CD44和CD24标记的宫颈癌细胞亚群特性

目的 考察Siha细胞系中的CD44^+/CD24⁺能否富集宫颈癌干细胞。方法 用流式细胞仪分选出CD44⁺/CD24⁺Siha细胞,用无血清悬浮培养观察成球能力、裸鼠移植瘤实验观察成瘤能力、透射电子显微镜观察辐射前后两组细胞形态变化,并通过Transwell侵袭实验比较细胞侵袭能力的差异。结果 耐放疗细胞中CD44^+/CD24⁺Siha细胞比例明显高于其在亲代Siha细胞中的比例;无血清培养CD44⁺/CD24⁺Siha细胞组可以形成致密且体积较大的细胞球,CD44^+/CD24⁺Siha细胞组致瘤时间早,成瘤率高;辐射后CD44⁺/CD24⁺Siha细胞较亲代Siha细胞更抗凋亡;CD44⁺/CD24⁺Siha细胞组的迁移细胞数明显高于亲代Siha细胞组,所有数据均具有统计学意义。结论 CD44⁺/CD24⁺Siha具备部分干细胞特性,CD44⁺/CD24⁺可能成为宫颈癌干细胞特异性表面标志物。     

CD44+/CD24-/low在乳腺癌中的表达及其与含蒽环类药物化疗方案敏感度的关系

目的研究乳腺癌中CD44+/CD24-/low的表达及其与含蒽环类药物化疗方案敏感度的关系。方法 91例乳腺癌接受含蒽环类药物的术前新辅助化疗,2～4个疗程后进行效果评价;采用双染免疫组织化学方法检测化疗前后CD44+/CD24-/low的表达,t检验分析化疗前后CD44+/CD24-/low细胞比例的变化,χ2检验分析乳腺癌CD44+/CD24-/low表型与乳腺癌临床病理参数及化疗疗效的关系。结果乳腺癌中CD44+/CD24-/low阳性表达率为39.6%(36/91),在接受含蒽环类药物的新辅助化疗后乳腺癌中CD44+/CD24-/low细胞比例较化疗前明显增加(P=0.028)。CD44+/CD24-/low阳性组ER阳性率明显低于CD44+/CD24-/low阴性组(25.0%vs.47.3%,P=0.033)。三阴性乳腺癌中CD44+/CD24-/low阳性率明显高于非三阴性乳腺癌(61.9%vs.32.9%,P=0.017)。CD44+/CD24-/low表型与年龄、肿瘤大小、临床分期、病理类型、组织学分级等乳腺癌临床病理参数无明显关系(P>0.05)。CD44+/CD24-/low阳性组的总有效率高于CD44+/CD24-/low阴性组,但两组之间差异无统计学意义(75%vs.69.1%,P=0.542);CD44+/CD24-/low阳性组的病理完全缓解率明显高于CD44+/CD24-/low阴性组(38.9%vs.18.2%,P=0.028)。结论 CD44+/CD24-/low细胞仅存在于部分乳腺癌,CD44+/CD24-/low表型与ER(﹣)、三阴性乳腺癌相关,CD44+/CD24-/low表型与乳腺癌对含蒽环类药物化疗方案的敏感度相关,CD44+/CD24-/low表型可能成为乳腺癌临床化疗疗效的预测指标之一。     

Phase I study of salazosulfapyridine in combination with cisplatin and pemetrexed for advanced non‐small‐cell lung cancer

