人前列腺癌靶向超声造影剂对荷瘤裸鼠靶向显像的研究

目的：探讨携前列腺特异性膜抗原（PSMA）抗体和携血管内皮生长因子（VEGF）抗体的靶向超声造影剂对荷人前列腺癌裸鼠的特异增强显像效果。方法：人前列腺癌细胞株LNCaP体外无菌条件下培养传代，BALB／c-nu雄性裸鼠皮下接种，建立荷人前列腺癌裸鼠动物模型。分别经荷瘤裸鼠尾静脉推注普通造影剂和携PSMA抗体及携VEGF抗体的靶向造影剂12min后采用能量多普勒成像，测量并计算肿瘤截面内血流信号所占的面积比，并分析比较普通造影剂和靶向造影剂对前列腺癌组织能量多普勒信号的增强效果。结果：成功建立了荷人前列腺癌裸鼠动物模型，推注携带VEGF抗体的靶向造影剂超声造影后行能量多普勒显像，可见前列腺癌组织能量多普勒信号显著增强，与普通造影剂和携带PSMA抗体的靶向造影剂相比较，差异有统计学意义（P〈0．01）。结论：VEGF抗体介导的靶向超声造影剂在体内可以有效特异性地与前列腺肿瘤新生血管内皮细胞结合，可作为诊断前列腺癌的靶向造影剂。能量多普勒延迟显像能有效显示靶向造影后肿瘤组织内的能量多普勒增强信号。     

前列腺癌组织中前列腺特异性膜抗原基因表达的临床意义

目的检测前列腺特异性膜抗原(PSMA)基因在前列腺癌组织中的表达,并探讨其意义。方法利用半定量(RT-PCR)法检测52例前列腺癌组织及35例前列腺增生组织中PSMA的表达。结果 (1)PSMA在前列腺癌及前列腺增生组织中的阳性表达率分别为84.6%(44/52)及68.6%(24/35),虽然前列腺癌组织中PSMA表达阳性率高于前列腺增生组织,但两者差异无统计学意义(P>0.05)。PSMA mRNA半定量结果在两组间存在差异,癌组织中PSMA的表达量明显高于增生组织中的表达,差异有统计学意义(P<0.05)。(2)PSMA基因表达量与肿瘤的分化程度有关,分化程度越低,表达量越高,高分化前列腺癌与低分化前列腺癌间差异有统计学意义(P<0.05)。结论 PSMA在前列腺癌中的表达与肿瘤的分化程度有关,且高于前列腺增生组织,提示PSMA可作为判断预后及分化程度的指标。     

68Ga-PSMA-11与18F-PSMA-1007在前列腺癌中的应用

前列腺癌的发病率在全球男性恶性肿瘤中排名第2,其发病率及病死率呈快速上升趋势。前列腺特异性膜抗原（PSMA）PET/CT 在前列腺癌的检测方面具有高灵敏度和特异度,可用于原发性前列腺癌的定位与分期、复发性前列腺癌的监测。目前,以PSMA为靶点的显像剂主要是放射性核素⁶⁸Ga和¹⁸F的化合物。该文综述了临床常用的小分子抑制剂⁶⁸Ga-PSMA-11和¹⁸F-PSMA-1007的特点,以及其在前列腺癌显像中的优缺点,为临床诊断提供依据,并指导其在不同患者中的临床应用。     

携载PSMA单抗靶向前列腺癌的纳米级脂质微泡的制备及体外寻靶能力

目的制备携载PSMA单抗靶向人前列腺癌的纳米级脂质微泡,并观察其体外寻靶能力。方法利用静电吸附法制备携载PSMA单抗靶向前列腺癌的脂质纳米级微泡,检测其一般特性;免疫荧光法检测抗体与纳米级微泡的结合情况;用细胞免疫荧光分析鉴定PSMA的表达及其在细胞膜上的定位;普通光镜下观察靶向微泡对前列腺癌细胞的寻靶能力,同时以胃癌细胞作对照。结果携载PSMA单抗的靶向纳米级微泡分布均匀,平均粒径为(623.70±66.05)nm,免疫荧光法显示微泡表面可见绿色荧光,细胞免疫荧光检测前列腺癌细胞膜高表达PSMA,体外寻靶实验显示该靶向微泡可与人前列腺癌LNCAP及C4-2细胞牢固结合,而与胃癌MKN45细胞不结合。结论本实验成功制备的携载PSMA单抗靶向人前列腺癌的纳米级脂质微泡具有较强的体外寻靶能力,能特异性地与前列腺癌细胞结合。     

