Epirubicin-[Anti-HER2/neu] Synthesized with an Epirubicin-(C13-imino)-EMCS Analog: Anti-Neoplastic Activity against Chemotherapeutic-Resistant SKBr-3 Mammary Carcinoma in Combination with Organic Selenium

Purpose: Discover the anti-neoplastic efficacy of epirubicin-(C13-imino)-[anti-HER2/neu] against chemotherapeutic- resistant SKBr-3 mammary carcinoma and delineate the capacity of selenium to enhance it’s cytotoxic anti-neoplastic potency. Methods: In molar excess, EMCH was combined with epirubicin to create a covalent epirubicin-(C13-imino)-EMCH-maleimide intermediate with sulfhydryl-reactive properties. Monoclonal immunoglobulin selective for HER2/neu was then thiolated with 2-iminothiolane at the terminal ε-amine group of lysine residues. The sulfhydryl-reactive epirubicin-(C13-imino)-EMCH intermediate was then combined with thiolated anti-HER2/neu monoclonal immunoglobulin. Western-blot analysis was utilized to characterize the molecular weight profiles while binding of epirubicin-(C13-imino)-[anti-HER2/neu] to membrane receptors was determined by cell-ELISA utilizing populations of SKBr-3 mammary carcinoma that highly over-expresses HER2/neu complexes. Anti-neoplastic potency of epirubicin-(C13-imino)-[anti-HER2/ neu] between the epirubicin-equivalent concentrations of 10–12 M and 10–7 M was determined by vitality staining analysis with and without the presence of selenium (5 μM). Results: Epiribucin-(C13-imino)-[anti-HER2/neu] between epirubicin-equivalent concentrations of 10–8 M to 10–7 M consistently evoked higher anti-neoplastic potency than “free” non- conjugated epirubicin which corresponded with previous investigations utilizing epirubicin-(C3-amide)-[anti-HER2/neu] and epirubicin-(C3-amide)-[anti-EGFR]. Selenium at 5 mM consistently enhanced the cytotoxic anti-neoplastic potency of epirubicin-(C13-imino)-[anti-HER2/neu] at epirubicin equivalent concentrations (10–12 to 10–7 M). Conclusions: Epirubicin-(C13-imino)-[anti-HER2/neu] is more potent than epirubicin against chemotherapeutic-resistant SKBr-3 mammary carcinoma and selenium enhances epirubicin-(C13-imino)-[anti-HER2/neu] potency. The methodology applied for synthesizing epirubicin-(C13-imino)-[anti-HER2/neu] is relatively time convenient and has low instrumentation requirements.

Phytochemical analysis and anticancer capacity of Shemamruthaa,a herbal formulation against DMBA-induced mammary carcinoma in rats

Objective:To investigate the bioactive-constituents of Shemamruthaa(SM),a herbal combination and its therapeutic effects on the mitochondrial functions with reference to lipid peroxidation(LPO),antioxidant status,citric acid cycle enzymes and electron transport chain enzymes in mammary tissues of 7,12-dimethylbenz(a)-anthracene(DMBA)induced mammary carcinoma in rat model.Methods:Adult Female Sprague-Dawley rats were used for the study and were divided into four groups.CroupⅠserved as control and CroupⅡrats were induced mammary carcinoma by administration of DMBA(25 mg/kg b.w.)orally.The normal and cancer-induced rats(GroupⅢ)were treated with SM(400 mg/kg b.w./day)orally by gastric incubation for 14days.CroupⅣrats served as SM-treated control animals.Results:Cancer-induced rats showed a considerably increased level of LPO with concomitant decreased levels of antioxidants,citric acid cycle enzymes,electron transport chain enzymes and cytochrome contents in the mammary tissue.Treatment with SM brought back the aforementioned biochemical parameters to near normal.Conclusions:From the results,it can be inferred that Shemamruthaa possesses significant anticancer effect through its role in attenuation of LPO,prevention of membrane damage and restoring membrane integrity.

