Co‐expression of TLR‐9 and MMP‐13 is associated with the degree of tumour differentiation in prostate cancer

In vitro experiments demonstrated that stimulation of Toll‐like receptor 9 (TLR‐9) by synthetic TLR‐9 ligands induces the invasion of TLR‐9‐expressing prostate cancer cells through matrix metalloproteinase 13 (MMP‐13). However, the clinical value of TLR‐9 and MMP‐13 co‐expression in the pathophysiology of the prostate is unknown. In the study, we evaluated the expression levels and clinical significance of the TLR‐9 and MMP‐13 in a series of prostate tissues. One hundred and eighty prostate tissues including prostate cancer (PCa) (n = 137), high‐grade prostatic intraepithelial neoplasia (HPIN) (n = 18) and benign prostatic hyperplasia (BPH) (n = 25) were immunostained for the TLR‐9 and MMP‐13 markers. Subsequently, the correlation between the TLR‐9 and MMP‐13 staining scores and clinicopathological parameters was obtained. Higher expressions of TLR‐9 and MMP‐13 were found in PCa and high‐grade prostatic intraepithelial neoplasia compared to benign prostatic hyperplasia tissues. Among PCa samples, a positive relationship was revealed between the MMP‐13 expression and Gleason score (P < 0.001). There was a significant correlation between TLR‐9 expression and regional lymph node involvement (P = 0.04). The expression patterns of TLR‐9 and MMP‐13 markers demonstrated a reciprocal significant correlation between the two markers in the same series of prostate samples (P < 0.001). Furthermore, the Gleason score of TLR‐9^high/MMP‐13^high and TLR‐9^low/MMP‐13^low phenotypes showed a significant difference (P = 0.002). Higher expressions of TLR‐9 and MMP‐13 can confer aggressive behaviour to PCa. Therefore, these markers may be used as a valuable target for tailored therapy of PCa.     

Androgen receptor expression in breast cancer patients tested for BRCA1 and BRCA2 mutations

Pristauz G, Petru E, Stacher E, Geigl J B, Schwarzbraun T, Tsybrovskyy O, Winter R & Moinfar F
(2010) Histopathology 57, 877–884 Androgen receptor expression in breast cancer patients tested for BRCA1 and BRCA2 mutations Aim: To assess the expression of receptors for androgen (AR), oestrogen (ER) and progesterone (PR) as well as human epidermal growth factor receptor type 2 (Her‐2/neu) status of breast carcinomas in breast cancer susceptibility gene (BRCA) BRCA1/2 mutation carriers and BRCA1/2 negative tested women. Methods: One hundred and thirty‐five breast cancers in women tested for BRCA1/2 mutations. Screening for BRCA1 and BRCA2 mutations was performed by direct sequencing of all BRCA1 and BRCA2 exons as well as the surrounding intronic sequences. Additionally, BRCA genes were analysed with multiplex ligation‐dependent probe amplification. Consecutive paraffin sections were examined immunhistochemically for AR, ER, PR and Her‐2/neu. Results: Of the 135 tumours, 43 (32%) were BRCA1‐related, 18 (13%) were BRCA2‐related and 74 (55%) were BRCA1/2‐negative. Seventy‐two per cent of the BRCA1‐related, 22% of the BRCA2‐related and 12% of the BRCA1/2‐negative tumours were triple (ER, PR, Her2neu)‐negative. Eighty‐four per cent of BRCA1 mutated cancers were high‐grade (G3) tumours. ARs were expressed in 30% (13 of 43) of BRCA1‐related, in 78% (14 of 18) in BRCA2‐related tumours and in 76% (56 of 74) in BRCA1/2 negative tumours. Twenty‐one per cent of ER‐negative BRCA1‐related tumours expressed androgen receptors. Conclusion: Approximately one in five BRCA1 mutated breast cancers negative for ER and PR express androgen receptors. Modulation of AR might open a new avenue for treating these high‐risk cancers.     

Does family history predict the age at onset of new breast cancers in BRCA1 and BRCA2 mutation‐positive families?

