上海地区妇女乳腺癌组织PIK3CA基因热点突变区域研究

目的:探讨乳腺癌组织中磷脂酰肌醇一3-激酶催化亚单位α(PIK3CA)基因2个热点突变区域第9和第20外显子的突变,以及该突变与乳腺癌临床病理特征之间的关系.方法:45例乳腺癌标本提取DNA后.对第9及第20外显子进行PCR扩增,产物纯化后测序.另取10例乳腺良性肿瘤组织标本作为对照.结果:在45例乳腺癌组织标本中共发现14例(31%)PIK3CA基因突变,其中第9外显子8例(57%),第20外显子6例(43%),在第9外显子突变中,1例为C1616G(P539R),1例为G1633A(E545K),6例为A1634C(E545A).第20外显子的6例突变均为A3140G(H1047R).10例良性肿瘤对照中均未发现突变.第9与第20外显子突变在浸润性导管癌和浸润性小叶癌中的分布差异无统计学意叉,P>0.05.PIK3CA基因突变与患者年龄、肿瘤大小、淋巴结状态、ER/PR状态、HER-2表达和p53表达之间无明显相关性,P>0.05.结论:PIK3CA基因可能与乳腺癌的发生、发展及预后相关.但PIK3CA基因突变与乳腺癌临床病理特征之间无明显相关性.     

中国人群结直肠癌中PIK3CA基因突变分析

目的:PIK3CA基因编码ⅠA型磷脂酰肌醇-3-激酶(PI3Ks)的p110催化亚基,致癌性的PIK3CA突变可通过激活P13K通路参与结直肠癌的发生发展。PIK3CA在西方结直肠癌患者中有较高的突变频率,但其在中国人结直肠癌中的突变情况尚不明确。本研究旨在探讨中国人结直肠癌中PIK3CA基因的突变频率、分布特点及其与结直肠癌的关系。方法:采用PCR产物直接测序法,对79例中国结直肠癌患者肿瘤标本中PIK3CA基因外显子9和外显子20中的突变进行检测分析。结果:在79例肿瘤标本中检出PIK3CA基因突变率为8.9%(7/79),其中外显子9突变率为6.3%(5/79),外显子20突变率为2.5%(2/79),其突变热点分布与既往文献报道基本相符。结论:中国人结直肠癌中存在一个PIK3CA基因突变的亚群,其E542K、E545K和H1047R突变,与以往报道的该基因的致癌性突变相一致,可能是这部分结直肠癌中PI3K信号通路激活的原因。     

PI3KCA基因高频突变位点与子宫颈鳞癌关系的初步研究

目的:探讨宫颈鳞癌与PIK3CA基因高频突变点E542K、E545k和H1047R的关系.方法:随机选择50例病理确诊宫颈鳞癌组织作为实验组,患者本人的外周血作为对照组,提取其基因组DNA,用巢式-PCR方法扩增PIK3CA基因exon9和exon20基因片段并测序分析.结果:实验组均发现exon9 E542K位点突变,相应外周血DNA检测也发现相同突变,PI3KCA基因与定位染色体22q11.2的Cat Eye Syndrome基因片段具有高度重合现象.改变引物设计发现50例样本中E542K和E545k位点分别发现2例突变,H1047R位点未发现突变.对照组未见突变.结论:宫颈鳞癌组织中PIK3CA基因exon9(E542K和E545k)和exon20 H1047R位点在其它实体肿瘤中具有较高突变频率的PI3KCA基因,而在宫颈鳞癌中的突变率较小.     

