Viral shedding and biodistribution of G207, a multimutated, conditionally replicating herpes simplex virus type 1, after intracerebral inoculation in aotus.

G207 is a multimutated, conditionally replicating herpes simplex virus type 1 (HSV-1) that is currently in clinical trial for patients with malignant glioma. G207 exhibits an efficient oncolytic activity in tumor cells, yet minimal toxicity in normal tissue when injected into the brains of HSV-susceptible mice or nonhuman primates. In this study, we evaluated the shedding and biodistribution of clinical-grade G207 after intracerebral inoculation (3 x 10(7) pfu) in four New World owl monkeys (Aotus nancymae). Using PCR analyses and viral cultures, neither infectious virus nor viral DNA was detected from tear, saliva, or vaginal secretion samples at any time point up to 1 month postinoculation. Analyses of tissues obtained at necropsy at 1 month from two of the four monkeys, plus one monkey inoculated with laboratory-grade G207 (10(9) pfu) 2 years earlier, showed the distribution of G207 DNA restricted to the brain, although infectious virus was not isolated. Histopathology revealed normal brain tissues including the sites of inoculation. A measurable increase of serum anti-HSV antibody titer was observed in all monkeys, as early as 21 days postinoculation. The results ascertain the safety of G207 in the brain and indicate that strict biohazard management may not be required for G207-treated patients.

Intraoperative computed tomography for complex craniocervical operations and spinal tumor resections.

OBJECTIVE: To improve intraoperative observation of unexposed anatomic features and to verify surgical correction, a mobile computed tomographic (CT) scanner has been introduced into the operating room. To date, intraoperative CT scanning has been used predominantly for intracranial procedures. We report on the expanded use of intraoperative CT scanning for spinal surgery, because CT scanning provides excellent observation of osseous pathological features. We report on our first 17 cases, which involved complex craniocervical operations and spinal tumor resections. METHODS: The Tomoscan M CT scanner (Philips Medical Systems, Eindhoven, The Netherlands) is mobile and consists of a translatable gantry, a translatable table, and an operator's workstation. In the operating room, the patient is placed on the CT table and prepared in the usual manner. The aperture of the gantry is covered with sterile plastic drapes. The gantry is docked to the table for intraoperative CT scanning as needed for navigation and verification during surgery. Each series of scans requires approximately 15 to 20 minutes. RESULTS: Our initial experience with neurosurgical spinal cases demonstrated that the use of intraoperative CT scanning changed the course of surgery in 6 of 17 cases. CT scanning was beneficial in facilitating adequate ventral clival and craniocervical decompressions, promoting more complete tumor resections, and verifying correct graft and instrument placement before surgical closing. Other settings in which we have found the mobile CT scanner useful include the neurointerventional suite and the intensive care unit; it is also useful for radiotherapy planning. CONCLUSION: On the basis of findings for our first 17 spinal surgery cases, we conclude that intraoperative CT scanning of the spine is both feasible and beneficial for select complex spinal procedures from the craniocervical junction to the sacrum.

Conditionally replicating herpes simplex virus mutant, G207 for the treatment of malignant glioma: results of a phase I trial.

G207 is a conditionally replicating derivative of herpes simplex virus (HSV) type-1 strain F engineered with deletions of both gamma(1)34.5 loci and a lacZ insertion disabling the UL39 gene. We have demonstrated the efficacy of G207 in treating malignant glial tumors in athymic mice, as well as the safety of intracerebral G207 inoculation in mice and in Aotus nancymai. We sought to determine the safety of G207 inoculation into cerebral malignant glial tumors in humans. Criteria for inclusion into this dose-escalation study were the diagnosis of histologically proven malignant glioma, Karnofsky score > or = 70, recurrence despite surgery and radiation therapy, and an enhancing lesion greater than 1 cm in diameter. Serial magnetic resonance images were obtained for volumetric analysis. The trial commenced at a dose of 10(6) plaque forming units (p.f.u.) inoculated at a single enhancing site and was completed when the 21st patient was inoculated with 3x10(9) p.f.u. at five sites. While adverse events were noted in some patients, no toxicity or serious adverse events could unequivocally be ascribed to G207. No patient developed HSV encephalitis. We found radiographic and neuropathologic evidence suggestive of anti-tumor activity and long-term presence of viral DNA in some cases.

Hepatoma-specific antitumor activity of an albumin enhancer/promoter regulated herpes simplex virus in vivo.

Targeting viral vectors to appropriate cell types so that normal cells are not adversely affected is an important goal for gene therapy. Previously, we described a novel approach to viral gene therapy using a conditional, replication-competent herpes simplex virus (HSV), where replication and associated cytotoxicity are limited to a specific cell-type by the regulated expression of an essential immediate-early viral gene product. In this report we analyze the hepatoma-specific replication, cytotoxicity and anti-tumor effect of recombinant HSV G92A, regulated by the albumin enhancer/promoter. G92A efficiently replicated in vitro in two human hepatoma cell lines expressing albumin, but not in four human non-hepatoma, albumin-non-expressing tumor cell lines, while all cell lines were equally susceptible to a tissue nonspecific HSV recombinant, hrR3. In vivo, G92A replicated well in subcutaneous xenografts of human hepatoma cells (Hep3B) in athymic mice, but not in non-hepatoma subcutaneous tumors (PC3 and HeLa), whereas, hrR3 replicated well in both tumor types. Intratumoral inoculation of G92A inhibited the growth of established subcutaneous hepatoma tumors in nude mice, but not prostate tumors. Replication-competent viral vectors controlled by cell-specific transcriptional regulatory sequences provide a new therapeutic strategy for tumor therapy.