荧光定量PCR检测前列腺特异膜抗原及其变异体DNA的方法学

目的拟建立以荧光定量逆转录聚合酶链反应（FQ-RT-PCR）检测前列腺特异性膜抗原（PSMA）及其变异体5（PSMA5）DNA的定量方法。方法自行设计三对引物,自制PC3.0-T-PSMA及PC3.0-T-PSMA5质粒作为标准品,采用FQ-RT-PCR检测前列腺癌LNCap细胞中PSMA及PSMA5 DNA的含量。结果标准曲线的斜率为-0.29,相关系数为0.995,成功地扩增出前列腺癌LNCap细胞中PSMA及PSMA5基因的目标DNA片段。结论应用自行设计的引物及标准品的FQ-RT-PCR能对前列腺癌细胞株中PSMA及变异体PSMA5的DNA进行定量PCR检测。     

超声分子影像组学评估前列腺癌前列腺特异膜抗原表达：实验研究

目的 观察超声分子影像组学评估前列腺癌(PCa)前列腺特异膜抗原(PSMA)表达的价值。方法 分别于裸鼠皮下接种22RV1细胞(PSMA阳性表达,n=9)或PC-3细胞(PSMA阴性表达,n=10),构建不同PSMA表达水平人前列腺癌细胞移植瘤裸鼠模型。制备靶向PSMA的超声纳米泡(PSMA-NB),进行表征检测及体内、外超声分子成像。以7 ∶ 3比例将裸鼠随机分为训练集及测试集,提取训练集纹理特征,以组内相关系数、单因素分析、最小绝对收缩和选择算子等方法筛选特征,构建超声分子影像组学模型,并评估模型诊断PSMA阳性PCa的效能。结果 所制备PSMA-NB平均粒径为(392.2±31.56)nm,Zeta电位为(-29.3±0.95)mV,体外显像效果理想,稳定性好。共基于训练集数据提取1 561个纹理特征,最终筛选出3个特征,包括exponential_glszm_ZoneVariance、wavelet_HHH_gldm_SmallDependenceLowGrayLevelEmphasis及logarithm_gldm_DependenceVariance,以之构建的超声分子影像组学模型诊断训练集、测试集PSMA阳性表达PCa的曲线下面积分别为0.950、0.875。结论 基于靶向PSMA超声分子影像组学模型可用于评估PCa裸鼠PSMA表达。     

RT-PCR检测前列腺癌患者PSMA mRNA表达的临床意义

目的应用巢氏逆转录聚合酶链反应（RT-PCR）检测前列腺癌（PCa）患者外周血前列腺特异性膜抗原mRNA（PSMA mRNA）的表达并探讨其临床应用意义。方法采用巢氏RT—PCR检测28例PCa患者、20例前列腺增生（BPH）患者和10例健康男性外周血中PSMA mRNA的表达情况。结果28例PCa患者中PSMA mRNA表达阳性18例。20例BPH患者和10例健康男性均为阴性。检测灵敏度为64．29％,特异性为100．00％。18例接受内分泌治疗的患者中PSMA mRNA表达阳性13例,阳性率为72．22％,未接受内分泌治疗的患者阳性率为40．oo％。PSMA mRNA与前列腺特异性抗原（PSA）水平具有明显相关性,随着PSA水平的升高PSMA mRNA阳性率也随之升高,各组间比较差异有统计学意义（P〈0．05）。PSMA mRNA与PCa的临床分期也有明显的相关性,D期PCa患者的PSMA mRNA阳性率明显高于B、C期,差异有统计学意义（P〈0．05）。PSMA mRNA与PCa的病理分期无明显相关性。PSMA mRNA与PCa的转移有明显的相关性,己出现骨转移者PSMA mRNA阳性率明显升高。差异有统计学意义（P〈0．05）。结论巢氏RT—PCR检测PCa患者外周血中PSMA mRNA表达的方法是一种十分有应用价值的诊断PCa及监测PCa微转移的方法,特别是对接受过内分泌治疗的患者,检测PSMA mRNA要比PSA mRNA更有意义。     

