A large percentage of human hepatomas produce a-fetoprotein (AFP), but the levels of AFP expression vary greatly among hepatomas. To understand the molecular basis for this variation, we analyzed tanscriptional regulatory activities associated with the 5'-flanking region of the AFP gene in two human hepatoma cell lines, HuH-7 and huH-1/cl-2, which produce a high and a low level of AFP, respectively. We found that the low level of AFP production in huH-1/cl-2 is due to the action of at least two silencer regions located between the enhancer and the promoter of the AFP gene. In contrast, no silencer activity is expressed in HuH-7. We identified 5'-CTTCATAACTAATACTT-3' to be a core sequence responsible for the negative regulatory activity. This sequence is repeated four times in a strong, distal silencer region, Sd, whereas one copy is present in a weak, proximal silencer region, Sp. The silencer reduces transcriptional initiation by blocking enhancer activation of the AFP promoter in a position-dependent manner. The silencer functions in the presence of positive transcription factors and may play a key role in developmental repression as well as variable expression of the AFP gene in hepatomas.The a-fetoprotein (AFP) gene is a developmentally regulated gene whose activity decreases rapidly after birth and becomes hardly detectable in adult life. However, the AFP gene is often reactivated to a high level in hepatocellular carcinoma (HCC) and teratocarcinomas (1, 4, 36). In contrast, the expression of the albumin gene, which is evolutionarily related to the AFP gene, steadily increases during normal development and stays high throughout adult life.Elevated levels of AFP have been observed in more than 70% of HCC patients, making AFP as a reliable diagnostic and prognostic marker of HCC. The AFP levels among AFP-positive patients, however, vary widely. Analysis of mechanisms underlying the variation of AFP production in HCC is of importance from clinical as well as basic biological standpoints.HuH-7 and huH-1/cl-2 are two human hepatoma cell lines that exhibit various liver-specific functions (23-25). HuH-7 produces a large amount of AFP and a moderate level of albumin. huH-1/cl-2, on the other hand, produces only a detectable amount of AFP but a high level of albumin. These cell lines provide useful model systems for the study of the molecular basis for different levels of expression of the AFP and albumin genes in hepatoma.We have shown previously that in HuH-7 cells, AFP gene expression is regulated positively by the enhancer, promoter, and glucocorticoid-responsive element (27,32,39). We have not so far detected DNA elements that suppress AFP gene transcription in this cell line. In this study, we analyzed mechanisms responsible for a low level of AFP gene expression in huH-1/cl-2. Two possibilities were considered: (i) the reduced activity of positive regulatory elements and (ii) the activation of negative control elements. We present evidence that the latter mechanism is largely * Corresponding author. t Presen...

It is the authors' impression that the use of the endoscope in the microsurgical management of cerebral aneurysms enhanced the safety and reliability of the surgery. However, there is a prerequisite for the surgeon to be familiar with this instrumentation and fully prepared for the risks and inconveniences of endoscopic procedures.

The "bystander effect" describes the killing of nearby unmodified cells and herpes simplex thymidine kinase (HTK)-transduced cells by ganciclovir (GCV) treatment. This effect is thought to be produced by contact between these cells. In this study, we showed that injected glioma cells migrated rapidly to a place distant from the injection point whereas injected virus-producing fibroblast cells did not migrate in a murine brain model. Moreover, the initially injected glioma cells and glioma cells injected at a later time mix very well, even at a place distant from the injection point. This suggested that glioma cells migrating after injection could be targeted by HTK-modified glioma cells introduced in a second injection and be killed together by GCV treatment. Therefore, we injected HTK-modified glioma cells 3 days after injection of wild glioma cells to investigate whether wild-type glioma cells that migrated to a place distant from the injection point could also be killed by GCV treatment. Tumor growth was suppressed after the GCV treatment. Suppression of tumor growth of wild glioma cells is not solely mediated by the immune response, which may be triggered by the killing of HTK-modified glioma cells with GCV, because inoculation of HTK-modified glioma to the contralateral side followed by GCV treatment did not cure the initial wild glioma. Moreover, the migration of the second inoculum of glioma cells is necessary for effective killing, because early administration of GCV resulted in insufficient killing.

Implantation of retrovirus-producing cells within a tumor has been demonstrated to eliminate malignant brain tumors effectively in animal models. In our previous study, the implantation of high-titer retrovirus-producing fibroblasts into tumors resulted in highly efficient transduction in vivo. The transduced glioma cells migrated far from the implantation site, potentiating the induction of the remarkable bystander effect. It is also possible, however, that the implantation of murine fibroblast-derived virus-producing cells may induce an immune response in patients. In this study, we prepared retroviruses carrying the herpes simplex virus thymidine kinase (HTK) gene with titers of 1.4-2.5 × 10 11 colony-forming units (c.f.u.)/ml, and stereotactically inoculated only 3 l of

An adoptive immunotherapy of 6 patients with medulloblastoma by lymphokine-activated killer (LAK) cells is described. They were from 2 to 9 years in age and had cerebrospinal fluid (CSF) dissemination of the tumours. All patients underwent the whole-neuraxis irradiation and chemotherapy. After the usual treatments, they were submitted to an adoptive transfer of one-haplotype identical LAK cells. The LAK cells were induced from peripheral blood lymphocytes (PBL) of their relatives with human recombinant interleukin-2 (rIL-2). 3 - 15 x 10(9) LAK cells were transferred intrathecally in 2-3 months. In 3 of 6 patients, neurological signs were improved and malignant cells had never been detected on CSF cytology after the adoptive immunotherapy. One among these 3 patients showed complete response in 20 months. Thus, this is an attractive approach for the treatment of medulloblastoma with CSF dissemination of the tumour which current therapeutic intervention can not cure.