放射免疫结合外照射对实体瘤的实验研究

目的 探讨放射免疫结合外照射对实体瘤的外照射剂量、二者最佳的应用时机与顺序。方法 TS1是一种抗细胞角蛋白8的单克隆抗体，它可以与肿瘤坏死区域中的细胞角蛋白(CKs)8特异地结合，用^131I标记单克隆抗体形成放射性螫合物。60只荷瘤(人宫颈鳞癌)小鼠用于研究：24只做预实验，观察肿瘤的退缩情况，决定合适的外照射剂量；36只做正式实验，分为6个组，分别给不同的治疗方案后比较肿瘤中放射活度比、平均累积吸收剂量、累积吸收总量以及全身放射性螯合物药代动力学。结果 预实验中，当肿瘤接受20Gy外照射时生长明显受抑，增加剂量肿瘤受抑幅度减小。正式实验中，给药前进行外照射组，肿瘤放射活度比、平均累积吸收剂量、累积吸收总量最高，与其他组相比有显著性差异。结论 ^131I-TS1放射免疫导向结合外照射治疗实体瘤，在给药前行20Gy的外照射组中，肿瘤放射活度比、平均累积吸收剂量、累积吸收总量最高，疗效最佳。     

核素标记不同性质单抗瘤内注射治疗大肠癌的研究

目的：探讨核素标记抗细胞膜抗原单克隆抗体联合核素标记抗细菌核单克隆抗体瘤内注射治疗荷人大肠癌裸鼠移植瘤的可行性和优越性,为临床应用核素标记多种不同性质的单克隆抗体进行放射免疫治疗提供实验依据。方法：在裸鼠荷人大肠癌Lovo移植瘤生长至1cm左右时,瘤内分别或同时注射^131I标记的抗大肠癌细胞膜抗原CL3单克隆抗体（^131I-CL3)和^131I标记人／鼠嵌合抗细胞核的单克隆抗体chTNT(^131I-chTNT),治疗后行SPECT显像观察标记抗体在肿瘤内的浓聚,并进行疗效观察。结果：研究发现2种标记抗体联合应用组的肿瘤抑制率为82．3％,明显高于131I－CL3－IT组的57．9％或131I－chTNT组的53．1％,联合应用组标记抗体在肿瘤组织／非肿瘤组织内的放射活度比值（T／NT值）大于单独应用组。结论：131I－CL3联合131I－chTNT瘤内注射放射免疫治疗可增加标记抗体在肿瘤组织内的浓聚,并能提高放射免疫治疗效果。     

^131I—chTNT单克隆抗体瘤体内注射治疗肺癌疗效观察

目的：观察^131I-chTNT(^131I标记的肿瘤标记细胞核单克隆抗体）瘤体内注射治疗肺癌的疗效和毒性。方法：25例肺癌患者采用^131I-chTNT注射液,每人2次,每次0.8mCi/kg瘤体内直接注射。结果：有效率48％,CR：8％,PR：40％。仅2例发生气胸,骨髓抑制0－Ⅱ度。结论：^131I-chTNT瘤体内注射疗效高,毒副反应轻,值得进一步应用观察。     

^131I标记人源抗HBsAg Fab对荷人肝癌裸鼠的放射免疫治疗

目的 ^131I标记人源抗HBsAg Fab经腹腔注射治疗荷人肝癌裸鼠移植瘤,与^131I-S102比较,评价其作为肝癌放射免疫治疗（RIT）的可能性。方法 荷瘤裸鼠分为4组分别经腹腔注射不同放射剂量的^131I-抗HBsAg Fab、^131I-S102、^131I-无关Fab及PBS。按一定时间间隔作组织分布和血液清除速率测定,继续观察4周,以外周血白细胞和血小板数量变化行毒性分析,计算各组肿瘤生长抑制率。结果 ^131I-抗HBsAg Fab被肿瘤区的摄取和在血液中清除速率均明显快于^131I-S102,同等剂量下,前者抗肿瘤疗效略低,但血液毒性明显减轻。结论 人源抗HBsAg Fab具有被肿瘤快速摄并在体内正常组织中快速清除,毒性减低,具有良好的抗肿瘤疗效,是原发性肝癌RIT的理想载体。     