乳腺癌PIK3CA基因热点突变的研究

目的:采用突变敏感性分子开关技术,检测乳腺癌组织中PIK3CA基因热点突变的突变频率,并分析乳腺癌PIK3CA基因热点突变与乳腺癌临床病理特征之间的关系.方法:用突变敏感性分子开关技术对90例乳腺癌组织和10例乳腺纤维腺瘤组织DNA进行检测,并利用基因测序进一步验证.结果:在90例乳腺癌患者中检测出18例PIK3CA基因热点突变(突变率20％,18/90),包含A3140G突变14例(突变率15.6％,14/90)、G1633A突变3例(突变率3.3％,3/90)、GI624A突变2例(突变率2.2％,2/90),其中1例患者同时发生A3140G和G1624A突变.10例乳腺纤维腺瘤中未发现以上热点突变.PIK3CA基因突变与HER-2表达有关(x2=4.119,P=0.048),过表达者中PIK3CA基因突变率低,而与年龄分布(x2 =0.238,P=0.602)、肿瘤大小(P=1.000)、淋巴结状态(x2=3.689,P＝0.056)及ER(x2=0.957,P=0.328)、PR(x2 =0.012,P=0.914)和EGFR(x2 =0.011,P=0.916)是否表达无相关性.结论:PIK3CA基因热点突变与患者的HER-2过表达呈负相关.     

PIK3CA基因突变与乳腺癌的关系

PIK3CA基因在细胞生长、凋亡及肿瘤的形成过程中起重要的作用.功能及遗传学研究显示,PIK3CA基因是一个癌基因.近期研究显示在多种人类实体瘤中存在PIK3CA基因的突变.乳腺癌中PIK3CA基因突变的频率约为8%～40%.其突变80%～90%聚集在该基因的第9外显子和第20外显子.研究表明,PIK3CA基因突变与乳腺癌生物学特性及临床预后等有关.     

C-KIT基因在鼻咽癌患者中的突变

背景与目的:C-KIT基因功能获得性突变是应用伊马替尼治疗胃肠间质瘤有效的真正分子标记,鼻咽癌是否存在C-KIT基因突变尚不清楚.我们检测了鼻咽癌患者C-KIT基因的突变情况,期望为伊马替尼治疗鼻咽癌提供初步依据.方法:采用直接测序法检测34例鼻咽癌组织中C-KIT基因9至21号外显子序列.结果:在34例鼻咽癌患者的442个外显子中,发现10个外显子存在突变,其中3个是点突变,1个是缺失突变.9、10、18、21号外显子的总突变频率分别是2.9%(1/34),5.996(2/34),17.6%(6/34),2.9%(1/34).结论:鼻咽癌组织中C-KIT基因9、10、18、21号外显子存在突变,为进一步研究伊马替尼在鼻咽癌中的治疗价值提供了初步依据.     

p53基因在鼻咽癌中的热点突变

采用ＤＮＡ测序和免疫组织化学技术对２３例鼻咽癌标本ｐ５３基因第７、８外显子的突变和该基因突变后的表达情况进行了检测，发现６５．２％（１５／２３例）的标本存在ｐ５３基因突变，且突变点均位于第２７３密码子第２个碱基，均为Ｇ：Ｃ→Ａ：Ｔ颠换，所编码的精氨酸为组氨酸所取代。免疫组化结果显示，除上述１５例突变标本外，另有１例存在ｐ５３过量表达。研究提示：鼻咽癌中ｐ５３基因突变率较高，该基因的突变和表达可能在鼻咽癌的发病中起着重要的作用；ｐ５３基因突变以第２７３密码子为突变热点，可能是由于患者接触了环境中目前尚未确知的某种特殊致癌物所致，或该点突变为鼻咽癌发病过程中的必要因素。     