Chinese Herbal Medicine Combined Conventional Chemotherapy Regimens in Advanced Recurrent Breast Mammary Carcinoma

Purpose: Aiming at starting the ball rolling and contributing humble effort to promoting traditional Chinese medicine, we report one successful case of “GP” regimen of combining chemotherapy with traditional Chinese medicine decoction (“lung description”) for breast cancer with lung and bone metastases. Patients and Methods: A second-line chemotherapy regimen of GP (cisplatin + gemcitabine) was applied. In the meantime, we administered Chinese herbal medicine (Fei Decoction, mixed with a variety of effective herbal components) to help her recover from the poor condition. Results: The tumor markers (CEA, CA15-3) had dramatically decreased to the normal range. Both lung and bone metastatic sites reduced according to CT and ECT imaging, and this patient felt free from the complaint of pulmonary and cardiac discomfort. The quality of life has been greatly improved, and the PFS (Progression-Free-Survival) and TTP (Time-to-Progression) have been prolonged from the onset to date. Conclusions: In the course of this combined treatment, it has been shown that Chinese herbal medicine played an important role in the therapy of breast mammary carcinoma. Therefore, Chinese herb may turn out to be an additional choice with its better benefits and tolerability in the treatment of recurrent breast mammary carcinoma.

Methylation Abnormalities in Mammary Carcinoma: The Methylation Suicide Hypothesis

Promoter silencing by ectopic?de novo?methylation of tumor suppressor genes has been proposed as comparable or equivalent to inactivating mutations as a factor in carcinogenesis. However, this hypotheses had not previously been tested by high resolution,?high-coverage whole-genome methylation profiling in primary carcinomas. We have determined the genomic methylation status of a series of primary mammary carcinomas and matched control tissues by examination of more than 2.7 billion CpG dinucleotides. Most of the tumors showed variable losses of DNA methylation?from all sequence compartments, but increases in promoter methylation were infrequent, very small in extent, and were observed largely at CpG-poor promoters. De novo methylation at the promoters?of proto-oncogenes and tumor suppressor genes occurred at approximately the same frequency. The findings indicate that tumor suppressor silencing by?de novo?methylation is much less common than currently believed. We put forward a hypothesis under which the demethylation commonly observed in carcinomas is a manifestation of a defensive system that kills incipient cancer cells.

Mammary Analogue Secretory Carcinoma of the Soft Palate: A Report of Two Cases

Mammary analogue secretory carcinoma (MASC) is a recently identified salivary tumor that shares morphological features of secretory carcinoma of the breast, salivary acinic cell carcinoma, low grade cystadenocarcinoma and mucoepidermoid carcinoma. To date, a majority of cases have been reported in the major salivary glands with very few seen in the palate. We present 2 single institution cases of MASC of the soft palate to help further characterize this novel tumor.

Differential Expression of Glu-Tubulin in Basal-Like Phenotype Carcinoma of the Mammary Gland

High levels of tubulin expression have been described in a variety of human malignant tumors, and glu-tubulin, in which the C-terminal tyrosine of α-tubulin is removed by tubulin carboxypeptidase. Over-expression has been reported in the malignant tumors of the mammary gland and correlated with poor prognosis immunohistochemically. Furthermore, Nielsen et al. proposed that the use of a panel of four markers (ER, HER 2, CK 5/6, and EGFR) could accurately identify basal-like phenotype carcinoma (BPC) with widely available standard pathologic tools. In our study, major prognostic factors such as patient age, tumor size, histological grade, axillary lymph node metastasis, vessel invasion, and local recurrence in BPC were not significantly different from non BP carcinoma (NBPC). However, the BPC group showed a higher ratio of distant metastasis than that of the NBPC group. In triple-negative carcinoma (TNC) cases, staining for glu-tubulin was observed in 46 cases (63.8%), which consisted of 42 of the 58 BPC patients (72.4%) and 4 of the 14 NBPC patients (28.6%). A significant association was found between the expression of glu-tubulin and BPC, but not NBPC. It seems that our findings also agree with the observation that BPC exhibits aggressive biological behavior and increases the content of glu-tubulin, which plays a greater role in migration and invasion.