131^I标记抗人成骨肉瘤单抗荷瘤裸鼠体内分布和放射免疫显像研究

目的：研究131^I标记抗人成骨肉瘤单克隆抗体在荷人成骨肉瘤裸鼠体内分布和放射免疫定位显像。方法：采用Iodogen固相法标记制备131^—HOS McAb。25只荷瘤裸鼠随机分为5组，分别腹腔注射131^I—HOS McAb后，于6、12、24、48和72 h 5个时间段进行裸鼠的体内分布研究；对5只荷瘤裸鼠分别腹腔注射131^I—HOS McAb后，于6、12、24、48和72h 5个时间段进行荷瘤裸鼠的放射免疫定位显像研究。结果：在荷人成骨肉瘤裸鼠腹腔注射131^I—HOS McAb后，12h肿瘤与血的T／NT比值为1．37，24h为3．75，48h达到最高为5．24。腹腔注射131^I—HOS McAb后，12h肿瘤部位即可见明显放射性浓聚，48h本底明显降低，肿瘤呈放射性热区。结论：131^I—HOS McAb对成骨肉瘤定向性较好，对放射免疫定位显像有利，为进一步放射免疫治疗研究提供了理论基础。     

^131I标记抗CD20单克隆抗体在荷瘤裸鼠体内的放射免疫显像

目的探讨^131I-anti-CD20McAb经瘤内注射后在荷人Burkitt’s淋巴瘤细胞系Raji细胞移植瘤裸鼠体内的放射免疫显像。方法^131I标记物的标记采用IODO-GEN碘化标记；注射标记物后第1、3、7、15天将荷瘤裸鼠SPECT显像后活杀，定标器测量并计算瘤、血等12种器官或组织的％ID／g值，根据MIRD委员会推荐的公式计算肿瘤累积吸收剂量。结果^131I-anti-CD20McAb瘤内注射组的SPECT显像结果优于腹腔注射组和^131I-IgG瘤内注射组，该组肿瘤％ID／g值在给药后第1、3和7天分别为后两组的1．4～17倍和1．7～3．7倍，肿瘤累及吸收剂量在给药后第3、7和15天分别为后两组的1．5～2．5倍和6．0～12．6倍。结论^131I-anti—CD20McAb经瘤内途径给药可以使肿瘤获得最高的放射性药物摄取率．为下一步运用该途径进行放射免疫治疗提供了实验依据。     

^131I照射对分化型甲状腺癌细胞摄碘水平及NIS mRNA表达的影响

目的：探讨^131I不同时间照射后分化型甲状腺癌细胞摄取放射性碘（^131I）水平的变化以及检测是否影响NIS mRNA表达。方法：分化型乳头状甲状腺癌细胞株（GTHW3）培养在DMEM培养基中,应用同一活度（20μCi）的^131I对培养癌细胞进行不同时间（6h、12h、24h、48h）的照射。γ计数仪测定放射性碘（^131I）摄取率。采用RT—PCR法检测甲状腺癌细胞的NIS mRNA表达。结果：不同时间的放射性^131I照射使甲状腺癌细胞摄取^131I降低;凝胶电泳显示放射性^131I照射24h后GTHW3细胞NIS mRNA表达降低;照射组各时间点的^131I摄取率及24h和48h的NIS mRNA表达与未照射对照组比较均有显著性差异。结论：^131I照射可使DTC细胞的NIS mRNA表达降低,提示^131I照射致DTC细胞失分化及其摄碘能力降低可能与DTC细胞NIS mRNA表达降低有关。     