PIK3CA和C-KIT基因突变与伴腋窝淋巴结转移乳腺癌临床病理特征及预后的关系

目的 探讨PIK3CA和C-KIT基因突变与乳腺癌伴腋窝淋巴结转移患者临床病理特征及预后的关系。方法 收集84例乳腺癌伴淋巴结转移患者术后存档癌组织石蜡标本,采用Ion Torrent测序技术检测其PIK3CA、C-KIT基因的突变情况（突变率及突变位点分布）;分析两种基因突变与临床病理特征（年龄、月经、淋巴结转移数、癌栓、肿瘤大小、临床分期、ER／PR和HER-2表达情况）的关系;根据突变情况分组（突变组和未突变组）,随访3年无瘤生存率。结果 PIK3CA、C-KIT基因突变率分别为63.1％（53／84）和15.5％（13／84）,均为错义突变;其中PIK3CA第9、13和20号外显子突变依次有7、8和24例,第13、20号外显子双突变共14例,而C-KIT基因突变均位于第10号外显子。两基因突变均与HER-2表达有关,仅PIK3CA与临床分期和ER／PR表达有关,但其各外显子突变与临床病理特征无关。PIK3CA基因突变组的3年无瘤生存率为75.5％,未突变组为48.4％;而C-KIT基因突变组与未突变组分别为76.9％和62.0％,两种基因不同突变情况的差异均无统计学意义（P＞0.05）。年龄、月经、淋巴结转移数、肿瘤大小、临床分期、ER／PR表达、HER-2表达、PIK3CA基因突变和C-KIT基因突变均不是影响乳腺癌伴淋巴结转移患者预后的独立因素。结论 乳腺癌伴腋窝淋巴结转移患者PIK3CA的基因突变率较高,与临床分期、ER／PR及HER-2表达有关;C-KIT仅与HER-2表达有关,两种基因突变可能在乳腺癌的发生发展及腋窝淋巴结转移中起一定作用。     

结直肠癌Ras、BRAF和PIK3CA基因突变分析及与临床病理特征的关系

目的 分析结直肠癌患者中Ras（K-Ras／N-Ras）、BRAF和PIK3CA基因突变情况及其与临床病理特征的关系。方法 回顾性分析2013年12月至2014年10月于北京大学肿瘤医院消化肿瘤内科接受诊治的200例结直肠癌患者的肿瘤组织标本,采用PCR扩增-直接测序法检测Ras,包括K-Ras（第2、3、4外显子）、N-Ras（第2、3、4外显子）、BRAF（第15外显子）及PIK3CA（第9、20外显子）基因的突变状态,分析其与结直肠癌临床病理特征的关系。结果 200例患者中存在Ras基因突变92例（46.0％）,其中K-Ras基因突变87例（43.5％）,主要发生在外显子2,N-Ras基因突变5例（2.5％）;其中1例患者存在K-Ras、N-Ras基因同时突变。存在BRAF基因突变15例（7.5％）,突变类型均为V600E,且与K-Ras突变存在排他性。存在PIK3CA基因突变9例（4.5％）,可与Ras或BRAF基因突变共存。Ras（K-Ras／N-Ras）基因在年龄≥65岁患者中的突变率明显高于＜65岁者（P＜0.05）,其表达与性别、原发部位、组织学类型、分化程度、TNM分期、区域淋巴结转移、远处转移、术后复发转移均无关（P均＞0.05）。BRAF、PIK3CA基因在原发部位为右半结肠患者中的突变率明显升高（P＜0.05）,但与年龄、性别、组织学类型、分化程度、TNM分期、区域淋巴结转移、远处转移、术后复发转移均无关（P＞0.05）。结论 N-Ras、PIK3CA基因在中国结直肠癌患者中的突变率较低。K-Ras、N-Ras基因突变与年龄相关,BRAF、PIK3CA基因与肿瘤原发部位相关;对结直肠癌患者进行Ras（K-Ras／N-Ras）、BRAF及PIK3CA基因的联合检测将会为提高临床靶向治疗的疗效提供更加可靠的依据。     

耳、鼻、咽、喉（071199～071246）

071199 治疗13例外耳道癌和中耳癌的临床经验；071200 鼻咽癌组织中PIK3CA基因热点突变区的突变筛查；071201 鼻咽癌组织中GNAT1基因的表达、杂合性丢失及甲基化分析；071202 建立人鼻咽癌裸鼠皮下移檀瘤模型不同方法的比较；071203 诱导型一氧化氮合酶在鼻咽癌中的表达及意义……[第一段]     

Absence of PIK3CA hotspot mutations in hepatocellular carcinoma in Japanese patients

