Adoptive immunotherapy of human meningeal gliomatosis and carcinomatosis with LAK cells and recombinant interleukin-2

Previous in vitro studies have demonstrated that peripheral blood lymphocytes activated with recombinant interleukin-2 (rIL-2) generated cells that were lytic for fresh autologous tumor cells but not for normal lymphocytes or lymphoblasts. Adoptive transfer of autologous lymphokine-activated killer (LAK) cells induced in vitro with rIL-2 was used in two patients: one with meningeal gliomatosis and the other with meningeal carcinomatosis. The adoptive transfer of LAK cells was very effective in reducing the clinical symptoms and signs, and in eliminating the malignant cells from the cerebrospinal fluid. Thus, this therapy is an attractive approach for the treatment of malignant tumors that have poor immunogenicity and are insensitive to several anti-cancer agents, and for patients with severe immunosuppressive conditions induced by repeated radiation therapy or chemotherapy.     

Lymphokine-activated killer (LAK) therapy for advanced renal cell carcinoma: clinical study on arterial LAK therapy and experimental study on LAK cell activity]

Experimental and clinical studies were conducted on lymphokine-activated killer (LAK) cell therapy for advanced renal cell carcinoma (RCC). The traffic assay using radiolabeled LAK cells indicated short-term but appreciable accumulation of LAK cells in the tumor site when trans-arterially infused. By contrast, systemically infused LAK cells were localized not to the tumor tissue but to the lung. Therefore, we began treatment of the patients with extrapulmonary metastases by means of regional arterial administration of LAK cells and those who had pulmonary metastases by a systemic LAK therapy. Regimen of Interleukin-2 (IL-2) administration was bolus infusion of 5 x 10 U IL-2 twice daily. Frequency of LAK cell administration varied from one to three times a week depending upon the patient's condition. Eight out of 16 metastases, such as bone, muscle, and lymph node metastases, in 9 patients treated by arterial LAK therapy showed regression. Side effects during LAK therapy were not serious. Past history of having chemotherapy was an unfavorable factor that could reduce the sensitivity to LAK therapy. Our laboratory study showed the production of Interferon (IFN)-gamma and Tumor Necrosis Factor (TNF)-alpha by LAK cells when preincubated with RCC cells, which may indicate the mechanism of LAK cell-mediated antitumor activity in vivo. The study also showed that LAK cells as well as monocytes preincubated with the supernatant of LAK cells damaged normal endothelial cells in vitro, which suggested the possibility that LAK therapy risks increasing the frequency of brain metastasis by damaging the blood-brain barrier.(ABSTRACT TRUNCATED AT 250 WORDS)     

自体LAK细胞和rIL－2治疗晚期肾癌的实验与临床研究

应用自体ＬＡＫ细胞和重组白细胞介素－２（ｒＩＬ－２）治疗２０例晚期肾癌患者。自患者周围血分离到的单个核细胞（ＰＢＭＣ）体外经ｒＩＬ－２短期培养，其ＮＫ、ＬＡＫ活性明显增强并于第５、７天达高峰。当这些ＬＡＫ细胞与ｒＩＬ－２过继回输给同一患者后，病人周围血ＮＫ、ＬＡＫ活性明显增加（Ｐ＜０．０ｌ），ＮＫ比率、ＩＬ－２受体表达明显增加（Ｐ＜０．０５），提示对肾癌患者的免疫调节作用。本组病人获部分缓解（ＰＲ）ｌ例，轻度缓解（ＭＲ）３例，平均缓解期５个月。毒副作用主要表现为发热、寒战，病人能耐受，表明ＬＡＫ／ｒＩＬ－２疗法是安全的方法。     

IL－2和LAK细胞过继免疫疗法在肾癌的应用

１９９２～１９９３年用ＩＬ－２和ＬＡＫ细胞输注治疗肾细胞癌１４例，全组Ｔ（２～４）期者１１例，５例有远处转移。治疗后获２２个月以上随访，除４例死亡外余尚存活。认为过继免疫疗法对中、晚期肾癌有一定效果，但ＬＡＫ细胞在体内聚集、活化、归转、监测等问题，尚需继续探讨。     

