乳腺癌基因甲基化的研究进展

乳腺癌的发生的分子机制仍然不明确。研究显示表观遗传学改变（Epigenetics）所导致的基因表达异常也是乳腺癌发生、发展的重要原因。表观遗传学改变是基因的核苷酸序列不发生改变的情况下基因表达的可遗传的变化,包括DNA甲基化、组蛋白乙酰化、染色质重塑、基因组印记以及非编码RNA等。乳腺腺的DNA甲基化是常见的分子事件,具有独特的DNA甲基化特征,既往的研究发现DNA甲基化可以作为乳腺癌早期诊断、分型、监测药物治疗效果和预后的分子标志物。本文将对DNA甲基化在乳腺癌的研究进展进行综述。     

乳腺癌组织中AXIN2基因启动子区甲基化表达及临床意义

目的本研究旨在通过检测乳腺癌组织中AXIN2基因启动子区甲基化状态,分析其与乳腺癌相关危险因素、临床病理特征及mRNA表达间的关系,探讨其在临床应用中的价值。方法选取2018年1月至2019年12月在山东第一医科大学附属肥城市人民医院普外科经手术切除治疗的84例女性乳腺癌患者的乳腺癌组织和癌旁组织,检测组织的AXIN2基因启动子的甲基化状态和mRNA表达量,使用不同浓度的甲基转移酶抑制剂(5-Azacytidine,5-Aza)处理人乳腺癌ZR-75-1细胞,且进行乳腺癌相关危险因素和临床病理特征的收集与分析。结果乳腺癌组织的AXIN2基因启动子甲基化阳性率(40.5%)显著高于癌旁组织的甲基化阳性率(19.0%)(χ2=10.401,P=0.001)。AXIN2甲基化阳性率与乳腺癌患者的初产年龄(χ2=5.467,P=0.019)、是否有人工流产史(χ2=8.372,P=0.006)有关;AXIN2甲基化阳性率与是否发生淋巴结转移(χ2=5.338,P=0.021)、ER表达状态(χ2=9.141,P=0.002)及PR表达状态(χ2=6.918,P=0.009)有关。此外,乳腺癌组织的AXIN2基因mRNA相对表达量(00.22±0.03)显著低于癌旁组织的mRNA相对表达量(0.46±0.06)(t=3.527,P<0.001);甲基化阳性乳腺癌组织的mRNA相对表达量(0.13±0.02)显著低于甲基化阴性乳腺癌组织的mRNA相对表达量(0.29±0.04)(t=3.616,P<0.001)。ZR-75-1细胞使用5-Aza处理后,AXIN2基因mRNA相对表达量显著升高(t=3.824,P<0.001)。结论AXIN2基因启动子区DNA甲基化与乳腺癌发生机制有关,在临床中有一定应用价值。     

肝癌组织中HGFAC表达与DNA甲基化及预后的相关性

目的:探讨肝癌手术患者癌组织中肝细胞生长因子激活因子(HGFAC)表达与DNA甲基化及预后的相关性研究。方法:选取延安大学附属医院普通外科行手术治疗的肝癌患者46例为肝癌组,肝外伤手术治疗患者20例为对照组。两组手术前后分别采用酶联免疫定量法、蛋白质免疫印迹法、免疫组织化学法检测患者手术后肝组织中HGFAC表达,采用Kaplan-Meier法计算生存率并进行生存分析。结果:HGFAC表达与血清甲胎蛋白、总胆红素、谷丙转氨酶指标具有相关性(P<0.05);肝癌组术后2、6、12、24 h各时间段血清HGFAC水平均低于对照组(P<0.05);免疫组织化学结果显示,对照组中HGFAC的表达水平明显高于肝癌组(P<0.05);HGFAC低表达患者的生存率明显低于HGFAC高表达患者,差异有统计学意义(P=0.003)。结论:HGFAC在肝癌组织中的表达降低与不良生存结局相关,DNA高甲基化导致HGFAC表达下降,HGFAC可能是预测肝癌预后的一种生物标志物。     

循环游离DNA甲基化在乳腺癌中的研究进展

DNA基因启动子甲基化是重要的表观遗传学机制之一,是导致抑癌基因表达沉默的重要分子生物学基础.乳腺癌患者外周血中循环游离DNA含量明显高于健康人,且具有与肿瘤原发灶一致的基因异常改变.研究显示,乳腺癌患者外周血循环游离DNA中存在多种基因甲基化异常,且与患者的临床病理特征存在不同程度的相关性,在乳腺癌的早期诊断、预后判断、疗效预测和复发监测等方面具有重要价值,本文就循环游离DNA甲基化在乳腺癌中的相关研究进展作一综述.     