Parotid mammary analogue secretory carcinoma:A case report and review of literature

BACKGROUND Mammary analogue secretory carcinoma(MASC)is a rare low-grade malignant salivary gland tumor.The morphological and immunohistochemical features of MASC closely resemble those of breast secretory carcinoma.The key characteristics of the lesion are a lack of pain and slow growth.There is no obvious specificity in the clinical manifestations and imaging features.The diagnosis of the disease mainly depends on the detection of the MASC-specific ETV6-NTRK3 fusion gene.CASE SUMMARY This report describes a rare case of a 32-year-old male patient who presented with a gradually growing lesion that was initially diagnosed as breast-like secretory carcinoma of the right parotid gland.Imaging and histological investigations were used to overcome the diagnostic difficulties.The lesion was managed with right parotidectomy,facial nerve preservation,biological patch implantation to restore the resulting defect,and postoperative radiotherapy.On postoperative follow-up,the patient reported a mild facial deformity with no complications,signs of facial paralysis,or Frey’s syndrome.CONCLUSION The imaging and histological diagnostic challenges for MASC are discussed.

THE EFFECT OF THE TRANSFER OF LAK CELLS, COMBINED WITH ALLOIMMUNIZATION, ON THE PULMONARY METASTASES OF RAT MAMMARY CARCINOMA

Alloimmunization was combined with lympho-kine activated killer (LAK) cells to assess its effect on mammary carcinoma in rats. The animals were injected with both irradiated allosplenocytes and syngeneic LAK cells. Metastatic lung nodules were markedly reduced using combined therapy when compared with the transfer of LAK cells or alloimmuni-zation alone. IL-2 activity in the serum of alloim-munized rats could be detected. This activity, maintained in vivo for one week, may be responsible for enhancing the antitumor effect of transferred LAK cells.

A STUDY OF THE RADIOSENSITIVE EFFECTS ON MAMMARY CARCINOMA IN MICE BY CHINESE MEDICINE (SALVIA PLUS ASTRAGALUS) AND ASPIRIN

The effect of treatment with Chinese medicine including Salvia miltiorrhizan and Astragalus, and the nonsteroid anti-inflammatory drug aspirin alone or combining radiotherapy, was investigated in 6-8 week-old TA2 mice being inoculated mammary carcinoma. The date indicated the following conclusions: the tumor growth could be inhibited by aspirin alone (p<0.01) but Chinese medicine group was observed. Mice treated with radiotherapy together with medicine both Chinese medicine and aspirin, had a statistically significant tumor inhibiting (p<0.01) as compared to mice treated with radiotherapy alone. The function to prevent the normal tissues from radiation by these two medicine groups were observed simultaneously. In addition, blood-flow volume of microcirculation, immune function and lymphocyte micronucleus were examined, which were used to analyse potential mechanism of sensitizing enhancement for Chinese medicine and aspirin.

Heparanase antisense oligodeoxynucleotide inhibits angiogenesis and metastasis of human mammary carcinoma cell xenografts in nude mice

Objective:To evaluate the inhibitory effect of heparanase antisense oligodeoxynucleotide (AS-ODN) on the angiogenesis and metastasis of human mammary carcinoma cell xenografts in nude mice. Methods:The AS-ODN complementary to the start codon region of heparanase mRNA and its control, scrambled nonsense oligodeoxynucleotide (NS-ODN) were designed and synthesized.A subcutaneous growth model and an acute hematogenous metastasis model of human mammary carcinoma were estab- lished in nude mice and were treated with ODNs.The heparanase expression in tumor was evaluated by RT-PCR and Western blot.The microvessel density (MVD) was measured by immunohistochemistry for factorⅧ.The tumor volume was calculated and lung metastatic nodules were counted.Results:The hep- aranase expression,MVD,tumor volume and lung metastatic nodules in AS-ODN treated group were sig- nificantly decreased compared with that in NS-ODN treated group and that in PBS group (P＜0.01).Con- clusion:Heparanase AS-ODN has significant inhibitory effect on the angiogenesis and metastasis of human mammary carcinoma cell xenografts in nude mice.