188Re-CL58、131I-chTNT放射免疫治疗的临床研究

目的：比较多次小剂量与大剂量放射免疫治疗(RIT)对实体瘤的初步临床疗效及不良反应。方法：采用多次小剂量^188Re—CL58及大剂量^131I-chTNT对临床病例治疗。用脱氧葡萄糖(FDG)—正电子发射断层扫描(PET)显像方法及动态观察血清中肿瘤标志物变化监测疗效。用间接免疫荧光法评价RIT对T淋巴细胞亚群的影响观察其对造血系统和肝、肾等器官的不良反应。结果：多次小剂量^188Re—CL58治疗后病灶部位的FDG摄取SUV值(Standard uptake value)均下降，对于小转移灶SUV下降更明显，且血清CEA等水平均下降，而大剂量^131I—chTNT组CEA下降不及多次小剂量组明显。多次小剂量RIT后外周血CD4^ 细胞升高、CD8^ 细胞减少以及CD4^ ／CD8^ 比例升高，而大剂量RIT后CD4^ ／CD8^ 比例降低。大剂量组比多次小剂量组时白细胞、血小板、血色素的抑制明显，结论：多次小剂量^188Re—CL58治疗效果优于大剂量^131I—chTNT，且能解除肌体的免疫抑制：多次小剂量^188Re—CL58治疗对造血系统抑制小于大剂量^131I—chTNT。     

放射性碘标记热休克蛋白90α单克隆抗体用于肿瘤放射免疫治疗的研究

[目的]用放射性碘Na¹³¹I标记热休克蛋白90α单克隆抗体,研究其在胶质瘤荷瘤小鼠模型的生物分布以及放射免疫治疗效果。[方法]热休克蛋白90α单克隆抗体的放射性¹³¹I标记采用氯胺T法进行。标记抗体经尾静脉注射到U87细胞荷瘤小鼠模型体内,在注射体内后6、12、24h处死小鼠,测量其在各组织器官内的分布。放免实验分为实验组、对照组和未标记组,每组6只小鼠,注射药物后每5天测量肿瘤体积,连续40天。并记录全部小鼠注射后的生存时间。[结果]热休克蛋白90α单克隆抗体标记效率达到56%,纯化后放射化学纯度超过95%。分布数据表明¹³¹I-Hsp90α单克隆抗体主要累积在肝脏、脾脏和肿瘤。治疗组肿瘤体积明显小于其他两组,抑瘤率高于其余两组且生存时间更长。[结论]热休克蛋白90α单克隆抗体可以用氯胺T法成功标记¹³¹I。¹³¹I标记热休克蛋白90α单克隆抗体可以被U87荷瘤小鼠的肿瘤组织特异性摄取,其用于放免治疗有一定的研究价值和临床应用前景。     

放射性^131I照射与分化型甲状腺癌细胞摄碘（^125I）率关系的实验研究

目的：探讨分化型甲状腺癌培养细胞接受放射性碘（^131I）照射剂量与摄取放射性碘间的相关性,为放射性碘去除治疗甲状腺癌提供新的思路.方法：设两个实验组,对分化型甲状腺癌细胞进行培养,其一应用同一活度（20μCi）的放射性^131I对培养细胞进行不同时间的照射,另组应用不同活度的放射性^131I对培养细胞进行相同时间（12h）的照射,分别测定两组甲状腺癌细胞摄取放射性^125I水平.结果：不同活度或不同时间的放射性^131I照射使甲状腺癌细胞摄取碘的水平降低,受照射组与未照射对照组细胞摄碘率比较有显著性差异（P＜0.01）.结论：放射性^131I照射可以使分化型甲状腺癌细胞摄碘率显著降低.     