前列腺特异性膜抗原PET/MRI在前列腺癌中的应用现状与前景

前列腺癌(prostate cancer, PCa)是常见的老年男性恶性肿瘤,并是其主要致死原因之一。前列腺特异性膜抗原(prostate specific membrane antigen, PSMA)是一种PCa组织特异性高表达的跨膜蛋白,这使得PSMA成为PCa良好的特异性分子影像靶点。目前,PSMA PET/CT在PCa诊断、分期等方面的价值已被广泛认可。随着PET/MRI逐步进入临床,PSMA PET与MRI两种诊断PCa的重要影像学方法的有机结合成为可能。本文就PSMA PET/MRI在PCa中的应用现状与前景进行阐述,以期为临床诊疗提供借鉴。     

前列腺特异膜抗原(PSMA)的检测及临床应用进展

前列腺特异膜抗原 (PSMA)是一种新近发现的前列腺癌相关抗原 ,由于其较高的前列腺特异性和膜结合特性 ,使其在前列腺癌的临床诊断中 ,特别是在判断肿瘤的转移和进展方面 ,是一种较PSA更加敏感、特异的CaP肿瘤标记物。     

前列腺特异膜抗原（PSMA）的检测及临床应用进展

前列腺特异膜抗原（PSMA）是一种新近发现的前列腺癌相关抗原,由于其较高的前列腺特异性和膜结合特性,使其在前列腺癌的临床诊断中,特别是在判断肿瘤的转移和进展方面,是一种较PSA更加敏感,特异的CaP肿瘤标记物。     

Synthesis and in vitro evaluation of MR molecular imaging probes using J591 mAb‐conjugated SPIONs for specific detection of prostate cancer

Carcinoma of the prostate is the most frequent diagnosed malignant tumor in men and is the second leading cause of cancer‐related death in this group. The cure rate of prostate cancer is highly dependent on the stage of disease at the diagnosis and early detection is key to designing effective treatment strategies. The objective of the present study is to make a specific MR imaging probe for targeted imaging of cancer cells. We take advantage of the fact that many types of prostate cancer cells express high levels of prostate‐specific membrane antigen (PSMA) on their cell surface. The imaging strategy is to use superparamagnetic iron oxide nanoparticles (SPIONs), attached to an antibody (J591) that binds to the extracellular domain of PSMA, to specifically enhance the contrast of PSMA‐expressing prostate cancer cells. Conjugation of mAb J591 to commercial SPIONs was achieved using a heterobifunctional linker, sulfo‐SMCC. Two types of prostate cancer cell lines were chosen for experiments: LNCaP (PSMA+) and DU145 (PSMA?). MRI and cell uptake experiments demonstrated the high potential of the synthesized nanoprobe as a specific MRI contrast agent for detection of PSMA‐expressing prostate cancer cells. Copyright © 2012 John Wiley & Sons, Ltd.     

In vitro and in vivo responses of advanced prostate tumors to PSMA ADC, an auristatin-conjugated antibody to prostate-specific membrane antigen