Panchal S, Bordeleau L, Poll A, Llacuachaqui M, Shachar O, Ainsworth P, Armel S, Eisen A, Sun P, Narod SA. Does family history predict the age at onset of new breast cancers in BRCA1 and BRCA2 mutation‐positive families? Women who carry BRCA mutations are advised to begin breast cancer screening based on the age‐specific risks of breast cancer development. It is not clear to what extent the family history of breast cancer influences age of onset. We evaluated the use of family history to predict the age of breast cancer onset in BRCA mutation carriers. Pedigrees from an Ontario‐based registry were reviewed to identify the index case of breast cancer (most recent diagnosis) and other family cases of breast cancer. The youngest age of breast cancer diagnosis and mean age at breast cancer diagnosis in the other family cases were compared to the age of onset in the index case. The 260 BRCA1 and 213 BRCA2 pedigrees were reviewed. In BRCA2 families, the index case was diagnosed on average at 44.4 years when the youngest reported family case was less than or equal to 35 years, compared to 51.9 years when the earliest cases were diagnosed after age 50 (p = 0.04). A modest trend was seen for BRCA1 carriers, but this was not statistically significant. To a small extent, the onset of breast cancer in a BRCA2 mutation carrier can be predicted from her family history of cancer, however, the trend is modest and should not alter clinical recommendations regarding initiation of screening.     

The risk of breast cancer in BRCA1 and BRCA2 mutation carriers without a first‐degree relative with breast cancer

The objective of this study was to estimate the lifetime risk of breast cancer in women with a BRCA1 or BRCA2 mutation with and without at least 1 first‐degree relative with breast cancer. A total of 2835 women with a BRCA1 or BRCA2 mutation were followed. Age‐ and gene‐specific breast cancer rates were calculated. The relative risks of breast cancer for subjects with a family history of breast cancer, compared to no family history were calculated. The mean age at baseline was 41.1 years, and they were followed for a mean of 6.0 years. The estimated penetrance of breast cancer to age 80 years was 60.8% for BRCA1 and 63.1% for BRCA2. For all BRCA carriers, the penetrance of breast cancer to age 80 for those with no first‐degree relative with breast cancer was 60.4% and 63.3% for those with at least 1 first‐degree relative with breast cancer. The risk of breast cancer for BRCA carriers with no first‐degree relative with breast cancer is substantial, and as a result, clinical management for these women should be the same as those for women with an affected relative.     

Prevalence of founder mutations in the BRCA1 and BRCA2 genes among unaffected women from the Bahamas

Population‐based testing for BRCA1/2 mutations detects a high proportion of carriers not identified by cancer family history‐based testing. We sought to determine whether population‐based testing is an effective approach to genetic testing in the Bahamas, where 23% of women with breast cancer carry one of seven founder mutations in the BRCA1 or BRCA2 gene. We determined the prevalence of founder BRCA mutations in 1847 Bahamian women without a personal history of breast or ovarian cancer, unselected for age or family history. We found that 2.8% (20/705) of unaffected women with a family history of breast/ovarian cancer and 0.09% (1/1089) of unaffected women without a family history carry a BRCA mutation. A total of 38% of unaffected women with a known mutation in the family were found to carry the familial mutation. We previously suggested that all Bahamian women with breast or ovarian cancer be offered genetic testing. These current data suggest that additionally all unaffected Bahamian women with a family history of breast/ovarian cancer should be offered genetic testing for the founder BRCA mutations.     

Reelin expression in human prostate cancer: a marker of tumor aggressiveness based on correlation with grade.

