Expression of two cytochromes P450 involved in carcinogen activation in a human colon cell line

Cytochrome P450 is known to cause carcinogen activation and correspondingly increased cancer risk in animal models. In order to determine whether P450 in the colon may be involved in cancer development in the human, the human colon cell line LS174T was examined for the presence of various cytochromes P450. Two isozymes of P450 were identified in the human cell line. Expression of P450IAl or IA2 was increased by treatment of the cell line with benzanthracene; the induction was demonstrated by an increase in RNA hybridizing to a probe for P4501Al and by ethoxyresorufin deethylation activity. Western analysis of microsomes isolated from human colon tissue also demonstrated the presence of P4501A1, as well as a form which cross-reacted to an antibody to human P450IIC9. Another isozyme, P450IIE1, was identified by polymerase chain reaction amplification of RNA from LS174T cells. These results underscore the presence of cytochromes P450 in colonic tissue and provide a basis for the involvement of isozyme-specific P450 mediated reactions in carcinogenesis of the colon.Some of the data presented here were taken from a thesis submitted by D.K.H. in partial fulfillment of the requirements for the Ph.D. degree in the University of Texas Graduate School of Biomedical Sciences.     

In situ activation of mouse macrophages and therapy of spontaneous renal cell cancer metastasis by liposomes containing the lipopeptide CGP 31362

Summary We determined whether the intravenous administration of multilamellar vesicle liposomes (MLV) containing a lipopeptide analogue of a fragment from the cell wall of gram-negative bacteria (CGP 31 362) can render BALB/c mouse alveolar macrophages tumoricidal in situ and reduce the incidence of spontaneous lung metastasis of syngeneic renal carcinoma (RENCA) cells. Alveolar macrophages (a) incubated in vitro with MLV containing CGP 31 362 (MLV-31 362) and (b) harvested from mice injected i.v. with MLV-31 362 were rendered cytotoxic against the RENCA cells. Maximum cytotoxic activity of the macrophages was induced by injecting 5 µmol MLV consisting of 250 mg phospholipids and 0.5 mg CGP 31 362. The single i.v. injection of 5 µmol MLV-31 362 produced activation of macrophages that lasted for up to 4 days. Repeated i. v. injections of MLV-31 362 produced a continuous antitumor activity in alveolar macrophages. To study the lipopeptide's effects on metastasis, we injected the left kidneys of BALB/c mice with RENCA cells. The kidney with growing tumor was resected 10 days later and, after a further 2 days, groups of mice were injected i.v. with MLV-31 362 or with MLV-HBSS (twice weekly for 3 weeks). Treatment with MLV-31 362 significantly decreased the median number of spontaneous lung metastases. These data demonstrate that the systemic administration of MLV-31 362 can activate murine lung macrophages in situ and reduce the incidence of spontaneous RENCA lung metastases.     

Two monoclonal antibodies against small-cell lung cancer show existence of synergism in binding

Summary Murine IgG1 monoclonal antibodies (mAbs), ITK-2 and ITK-3, were generated against a small-cell lung cancer (SCLC) cell line. Enzyme-linked immunosorbent assay using a variety of established cell lines as substrates, immunoperoxidase staining of freshly frozen tissue sections, and fluorescence-activated cell sorter analysis of peripheral blood leukocytes showed that these mAbs recognize a part of the SCLC-associated cluster 1 antigen. In immunoprecipitation studies, both ITK-2 and ITK-3 bound to a 145-kDa glycoprotein of SCLC cell membrane extracts, as did MOC-1 and NKH-1, which both recognize the cluster 1 antigen. However, because the binding of¹²⁵I-labeled ITK-2 to SCLC cells was not inhibited by MOC-1 or NKH-1, the binding site of ITK-2 on SCLC cells appeared to be different from that of either MOC-1 or NKH-1. Unexpectedly, binding of¹²⁵I-labeled ITK-2 to SCLC cells increased in the presence of ITK-3. This ITK-3-induced increase in ITK-2 binding was due partly to an increase in the number of binding sites for ITK-2 on SCLC cells. Addition of ITK-3 may, therefore, improve the effectiveness of ITK-2-based tumor detection or therapy.     