人脑胶质瘤白介素13受体基因表达与肿瘤增殖活性的关系研究

目的探讨人脑胶质瘤白介素13受体（IL-13R）基因表达与肿瘤增殖活性的关系。方法对6例正常脑组织，50例人脑胶质瘤和2个脑瘤体外细胞系采用RT—PCR法和免疫组织化学法检测IL-13R。结果人脑胶质瘤组织IL-13RαmRNA总阳性表达率70％，正常脑组织中仅1例有极弱的表达；2例恶性胶质瘤体外细胞系均高表达。IL-13RαmRNA表达率和表达丰度与胶质瘤分级（前者rs=0．87，P〈0．01；后者rs=0．69，P〈0．01）、肿瘤增殖活性Ki-67LI（r=0．64，P〈0．01）呈正相关，即胶质瘤恶性程度越高，IL-13RαmRNA表达率和表达水平越高。结论IL-13Rα基因在人脑胶质瘤中表达上升，与肿瘤的分级和肿瘤增殖活性呈正相关，可作为预测某些肿瘤治疗效果及监测复发的指标之一。     

Two cases of renal cell carcinoma with ascites after interleukin-2 therapy

One of the most common and problematic side effects of interleukin-2 therapy (IL-2) is vascular leak syndrome (VLS). VLS is characterized by an increase in vascular permeability accompanied by extravasation of fluids and proteins from the capillary vessels into the tissues resulting in interstitial edema, weight gain, pleural effusion, ascites in severe form, multiple organ failure. We, herein, report two cases of VLS, which occurred following IL-2 therapy against metastatic renal cell carcinoma. One of them died of multiple organ failure one month after the cessation of IL-2 therapy.     

Lymphokine-activated killer (LAK) therapy for metastatic renal cell carcinoma]

Fifteen patients with metastatic renal cell carcinoma (RCC) were treated by administration of autologous lymphokine-activated killer (LAK) cells given together with systemic administration of interleukin-2 (IL-2). Pulmonary metastases alone were found in 10 cases, pulmonary and mediastinal nodal metastases in 3, and pulmonary and bone metastases in 2. LAK cells, generated by incubation in 700 units/ml of IL-2 for 3-4 days, were intravenously administered once a week. In addition, beginning on the day of the first LAK cell infusion, 3.5 x 10(5) units of IL-2 were intravenously infused once or twice a day with occasional supplementation of 3.5 x 10(5) units of IL-2 on each day of LAK cell infusion. The total number of LAK cells and total amount of IL-2 administered per patient in this study ranged from 0.8 x 10(10) to 6.9 x 10(10) cells and from 10.2 x 10(6) to 74.9 x 10(6) units, respectively. As toxic effects caused by the infusion of LAK cells, headache, shaking chills, fever and leukocytosis were found in all cases. Side effects possibly induced by IL-2 infusion were tolerable fever, fluid retention (body weight gain of 2-3 kg) and eosinophilia. Out of 15 patients, a partial response was observed in 4 patients who had pulmonary metastases alone. One of the 4 patients with a partial response was clinically free of disease after undergoing a thoracotomy for resection of residual lesions, but a brain metastasis was detected 10 months after the thoracotomy. The remaining 3 patients are being closely followed up at present.(ABSTRACT TRUNCATED AT 250 WORDS)     