Reelin is a glycoprotein that plays a critical role in the regulation of neuronal migration during brain development and, since reelin has a role in the control of cell migration, it might represents an important factor in cancer pathology. In this study, 66 surgical specimens of prostate cancer were analyzed for reelin expression by immunohistochemical method. The reelin expression was correlated with Gleason score and individual Gleason patterns. Reelin expression was found in 39% prostate cancers. Stromal tissues, normal epithelial cells and prostate intraepithelial neoplasia (PIN) of any grade around and distant from cancer were always negative for reelin. Reelin was found in malignant prostatic epithelial glands of 50% cases Gleason score 10, 52% Gleason score 9, 56% Gleason score 8, 18% Gleason score 7, while no sample of prostate cancers with Gleason score 6 showed reelin expression (P=0,005). As reelin staining is frequently found in high Gleason score prostate cancers, we explored whether reelin expression is influenced by single Gleason patterns. While Gleason 3 pattern did not show reelin immunoreactivity, reelin expression was found in 35% Gleason 4 patterns and 45% Gleason 5 patterns (P<0.001). Our results demonstrated for the first time that reelin is expressed in prostate cancer and not in benign prostate tissue and its expression occurs in higher Gleason score and correlates significantly with increasing of single Gleason patterns. This suggests reelin may behave as a specific histological marker and may represent a useful biomarker to predict aggressive phenotypic behavior of prostatic cancer cells.     

The length of positive surgical margins correlates with biochemical recurrence after radical prostatectomy

van Oort I M, Bruins H M, Kiemeney L A L M, Knipscheer B C, Witjes J A & Hulsbergen‐van de Kaa C A
(2010) Histopathology 56 , 464–471 The length of positive surgical margins correlates with biochemical recurrence after radical prostatectomy Aims: To evaluate the prognostic role of the length of a positive surgical margin (+SM) for biochemical recurrence (BCR) after radical prostatectomy (RP) for prostatic cancer. Methods and results: Consecutive RP specimens (n = 267) with +SM were analysed. All RP specimens were sectioned at 4‐mm intervals and completely embedded. Data were analysed using Kaplan–Meier survival analysis and proportional hazards models. In 267 patients the length of +SM ranged from 0.4 to 174.5 mm (median 11.2 mm) and correlated with preoperative prostate specific antigen (PSA) (P < 0.001), pathological stage (P < 0.001), tumour volume (P = 0.001), number of +SM (P < 0.001), Gleason grade at +SM (P < 0.001) and Gleason score (P = 0.015). Patients with detectable postoperative PSA levels (n = 34) or adjuvant therapy (n = 59) were excluded from BCR analysis. In the remaining 174 patients the 5‐year risk of BCR was 29%; in patients with +SM ≤10 mm and >10 mm this was 21% and 39%, respectively. On multivariable analysis BCR was associated with an increasing length of +SM (≤10 mm versus >10 mm; hazard ratio 2.15; 95% confidence interval 1.12, 4.15; P = 0.022). Conclusions: The length of +SM is an independent prognostic factor for BCR in patients with undetectable PSA after RP.     

Oestrogen‐mediated phosphorylation and stabilization of BRCA2 protein in breast

Disease‐associated BRCA2 mutations typically result in protein truncations that delete the phosphorylation‐regulated S3291 BRCA2 domain that interacts with Rad51. BRCA2 hereditary breast cancers are usually ER⁺, differing from BRCA1 hereditary cancers, which are usually ER^?. We studied BRCA2 protein expression and S3291 phosphorylation in normal breast tissues and in sporadic breast cancers and observed that BRCA2 is expressed and phosphorylated in normal breast and 10 ER⁺ breast cancers but not in 10 ER^? breast cancers. In order to study this correlation between ER and BRCA2 expression, we studied ER⁺ breast cancer cell lines. We found that a rapid increase in BRCA2 S3291 phosphorylation occurs following 17‐β‐oestradiol (E2) treatment. This increase seen in BRCA2 total and phospho‐S3291 protein levels was found to be unaffected with cycloheximide pre‐treatment, but decreased following tamoxifen, ICI 182,780 or roscovitine treatment. This suggests a requirement for ER and cdk (cyclin‐dependent kinase) in mediating the increased protein levels. MCF7 cell cycle distribution analysis following E2, in both the presence and absence of roscovitine (a cdk inhibitor), did not demonstrate any changes during an 8 h period, which further supports our hypothesis that mitogenic effects of E2 are not predominant at early time points. Studies with MG132 proteasome inhibitor and siRNA to skp2 support a model in which skp2‐mediated proteasomal degradation of BRCA2 rapidly degrades BRCA2 protein in the absence of hormone treatment, which likely inhibits this pathway. E2 was shown to improve survival of MCF7 cells upon radiation treatment and roscovitine partially reversed this effect. We have demonstrated that BRCA2 protein is specifically expressed in ER⁺ breast cancers and are investigating a pathway that may show a link between E2 action and BRCA2 protein function in breast cancer. Copyright © 2008 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Hereditary breast cancer: syndromes,tumour pathology and molecular testing