131Ⅰ—chTNT放射免疫治疗晚期肺癌的疗效观察

目的：观察131Ⅰ肿瘤细胞核人鼠嵌合单克隆抗体注射液（131Ⅰ—chTNT）对晚期肺癌的临床疗效，探讨晚期肺癌治疗的新方法。方法：选取2006年6月-2007年6月我院收治的13例经病理证实的晚期肺癌，予以131Ⅰ—chTNT抗体放射免疫治疗。每次静滴131Ⅰ—chTNT抗体50mCi，3周-4周后再用1次，每次治疗10天后行ECT检查131Ⅰ—chTNT在体内的分布。对照组为16例行普通化疗的患者。结果：完全缓解（CR）1例，部分缓解（PR）3例，无变化（NR）8例，进展（PD）1例，总有效率30．76％。与对照组有显著性差异（P〈0．05）。结论：131Ⅰ—chTNT作为一种新型放射免疫治疗药物，毒副作用较小，对晚期难治性肺癌治疗有一定的疗效。     

Pivotal study of iodine-131-labeled chimeric tumor necrosis treatment radioimmunotherapy in patients with advanced lung cancer.

PURPOSE: Tumor necrosis treatment (TNT) uses degenerating tumor cells and necrotic regions of tumors as targets for radioimmunotherapy. Previous studies in animal tumor models and clinical trials have demonstrated that when linked to the therapeutic radionuclide iodine-131, recombinant chimeric TNT antibody ((131)I-chTNT) can deliver therapeutic doses to tumors regardless of the location or type of malignancy. Therapeutic efficacy and toxicity of (131)I-chTNT in advanced lung cancer patients were studied in this pivotal registration trial. PATIENTS AND METHODS: Patients with advanced lung cancer were treated with systemic or intratumoral injection of (131)I-chTNT in eight oncology centers in China. The objective response rate (ORR) was assessed as the primary end point. RESULTS: All 107 patients who were entered onto the study and completed therapy had experienced treatment failure after prior radiotherapy or chemotherapy a mean of three times. The results showed an ORR of 34.6% (complete response, 3.7%; partial response, 30.8%; no change, 55.1%; and progressive disease, 10.3%) in all patients and 33% in 97 non-small-cell lung cancer patients. A biodistribution study demonstrated excellent localization of the radioactivity in tumors in both systemically and intratumorally injected patients. The most obvious adverse side effect was mild and reversible bone marrow suppression. CONCLUSION: Radioimmunotherapy with (131)I-chTNT was well tolerated and can be used systemically or locally to treat refractory tumors of the lung.     

Pharmacokinetics and tumor localization of 131I-labeled anti-tenascin monoclonal antibody 81C6 in patients with gliomas and other intracranial malignancies

We previously have reported that radioiodinated anti-tenascin monoclonal antibody 81C6 exhibits therapeutic potential against both s.c. and intracranial human glioma xenografts in athymic mice and rats. Herein we report the selective tumor localization of 131I-labeled 81C6 in patients with gliomas and other intracranial malignancies. Nine patients were simultaneously administered 5-50 mg of 131I-labeled 81C6 and 1-2 mg of 125I-labeled 45.6, an isotype-matched control monoclonal antibody. The blood clearance half-time for 81C6, normalized to that of 45.6 in the same patient, appeared to decrease with 81C6 protein dose. Gamma camera images obtained at 1 to 3 days exhibited increased uptake of 131I in regions corresponding to tumor with varying degrees of contrast to surrounding normal brain. Biopsy specimens of tumor and normal brain were obtained and analyzed histologically for tumor content. The average uptake of 81C6 in tumor ranged from 0.6 to 4.3 x 10(-3)% of the injected dose per gram. In patients receiving 20-50 mg of 81C6, the average tumor-to-normal-brain ratio was 25:1 with ratios as high as 200:1 seen in some samples. Localization indices were calculated by normalizing the uptake of 81C6 per gram tumor to the uptake of 81C6 per gram blood and dividing by the same ratio for 45.6 control monoclonal antibody. Localization indices for muscle and brain were about 1, in contrast to up to five for tumor. These studies demonstrate that the tumor uptake of 131I-labeled 81C6 in patients with gliomas and other intracranial malignancies is due to specific processes.     