Spontaneous Mammary Carcinomas in Female Dogs:Association between the Immunohistochemical Degree of Aggressiveness of Tumors,Intensity of DNA Damage and Residues of Pyrethroids

Diagnosis and biological behavior of breast cancer of female dog represent one of the biggest challenges facing the Veterinarian in recent years. Due to its exponential growth and the degree of aggressiveness, the exact cause of this tumor is probably multifactorial and it is believed that may suffer influence from environmental factors. Among the suspected environmental contaminants are the pyrethroids. Aiming to investigate the participation of pyrethroids in tumorigenesis in female dogs, a study was conducted using 50 female dogs, 22 were positive for simple breast carcinoma (Group I), 18 with a diagnosis of complex breast carcinoma (Group II) and 10 negative (Group III) for breast cancer. In order to detect DNA damage, the Comet assay was performed on mammary samples of these animals, which also had samples submitted to the technique of High Performance Liquid Chromatography (HPLC), which aimed to quantify the concentration of pyrethroids. The results of HPLC of each animal were compared with those obtained by the Comet assay analysis of variance and the means were compared by the test groups “Student T” at the significance level of p ￡ 0.05. Despite presenting correlation between the amount of DNA damage and tumor aggressiveness, no statistical differences were found in the DNA damage of different histologic types of breast carcinoma. As for pyrethroids, even these were detected in 22% of tumor tissues and peritumoral fat, there was no difference in DNA damage between cells exposed and not exposed to environmental contaminant.

3例犬炎性乳腺癌的病理学诊断和组织学分级评价

为了对3例临床疑似犬炎性乳腺癌的病例进行确诊,本试验采用苏木精-伊红(H.E.)染色对送检肿物进行组织病理学检查,应用全角蛋白(Pan ck)抗体、角蛋白8(CK8)抗体和血管性血友病因子(VWF)抗体进行免疫组织化学染色以判断免疫表型。结果显示,3例患犬肿瘤内部形成管状癌、致密癌、筛状癌和粉刺性癌,并伴有淋巴管扩张、肿瘤栓子浸润淋巴管等现象,符合犬炎性乳腺癌的特征;免疫组织化学染色结果证实,肿瘤为腺上皮起源,且淋巴管、血管内存在大量肿瘤栓子。通过病理学分析结合临床表现,3例病例均确诊为犬炎性乳腺癌。根据Pena分级标准,将病例1判定为Ⅲ级炎性乳腺癌,病例2和3判定为Ⅱ级炎性乳腺癌。本试验结果不仅可为犬炎性乳腺癌的病理诊断提供参考,而且为犬炎性乳腺癌临床病例治疗方案的制定和预后评估提供理论依据。

Reversal of Adriamycin Resistance in Human Mammary Cancer Cells by Small Interfering RNA of MDR1 and MDR3 Genes