Hereditary factors account for a significant proportion of breast cancer risk. Approximately 20% of hereditary breast cancers are attributable to pathogenic variants in the highly penetrant BRCA1 and BRCA2 genes. A proportion of the genetic risk is also explained by pathogenic variants in other breast cancer susceptibility genes, including ATM, CHEK2, PALB2, RAD51C, RAD51D and BARD1, as well as genes associated with breast cancer predisposition syndromes – TP53 (Li–Fraumeni syndrome), PTEN (Cowden syndrome), CDH1 (hereditary diffuse gastric cancer), STK11 (Peutz–Jeghers syndrome) and NF1 (neurofibromatosis type 1). Polygenic risk, the cumulative risk from carrying multiple low-penetrance breast cancer susceptibility alleles, is also a well-recognised contributor to risk. This review provides an overview of the established breast cancer susceptibility genes as well as breast cancer predisposition syndromes, highlights distinct genotype–phenotype correlations associated with germline mutation status and discusses molecular testing and therapeutic implications in the context of hereditary breast cancer.     

BARD1 homozygous deletion,a possible alternative to BRCA1 mutation in basal breast cancer

Hereditary breast cancers (BCs) are incompletely explained by BRCA genes abnormalities, as ～70% of them are not associated with known genetic alterations. Array‐based comparative genomic hybridization (aCGH) of tumors provides an opportunity for identifying new BC susceptibility genes. By analyzing our database of high‐resolution aCGH profiles of 330 BCs, we identified a case with homozygous deletion of the entire BARD1 gene. The BARD1‐deleted case displayed a familial history of BC and other clinico‐pathological features of BRCAness, and a 17% probability of BRCA1/2 mutation. Analysis of constitutional DNA showed a BARD1 germline heterozygous deletion without BRCA1/2 mutation. Gene expression analysis using DNA microarrays classified the tumor as basal‐like, with very low BARD1 and ID4 expression, but high expression of BRCA1, RAD51, PARP1, CHEK1, and FANCA. The tumor displayed a BRCA1‐mutated expression profile. This is the first report of a non‐BRCA1/2‐mutated BC with somatic homozygous and germ‐line heterozygous deletion of the entire BARD1 gene. This observation suggests that BARD1 might be a BC susceptibility gene that follows the Knudson rule. Identification of BARD1 deletion could have clinical applications including screening for hereditary forms. © 2010 Wiley‐Liss, Inc.     

Prevalence of BRCA1 and BRCA2 mutations in unselected breast cancer patients from Peru

The prevalence of BRCA1 and BRCA2 mutations among breast cancer patients in Peru has not yet been explored. We enrolled 266 women with breast cancer from a National cancer hospital in Lima, Peru, unselected for age or family history. DNA was screened with a panel of 114 recurrent Hispanic BRCA mutations (HISPANEL). Among the 266 cases, 13 deleterious mutations were identified (11 in BRCA1 and 2 in BRCA2), representing 5% of the total. The average age of breast cancer in the mutation‐positive cases was 44 years. BRCA1 185delAG represented 7 of 11 mutations in BRCA1. Other mutations detected in BRCA1 included: two 2080delA, one 943ins10, and one 3878delTA. The BRCA2 3036del4 mutation was seen in two patients. Given the relatively low cost of the HISPANEL test, one should consider offering this test to all Peruvian women with breast or ovarian cancer.     