Spliced variant isoforms of CD44 (CD44v) are a marker of cancer stem cells in solid tumors. They stabilize the xCT subunit of the transporter system xc(–) and thereby promote synthesis of the antioxidant glutathione. Salazosulfapyridine (SASP) is an inhibitor of xCT and suppresses the proliferation of CD44v‐positive cancer cells. Chemotherapy‐naïve patients with advanced non‐squamous non‐small‐cell lung cancer were enrolled in a dose‐escalation study (standard 3 + 3 design) of SASP in combination with cisplatin and pemetrexed. The primary end‐point was the percentage of patients who experience dose‐limiting toxicity. Fifteen patients were enrolled in the study. Dose‐limiting toxicity was observed in one of six patients at a SASP dose of 1.5 g/day (elevation of aspartate and alanine aminotransferase levels, each of grade 3), two of five patients at 3 g/day (hypotension or pneumonitis, each of grade 3), and two of three patients at 4.5 g/day (anorexia of grade 3). The maximum tolerated dose was thus 3 g/day, and the recommended dose was 1.5 g/day. The overall response rate was 26.7% and median progression‐free survival was 11.7 months, much longer than that for cisplatin–pemetrexed alone in previous studies. Exposure to SASP varied markedly among individuals according to ABCG2 and NAT2 genotypes. The serum concentration of free CD44v protein was increased after the first cycle of treatment, possibly reflecting death of cancer stem cells. Salazosulfapyridine was thus given safely in combination with cisplatin–pemetrexed, with the addition of SASP tending to prolong progression‐free survival. This trial is registered in the UMIN Clinical Trials Registry as UMIN000017854.     

Application of Stem Cells in Targeted Therapy of Breast Cancer: A Systematic Review

Background: The aim of this systematic review was to investigate whether stem cells could be effectivelyapplied in targeted therapy of breast cancer. Material and Method: A systematic literature search was performedfor original articles published from January 2007 until May 2012. Results: Nine studies met the inclusioncriteria for phase I or II clinical trials, of which three used stem cells as vehicles, two trials used autologoushematopoetic stem cells and in four trials cancer stem cells were targeted. Mesenchymal stem cells (MSCs)were applied as cellular vehicles to transfer therapeutic agents. Cell therapy with MSC can successfully targetresistant cancers. Cancer stem cells were selectively targeted via a proteasome-dependent suicide gene leadingto tumor regression. Wnt/β-catenin signaling pathway has been also evidenced to be an attractive CSC-target.Conclusions: This systematic review focused on two different concepts of stem cells and breast cancer markinga turning point in the trials that applied stem cells as cellular vehicles for targeted delivery therapy as well asCSC-targeted therapies. Applying stem cells as targeted therapy could be an effective therapeutic approach fortreatment of breast cancer in the clinic and in therapeutic marketing; however this needs to be confirmed withfurther clinical investigations.     

胃癌干细胞中差异表达microRNA对其生物学特性的影响

[目的]探讨胃癌干细胞中差异表达microRNA及其对胃癌干细胞生物学特性(自我更新、侵袭、耐药能力)的调控作用.[方法]采用无血清悬浮培养法和干细胞标志物流式分选术分离人胃癌干细胞;分别采用胃癌细胞球形成实验、体外侵袭、耐药实验鉴定胃癌干细胞的生物学特性;microRNA表达谱芯片检测人胃癌干细胞中差异表达microRNA;采用定量逆转录PCR(qRT-PCR)技术检测相关差异表达microRNA的表达情况,验证microRNA芯片结果;相关microRNA抑制剂干扰后,分别进行胃癌细胞球形成实验、体外侵袭与耐药实验检测microRNA对人胃癌干细胞自我更新、侵袭、耐药能力的影响.[结果]成功分离得到人胃癌干细胞CD44(+)亚群,其具有典型的肿瘤干细胞生物学特性:较强的自我更新能力、侵袭和耐药能力.与CD44(-)细胞亚群相比,人胃癌干细胞CD44(+)细胞亚群高表达miRNAs共有17个;低表达miRNAs共有33个.采用qPCR技术验证microRNA表达谱芯片相关结果,得到:CD44(+)亚群中miR-196a-5p、miR-155-5p、miR-30a、miR-30b、miR-30c表达上调,miR-1246、miR-195-5b表达下调.抑制相关高表达microRNA(miR-196a-5p、miR-155-5p,miR-30a,miR-30b、miR-30c)表达后,肿瘤干细胞的自我更新能力、侵袭和耐药能力均下降.[结论]胃癌干细胞中具有多种差异表达microRNA,microRNA对胃癌干细胞生物学特性具有调控作用.     