Suberoylanilide hydroxamic acid (SAHA) combined with bortezomib inhibits renal cancer growth by enhancing histone acetylation and protein ubiquitination synergistically

What's known on the subject? and What does the study add? The treatment modality for advanced renal cancer is limited and new treatment approaches are urgently needed. Beneficial effects of bortezomib combined with SAHA have recently been reported. However, there are no previous reports of this combination being tested against renal cancer and its further mechanisms of action should be clarified. This study examined the combined effects of these two clinically feasible drugs and showed that the combination inhibits renal cancer cell proliferation by enhancing both protein ubiquitination and histone acetylation synergistically.

OBJECTIVE

• ? To investigate the combined effect of two clinically feasible drugs, the proteasome inhibitor bortezomib and the histone deacetylase (HDAC) inhibitor suberoylanilide hydroxamic acid (SAHA), on human renal cancer cells in vitro and in vivo.

MATERIALS AND METHODS

• ? The effectiveness of the combination of bortezomib (10–20 nm ) and SAHA (1–5 µm ) on renal cancer cells (Caki‐1, ACHN, A‐498, 786‐O, 769‐P) was assessed by MTS assay, colony formation assay, cell cycle analysis, and apoptosis assay.
• ? In vivo efficacy was evaluated using murine subcutaneous (s.c.) tumour models.
• ? Protein ubiquitination, unfolded protein response, histone acetylation, and changes in the expression of HDAC were evaluated by western blotting.

RESULTS

• ? The combination of SAHA and bortezomib induced apoptosis and inhibited cancer cell proliferation synergistically (combination indices <1) and colony formation significantly (P < 0.05).
• ? In s.c. tumour models a 10‐day treatment with a combination of SAHA (50 mg/kg) and bortezomib (60 µg/kg) inhibited tumour growth significantly (P < 0.05).
• ? Mechanistically, SAHA combined with bortezomib enhanced protein ubiquitination synergistically and enhanced histone acetylation by inhibiting the expression of HDACs.

CONCLUSION

• ? SAHA combined with bortezomib inhibits the proliferation of renal cancer cells in vitro and in vivo, and the effectiveness of the combination is due to its synergistic enhancement of histone acetylation and protein ubiquitination.

     

DNA甲基化与胃癌抑癌基因研究进展

DNA甲基化是基因表达调控的一种方式,抑癌基因启动子高甲基化可以使其表达失活,最后导致肿瘤发生。胃癌的发生与多基因异常表达密切相关,其中抑癌基因甲基化是胃癌发生、发展的重要机制之一。     

叶酸缺乏导致人正常结肠黏膜上皮细胞DNA甲基化状态的改变与结直肠癌的检测关系

目的 通过甲基化芯片对叶酸缺乏导致人正常结肠黏膜上皮细胞(NCM460)的DNA甲基化状态变化进行筛查,探讨相关位点与散发性结直肠癌发生的关系.方法 借助NimbleGen公司的甲基化芯片,检测不同叶酸浓度(高叶酸浓度为150 nmol/L,低叶酸浓度为25 nmol/L)培养的NCM460细胞中基因启动子区CpG岛的甲基化状态,并通过结合重亚硫酸盐的测序方法在细胞和15例(男∶8,女∶7)结直肠癌患者的组织样本中对芯片的结果进行验证.结果 从高叶酸浓度培养的NCM460细胞中筛选到了2680个发生了甲基化的CpG岛,从低叶酸组筛选到了3073个.在21号染色体上,17个基因启动子区CpG岛发生了超甲基化或去甲基化的改变,其中8个基因甲基化状态的改变可能与结直肠癌的发生相关.结论 叶酸缺乏导致人NCM460的DNA甲基化状态改变可能与结直肠癌的发生相关.     

The effect of neoadjuvant chemotherapy on estrogen and progesterone receptor expression and hormone receptor status in breast cancer

BACKGROUND: Neoadjuvant chemotherapy may decrease tumor volume to allow breast conservation surgery. Its effect on estrogen and progesterone receptor (ER/PR) expression and hormone receptor (HR) status is controversial. METHODS: From February 2001 to July 2002, 56 breast cancer patients treated with neoadjuvant chemotherapy and 56 non-neoadjuvant therapy (control) patients with adequate tissue samples were identified. Quantitative ER/PR expression was analyzed in preneoadjuvant or preoperative core biopsies and final surgical specimens. Changes between the two groups were compared to determine if alterations were due to neoadjuvant chemotherapy or tissue sampling. RESULTS: The ER/PR expression changed in 34 (61%) neoadjuvant chemotherapy patients and 27 (48%) control patients. These expression changes resulted in HR status (positive/negative) alterations in 3 patients (5%) in both groups. Age, histology, chemotherapy regimen, and neoadjuvant response did not predict change. CONCLUSIONS: Hormone receptor status changed in 5% of neoadjuvant chemotherapy and control groups due to tissue sampling. As these changes may impact treatment, HR expression reanalysis in final surgical specimens is recommended.     