Prostate-specific membrane antigen (PSMA) is a membrane protein that is overexpressed manifold in prostate cancer and provides an attractive target for therapy. PSMA ADC is an antibody-drug conjugate (ADC) that consists of a fully human anti-PSMA monoclonal antibody conjugated to monomethylauristatin E through a valine-citrulline linker. In this study, the antitumor activity of PSMA ADC was evaluated against a panel of prostate cancer cell lines in vitro and in a novel in vivo model of taxane-refractory human prostate cancer. In vitro cell killing was efficient for cells with abundant PSMA expression (>10(5) molecules/cell; IC(50) ≤ 0.022 nmol/L) and 1,000-fold less efficient for cells with undetectable PSMA (IC(50) > 30 nmol/L). Intermediate potency (IC(50) = 0.80 nmol/L) was observed for cells with approximately 10(4) molecules of PSMA per cell, indicating a threshold PSMA level for selective cell killing. Similar in vitro activity was observed against androgen-dependent and -independent cells that had abundant PSMA expression. In vitro activity of PSMA ADC was also dependent on internalization and proper N-glycosylation/folding of PSMA. In contrast, less potent and nonselective cytotoxic activity was observed for a control ADC, free monomethylauristatin E, and other microtubule inhibitors. PSMA ADC showed high in vivo activity in treating xenograft tumors that had progressed following an initial course of docetaxel therapy, including tumors that were large (>700 mm(3)) before treatment with PSMA ADC. This study defines determinants of antitumor activity of a novel ADC. The findings here support the clinical evaluation of this agent in advanced prostate cancer.     

Targeting prostate cancer: Prostate-specific membrane antigen based diagnosis and therapy

The high incidence rates of prostate cancer (PCa) raise demand for improved therapeutic strategies. Prostate tumors specifically express the prostate-specific membrane antigen (PSMA), a membrane-bound protease. As PSMA is highly overexpressed on malignant prostate tumor cells and as its expression rate correlates with the aggressiveness of the disease, this tumor-associated biomarker provides the possibility to develop new strategies for diagnostics and therapy of PCa. Major advances have been made in PSMA targeting, ranging from immunotherapeutic approaches to therapeutic small molecules. This review elaborates the diversity of PSMA targeting agents while focusing on the radioactively labeled tracers for diagnosis and endoradiotherapy. A variety of radionuclides have been shown to either enable precise diagnosis or efficiently treat the tumor with minimal effects to nontargeted organs. Most small molecules with affinity for PSMA are based on either a phosphonate or a urea-based binding motif. Based on these pharmacophores, major effort has been made to identify modifications to achieve ideal pharmacokinetics while retaining the specific targeting of the PSMA binding pocket. Several tracers have now shown excellent clinical usability in particular for molecular imaging and therapy as proven by the efficiency of theranostic approaches in current studies. The archetypal expression profile of PSMA may be exploited for the treatment with alpha emitters to break radioresistance and thus to bring the power of systemic therapy to higher levels.     

Changing the Goal Posts: Prostate-specific Membrane Antigen Targeted Theranostics in Prostate Cancer

ObjectiveProstate-specific membrane antigen (PSMA) theranostics is changing the face of prostate cancer diagnosis and therapy. PSMA, a transmembrane protein over-expressed in many prostate cancers, is a promising target for theranostics. Theranostics is the concept of small molecule proteins that are labelled to different radionuclides and can be used for either diagnosis or therapy, dependent on whether they are labelled with an imaging or therapy radionuclide. By directly targeting the cancer cells with imaging and then for therapy, this approach embodies the philosophy of precision medicine – right drug, right time, right dose. The question is how to best utilise these new imaging and therapy agents in clinical practice. This review will evaluate the importance of PSMA in prostate cancer, its role in diagnostic imaging, and its potential as a therapy of advanced prostate cancer.Data SourcesElectronic databases including MEDLINE, Scopus, professional websites were searched.ConclusionPSMA-directed theranostics has an expanding role in prostate cancer because of its utility as a sensitive diagnostic tool that can be coupled with efficacious and low-toxicity therapeutic options. Ongoing research is required to determine how to use this effective tool for best patient care.Implications for Nursing PracticePSMA theranostics is rapidly being incorporated into the routine care of men with prostate cancer. Understanding its strengths, its limitations, and where it may be valuable in clinical care is important in undertaking best patient practice.     