A recent study revealed that the p110alpha (PIK3CA), catalytic subunit of phosphatidylinositol 3-kinase (PI3K), is somatically mutated in many types of cancer. For example, PIK3CA is mutated in an estimated 35.6% of hepatocellular carcinoma (HCC) cases. To measure the frequency of PIK3CA hotspot mutations in Japanese HCC patients, exons 9 and 20 of the PIK3CA gene were sequenced in 47 clinical HCC samples. Contrary to expectations, no hotspot mutations were found any of the HCC samples. In addition, we found abnormally migrating waves near the end of exon 9 in the PCR chromatograms from 13 of the 47 samples. PCR amplification and subsequent cloning and sequencing revealed that these chromatograms contained two distinct sequences, the wild-type p110alpha sequence and a different sequence found on human chromosome 22q11.2, the Cat Eye Syndrome region, which contains a putative pseudogene of PIK3CA. These abnormally migrating waves were also found in noncancerous liver tissue, indicating that this was not a result of HCC-associated mutations. Therefore, it is likely that the percentage of hotspot mutations in the PIK3CA gene of Japanese HCC patients is lower than was previously reported.     

PIK3CA mutations in head and neck squamous cell carcinoma.

PURPOSE: Recent studies have reported high frequencies of somatic mutations in the phosphoinositide-3-kinase catalytic alpha (PIK3CA) gene in several human solid tumors. Although gene amplifications of PIK3CA have been reported in head and neck squamous cell carcinoma (HNSCC), small mutation of the gene has not been evaluated in HNSCC previously. In this study, we examined the mutation frequency of PIK3CA in HNSCC. EXPERIMENTAL DESIGN: More than 75% of the somatic mutations of PIK3CA are clustered in the helical (exon 9) and kinase domains (exon 20). To investigate the possible role of PIK3CA in HNSCC tumorigenesis, exons 1, 4, 5, 6, 7, 9, and 20 of the gene were analyzed by direct genomic DNA sequencing in 38 HNSCC specimens. RESULTS: We identified four missense mutations in the seven exons of PIK3CA from 38 HNSCC specimens (11%). Three of the four mutations (i.e., H1047R, E542K, and E545K) have been previously reported as hotspot mutations. The remaining novel mutation, Y343C, is identified at exon 4 nucleotide 1028 A --> G. Three of the four mutations were shown to be somatic, whereas the fourth mutation (H1047R) was identified in a cell line. Interestingly, three of the four mutations identified were in pharyngeal cancer samples. CONCLUSIONS: These data provide evidence that oncogenic properties of PIK3CA contribute to the carcinogenesis of human head and neck cancers, especially in pharyngeal cancer. A specific kinase inhibitor to PIK3CA may potentially be an effective therapeutic reagent against HNSCC or pharyngeal cancer in particular.     

A novel PIK3CA hotspot mutation in Isfahanian breast cancer patients

Somatic mutations of phosphatidylinositol 3-kinase (PI3K) catalytic subunit (PIK3CA) play an important role in tumorigenesis. Using PCR-SSCP, followed by sequencing, we examined the PIK3CA gene over the previously identified mutational hotspots in a panel of 50 breast cancer and 5 normal breast tissue, as well as 50 normal blood samples isolated from a population of Iranian patients. In the present study, the frequency of PIK3CA mutation was 14%. However, the previously identified hotspot mutations in exons 20 were completely absent in these breast cancer samples. Instead, we found a new hotspot mutation (3 of 50 samples) in exon 20.     

PIK3CA mutation status in Japanese lung cancer patients

Somatic mutations of the PIK3CA (phosphatidylinostitol 3-kinase catalytic subunit) gene have been found in human cancer patients. Previous reports suggested that about 4% of lung cancers harbored PIK3CA gene mutations. However, the clinico-pathological background for PIK3CA gene mutations has not yet been investigated in lung cancer. We have genotyped the PIK3CA gene in Japanese lung cancer patients. The study included 235 lung cancer cases surgically removed in Nagoya City University Hospital. The two PIK3CA mutation hot spots (exon 9 and exon 20) were analyzed by real time polymerase chain reaction (PCR)-based assay. The data were confirmed by direct sequencing. In exon 9, somatic mutation was found in eight patients (3.4%). The mutation included three E542K (G1624A), three E545K (G1633A), one E542Q (G1624C), and one Q546K (C1636A). However, in exon 20, there was no mutation in our lung cancer patients. PIK3CA mutations were not correlated with gender (women versus men, p=0.4162), age (< or =60 versus >60, p=0.8027), or smoking status of the lung cancers (never versus smoker, p=0.5666). PIK3CA mutation incidence was significantly lower in adenocarcinoma (2/135, 1.5%) than in squamous cell carcinoma (5/77, 6.5%, p=0.0495). Among eight patients with a PIK3CA mutation, three patients also harbored an EGFR somatic mutation. PIK3CA gene mutations were rare in lung cancer; rarer in adenocarcinoma. Further functional analyses of the PIK3CA mutations are warranted to study if they could be the target of therapy for the lung cancer.     