Imbalances in T cell subpopulations in human gliomas

The quantitation of cells bearing CD3, CD4, CD8, and B cell phenotypic markers, as well as an estimation of serum immunoglobulin (Ig)G, IgA, and IgM, was carried out in a group of 39 glioma patients with different grades of malignancy. The findings were compared with those obtained from 21 normal healthy control subjects. The analysis revealed a significant decrease both in the absolute numbers and in the percentages of circulating CD3+ (p less than 0.001) and CD4+ (p less than 0.001) cells, while the CD8+ and Pan B+ cells remained within the normal range irrespective of the type and grade of tumor. The CD4+:CD8+ ratio was significantly decreased in all categories of patients. The CD4 lymphopenia was also evident in 10 patients who had no history of previous immunosuppressive drug therapy (steroids and anticonvulsants) until the commencement of the study. The Ig levels were within the normal range in patients with malignant astrocytoma and glioblastoma multiforme, whereas a three- and fourfold increase in the IgM level was observed in patients with astrocytoma. It is suggested that T cell lymphopenia in glioma patients could mainly be due to a selective depletion of CD4+ cells and that it occurs principally as a reaction to the tumor.     

Adenovirus-mediated p53 gene therapy for human gliomas.

OBJECTIVE: The rationale and current evidence for using p53 gene replacement as a potential treatment for human gliomas are reviewed. The possible benefits of and obstacles to this approach are delineated. METHODS: One approach to overcoming the poor outcomes associated with conventional glioma therapies involves the replacement of tumor suppressor genes that are fundamental to glioma development. The p53 gene is one of the most frequently mutated genes in human gliomas, and loss of p53 function is critical to the development of glial neoplasms. Consequently, replacement of the p53 gene using viral vectors may be a potential treatment for human gliomas. RESULTS: In vitro studies demonstrate that adenovirus-mediated p53 gene transfer into gliomas with mutant p53 results in massive apoptosis. Similarly, transfer of p53 inhibits tumor growth in vivo. In contrast to mutant p53 gliomas, wild-type p53 glioma cells are resistant to the apoptotic effects of p53 transfer, but this resistance can be overcome by the addition of deoxyribonucleic acid-damaging agents such as ionizing radiation or chemotherapy. The main obstacle to p53 gene therapy involves the limitations associated with current modes of delivery. CONCLUSION: Preclinical data strongly support the use of p53 gene transfer as a potential treatment for human gliomas.     

白细胞介素2基因工程人大肠癌细胞瘤苗的建立及鉴定

目的 建立高表达白细胞介素2的基因工程人大肠癌细胞瘤苗，并完成相关的实验室鉴定。方法 采用重组逆转录病毒将人白细胞介素2基因转导至三株人大肠癌细胞，G418筛选并挑选单克隆，ELISA检测各单克隆24h白细胞介素2表达量，用Student Newman Keuls检验三株细胞白细胞介素2的表达差异，PCR、Southern blot检测白细胞介素2基因整合，RT-PCR、Northern blot检测白细胞介素2基因转录水平。结果 获得72株G418抗性单克隆，其中表达量最高者为COL0205达138.01ng/L×10^7/24h，统计显示COLO205与SW1116、HT-29组间相比差异有统计学意义（P〈0.05），白细胞介素2基因整合至细胞基因组中，未发生重组或部分缺失，且获得成功表达。结论 本研究成功建立了高表达白细胞介素2的基因工程人大肠癌细胞瘤苗。     

LAK cell adoptive immunotherapy and its problems

Clinical efficacy of lymphokine-activated killer (LAK) cell adoptive immunotherapy (AIT) in combination with plasma exchange and interleukin (IL-2) was investigated in 24 patients with advanced cancer. Partial response (PR) was found in 4 patients (20%), including 1 primary liver tumor, 1 metastatic lung tumor from renal cancer and 2 malignant pleural effusions from gastric and lung cancer. Based on these results new AIT in combination with plasma exchange, OK-432, IL-2 and cyclophosphamide was designed to target liver and lung tumors, in which LAK cells and other drugs were administered through the catheter located in the feeding artery of the tumor. Out of ten patients treated, 1 (10%) with metastatic liver tumor from gallbladder cancer was evaluated as PR. It is suggested that a strategy to enhance tumor accumulation and recognition of LAK cells should be attempted for development of gastrointestinal cancer therapy with AIT.     