N-glycosylation and microtubule integrity are involved in apical targeting of prostate-specific membrane antigen: implications for immunotherapy

Prostate-specific membrane antigen (PSMA) is an important biomarker expressed in prostate cancer cells with levels proportional to tumor grade. The membrane association and correlation with disease stage portend a promising role for PSMA as an antigenic target for antibody-based therapies. Successful application of such modalities necessitates a detailed knowledge of the subcellular localization and trafficking of target antigen. In this study, we show that PSMA is expressed predominantly in the apical plasma membrane in epithelial cells of the prostate gland and in well-differentiated Madin-Darby canine kidney cells. We show that PSMA is targeted directly to the apical surface and that sorting into appropriate post-Golgi vesicles is dependent upon N-glycosylation of the protein. Integrity of the microtubule cytoskeleton is also essential for delivery and retention of PSMA at the apical plasma membrane domain, as destabilization of microtubules with nocodazole or commonly used chemotherapeutic Vinca alkaloids resulted in the basolateral expression of PSMA and increased the uptake of anti-PSMA antibody from the basolateral domain. These results may have important relevance to PSMA-based immunotherapy and imaging strategies, as prostate cancer cells can maintain a well-differentiated morphology even after metastasis to distal sites. In contrast to antigens on the basolateral surface, apical antigens are separated from the circulation by tight junctions that restrict transport of molecules across the epithelium. Thus, antigens expressed on the apical plasma membrane are not exposed to intravenously administered agents. The ability to reverse the polarity of PSMA from apical to basolateral could have significant implications for the use of PSMA as a therapeutic target.     

Disulfide-constrained peptides that bind to the extracellular portion of the prostate-specific membrane antigen

The prostate-specific membrane antigen (PSMA) is a well-characterized surface antigen, overexpressed in the most advanced, androgen-resistant human prostate cancer cells. We sought to exploit PSMA cell surface properties as a target for short peptides that will potentially guide protein-based therapeutics, such as viral vectors, to prostate cancer cells. Two separate phage display peptide strategies were applied, in parallel, to purified PSMA protein bound to two separate substrates. We reasoned that peptide sequences common to both substrate selections would be specific binders of PSMA. Additionally, the design allowed for stringent cross-selections, where phage populations from one selection condition could be applied to the alternative substrate. These strategies resulted in a series of phage displayed peptides able to bind to PSMA by ELISA and direct binding assays, both with purified protein and in prostate cancer cells. Cell binding is competitively inhibited by purified PSMA. The synthesized peptides are capable of enhancing PSMA carboxypeptidase enzymatic activity, suggesting protein folding stabilization. The discovery of these peptides provides the foundation for subsequent development of peptide targeted therapeutics against prostate cancer.     

Generation and characterization of nanobodies targeting PSMA for molecular imaging of prostate cancer

Nanobodies show attractive characteristics for tumor targeting in cancer diagnosis and therapy. A radiolabeled nanobody binding the prostate‐specific membrane antigen (PSMA) could offer a noninvasive strategy to select prostate cancer patients eligible for PSMA‐targeted therapies. We here describe the generation, production and in vivo evaluation of anti‐PSMA nanobodies. Nanobodies were derived from heavy‐chain‐only antibodies, raised in immunized dromedaries. Binding characteristics were evaluated through ELISA and flow cytometry. Selected nanobodies were radiolabeled with ^99mTc at their hexahistidine tail, after which cell binding capacity and internalization were evaluated on PSMA^pos LNCaP and PSMA^neg PC3 cell lines. In vivo tumor targeting was analyzed in both LNCaP and PC3 xenografted mice through SPECT/microCT and tissue sampling. A panel of 72 generated clones scored positive on ELISA, all contributing to three nanobody groups, of which group 3 dominated with 70 clones. ELISA and FACS analysis led to the selection of two dominant nanobodies. ^99mTc‐labeled PSMA6 and PSMA30 both showed specific binding on LNCAP cells, but not on PC3 cells. ^99mTc‐PSMA30 internalized significantly more in LNCaP cells compared to ^99mTc‐PSMA6. Higher absolute tumor uptake and tumor‐to‐normal organ ratios were observed for ^99mTc‐PSMA30 compared with ^99mTc‐PSMA6 and a ^99mTc‐control nanobody in LNCaP but not in PC3 tumor‐bearing mice. PSMA30 nanobody has improved targeting characteristics both in vitro as well as in vivo compared with PSMA6 and the control nanobody, and was therefore selected as our in‐house‐developed lead compound for PSMA targeting. Copyright © 2014 John Wiley & Sons, Ltd.