PIK3CA amplification is predictive of poor prognosis in Tunisian patients with nasopharyngeal carcinoma

PI3Ks (phosphatidylinositol 3-kinases) are lipid kinases that regulate signalling pathways involved in cell proliferation, motility, and adhesion. Somatic mutations and amplification of the PIK3CA gene have been reported in various types of human cancers. However, little is known about the frequency and prognosis role of PIK3CA activation in nasopharyngeal carcinoma (NPC). This study was conducted with the aim to screen for PIK3CA mutations in the two hot spot regions (exons 9 and 20) and to investigate for the PIK3CA gene amplification combined with the expression analysis of the phosphorylated Akt (pAkt). We showed that among 88 specimens, none had mutation in the helical domain (exon 9) and only one (1.13%) had mutation in the kinase domain (exon 20). On the other hand, PIK3CA gene amplification was found in 21.6% of cases and was strongly associated with distant metastasis ( P  = 0.002), lymph node involvement ( P  = 0.032), and advanced tumor stage ( P  < 0.001). Moreover, patients with PIK3CA copy number gain have a significant reduced overall survival time (P log rank = 0.02). We concluded that PIK3CA gene amplification is frequent in NPC and occurs in the advanced stage of NPC. Moreover, our finding emphasizes the association of PIK3CA gene amplification with worse prognosis in nasopharyngeal carcinoma. ( Cancer Sci 2009); 00: 000–000)     

Detection of PIK3CA Gene Mutations with HRM Analysis and Association with IGFBP-5 Expression Levels in Breast Cancer

Breast cancer is the second most common cancer and second leading cause of cancer deaths inwomen. Phosphatidylinositol-3-kinase (PI3K)/AKT pathway mutations are associated with cancer andphosphatidylinositol-4, 5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) gene mutations have beenobserved in 25-45% of breast cancer samples. Insulin growth factor binding protein-5 (IGFBP-5) can showdifferent effects on apoptosis, cell motility and survival in breast cancer. We here aimed to determine theassociation between PIK3CA gene mutations and IGFBP-5 expressions for the first time in breast cancerpatients. Frozen tumor samples from 101 Turkish breast cancer patients were analyzed with high resolutionmelting (HRM) for PIK3CA mutations (exon 9 and exon 20) and 37 HRM positive tumor samples were analyzedby DNA sequencing, mutations being found in 31. PIK3CA exon 9 mutations (Q546R, E542Q, E545K, E542Kand E545D) were found in 10 tumor samples, exon 20 mutations (H1047L, H1047R, T1025T and G1049R)in 21, where only 1 tumor sample had two exon 20 mutations (T1025T and H1047R). Moreover, we detectedone sample with both exon 9 (E542Q) and exon 20 (H1047R) mutations. 35% of the tumor samples with highIGFBP-5 mRNA expression and 29.4% of the tumor samples with low IGFBP-5 mRNA expression had PIK3CAmutations (p=0.9924). This is the first study of PIK3CA mutation screening results in Turkish breast cancerpopulation using HRM analysis. This approach appears to be a very effective and reliable screening method forthe PIK3CA exon 9 and 20 mutation detection. Further analysis with a greater number of samples is needed toclarify association between PIK3CA gene mutations and IGFBP-5 mRNA expression, and also clinical outcomein breast cancer patients.     

Different prognostic roles of mutations in the helical and kinase domains of the PIK3CA gene in breast carcinomas.

