPV16 in 69% and 37% of the specimens, respectively. HPV L1 capsid proteins were detected in HSIL and LSIL for HPV hr in 33% and 44% and for HPV16 in 29% and 31% of the specimens, respectively. Expression of L1 capsid proteins was significantly reduced, by 59.6% for HPV hr L1 DNA-positive HSIL (P < .01) and by 40.4% for HPV 16 L1 DNA-positive HSIL (P < .01). In HPV 16 DNA-positive and HPV hr DNA-positive LSIL, no significant reduction of corresponding L1 capsid protein expression could be demonstrated. CONCLUSION: These data suggest a disturbed viral cellular interaction in HPV 16 and HPV hr-infected HSIL, with loss of viral L1 capsid antigen. In this context there is a possible role of T31 and T16 as prognostic markers to predict the prognosis of CIN.