Attenuated, replication-competent herpes simplex virus type 1 mutant G207: safety evaluation of intracerebral injection in nonhuman primates.

This study examined the safety of intracerebral inoculation of G207, an attenuated, replication-competent herpes simplex virus type 1 (HSV-1) recombinant, in nonhuman primates. Sixteen New World owl monkeys (Aotus nancymae [karyotype 1, formerly believed to be A. trivirgatus]), known for their exquisite susceptibility to HSV-1 infection, were evaluated. Thirteen underwent intracerebral inoculation with G207 at doses of 10(7) or 10(9) PFU, two were vehicle inoculated, and one served as an infected wild-type control and received 10(3) PFU of HSV-1 strain F. HSV-1 strain F caused rapid mortality and symptoms consistent with HSV encephalitis, including fever, hemiparesis, meningitis, and hemorrhage in the basal ganglia. One year after G207 inoculation, seven of the animals were alive and exhibited no evidence of clinical complications. Three deaths resulted from nonneurologic causes unrelated to HSV infection, and three animals were sacrificed for histopathologic examination. Two animals were reinoculated with G207 (10(7) PFU) at the same stereotactic coordinates 1 year after the initial G207 inoculation. These animals were alive and healthy 2 years after the second inoculation. Cerebral magnetic resonance imaging studies performed both before and after G207 inoculation failed to reveal radiographic evidence of HSV-related sequelae. Despite the lack of outwardly observable HSV pathology, measurable increases in serum anti-HSV titers were detected. Histopathological examination of multiple organ tissues found no evidence of HSV-induced histopathology or dissemination. We conclude that intracerebral inoculation of up to 10(9) PFU of G207, well above the efficacious dose in mouse tumor studies, is safe and therefore appropriate for human clinical trials.

Spondyloptotic fracture of the cervical spine in a pregnant, anemic Jehovah's Witness: technical and ethical considerations. Case report.

The authors present the case of a 15-year-old Jehovah's Witness with a hematocrit level of 19% who was 4 months pregnant and had a two-level spondyloptotic cervical spine fracture. The patient was transferred to Georgetown University Medical Center with C-5 quadriplegia 3 weeks after having been injured in an automobile collision. The neurosurgical issues in this case included addressing a rare cervical spine injury, assessing and treating a vertebral artery injury, and performing surgery on a pregnant minor with severe anemia who held strong Jehovah's Witness beliefs. An ethics consultation was convened to determine the validity of a pregnant minor's refusal to undergo transfusion on the grounds of her religious beliefs. This case illustrates the potential benefits of thorough technical and ethical evaluations and reveals how they may contribute to the delivery of care in complex neurosurgical cases. To the authors' knowledge, this is the first two-level spondyloptotic cervical spine fracture dislocation to be reported in the literature. The added complexities of the case, given that the patient was an anemic, adolescent, pregnant Jehovah's Witness who refused blood transfusion, also appear to be unprecedented.

Human T-lymphotropic virus type II in Panamanian Guaymi Indians.

BACKGROUND: Currently, human T-lymphotropic virus type II (HTLV-II) is the most prevalent human retrovirus, detected in persons presenting to donate blood in the United States. Only scant information is available with which to counsel HTLV-II-seropositive deferred donors or other persons about the ways in which they may spread HTLV-II to others. STUDY DESIGN AND METHODS: To increase understanding of the modes of transmission of HTLV-II, a seroepidemiologic study was conducted among Panamanian Guaymi Indians, a recently identified focus of endemic HTLV-II infection. Subjects were tested for serologic evidence of infection by HTLV-II, HTLV type I, hepatitis B virus, and nine other infectious agents by enzyme immunoassays and specific confirmatory tests. RESULTS: Nine (8.3%) of 109 persons tested HTLV-II-seropositive. HTLV-II seropositivity was more likely in persons with serologic evidence of prior hepatitis B virus infection. Sexual contact with HTLV-II-seropositive partners, but neither parenteral exposure nor breast-feeding, was identified as a risk factor for HTLV-II. CONCLUSION: In Guaymi Indians, HTLV-II appears to be spread primarily through sexual transmission.

Mutational analysis of a predicted zinc-binding motif in the 26-kilodalton protein encoded by the vaccinia virus A2L gene: correlation of zinc binding with late transcriptional transactivation activity.

Transient transfection assays indicated that A2L is one of three vaccinia virus intermediate genes that are required for the transcriptional transactivation of viral late genes. We have expressed the A2L open reading frame in Escherichia coli and shown by blotting experiments that the 26-kDa protein binds zinc, a property predicted by the presence of a CX2CX13CX2C zinc finger motif. The specificity for zinc binding was demonstrated by competition with other metals. The role of the sequence motif in zinc binding was established by analysis of a series of mutations, including truncations and conservative single amino acid substitutions. Mutations that reduced zinc binding in vitro prevented the ability of A2L to transactivate late genes in vivo.