PURPOSE: In breast cancer, the PIK3CA gene is frequently mutated at "hotspots" in exons 9 and 20, corresponding to the helical and kinase domains, respectively. We decided to investigate the association of PIK3CA mutations with pathologic features and clinical outcome in a large series of patients with breast cancer. EXPERIMENTAL DESIGN: Frozen samples from 163 consecutive patients were analyzed for PIK3CA mutations using PCR single-strand conformation polymorphism and sequence analyses. RESULTS: We identified 46 missense mutations, 24 (53%) in exon 9, and 21 (47%) in exon 20. Twelve (50%) of the 24 mutations in exon 9 were of the E542K type and 11 (46%) were of the E545K type. Twenty (95%) of the 21 mutations in exon 20 were H1047R substitutions. Mutations in exon 9 were more frequent in lobular carcinomas (42% of cases) than in ductal carcinoma (11% of cases; P = 0.002). At univariate survival analysis, PIK3CA exon 20 mutations were associated with prolonged overall and disease-free survival, whereas mutations in exon 9 were associated with significantly worse prognosis. At multivariate analysis, exon 9 PIK3CA mutations were the strongest independent factor to predict poor prognosis for disease-free survival (P = 0.0003) and overall survival (P = 0.001). CONCLUSION: Our data show that exon 9 PIK3CA mutations are typical of infiltrating lobular carcinomas. In addition, they indicate that PIK3CA mutations in different exons are of different prognostic value: exon 9 mutations are independently associated with early recurrence and death, whereas exon 20 PIK3CA mutations are associated with optimal prognosis.     

胃癌患者错配修复缺陷和PIK3CA基因突变与临床病理特征的关系

目的:分析胃癌中错配修复缺陷(dMMR)和PIK3CA基因突变分别与临床病理特征的关联,以及两者之间的相关性。方法:选取本中心554例胃癌肿瘤组织标本,收集临床病理参数。免疫组化检测错配修复蛋白MSH2、MSH6、MLH1和PMS2,评判是否为dMMR。ARMS-PCR检测PIK3CA基因突变。应用统计学方法,分析dMMR和PIK3CA突变与临床病理特征的关系,以及dMMR与PIK3CA突变的关联。结果:554例胃癌患者中,dMMR 30例(5.4%)。其中,MLH1和PMS2双缺失27例(90.0%),MSH2和MSH6双缺失2例(6.7%),PMS2缺失1例（3.3%）。PIK3CA突变34例（6.1%）,其中15例E545 K (44.1%), 7例E542 K (20.1%),1例E545D(2.9%),6例H1047R(17.6%),4例H1047L(11.8%),1例E545K和E542K双突变（2.9%）。9号外显子突变24例（70.6%）,20号外显子突变10例（29.4%）。单因素分析发现,dMMR与女性,年龄> 65岁,肿瘤直径> 5 cm,肿瘤未侵及浆...     

PIK3CA mutations in HER2-positive Breast Cancer Patients; Frequency and Clinicopathological Perspective in Egyptian Patients

Missense mutations in PIK3CA are common in breast cancers. They mostly involve exons 9 and 20 which encode kinase and helical domains of the protein and may result in its activation. PIK3CA activating mutations were previously shown to predict lower pathologic complete response (pCR) in HER2-positive breast cancer cases undergoing neoadjuvant human epidermal growth factor receptor 2-targeting therapy. Hence, the present work was conducted to estimate the mutation frequency in PIK3CA in 51 HER2-positive patients by direct sequencing. Our results showed 8 out of 51 (15.7%) to harbor PIK3CA mutations in either exon 9 or 20, or both. Three patients had mutations in both exons 9 and 20. Seven (13.7%) possess missense mutations in exon 20 which changed the amino acid sequence of the protein (H1047R, M1040I, and G1049G). Only four cases harbored mutations in exon 9, changing the codon sequences (E545K E545A, and R524K). Taking the clinicopathological data to account, the mutation frequency was greater in ductal than lobular carcinomas, in grade II rather than III and in lymph node positive lesions, with a higher HER2 score and which are ER/PR negative. However, none of the correlations proved statistically significant. In conclusion, to the best of our knowledge, the PIK3CA mutation frequency in this study is the first report regarding HER2-positive breast cancer patients in Egypt. Hereby, we highlight a moderate frequency which could be useful in the future as a predictive marker for anti-HER2 therapy.