The purpose of this paper is to investigate the reversal effect of small interfering RNA （siRNA） targeting MDRI and MDR3 genes on the resistance of MCF-7/ADR cells to adriamycin. siRNA plasmid vector targeting MDRI and MDR3 genes was transfected into MCF-7/ADR cells, and then was stained with Annexin-V FITC （fluorescein isothiocyanate conjugated） to detect the early stage cell apoptosis by flow cytometry （FCM）. 50 % inhibition concentration （IC50） of adriamycin for MCF-7/ADR cells was determined by MTT method. MDRI and MDR3 mRNA was assessed by RT-PCR. Treatment of MCF-7/ADR cells with the two kinds of siRNAs resulted in a reversal of adriamycin resistance of MDR to different extents. 1） The apoptosis efficiency of MDRI and MDR3 siRNA vector after transfection was （18.21 ± 1.65） % and （9.07±2.16） % respectively （P〈0.05）, and there was significant differences in the apoptosis efficiency between pSuppressor Neo vector and the MDRIsiRNA or MDR3 siRNA vector （P〈0.01）; 2） The reversal effect of MDRIsiRNAis higher than that of MDR3 siRNA （P〈0.05）; 3 ） The expression of MDRI and MDR3 mRNA can be restrained by pSuppressor Neo MDRI and MDR3 siRNA respectively, and the reduction in the mRNA level was in a time-dependent manner （P〈0.01）. MDRI and MDR3 gene silencing can enhance intracellular adriamycin accumulation in MCF-7/ADR cells, improve sensitivity of MCF-7/ADR cells to adriamycin, and induce cell apoptosis. The reversal effect of adriamycin resistance by siRNA of MDR1 was more effective than that of MDR3.

Secretory carcinoma——impact of translocation and gene fusions on salivary gland tumor

Secretory carcinoma（SC）, previously described as mammary analogue secretory carcinoma（MASC）, is a recently described salivary gland tumor which morphologically resembles mammary secretory carcinoma. The first description of SC/MASC, reported by Skálová et al. in 2010, was as a rare salivary carcinoma imitating secretory carcinoma of the breast. SC/MASC is a unique salivary gland tumor with morphological overlap with acinic cell carcinoma（Aci CC）, mucoepidermoid carcinoma（MEC）, and adenocarcinoma not otherwise specified（ADCNOS）. SC/MASC shares similar clinicopathological features with Aci CC. As a critical difference between SC/MASC and Aci CC, SC/MASC characteristically has the chromosomal translocation t（12;15）（p13;q25） which leads to a fusion gene between the ETV6 gene on chromosome 12 and the NTRK3 gene on chromosome 15. This genetic background is an important differential diagnostic finding for excluding other salivary gland tumors and may be a critical factor determining the prognosis for patients with SC/MASC. Research in recent years has provided a large body of new data on SC/MASC and suggests the possibility that the ETV6-NTRK3 translocation could be a therapeutic target. Here, we review the morphological and clinicopathological features of SC/MASC and discuss new directions for therapy.

经络辨证逆针灸对二甲基苯蒽诱导的大鼠乳腺肿瘤发生过程的影响

目的 观察经络辨证逆针灸对二甲基苯蒽[7,12-dimethylbenz-(a)anthranene, DMBA]诱导的乳腺肿瘤模型大鼠肿瘤发生过程的影响。方法 选取8周龄清洁级雌性SD未孕大鼠64只,随机分为空白对照组、模型组、他莫昔芬组和逆针灸组,每组各16只。采用DMBA单次灌胃(100 mg/kg)诱导自发性乳腺癌模型,空白对照组给予等量芝麻油灌胃。于造模后1周对治疗组进行干预,他莫昔芬组3 mg/kg灌胃,1次/d,持续16周;逆针灸组针刺“足三里、膻中、三阴交”,隔天1次,每次20 min,每周3次,干预16周。应用HE染色评估大鼠乳腺组织形态学变化;应用免疫组织化学法进行CK5/6、p63检测,观察其在不同类型病变乳腺组织中的分布及表达水平。结果 HE染色以及免疫组化染色CK5/6和p63表达结果显示,DMBA诱导的乳腺肿瘤模型大鼠肿瘤的发生发展途径为:正常→一般增生→非典型增生→原位癌→浸润性癌;对不同实验组乳腺组织不同类型病变的分布进行统计学分析,结果提示各组间组织病理学分布具有统计学意义(P<0.05)。结论 经络辨证理论指导下的逆针灸能够抑制DMBA诱导乳腺自发性肿瘤模型大鼠病变进展,有望应用于乳腺癌的临床防治。