Pharmacokinetic characteristics and biodistribution of radioiodinated chimeric TNT-1, -2, and -3 monoclonal antibodies after chemical modification with biotin

To improve the clinical potential of monoclonal antibodies (MAbs), new methods are required to augment antibody uptake in the tumor while minimizing binding in normal tissues. Our laboratory has pioneered the use of chemical modification to accomplish this goal. Using three chimeric MAbs, chTNT-1, chTNT-2, and chTNT-3, which target solid tumors by binding to common antigens found in the central necrotic core, we now demonstrate the potential of chemical modification to improve the pharmacokinetic characteristics of these unique MAbs. To identify optimal modification conditions, TNT MAbs were reacted with biotin at various ratios and tested by clearance and biodistribution analyses. The biodistribution results revealed that the numbers of biotin molecules per MAb yielding optimal tumor uptake were 3:1 for chTNT-1, 5:1 for chTNT-2, and 8:1 for chTNT-3. Biotinylated MAbs were found to have faster whole body clearance times and better biodistribution profiles compared to unmodified antibodies. Although chTNT-2 showed only a modest improvement after biotinylation, biodistribution results indicated that this MAb had the highest uptake in tumor. By reducing the charge of the antibody molecule, chemical modification appears to be a useful method for improving the pharmacokinetics and biodistribution of TNT antibodies directed to the necrotic region of solid tumors.     

Tumor targeting properties of indium-111 labeled genetically engineered Fab' and F(ab')2 constructs of chimeric tumor necrosis treatment (chTNT)-3 antibody

Genetic engineering techniques have allowed the construction of Fab' and F(ab')2 constructs of chimeric tumor necrosis treatment antibody (chTNT-3), a chimeric monoclonal antibody (MAb) that targets necrotic regions of solid tumors. The purpose of this study is to evaluate the in vitro and in vivo properties of Fab' and F(ab')2 constructs radiolabeled with indium-111 (111In) using diethylentriamine pentaacetic acid (DTPA) conjugation to develop a clinically useful imaging agent for the detection of necrosis in solid tumors. Optimization of the MAb-to-DTPA ratio showed that a 1:2 ratio gave the best immunoreactivity while providing good radiolabeling efficiency and high specific activity for all three DPTA conjugates. In addition, 111In-labeled Fab' and F(ab')2 conjugates were found to have faster whole body clearance times and better biodistribution profiles compared to parental 111In-labeled chTNT-3 in tumor-bearing mice. Although radiolabeled Fab' and F(ab')2 constructs showed lower tumor uptake than radiolabeled chTNT-3, biodistribution results showed that these constructs had significantly lower uptake in liver, spleen, and other normal organs (except the kidney), and therefore had higher tumor-to-organ ratios. In addition, a comparison of all derivatives showed that the F(ab')2 reagent gave the best results in tumor imaging studies. These results demonstrate that stable, a genetically engineered F(ab')2 construct can be successfully radiolabeled with 111In to produce potential imaging reagents for the imaging and monitoring of tumor necrosis.     

Limitations of radiolabeled monoclonal antibodies for localization of human neoplasms

Tumor-associated monoclonal antibodies were radiolabeled with 125I and 131I and given i.v. in pairs to 19 patients 1-26 days prior to surgical excision of primary and metastatic breast, ovarian, and gastrointestinal tumors. For individual patients each monoclonal antibody was designated as specific or nonspecific according to prior immunoperoxidase staining results on the appropriate target neoplastic tissues. Quantitation of antibody uptake was performed on resected normal and neoplastic tissues. Although good tumor:non-tumor ratios were obtained with the specific antibodies (maximal tumor:blood ratio, 35.8:1 at 12 days postadministration), the absolute amount of radiolabel detected in tumors was small (mean value of 0.015% of total injected amount per g of tumor occurring 1 day postadministration). Furthermore, both specific and nonspecific antibodies accumulated in normal lymph nodes to a significant extent (mean value of 0.0026% of total injected amount per g of tissue occurring 1 day postadministration). Knowledge of such data is essential prior to considering therapeutic uses of radiolabeled monoclonal antibodies.     