Phase I trial of intravenous infusion of ex-vivo-activated autologous blood-derived macrophages in patients with non-small-cell lung cancer: Toxicity and immunomodulatory effects

Summary The purpose of this phase I study was to evaluate the toxicity and biological activity of autologous blood-derived macrophages activated ex-vivo with recombinant human interferon (rhuIFN) [monokine-activated killer (MAK) cells] and administered intravenously to 11 lung cancer patients once a week for 6 consecutive weeks. Peripheral blood monocytes were collected by leukapheresis and then purified by counterflow elutriation. The MAK cells were generated by culturing the purified monocytes in Teflon bags for 7 days and adding rhuIFN to the cultured cells for the last 18 h. These MAK cells expressed differentiation-associated surface antigen MAX1, and were cytotoxic in vitro against tumour cell line U937. The MAK cells were infused at dose levels from 1 × 10⁷ to 5 × 10⁸ on an intrapatient dose-escalating schedule. No severe adverse side-effects occurred. Toxicity was mild to moderate [primarly fever (75%) and chills (32%)], non-dose-dependent, and non-cumulative. No consistent change in haemostatic function, or liver or renal function was observed. Dose-limiting toxicity was not reached at 5 × 10⁸ cells (optimal dose reproduced for each patient). The maximum tolerated dose was not determined. The immunomodulatory activity of i.v. infused MAK cells was demonstrated both in vivo by significant increases in granulocyte count and neopterin level in the patients' peripheral blood postinfusion and in vitro by secretory products (IL-1. TNF, neopterin, and thromboplastin-like substance) in the culture supernatants. The in vivo traffic patterns of autologous MAK cells labelled ex-vivo with¹¹¹In oxine were studied in 7 patients. Gamma imaging showed an immediate but transient lung uptake (<24 h), and a progressive uptake of radioactivity in the liver and spleen was seen from 6 h to 72 h post-infusion. Our results indicate that the preparation of high numbers of autologous, blood-derived MAK cells is a feasible procedure, and their transfusion is safe for patients. This immunotherapeutic approach seems to be encouraging from the point of view of establishing an adjuvant therapeutic modality in cancer patients with minimal residual disease.This work was supported in part by a grant 6911 from the Association pour la Recherche contre le Cancer (ARC), grants from the Ligue Nationale contre le cancer and the Ligues Regionales (Bas-Rhin, Haut-Rhin) contre le cancer, and contract 891013 from the Institut National pour la Santé et la Recherche Médicale (INSERM), France     

正常中国人中枢运动系统传导时间的测定

本文应用高电压、低输出阻抗刺激器,经皮给予大脑皮层和脊髓电刺激(BSPES),同时在上肢鱼际(Thenar)和下肢胫骨前肌(Muscle tibialis anterior)上记录诱发肌肉动作电位,测定了64名正常健康中国人(男:46;女:18)的中枢运动系统传导时间。受试者年龄为20—67岁,身高为156—185cm。刺激大脑皮层出现反应的潜伏期与刺激脊髓出现反应的潜伏期之差为中枢运动传导时间(CMCT)。实验测得鱼际的 CMCT 为6.69±1.48ms;胫骨前肌的 CMCT 为12.90±1.59ms。经统计学处理证明,CMCT 与左右侧肢体、性别、年龄及身高无关。说明 CMCT 是无损伤测定与评价中枢运动系统功能的较精确的一种客观指标。本文根据所测数据,计算出脊髓内运动传导速度为71.34±10.89 m/s,与文献报道的锥体束传导速度50—70 m/s 相近。因此,CMCT 反映了锥体束的传导时间。     

Mesenchymal control of branching pattern in the fetal mouse lung

The effect of mesenchyme on specialization of respiratory epithelium in the fetal mouse was tested in organ cultures. Heterologous combinations were made between respiratory and non-respiratory lung epithelia and the corresponding mesenchymes. Isolated terminal respiratory buds of fetal mouse lungs were recombined with mesenchyme from chick lung parabronchi, mouse trachea or from the avascular, non-respiratory air sacs of chick lungs. Isolated non-branching chick air sacs were combined with mouse terminal bud mesenchyme or mesenchyme from the respiratory branches of chick lungs. Air sac epithelia branched in a pattern characteristic of the chick lung when combined with chick respiratory mesenchyme and in a pattern characteristic of mouse lung when combined with mouse terminal bud mesenchyme. Mouse terminal bud epithelia did not branch with either mouse tracheal mesenchyme or chick air sac mesenchyme but branched in a chick pattern with chick parabronchial mesenchyme. Electron microscopic examination of the cultures showed that all chick air sac epithelial cultures failed to produce surfactant (lamellar bodies) even when they branched. Control cultures of mouse terminal buds contained large numbers of lamellar bodies; mesenchyme which suppressed branching reduced the number of lamellar bodies to only a few in a small proportion of the cells. Culture medium supplemented with growth factors and hormones increased the number of lamellar bodies in heterologous mouse combinations but did not bring the number to control levels. Supplemented medium had no effect on lamellar body production by chick air sac epithelium. The results indicate that branching pattern is determined by the mesenchyme surrounding the epithelial primordium. However, the capacity to synthesize surfactant is determined by the source of the epithelium; mesenchyme may control the degree of expression but not the absolute presence or absence of the differentiated condition.     