Ribosome display and selection of single‐chain variable fragments effectively inhibit growth and progression of microspheres in vitro and in vivo

Distinguishing the surface markers of cancer stem cells (CSCs) is a useful method for early diagnosis and treatment of tumors, as CSCs may participate in tumorigenesis and metastasis by migrating into the circulatory system. However, the potential targets of CSCs are expressed at low levels in the natural state and are always changing. Thus, dynamic screening has been reported to be an effective measure for exploring CSC markers. In recent years, diverse single‐chain variable fragments (scFvs) have been widely used in immunotherapy. In this study, we determined that the scFvs, screened using RD, had a high affinity to microspheres and could inhibit their progression. We also observed that the selected scFvs underwent evolution in vitro, and antitumor‐associated proteins were successfully expressed. Combined with chemotherapy, the scFvs had a synergistic effect on the inhibition of the microspheres’ progression in vitro and in vivo, which could be ascribed to their high affinity for stem‐like cells and the inhibition of the microspheres’ collective behaviors. In addition, proteins inhibiting CD44⁺/CD24⁺ and MAPK were involved. Our data indicated that dynamic screening of the scFvs in a natural state was of great significance in the inhibition of the microspheres in vitro and in vivo.     

结肠癌干细胞与临床病理因素及survivin、Ki67表达的相关分析

 目的
探讨肿瘤干细胞标志物CD133在结直肠癌组织中的表达,及其与临床病理因素、survivin和Ki67的关系。方法应用免疫
组织化学方法检测60例手术切除的结直肠癌和30例癌旁组织及12例正常组织中CD133的表达、癌组织中survivin和
Ki67的表达,对各指标之间的相关关系及其与性别、年龄、肿瘤的部位和大小、病理分级、侵袭深度、淋巴结转移、
pTNM分期之间的关系进行分析。结果结直肠癌组织中CD133阳性表达率为50％(30/60),显著高于癌旁及正常组织
(P<0.01);CD133的表达与淋巴结转移和pTNM分期密切相关(P<0.01),与病理分级存在相关性(P<0.05),与其他临床病理
因素均无关;survivin的表达仅与肿瘤的病理分级相关(P<0.05),Ki67与各项临床病理参数均无相关性;CD133与
survivin、Ki67的表达呈正相关(P<0.05、P<0.01);survivin与Ki67的表达呈正相关(P<0.01)。结论结直肠癌组织中
存在CD133+肿瘤细胞,肿瘤干细胞在结直肠癌的发生、发展中起重要作用,检测肿瘤干细胞标志物CD133的表达可作为
一种判断肿瘤的恶性程度和预后的重要指标。     

Cancer Stem Cells in Head and Neck Squamous Cell Carcinoma: A Review

Research indicates that a small population of cancer cells is highly tumorigenic, endowed with the capacityfor self-renewal, and has the ability to differentiate into cells that constitute the bulk of tumors. These cells areconsidered the ‘‘drivers’’ of the tumorigenic process in some tumor types, and have been named cancer stemcells (CSC). Epithelial-mesenchymal transition (EMT) appears to be involved in the process leading to theacquisition of stemness by epithelial tumor cells. Through this process, cells acquire an invasive phenotype thatmay contribute to tumor recurrence and metastasis. CSC have been identified in human head and neck squamouscell carcinomas (HNSCC) using markers such as CD133 and CD44 expression, and aldehyde dehydrogenase(ALDH) activity. Head and neck cancer stem cells reside primarily in perivascular niches in the invasive frontswhere endothelial-cell initiated events contribute to their survival and function. Clinically, CSC enrichment hasbeen shown to be enhanced in recurrent disease, treatment failure and metastasis. CSC represent a novel targetof study given their slow growth and innate mechanisms conferring treatment resistance. Further understandingof their unique phenotype may reveal potential molecular targets to improve therapeutic and survival outcomesin patients with HNSCC. Here, we discuss the state-of-the-knowledge on the pathobiology of cancer stem cells,with a focus on the impact of these cells on head and neck tumor progression, metastasis and recurrence due totreatment failure.     