The relationship of estrogen receptor status to DNA ploidy in breast cancer

The relationship of estrogen receptor(ER) status to DNA ploidy was investigated in 121 patients with breast cancer who underwent surgery. Lymph node status was evaluated histologically and ER levels were determined by the dextran-coated charcoal method, with a level of 3 fmol/mg·protein being considered positive. Flow cytometric DNA content was analyzed using paraffin-embedded tissue blocks. Sixty-three per cent of the specimens were ER+, while 37 per cent were negative. Sixty-one patients (50.4 per cent) were diploid and 60 aneuploid. A statistically significant correlation between the ER positivity rate and diploid DNA was found. Higher ER levels were seen in the postmenopausal patients with diploid tumors than in those with aneuploid tumors and there was a significant tendency for ER levels to be higher in the diploid tumors. Nodal status was not correlated with ER positivity or ploidy pattern. The present results indicate that ER levels are correlated with DNA ploidy, and reflect the degree of functional differentiation.     

表观遗传学与乳腺癌

表观遗传学是基于非DNA序列改变而产生的基因表达的变化,与肿瘤的发生密切相关,具有可逆性和遗传性,主要包括DNA甲基化、组蛋白修饰和染色体重组,它们相互作用,影响基因表达。乳腺癌中,表观遗传调节改变了一些重要基因的表达,导致了肿瘤的产生和发展。表观遗传学的研究对乳腺癌的发生、发展、早期诊断、预后评估、治疗和复发监测产生了深远的影响。     

表观遗传学与乳腺癌

表观遗传学是基于非DNA序列改变而产生的基因表达的变化,与肿瘤的发生密切相关,具有可逆性和遗传性,主要包括DNA甲基化、组蛋白修饰和染色体重组,它们相互作用,影响基因表达.乳腺癌中,表观遗传调节改变了一些重要基因的表达,导致了肿瘤的产生和发展.表观遗传学的研究对乳腺癌的发生、发展、早期诊断、预后评估、治疗和复发监测产生了深远的影响.     

胃癌组织中白细胞介素-8基因启动子甲基化及其mRNA表达的研究

目的 观察胃癌患者癌组织及与其配对的正常胃黏膜组织中白细胞介素-8(IL-8)基因启动子甲基化及其mRNA表达,探讨IL-8基因甲基化与胃癌的关系.方法 收集经病理确诊并行外科手术切除的胃癌组织及配对的正常胃黏膜组织各52例,用实时荧光定量聚合酶链反应(FQ-PCR)分别检测癌组织及正常组织中IL-8基因mRNA表达;用甲基化特异性PCR(MSP)检测上述两种组织中IL-8基因甲基化状态.结果 胃癌组织中IL-8基因mRNA的表达量是正常组织中的1.94倍(P＜0.01).胃癌组织中IL-8基因甲基化率为32.7％,而正常组织中的甲基化率为73.1％(P＜0.01);胃癌组织中IL-8基因mRNA表达及启动子的去甲基化与性别、年龄、肿瘤大小无明显相关(P＞0.05),而与肿瘤的分化程度、Borrmann分型、浸润程度及有无淋巴结转移均具有明显相关(P＜0.05);去甲基化的胃癌组织中IL-8 mRNA表达是甲基化胃癌组织的2.13倍(P ＜0.001).结论 IL-8基因mRNA在胃癌组织中过表达;胃癌组织中IL-8基因启动子区呈低甲基化状态;IL-8基因启动子区去甲基化促进了其在胃癌组织中高表达.     

Involvement of arginine methyltransferase CARM1 in androgen receptor function and prostate cancer cell viability

BACKGROUND: Androgen receptor (AR) may play a role in prostate cancer progression. Coactivator-associated arginine methyltransferase (CARM1) catalyzes methylation of histone H3 at Arg-17. METHODS: Immunohistochemistry of CARM1 was performed on primary prostate cancer specimens. CARM1 recruitment and histone methylation was analyzed by chromatin immunoprecipitation. The effect of CARM1 overexpression or CARM1 knockdown was assessed on reporter assays, cell proliferation, apoptosis, and endogenous androgen target gene expression. RESULTS: CARM1 expression was increased in the nucleus of castration-resistant, but not androgen-stimulated prostate cancer. Androgen stimulation led to CARM1 recruitment and methylation of histone H3 at androgen responsive enhancers. Overexpression of CARM1 stimulated and CARM1 knockdown inhibited AR reporter activity. CARM1 knockdown inhibited cell proliferation and induced apoptosis. CARM1 knockdown inhibited androgen-dependent prostate specific antigen (PSA) and hK2 mRNA expression. CONCLUSIONS: CARM1 is essential for AR function and may play a role in prostate cancer progression. CARM1 may represent a novel therapeutic target in prostate cancer.     