人体肝癌细胞系(SMMC-7721)的糖皮质激素受体以及糖皮质激素对酪氨酸转氨酶的诱导

本文以[~3H]Dex为配体,采用完整细胞GCR和核特异结合百分率的测定方法检测了人体肝癌细胞系(SMMC-7721)的GCR。实验表明,该细胞具有高亲和力、低容量、能与GC进行特异结合的Dex结合部位,该结合部位与[~3H]Dex结合后可向核内转位,因而具备了作为GCR的基本条件。但该细胞的GCR与正常细胞的GCR相比较亦有些差别,GC与受体结合后不能诱导TAT的生成。本文对上述改变的可能机理进行了讨论。     

甲基苄基亚硝胺对与人、大鼠和鸡肝脏或食管上皮细胞混合培养的V_(79)细胞的致突变作用

用人,雄性Wistar大鼠和雄性莱亨鸡的肝细胞,或食管上皮细胞与V_(79)细胞混合培养,观察甲基苄基亚硝胺(MBN)经肝细胞或食管上皮细胞代谢后,能否产生活性代谢产物,从而诱导V_(79)细胞突变及其它有关的生物学反应。研究结果初步表明,大鼠和鸡肝细胞可以明显代谢MBN产生活性代谢产物,诱导混合培养的V_(79)细胞SCE和微核增高以及6-TG抗性突变。人胎儿肝细胞也可以代谢这种亚硝胺并诱导V_(79)细胞6-TG抗性突变,但对SCE和微核诱导明显偏低。说明这三种肝细胞代谢MBN在性质上是比较近似,可能在代谢激活程度上有一定差异。在食管上皮细胞和V_(79)细胞混合培养的实验中,大鼠食管上皮可以明显代谢MBN诱导V_(79)细胞SCE和微核的增高及6-TG抗性突变。鸡食管上皮细胞未见有明显的代谢MBN引起V_(79)细胞突变的作用。26只雄性莱亨鸡喂以MBN实验最长者达975天,总剂量为1674毫克,结果未见一例鸡发生食管癌。结果表明鸡的食管上皮可能对某些挥发性亚硝胺的致癌作用是不敏感的,目前没有证据支持在林县高发区中所发现的鸡咽食管癌是由于已找到的挥发性亚硝胺引起的。人食管上皮细胞具有一定的代谢MBN产生有致突变作用的代谢产物的能力,然而人与人之间有明显的个体差异。和大鼠相比,人食管代谢MBN诱导V_(79)细胞突变的能力明显较低,有关亚硝胺在高发区人食管癌发生中的作用还有待进一步深入研究。     

抗人肺癌细胞单克隆抗体的制备及其特性的研究

用人肺鳞癌细胞LTEP-78细胞系免疫Blab/c小鼠获得3株抗人肺癌细胞的单克隆抗体杂交瘤系。其中BLTI-01株经六次克隆化培养,体外传代8个月以上。BLTI-01与白细胞抗原及血型抗原基本上无交叉反应;与骨髓细胞无交叉反应;与癌胚抗原和胎甲球蛋白不相关;与肺鳞癌、肺腺癌细胞系及部分其它肿瘤细胞呈阳性反应。     

癌症新型的诊疗靶点－APOBEC

APOBEC（“载脂蛋白质B mRNA编辑催化多肽”）是一类进化保守的胞苷脱氨酶家族。在人体内,已知含有保守的DNA胞嘧啶脱氨酶结构域的基因共有11种,包括AID、APOBEC1、APOBEC2、APOBEC3基因家族APOBEC3A、APOBEC3B、APOBEC3C、APOBEC3DE、APOBEC3F、APOBEC3G、APOBEC3H（分别称为A3A、A3B、A3C、A3D、A3F、A3G和A3H）和APOBEC4。APOBEC利用其脱氨酶活性通过与RNA和/或DNA结合,催化mRNA或使DNA中的胞嘧啶核苷酸转变为尿嘧啶,或者胞嘧啶核苷酸转变为胸腺嘧啶核苷酸,进而完成各自不同的功能。目前研究发现,AID及APOBEC3（A3s）的7种脱氨酶在人类的天然免疫和适应性免疫防御过程中发挥重要的作用,且在口腔癌,肺癌（腺癌和鳞状细胞癌）,结直肠癌和乳腺癌等的诊疗过程中具有重要的潜在应用价值。AID可以通过将胞嘧啶脱氨基成尿嘧啶,来启动SHM (体细胞超突变)和CSR (类别转换重组),进而在抗体多样性方面发挥作用。它的异常表达能够使B细胞淋巴瘤等恶性肿瘤的发病频率显著增加。而A3A、A3B通过胞嘧啶到尿嘧啶转换,以及自身表达量上调而在乳腺癌和肺癌诊疗中起作用。A3G通过APOBEC3G/miR 29/MMP2为了解结直肠癌肝转移和开发治疗晚期结肠癌的有效疗法开辟了新的途径。综上所述,本文将以AID,A3A,A3B,A3G为例子,对APOBEC在癌症诊断和治疗方面的应用进行综述,以期为进一步药物研究和临床应用等提供参考。