Radioimmunotherapy of human colon carcinoma by 131I-labelled monoclonal anti-CEA antibodies in a nude mouse model

A mixture of 3 MAbs directed against 3 different CEA epitopes was radiolabelled with 131I and used for the treatment of a human colon carcinoma transplanted s.c. into nude mice. Intact MAbs and F(ab')2 fragments were mixed because it had been shown by autoradiography that these 2 antibody forms can penetrate into different areas of the tumor nodule. Ten days after transplantation of colon tumor T380 a single dose of 600 microCi of 131I MAbs was injected i.v. The tumor grafts were well established (as evidenced by exponential growth in untreated mice) and their size continued to increase up to 6 days after radiolabelled antibody injection. Tumor shrinking was then observed lasting for 4-12 weeks. In a control group injected with 600 microCi of 131I coupled to irrelevant monoclonal IgG, tumor growth was delayed, but no regression was observed. Tumors of mice injected with the corresponding amount of unlabelled antibodies grew like those of untreated mice. Based on measurements of the effective whole-body half-life of injected 131I, the mean radiation dose received by the animals was calculated to be 382 rads for the antibody group and 478 rads for the normal IgG controls. The genetically immunodeficient animals exhibited no increase in mortality, and only limited bone-marrow toxicity was observed. Direct measurement of radioactivity in mice dissected 1, 3 and 7 days after 131I-MAb injection showed that 25, 7.2 and 2.2% of injected dose were recovered per gram of tumor, the mean radiation dose delivered to the tumor being thus more than 5,000 rads. These experiments show that therapeutic doses of radioactivity can be selectively directed to human colon carcinoma by i.v. injection of 131I-labelled anti-CEA MAbs.     

Localization of human breast tumors grafted in nude mice with a monoclonal antibody directed against a defined cell surface antigen of human mammary epithelial cells

Summary A mouse monoclonal antibody (BLMRL-HMFG-Mc5) prepared against a defined cell surface antigen of human mammary epithelial cells, non-penetrating glycoprotein (NPGP), was used in imaging and distribution studies in athymic nude mice grafted with human breast tumors. Forin vivo tissue distribution studies,¹²⁵I-labeled monoclonal antibody was injected into nude mice carrying simulated metastases of human tumors (breast and colon carcinomas). After 22–24 hr the amount of radioactivity per gram of tissue was 3–4 times higher in the breast tumor than in liver, brain, lung, muscle, or spleen. In contrast, colon carcinoma tissue, grafted and treated likewise, did not show higher accumulation of radioactivity relative to other tissues. At 4 days, the incorporation in breast tumors remained almost as high, while the circulating radioactive tracer and the incorporation in tissues other than breast had fallen significantly.In tumor imaging studies, breast tumor masses as small as 4 mm in diameter were clearly localized on a whole body scan using¹³¹I-labeled BLMRL-HMFG-Mc5 antibodies with a High-Purity germanium gamma camera. Normalization of¹³¹I-distribution to that of^99mTc-pertechnetate increased the specificity of this imaging methodology. The quantitative density of¹³¹I-label was 2–3 fold higher over the breast tumor than over comparable areas of the mouse. No positive localization images were obtained for similar implants of colon and lung carcinomas or melanomas after injections of¹³¹I-labeled BLMRL-HMFG-Mc5. Localization of human breast tumors in this model can be achieved with¹³¹I-labeled anti-breast epithelial monoclonal antibodies.Abbreviations NPGP non-penetrating glycoprotein - CEA carcinoembryonic antigen