Adoptive immunotherapy with LAK cell and IL-2 against primary lung cancer

Adoptive immunotherapy using LAK cells and IL-2 was performed against stage III and IV primary lung cancer patients after surgery. A randomized controlled study consisting of control group A, chemotherapy (CDDP + VDS) group B and chemo-immunotherapy (CDDP+VDS, IL2+LAK cells) group C suggested better survival rate in the group C. Direct effects were studied against 8 recurrent or inoperable lung cancer cases. Complete response was obtained against a pleuritis and pericarditis carcinomatosa case when in vitro stimulated LAK cells (St-LAK) were administered locally. Partial response was observed against a inoperable case when LAK-BAI (bronchial arterial infusion) was combined with radiation therapy.     

Metastatic type-2 papillary renal cell carcinoma responded to interleukin-2 therapy: case report

This report documents a case of metastatic papillary renal cell carcinoma (PRCC) which successfully responded to interleukin-2 (IL-2) therapy. A 59-year-old male presented with a left renal mass measuring 3.0 cm in diameter and a right adrenal mass measuring 5.0 cm in diameter. He underwent a left partial nephrectomy and a right adrenalectomy. The histological findings revealed pT1bN1M1 type-2 PRCC and metastatic renal cell carcinoma in the right adrenal gland. The patient was given interferon-α (IFN-α) after the operation for 3 months. A CT scan revealed a metastatic nodule measuring 6.0 cm in diameter near the surface of the liver at 4 months after the opereation. The patient was given interleukin-2 (IL-2), 7 × 10⁵ units/day intravenously, for 3 days per week. A CT scan revealed this hepatic nodule to have decreased in size from 6.0 to 4.0 cm after 4 months of IL-2 therapy. However, a new metastatic nodule measuring 6.0 cm in diameter was found which came in contact with the spleen. Next, the patient was given an increased dose of IL-2 from 7 × 10⁵ to 1.4 × 10⁶ units/day intravenously, for 3 days per week. At 9 months of follow-up after the dose escalation, a CT scan revealed a dramatic decrease in the size of these two metastatic nodules to 1.5 and 0.5 cm, respectively. This is a very rare case in that it represents a type-2 PRCC which dramatically responded to low-dose IL-2 therapy.     

腺病毒介导的HSV-tk基因治疗人脑胶质瘤SHG44的研究

目的　探讨带有ＨＳＶｔｋ 基因的重组腺病毒（Ａｄ．ｔｋ） 结合ＧＣＶ 对国人脑胶质瘤ＳＨＧ４４ 的治疗作用。方法　Ｘｇａｌ 染色比较带有报告基因ＬａｃＺ的重组腺病毒（Ａｄ．ＬａｃＺ） 转染ＳＨＧ４４ 细胞和大鼠脑胶质瘤Ｃ６ 细胞的效率,ＭＴＴ法比较Ａｄ．ｔｋ／ＧＣＶ 系统对ＳＨＧ４４ 和Ｃ６ 细胞的敏感性,ＤＮＡ片段分析细胞杀伤机制;在活体用Ａｄ．ｔｋ／ＧＣＶ 治疗ＳＨＧ４４ 移植瘤。结果　ＭＯＩ为５０ 时,Ａｄ．ＬａｃＺ感染ＳＨＧ４４ 细胞的效率近１００％ ,较Ｃ６ 细胞高;ＭＯＩ为１００ 时,Ａｄ．ｔｋ 感染的ＳＨＧ４４ 细胞对ＧＣＶ 的敏感性较Ｃ６ 细胞高,其半致死剂量（ＩＣ５０） 分别为０ ．１２ μｍｏｌ／Ｌ 和０ ．２６ μｍｏｌ／Ｌ,有凋亡机制参与其细胞杀伤;５ ×１０８ＰＦＵ 的Ａｄ．ｔｋ 肿瘤原位注射结合ＧＣＶ治疗,有效抑制ＳＨＧ４４ 移植瘤生长,肿瘤组织坏死,出血伴有纤维组织增生;与Ａｄ ．ｔｋ／ＰＢＳ组、Ａｄ．ＬａｃＺ／ＧＣＶ 组、ＰＢＳ组相比,肿瘤抑制效率分别为９２．５ ％ 、９２．６％ 、８９ ．５％ （ Ｐ＜０ ．０１） 。结论　Ａｄ．ｔｋ／ＧＣＶ系统对人脑胶质瘤细胞的感染效率和敏感性较高,治疗国人脑胶质瘤效果显著。     