Retracted: Promoter Methylation of the RASSF1A Gene may Contribute to Colorectal Cancer Susceptibility: A Meta‐Analysis of Cohort Studies

This meta‐analysis of published cohort studies was conducted to evaluate whether promoter methylation of the RASSF1A gene contributes to colorectal cancer (CRC) susceptibility. A range of electronic databases were searched without language restrictions. Meta‐analysis was conducted using the STATA 12.0 software. Crude risk differences (RD) with their 95% confidence intervals (95%CI) were calculated. In this meta‐analysis, 11 clinical cohort studies with a total of 630 CRC patients were included. The pooled results revealed that the frequency of RASSF1A gene methylation in cancer tissues was significantly higher than that in benign, adjacent, and normal tissues (cancer tissues vs. benign tissues: RD = 0.25, 95%CI = 0.13–0.38, P < 0.001; cancer tissues vs. adjacent tissues: RD = 0.32, 95%CI: 0.20–0.45, P < 0.001; cancer tissues vs. normal tissues: RD = 0.38, 95%CI: 0.26–0.50, P < 0.001; respectively). Subgroup analysis by ethnicity demonstrated that RASSF1A promoter methylation also exhibited a higher frequency in cancer tissues among both Asians and Caucasians (all P < 0.05). Our meta‐analysis has shown positive correlations between RASSF1A promoter methylation and CRC susceptibility. Thus, detection of RASSF1A promoter methylation may be utilized as a valuable diagnostic marker for CRC.     

Resistance to therapy in BRCA2 mutant cells due to loss of the nucleosome remodeling factor CHD4

Hereditary cancers derive from gene defects that often compromise DNA repair. Thus, BRCA-associated cancers are sensitive to DNA-damaging agents such as cisplatin. The efficacy of cisplatin is limited, however, by the development of resistance. One cisplatin resistance mechanism is restoration of homologous recombination (HR), which can result from BRCA reversion mutations. However, in BRCA2 mutant cancers, cisplatin resistance can occur independently of restored HR by a mechanism that remains unknown. Here we performed a genome-wide shRNA screen and found that loss of the nucleosome remodeling factor CHD4 confers cisplatin resistance. Restoration of cisplatin resistance is independent of HR but correlates with restored cell cycle progression, reduced chromosomal aberrations, and enhanced DNA damage tolerance. Suggesting clinical relevance, cisplatin-resistant clones lacking genetic reversion of BRCA2 show de novo loss of CHD4 expression in vitro. Moreover, BRCA2 mutant ovarian cancers with reduced CHD4 expression significantly correlate with shorter progression-free survival and shorter overall survival. Collectively, our findings indicate that CHD4 modulates therapeutic response in BRCA2 mutant cancer cells.     

The significance of strong histone deacetylase 1 expression in the progression of prostate cancer

Song Y H, Shiota M, Tamiya S, Kuroiwa K, Naito S & Tsuneyoshi M
(2011) Histopathology 58 , 773–780 The significance of strong histone deacetylase 1 expression in the progression of prostate cancer Aims: Histone deacetylases (HDACs) play important roles in many types of cancer. Recently, it has been reported that HDAC1 expression in prostate cancer is significantly higher than in benign prostate cell lines and tissues. The expression of HDAC1 in association with the clinicopathological data was investigated to define its functional and pathological roles in prostate cancer. Methods and results: HDAC1 expression was examined immunohistochemically in 148 patients with prostate cancer. Strong expression of HDAC1 in benign prostate glands, high‐grade prostatic intraepithelial neoplasia (PIN) and prostate cancer was observed in 17/148 (11%), 19/71 (27%) and 69/148 (47%) patients. Strong HDAC1 expression was correlated with high Gleason score (P = 0.025) and high pT stage (P = 0.012). Patients with strong HDAC1 expression had higher biochemical recurrence rates (P = 0.0010). Furthermore, strong HDAC1 expression had a significant impact on patient biochemical recurrence rates in multivariate analysis (P = 0.004). Conclusions: These results indicate that overexpression of HDAC1 contributes to progression and poor prognosis in prostate cancer. The findings may play an important role in the emergence of effective new approaches for therapy and prognostic markers of prostate cancer.     