Isolation and characterization of tumorigenic extrahepatic cholangiocarcinoma cells with stem cell‐like properties

Recent studies suggest that the ability to form and grow tumors specifically resides in a small cell population called cancer stem cells (CSCs). These studies were conducted mainly on various human cancers; however, isolation and characterization of stem cells from cholangiocarcinoma have not been attempted. The molecular markers CD24, CD44, CD34, and epithelial cell adhesion molecule (EpCAM) are widely used, individually or in combination, to characterize some types of CSCs. In this study, we used these markers to identify a subpopulation of cells in extrahepatic cholangiocarcinoma (ECC) with cancer stem/progenitor cell‐like properties. We found that CD24⁺CD44⁺EpCAM^high cells (0.39–2.27%) were present in human ECC tissues. The expression of a CD24⁺CD44⁺EpCAM^high subpopulation was consistent with primary cancers and could be duplicated during serial in vivo passaging in NOD/SCID mice. CD24⁺CD44⁺EpCAM^high cells isolated from 3 cholangiocarcinoma xenografts showed high tumorigenic potential compared with CD24^?CD44^?EpCAM^low/? cells. These tumorigenic ECC cells exhibited the stem cell properties of self‐renewal and ability to produce heterogeneous progeny. We report the identification of a CSC population in ECC characterized by CD24, CD44 and EpCAM phenotypes. Our findings could provide new insight into the tumorigenesis of cholangiocarcinoma and offer a potential target for anti‐cancer therapy.     

Cancer Stem Cells as Mediators of Treatment Resistance in Brain Tumors: Status and Controversies

Malignant primary brain tumors are characterized by a short median survival and an almost 100% tumor-related mortality. Despite the addition of new chemotherapy regimes, the overall survival has improved marginally, and radiotherapy is only transiently effective, illustrating the profound impact of treatment resistance on prognosis. Recent studies suggest that a small subpopulation of cancer stem cells (CSCs) has the capacity to repopulate tumors and drive malignant progression and mediate radio- and chemoresistance. This implies that future therapies should turn from the elimination of the rapidly dividing, but differentiated tumor cells, to specifically targeting the minority of tumor cells that repopulate the tumor. Although there exists some support for the CSC hypothesis, there remain many uncertainties regarding theoretical, technical, and interpretational aspects of the data supporting it. If correct, the CSC hypothesis could have profound implications for the way tumors are classified and treated. In this review of the literature, we provide original data and hypotheses supporting alternative explanations and outline some of the therapeutic implications that can be derived.     

CD44~+/CD24~-、CD44~-/CD24~+在乳腺良恶性病变中的分布特点和临床病理意义

[目的]探讨CD44+/CD24-细胞、CD44-/CD24+细胞在乳腺癌发生不同阶段中(乳腺良性增生、乳腺不典型增生、乳腺原位癌、乳腺癌)分布规律,及其与乳腺癌临床病理因素之间的关系。[方法]采用免疫组化双染色方法检测45例正常乳腺组织、41例良性增生乳腺组织、39例不典型增生乳腺组织、51例乳腺原位癌组织、121例乳腺癌组织中CD44、CD24的表达情况,分析CD44+/CD24-细胞、CD44-/CD24+细胞在乳腺癌发生不同阶段中的数量和分布特点。[结果]CD44+/CD24-细胞存在于20.0%(9/45)的正常乳腺组织中,在29.3%(12/41)的乳腺良性增生组织、35.9%(14/39)的乳腺不典型增生组织、43.1%(22/51)的乳腺原位癌组织、52.9%(64/121)的浸润性乳腺癌组织中检测到CD44+/CD24-细胞,随着病变的进展,CD44+/CD24-细胞的数量也增加,差异具有显著统计学意义。此外,在33.3%(15/45)的正常乳腺组织中有CD44-/CD24+表达,43.9%(18/41)的乳腺良性增生组织、46.2%(18/39)的乳腺不典型增生组织、68.6%(35/51)的乳腺原位癌组织、86.0%(104/121)的浸润性乳腺癌组织检测到CD44-/CD24+细胞,且阳性细胞百分率随病变程度升高。在浸润性乳腺癌组织中CD44+/CD24-的表达与肿瘤有无淋巴结转移相关(P<0.05),但与病人年龄、月经状态、肿瘤的组织学类型、病理分级、肿瘤大小、ER、PR及Her-2的表达无显著相关(P>0.05),CD44-/CD24+的表达率与乳腺癌患者的各项临床病理特征均无统计学意义(P>0.05)。[结论]CD44+/CD24-、CD44-/CD24+细胞在乳腺增生和癌变过程中可能起重要作用。     