Intratumoral BCG therapy of transplanted head and neck tumors in strain 2 guinea pigs

The generally unfavorable prognosis associated with advanced squamous cell carcinoma in the head and neck region in humans led us to immunotherapeutic experiments with bacillus Calmette-Guerin (BCG) in inbred guinea pigs with solid growing and lymphogenous metastasizing tumors. The injection of live BCG or BCG cell wall preparation (CWP) into the planum buccale in the guinea pig led to a pronounced local inflammatory reaction. If live BCG or BCG CWP were injected into the planum buccale together with line 10 tumor cells, no growth of the tumor could be observed. Animals treated in this manner developed a tumor-specific immunity. Guinea pigs treated intratumorally with live BCG or BCG CWP in established, 6-day-old tumors of the planum buccale showed regression of the primary tumor and prevention of the development of lymph node metastases. These cured animals also developed a tumor-specific immunity. Guinea pigs with 6-day-old tumors of the planum buccale remained tumor free after surgical removal of the primary tumor and the draining submandibular and cervical lymph nodes (radical operation) whereas locally operated animals (primary tumor excision) developed lymph node metastases. Animals that were tumor free after radical surgery did not develop tumor-specific immunity. Despite insufficiently clarified mechanisms for the induction and effect of BCG immunotherapy, we can state that, with intratumoral BCG therapy, a tumor experimentally placed in the head and neck region is eliminated, regionally manifest metastases are eliminated, and specific tumor immunity can be demonstrated in successfully treated animals.     

Surgical therapy of malignant gliomas

The authors have presented the current state of surgical therapy of malignant gliomas underlining the advantages and disadvantages of traditional and stereotactic surgery, preparation for surgery, mortality, morbidity, prognostic factors, and complications. Particular attention was placed on cell kinetic studies. Of the methods used in this study we would like to recall those of flow cytometry associated with administration of BUdR and utilization of the monoclonal antibody Ki-67 associated with immunohistochemical techniques.     

Influence of PGE2- and cAMP-modulating agents on human glioblastoma cell killing by interleukin-2-activated lymphocytes

Human glioblastoma cells secrete factors, such as prostaglandin E (PGE) and transforming growth factor beta type 2, which are capable of suppressing several immune functions. The present study investigated the effect of PGE2 and agents known to increase intracellular cyclic adenosine monophosphate (cAMP) levels on 1) the induction of lymphokine-activated killer (LAK) cell activity from the peripheral blood lymphocytes (PBL) of both normal and glioma patients and on 2) the cytolytic activities of tumor-infiltrating lymphocytes (TIL's) isolated from malignant gliomas after expansion in vitro with interleukin-2 (IL-2). Cytolytic activity was measured against autologous and allogeneic tumor cells and the natural killer-resistant Daudi cell line. The results demonstrate that PGE2 and agents known to increase intracellular cAMP levels can significantly suppress the IL-2-dependent generation of cytolytic activity from the PBL of normal and glioma patients and from glioblastoma-derived TIL's. The inhibitory effects of these agents could not be reduced by higher concentrations of IL-2 or by cyclic guanosine monophosphate. Although the suppressive effect of PGE2 was most significant during the early stages of LAK cell generation, an inhibitory effect was still evident when PGE2 was added directly to the cytotoxicity assay. Secretion of PGE2 by glioblastoma cells in vivo may regulate both the generation of an immune response and the effectiveness of adoptively transferred immune cells.