犬乳腺癌免疫治疗的研究进展

犬乳腺癌是老年母犬常见的肿瘤类型之一,手术是最常用的治疗方法,但单纯切除无法根治该种疾病,其较高的复发率和转移率使犬的预后差,另外,犬乳腺癌具有较强的异质性,常规的化疗方法往往伴随着副作用,额外的靶向和非靶向治疗可延长患犬的生命周期,故创新疗法可以发挥很大的作用。同时,犬乳腺癌常被作为研究人类乳腺癌良好的动物模型,故犬乳腺癌新疗法的研究是比较肿瘤学中一个很有前景的领域,引起了学者的广泛关注。本文对犬乳腺癌的免疫治疗方法进行了全面总结,为犬乳腺癌的个体化治疗打开新思路,同时为人乳腺癌新治疗方法的应用提供参考。

乳腺癌针吸细胞学诊断与病理组织学诊断对比分析

目的:探讨针吸细胞学诊断乳腺癌的准确性。方法:针吸细胞学检查乳腺癌306例,其检查结果与病理组织学检查结果对照。结果:306例中301例针吸细胞学诊断为乳腺癌与病理组织学诊断结果一致、3例针吸细胞学诊断为良性病变而病理组织学诊断为乳腺癌、2例细胞学诊断为乳腺癌而病理组织学诊断为良性病变,符合率为98.37%(301/306),假阴性0.98%(3/306),假阳性0.65%(2/306)。结论:本组资料显示针吸细胞学检查诊断乳腺癌的准确性较高,是乳腺癌的重要筛查和诊断方法。

乳腺癌针吸细胞体视学定量与细胞形态学分型的探讨

本文采用国内先进的MPIAS－500多媒体图像分析仪对50例乳腺癌的针吸细胞形态进行了多参数的体视学定量研究。并以细胞面积、核浆比值两项定量参数为分型标准、对50例乳腺癌进行了分型研究，共分四型、对各型之间细胞形态的体视学定量结果进行了对比分析，同时结合病理组织学分类进行对照分析。本文的研究结果建立了乳腺癌细胞形态体视学的量化指标及探讨了乳腺癌针吸细胞形态学的分型标准和方法，对鉴别诊断乳腺癌和判断乳腺癌的恶性程度以及指导临床观察乳腺癌的生物学特性、疗效及预后有重要意义。

槲皮素对实验性乳腺癌中血管生成抑制作用的研究

目的：探讨槲皮素对实验性乳腺癌中的肿瘤血管生成抑制作用。方法：本实验建立了DMBA诱导的乳腺癌动物模型。79只雌性SD大鼠随机分为四组：A组（DMBA），B组（DMBA＋TAM），C组（DMBA＋槲皮素），D组（空白对照）。喂养持续28周后，进行光镜观察、MVD计数，抗bFGF、VEGF及H－ras的免疫组化分析。结果：1）A组大鼠乳腺肿瘤发生率及癌灶平均直径（75．2％、2．37cm）明显高于B组（40．9％、1．82cm），C组（45．5％、1．71cm）及D组（0％、0cm），其差异均有显著意义（P＜0．05），B组和C组差异无显著意义（P＞0．05）；2）MVD、VEGF、bFGF及H－ras免疫组化分析显示：A组与B组和C组差异有显著意义（P＜0．05），B组和C组无显著性差异（P＞0．05）。结论：槲皮素有降低DMBA诱导的SD大鼠乳腺癌发生率及抑制其生长的作用，其机制可能与以下因素有关：1）抑制ras基因活性，从而抑制肿瘤细胞的增殖及血管生成；2）抑制VEGF，bFGF等血管生长因子的活性，肿瘤血管生成过程受阻，致乳腺癌组织内MVD降低。