Methylation profiles of hereditary and sporadic ovarian cancer

Bol G M, Suijkerbuijk K P M, Bart J, Vooijs M, van der Wall E & van Diest P J
(2010) Histopathology 57, 363–370
Methylation profiles of hereditary and sporadic ovarian cancer Aims: Tumour suppressor gene silencing through promoter hypermethylation plays an important role in oncogenesis. Carcinogenesis of hereditary cancers usually differs from that of their sporadic counterparts, but methylation has hardly been studied in hereditary ovarian cancer. The aim of this study was to investigate promoter methylation of a set of common tumour suppressor genes in BRCA1‐related ovarian cancer in comparison with sporadic ovarian cancer. Methods and results: Methylation‐specific multiplex ligation‐dependent probe amplification was used to assess the extent of promoter methylation of 24 different tumour suppressor genes in BRCA1‐associated (n = 25) and matched sporadic ovarian tumours (n = 50). A cumulative methylation index (CMI) was calculated and differences between individual genes were analysed. There was no significant difference in cumulative methylation between BRCA1‐associated and sporadic ovarian carcinomas (median CMI 108; CMI 110; P = 0.86). Also, methylation patterns of individual genes did not show distinct differences after correction for multiple comparisons. CDH13, GSTP1 and RASSF1 were frequently methylated in both sporadic and hereditary ovarian cancers. BRCA1 methylation occurred in 14% of sporadic tumours, but was not detected in BRCA1‐associated tumours. Conclusions: CDH13, GSTP1 and RASSF1 are frequently methylated in both sporadic and BRCA1‐associated ovarian cancers. Interestingly, methylation of BRCA1, while frequent in sporadic ovarian cancer, never occurred in the hereditary group. BRCA1‐associated ovarian cancers mimic their sporadic counterparts in extent and pattern of promoter methylation of several common tumour suppressor genes. This finding could have implications for future chemotherapy regimens based on epigenetic changes.     

Bi‐allelic alterations in DNA repair genes underpin homologous recombination DNA repair defects in breast cancer

Homologous recombination (HR) DNA repair‐deficient (HRD) breast cancers have been shown to be sensitive to DNA repair targeted therapies. Burgeoning evidence suggests that sporadic breast cancers, lacking germline BRCA1/BRCA2 mutations, may also be HRD. We developed a functional ex vivo RAD51‐based test to identify HRD primary breast cancers. An integrated approach examining methylation, gene expression, and whole‐exome sequencing was employed to ascertain the aetiology of HRD. Functional HRD breast cancers displayed genomic features of lack of competent HR, including large‐scale state transitions and specific mutational signatures. Somatic and/or germline genetic alterations resulting in bi‐allelic loss‐of‐function of HR genes underpinned functional HRD in 89% of cases, and were observed in only one of the 15 HR‐proficient samples tested. These findings indicate the importance of a comprehensive genetic assessment of bi‐allelic alterations in the HR pathway to deliver a precision medicine‐based approach to select patients for therapies targeting tumour‐specific DNA repair defects. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Expression of ERG oncoprotein is associated with a less aggressive tumor phenotype in Japanese prostate cancer patients

To investigate and characterize ERG oncoprotein expression in Japanese patients with prostate cancer (PCa), we evaluated 92 index tumors from Japanese patients who had undergone radical prostatectomy for PCa, using an antibody‐based detection method to determine ERG expression. Expression status was compared with clinicopathological findings including patient age, body mass index and preoperative prostate‐specific antigen (PSA) levels, specimen Gleason score, pathological stage, positive surgical margin, size of index tumor, prostatic volume, zonal origin of index tumor and biochemical failure. Overall, ERG protein was expressed in 16.3% (15/92) of the index tumors, but not in benign glands. Younger patient age, lower Gleason score and negative surgical margins were found to be independently associated with its expression in the multivariate analysis (P= 0.015, 0.003 and 0.038, respectively). ERG expression was not associated with biochemical failure. Though not statistically significant, ERG expression was more frequently observed in peripheral zone than in transition zone cancers (28.2% vs 10%, respectively). In conclusion, ERG protein was less frequently expressed in this Japanese PCa cohort compared with Western reports. ERG expression was associated with a less aggressive tumor phenotype, and its biological significance needs to be further investigated.