HER Story: The Next Chapter in HER‐2‐Directed Therapy for Advanced Breast Cancer

Untreated human epidermal growth factor receptor‐2 (HER‐2)‐positive advanced breast cancer (ABC) is an aggressive disease, associated with a poor prognosis and short overall survival. HER‐2‐directed therapy prolongs both time to disease progression and overall survival when combined with chemotherapy and has become the standard of care for those with HER‐2‐positive breast cancer in the early and advanced settings. Despite the remarkable therapeutic impact HER‐2‐directed therapy has had on disease outcomes, some patients with HER‐2‐positive disease will have primary resistant disease and others will respond initially but will eventually have progression, underscoring the need for other novel therapeutic options. This article reviews recent phase III trial data and discusses a practical approach to sequencing of HER‐2‐directed therapy in patients with HER‐2‐positive ABC. The significant cumulative survival gains seen in these trials are slowly reshaping the landscape of HER‐2‐positive ABC outcomes.     

Clinicopathological Significance of CD133 and ALDH1 Cancer Stem Cell Marker Expression in Invasive Ductal Breast Carcinoma

Background: Biomarkers in breast neoplasms provide invaluable information regarding prognosis and help determining the optimal treatment. We investigated the possible correlation between cancer stem cell (CSC) markers (CD133, and ALDH1) in invasive ductal breast carcinomas with some clinicopathological parameters. Aim: To assess the correlation between expression of cancer stem cell (CSC) markers (CD133, and ALDH1) and clinicopathological parameters of invasive ductal breast carcinomas. Materials and Methods: Immunohistochemical analysis of CD133 and ALDH1 was performed on a series of 120 modified radical mastectomy (MRM) specimens diagnosed as invasive ductal breast carcinoma. Results: Expression of both CD133 and ALDH1 was significantly changed and related to tumor size, tumor stage (TNM), and lymph node metastasis. A negative correlation between CD133 and ALDH1 was found. Conclusions: Detecting the expression of CD133 and ALDH1 in invasive ductal breast carcinomas may be of help in more accurately predicting the aggressive properties and determining the optimal treatment.     

紫杉醇联合重组人粒细胞集落刺激因子动员 乳腺癌患者外周血干细胞的效果及影响因素

 目的研究紫杉醇联合重组人粒细胞集落刺激因子（rhG-CSF）动员乳腺癌患者外周血干细胞（peripheral blood stem cell,PBSC）的效果及影响因素分析。方法2006年2月至2009年6月我科收治行紫杉醇动员的26例乳腺癌患者,紫杉醇（PTX,175 mg/m2 持续静脉滴注24 h）化疗后,白细胞降至1.0×109/L左右时使用rhG-CSF 5 μg /(kg·d) 动员至采集结束。并进一步分析患者年龄,化疗后白细胞最低数,采集前各类血细胞数,术后分期以及既往化疗等因素对采集单个核细胞（mononuclear cell,MNC）、CD34+细胞数的影响。结果白细胞计数于紫杉醇化疗后中位7d降至1.0×109/L 左右,皮下注射rhG-CSF中位4d进行外周造血干细胞采集,采集总MNC平均(7.89±1.45)×108/kg,采集总CD34+细胞平均(4.88±1.54)×106/kg。年龄与采集CD34+细胞数显著相关。而其他因素对MNC及CD34+细胞数均无显著影响（P>0.05）。所有患者均未出现严重不良反应。结论PTX（175 mg/m2 持续静脉滴注24h）联合rhG-CSF为转移性乳腺癌患者动员的有效安全方案。患者年龄显著影响CD34+细胞的采集数量。     

