RNA Biomarker Release with Ultrasound and Phase-Change Nanodroplets

Microbubbles driven by ultrasound are capable of permeabilizing cell membranes and allowing biomarkers or therapeutics to exit from or enter cancer cells, respectively. Unfortunately, the relatively large size of microbubbles prevents extravasation. Lipid-based perfluorobutane microbubbles can be made seven-fold smaller by pressurization, creating 430-nm nanodroplets. The present study compares microbubbles and nanodroplets with respect to their ability to enhance miR-21 and mammaglobin mRNA release from cultured ZR-75-1 cells. Mammaglobin mRNA and miR-21 release increased with escalating concentrations of nanodroplets up to, respectively, 25- and 42-fold with 2% nanodroplets (v/v), compared with pre-ultrasound levels, whereas cell viability decreased to 62.4%. Sonication of ZR-75-1 cells incubated with microbubbles or nanodroplets caused relatively similar levels of cell death and miR-21 release, suggesting that nanodroplets are similar to microbubbles in enhancing cell permeability, but may be more advantageous because of their smaller size, which may allow extravasation through leaky tumor vasculature.     

GATA3、MGB和GCDFP-15在乳腺癌中的病理诊断价值

目的探讨GATA结合蛋白3(GATA3)、乳腺珠蛋白(MGB)和巨囊性病液体蛋白-15(GCDFP-15)在乳腺癌中的病理诊断价值。方法采用免疫组化方法检测GATA3、MGB、GCDFP-15、雌激素受体(ER)、孕激素受体(PR)、人表皮生长因子受体2(HER2)、Ki67在乳腺癌中的表达,并根据组织来源及免疫表型将乳腺癌分为原发性/转移性乳腺癌、激素受体阳性/阴性乳腺癌及不同分子分型的乳腺癌,统计GATA3、MGB、GCDFP-15阳性率。结果GATA3在原发性/转移性乳腺癌中的阳性率(92.5%;94.25%)均显著高于MGB(42.11%;29.17%)和GCDFP-15(55.77%;31.34%)(P<0.01),对于评估原发灶不明来源的肿瘤,尤其是恶性积液的标本,GATA3具有更好的敏感性。此外,MGB和GCDFP-15在激素受体阴性和三阴型乳腺癌中的阳性率偏低,进一步加大了这两种转移性乳腺癌的诊断难度。而GATA3在激素受体阴性(90.38%)或三阴型乳腺癌中的阳性率(85.29%)显著高于MGB(20.00%;5.00%)和GCDFP-15(35.09%;33.33%)(P<0.01),提示其在激素受体阴性和三阴型转移性乳腺癌中具有潜在诊断价值,是一个优于MGB和GCDFP-15的特异性标记物。结论对于评估原发灶不明来源的肿瘤,尤其是恶性积液的标本,或免疫表型为激素受体阴性和三阴性的乳腺癌,GATA3具有比MGB和GCDFP-15更高的诊断价值。     

Mammaglobin: a candidate diagnostic marker for breast cancer

Mammaglobin, known for its mammary tissue specificity, has been discussed as a promising diagnostic marker in breast cancer for almost 10 years. In particular, the application of mammaglobin RT-PCR to detect disseminated breast cancer cells has been reported. More than 25 publications evaluate the detection of mammaglobin mRNA in lymph node, blood, and bone marrow specimens of breast cancer patients. Recently, structural details about the mammaglobin complex have been discovered, and these findings can be implemented to optimize detection of the secreted protein. This review summarizes the findings of almost 50 published studies and the current knowledge about the diagnostic utility of mammaglobin.     

Evaluation of expression based markers for the detection of breast cancer cells

SummaryIntroduction Genes that are expressed in a highly tissue- or disease-specific manner provide possible targets for therapeutics, early detection of cancer, and monitoring of disease burden during and after treatment. Further, genes of this type that code for secreted or shed proteins may allow for serum detection of the product facilitating our ability to specifically detect the cancer in all circumstances. To this end, we are working towards identification and characterization of such genes that are specifically expressed in breast epithelium. In the current study, we have measured the expression of two markers that emerged from a screen of the Incyte LifeSeq Database and were subsequently shown to be highly restricted to breast epithelium termed BU101 (also called Lipophilin B) and BS106 (small mucin-like protein). These two novel markers were compared with two other candidate markers, Mammaglobin and Cytokeratin 19 (CK19).Methods Utilizing quantitative real-time PCR, we compared the expression of these four genes in a series of 95 primary breast cancers, 9 lymph nodes from breast cancer patients, 13 lymph nodes from non-cancer patients and 10 normal breast tissues.Results Cytokeratin was shown to be highly sensitive in detecting all breast cancers, while BU101, BS106 and Mammaglobin were more restricted.Conclusion While no one of the these markers efficiently detects all breast cancers, a combination of two or more could achieve a very high sensitivity in assaying for circulating or occult breast cancer cells.These two authors contributed equally to the work     