散发性乳腺癌中BRCA1基因甲基化状态与其mRNA表达水平的相关研究

目的:探讨山东半岛地区散发性乳腺癌中BRCA1基因异常甲基化状态及其与mRNA表达的关系,进而研究其BRCA1基因异常的表观遗传学作用。方法:用甲基化特异性PCR和RT-PCR等方法研究乳腺癌组织、癌旁正常组织及乳腺良性肿瘤中BRCA1基因启动子甲基化状态及其mRNA表达水平。结果:在30例散发性乳腺癌中,BRCA1基因甲基化率为43%,BRCA1 mRNA表达下降率为33%。在13例BRCA1发生甲基化的癌组织标本中,BRCA1 mRNA的表达下降率为55%,而在17例BRCA1未发生甲基化的癌组织中,BRCA1 mRNA的表达下降率为18%。在肿瘤分级中,T3分级病人的肿瘤组织中BRCA1甲基化率(74%)和mRNA表达下降率(54%),均分别高于T1+T2分级病人肿瘤组织的BRCA1甲基化率(26%)和mRNA表达下降率(30%);有淋巴结转移病人的肿瘤组织中,BRCA1甲基化率(61%)和mRNA表达下降率(46%),均分别高于无淋巴结转移的甲基化率(17%)和mRNA表达下降率(16%)。结论:BRCA1基因表观遗传学改变是其mRNA表达下降乃至失活的重要原因,且与散发性乳腺癌的恶性程度及淋巴结转移的发生密切相关。     

Global and specific histone acetylation pattern in patients with Balkan endemic nephropathy,a worldwide disease

Background: Balkan endemic nephropathy (BEN) is a chronic tubulointerstitial nephropathy present in the Danube river regions in several Balkan countries. There appears to be a polygenic susceptibility to the disease in interaction with multiple environmental factors (aristolochic acid, ochratoxin A). In a previous study SEC61G, IL17RA, HDAC11 proved to be differently methylated throughout all patient-control pairs of BEN patients from Serbia and Bulgaria. Emerging connections between DNA methylation and histone acetylation prompted the present study on histone acetylation in patients with BEN. Methods: The study involved 39 patients with BEN, and 39 controls collected from non-endemic regions in Serbia. The EpiSeeker Histone H3 and H4 Total Acetylation Detection colorimetric Kits and specific acetylated at lysine 18 H3K18 and H3K36 acetylated at lysine 36 detection kits were used. Results: It was documented that total H4 histone acetylation level was increased significantly, while total H3 histone acetylation did not differ significantly. Specific histone structure and functional properties may be affected by the observed derangement of H3 histone acetylation pattern, since H3K36 site was significantly more acetylated, while H3K18 tended to be less acetylated than in control subjects. Multiple regression analysis revealed a statistically significant relationship between H4, H3T and H3K36 in BEN patients. Conclusion: This preliminary study suggests that the acetylation of histone lysine residues was detectable and found increased at specific sites of H3 and total H4 histones isolated from urothelial cells of patients with BEN. Having in mind a possible mechanism and biological role of epigenetic chromatin modification in urothelial tumor development they obtained results may open opportunity for selective therapeutic interventions in patients with BEN.     

A methyl-deficient diet modifies histone methylation and alters Igf2 and H19 repression in the prostate

BACKGROUND: Folate and methyl-group deficiency has been linked to prostate cancer susceptibility, yet the mechanisms underlying these observations are incompletely understood. The region of the genome containing the imprinted genes insulin-like growth factor 2 (Igf2) and H19, both of which display oncogenic functions, may be particularly sensitive to environmental influences. METHODS: To determine whether a methyl-deficient diet impacts epigenetic controls at the Igf2-H19 locus, we placed C57BL/6 mice containing a polymorphism at the imprinted Igf2-H19 locus on a choline and methionine deficient (CMD) diet. We interrogated this locus for expression and epigenetic changes in prostate tissues. RESULTS: A significant increase in both Igf2 and H19 expression was found in CMD prostate tissues compared to controls. These expression changes were reversible with shorter exposure to the CMD diet. Chromatin immunoprecipitation (ChIP) revealed significant decreases in repressive histone modifications (dimethyl-H3K9) within the H19 promoter, as well as Igf2 P2 and P3 promoters. DNA methylation within these promoters was not altered. No significant change in Igf2 or H19 imprinting was observed. CONCLUSIONS: These findings highlight the plasticity of the epigenome in an epithelial organ vulnerable to neoplastic change. They further suggest that chromatin modifications are more susceptible to methyl-deficient diets than DNA methylation at this locus.