乳腺癌干细胞上皮-间质转化标志物表达变化及意义

目的 探讨乳腺癌干细胞上皮．间质转化标志物表达变化及其临床意义。方法采用无血清悬浮培养法,从MCF-7细胞培养乳腺癌微球体细胞,应用流式细胞仪检测微球体细胞中CD44和CD24的表达,采用Westernblot方法检测微球体细胞中E-钙黏素、N-钙黏素、纤维连接蛋白、波形蛋白的表达水平,体外穿膜实验检测肿瘤细胞的迁移、侵袭能力。结果乳腺微球体富集了CD44＋CD24-的乳腺癌干细胞,并能在含血清培养基中增殖分化。该微球体细胞的上皮标志物E-钙黏素表达水平下调,而间质标志物N-钙黏素、纤维连接蛋白、波形蛋白的表达水平上调,同时乳腺癌干细胞的迁移、侵袭能力显著增强。结论乳腺癌干细胞具有上皮-间质转化的特征,具有显著增强的迁移、侵袭能力。     

SOX2在胰腺癌干细胞干性和化疗耐药中的作用及机制研究

[目的]探究性别决定相关基因簇2(SOX2)对胰腺癌干细胞干性与化疗耐药性的影响及影响机制。[方法]将培养至指数生长期胰腺癌干细胞随机分为SOX2抑制组、空白对照组与SOX2组,其中SOX2抑制组细胞与SOX2组细胞分别给予SOX2基因敲除与SOX2基因高表达处理,检测各组细胞SOX2 mRNA、八聚体结合转录因子4(OCT4)mRNA与谷胱甘肽过氧化物酶3(GPX3)mRNA表达水平;观察各组细胞增殖、侵袭及各组细胞对顺铂的耐药程度及在不同浓度顺铂处理下的凋亡情况。[结果] SOX2组细胞SOX2 mRNA与OCT4 mRNA表达水平明显高于SOX2抑制组与空白对照组细胞(P均=0.001);SOX2抑制组细胞增殖率与侵袭率明显低于空白对照组与SOX组(P<0.001);空白对照组细胞增殖率与侵袭率明显低于SOX2组(P<0.001);SOX2组细胞SOX2顺铂半数抑制浓度明显高于SOX2抑制组与空白对照组细胞(P<0.001);各浓度顺铂处理下的SOX2组细胞凋亡率均明显小于SOX2抑制组与空白对照组(P<0.001);SOX2组细胞GPX3 mRNA表达水平明显高于SOX2抑制组与空白对照组细胞(P<0.001)。[结论] SOX2能促进OCT4与GPX3表达,上调胰腺癌干细胞干性与化疗耐药性。     

Effects of Quinacrine on Expression of Hippo signaling Pathway Components (LATS1, LATS2, and YAP) in Human Breast Cancer Stem Cells

Objective: The Hippo signaling pathway has important role in the pathogenesis of some tumors. Breast cancer is the most prevalent cancer among females in the world. In recent years, various articles referred to inhibiting effect of quinacrine, a derivative of 9-aminoacridine, on the growth of several types of cancer cells. In this study, we evaluated the effect of quinacrine on expression of LATS1, LATS2, and YAP genes of the Hippo signaling pathway and YAP level in human breast cancer stem cells (MDA-MB 231 cell line). This cell line of breast cancer expresses the triple negative characteristics. Methods: MDA-MB 231 cells was treated with 0.5 µM of quinacrine for 3 days. The dose was selected using MTT assays. The expression of genes was quantified by Real-time PCR. The protein expression was performed by Western blotting. Significance of observations were checked by means of Mann-Whitney test using p