Apocrine Hidrocystoma of the Lower Lip: A Case Report and Literature Review

The hidrocystomas (HCs) are cystic forms of sweat gland resulting from proliferation of the apocrine secretory coil or eccrine duct. Apocrine -HCs are cystic lesions that arise from the apocrine secretory coil, while eccrine -HCs represent retention cysts of the eccrine duct. The commonest site for such lesions is around the eye, and they may also occur on the ears, scalp, chest, shoulders, or feet. However, HCs of the perioral region are uncommon. The differential diagnosis with minor salivary gland cyst or cystic neoplasms often poses a problem in this site. Here we report a rare case of apocrine -HC of the right lower lip for which excisional biopsy of the lesion was performed. Histopathologically, the lesion was a unilocular cyst lined by a double-layered epithelium of the apocrine secretory type. Immunohistochemically, the secretory epithelium was positive for mammaglobin, gross cystic disease fluid protein 15 (GCDFP-15), cytokeratin 7 (CK 7) and CK18, and the myoepithelium was positive for alpha-smooth muscle actin (α-SMA) and weakly positive for S100 protein. Here we present this very rare case of apocrine -HC of the lower lip, and discussed regarding differential diagnosis with minor salivary gland cystic lesion in the lip.     

A small subgroup of operable breast cancer patients with poor prognosis identified by quantitative real-time RT-PCR detection of mammaglobin A and trefoil factor 1 mRNA expression in bone marrow

Purpose The utility of three different epithelial mRNA markers to detect clinically significant, disseminated tumour cells in bone marrow (BM) was explored. Methods Mammaglobin A (hMAM), trefoil factor 1 (TFF-1) and prostate derived Ets factor (PDEF) mRNA were quantitated by real-time RT-PCR in BM samples from 192 breast cancer patients undergoing surgery (control group: 26 healthy women). Results During a median follow-up of 72 months, four of the five hMAM BM-positive and three of the seven TFF-1 BM-positive patients experienced a systemic relapse. Kaplan-Meier survival analyses demonstrated significantly shorter recurrence-free-, breast-cancer-specific- and overall survival for both hMAM and TFF-1 BM-positive patients. In contrast, PDEF mRNA quantitation did not reveal any significant differences in the survival analyses. Multivariate Cox regression demonstrated hMAM mRNA BM expression to be an independent predictor of both overall- (hazard ratio = 5.896), breast-cancer-specific- (hazard ratio = 10.208) and systemic-recurrence-free survival (hazard ratio = 14.304). TFF-1 status was related to hMAM status (P < 0.001). Conclusion Breast cancer patients with pre-operative elevated BM levels of hMAM and/or TFF-1 mRNA seem to constitute a small group of patients with a very poor prognosis. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Characterization of the role of CD8+T cells in breast cancer immunity following mammaglobin-A DNA vaccination using HLA-class-I tetramers

INTRODUCTION: Mammaglobin-A(mam-A) is expressed in over 80% of human breast tumors. We recently reported that mam-A DNA vaccination resulted in breast cancer immunity in a preclinical model. Here we investigated whether mam-A HLA-class-I tetramers could be used to monitor and define the role of CD8(+)cytotoxic T-lymphocytes(CTL) in mediating breast cancer immunity following mam-A DNA vaccination. STUDY DESIGN: Mam-A DNA vaccination was performed in HLA-A2(+)huCD8(+ )transgenic mice. HLA-A2 tetramers carrying the immunodominant mamA2.1 peptide were used to monitor CD8(+)CTL. Human breast cancer colonies were developed in immunodeficient SCID-beige mice. ELISPOT was used to correlate frequency of mamA2.1 tetramer(+)CD8(+)T cells and IFN-gamma production [spots per million cells (spm)] in human subjects. RESULTS: Vaccination of HLA-A2(+)huCD8(+) mice with mam-A DNA vaccine, but not empty vector, led to the expansion of mamA2.1 tetramer(+)CD8(+)T-cells in peripheral blood (<0.5% pre-vaccination compared to >2.0% post-vaccination). CD8(+)T cells from vaccinated mice specifically lysed UACC-812(HLA-A2(+)/mam-A(+), 25% lysis) but not MDA-MB-415(HLA-A2(-)/mam-A(+)) or MCF-7(HLA-A2(+)/mam-A(-)) breast cancer cells. Adoptive transfer of purified CD8(+)T cells from vaccinated mice into immunodeficient SCID-beige mice with established human breast cancer colonies led to tetramer(+)CD8(+ )T-cell infiltration with regression of UACC-812 but not MCF-7 tumors. HLA-A2(+) breast cancer patients revealed increased frequency of mamA2.1 tetramer(+)CD8(+ )T-cells compared to normal controls (2.86 +/- 0.8% vs. 0.71 +/- 0.1%, P = 0.01) that correlated with the IFN-gamma response to mamA2.1 peptide (48.1 +/- 20.9 vs. 2.9 +/- 0.8 spm, P = 0.03). CONCLUSIONS: CD8(+ )T-cells are crucial in mediating breast cancer immunity following mam-A DNA vaccination. Mam-A HLA-class-I tetramers can be effectively used to monitor development of CD8(+ )T-cells following mam-A vaccination.     

Morphology in Conjunction with Immunohistochemistry is Sufficient for the Diagnosis of Mammary Analogue Secretory Carcinoma

The recently described mammary analogue secretory carcinoma (MASC) is a low-grade salivary gland malignancy that harbors the recurrent cytogenetic abnormality t(12;15) (p13;q25) ETV6-NTRK3. Confirmation of this is currently considered the gold standard for diagnosis. Some have postulated that morphology together with supporting immunohistochemistry is sufficient to diagnose MASC. In this study we retrospectively review a series of 19 MASCs diagnosed based on histology in conjunction with immunohistochemistry; subsequently we performed in situ hybridization using an ETV6 break-apart probe. Immunohistochemistry for S100 protein and mammaglobin as well as fluorescence in situ hybridization using the Vysis ETV6 Dual Color Break-Apart FISH Probe Kit were performed on all cases. The 19 cases were from 12 females and 7 males with ages ranging from 16 to 76 years (mean = 45 years). Sixteen cases were from the parotid gland, 1 case was from a periparotid lymph node and 2 cases were from the submandibular gland. All 19 cases demonstrated moderate to strong expression of S100 protein. Eighteen cases demonstrated strong, diffuse expression of mammaglobin, while one case had only rare tumor cells that strongly expressed mammaglobin. Eighteen of 19 cases (95 %) demonstrated the ETV6 rearrangement by fluorescence in situ hybridization. Given that morphology together with immunohistochemistry is highly correlated with the ETV6 gene rearrangement, we conclude that molecular confirmation is not required to diagnose MASC.     

乳腺癌患者中乳腺球蛋白的表达及其临床意义

目的探讨乳腺球蛋白在乳腺癌患者的体内不同部位的表达情况及其对检测微小转移的临床意义。方法在本院普外科进行治疗的乳腺癌患者中选择29例，同时在乳腺良性增生的患者中选择29例进行研究，患者基本临床资料均相匹配。29例乳腺癌患者为观察组，29例良性乳腺增生患者为对照组，58例乳腺疾病患者术后的离体标本均保留在-70℃的环境下，以保证标本的有效性。乳腺癌患者在离肿瘤最近处进行皮肤组织取材，乳头乳晕复合体则经正中切面取材，取29例离体癌组织标本；良性乳腺增生的患者只在相同的位置进行皮肤取材即可同时取29例离体增生组织标本进行研究。本次试验采用逆转录-聚合酶链式反应技术来检测并比较两组患者相应组织中乳腺球蛋白的表达情况。结果观察组29例乳腺癌组织中乳腺球蛋白表达均为阳性；乳头乳晕复合体中阳性率为17．2％．皮肤组织阳性率为10．3％。对照组在增生的良性组织中及正常皮肤中乳腺球蛋白均为阴性。皮肤阳性率与皮肤橘皮样变及酒窝症等改变的程度有关；乳头乳晕复合体的阳性率与肿瘤的位置有关。结论乳腺球蛋白的阳性提示着者癌变微转移的出现，同时研究表明乳腺癌累及皮肤组织和乳头的机会比较小。在临床手术中可以考虑为患者实行皮肤及乳头保留性的乳腺癌根治术以减轻患者的痛苦，缓解患者的心理压力。     

Unmasking MASC: Bringing to Light the Unique Morphologic, Immunohistochemical and Genetic Features of the Newly Recognized Mammary Analogue Secretory Carcinoma of Salivary Glands

Mammary analogue secretory carcinoma (MASC) is a recently described salivary gland neoplasm that is characterized by its striking morphologic and molecular similarities to secretory carcinoma of the breast. This review highlights the characteristic clinical, histologic, immunophenotypic, and molecular features of MASC, and draws attention to the differential diagnosis of this increasingly recognized tumor.