A humanized form of a CD4-specific monoclonal antibody exhibits decreased antigenicity and prolonged plasma half-life in rhesus monkeys while retaining its unique biological and antiviral properties

Certain monoclonal antibodies (MAbs) directed against CD4 can efficiently block HIV-1 replication in vitro. To explore CD4-directed passive immunotherapy for prevention or treatment of AIDS virus infection, we previously examined the biological activity of a nondepleting CD4-specific murine MAb, mu5A8. This MAb, specific for domain 2 of CD4, blocks HIV-1 replication at a post-gp120-CD4 binding step. When administered to normal rhesus monkeys, all CD4+ target cells were coated with antibody, yet no cell clearance or measurable immunosuppression occurred. However, strong anti-mouse Ig responses rapidly developed in all monkeys. In the present study, we report a successfully humanized form of mu5A8 (hu5A8) that retains binding to both human and monkey CD4 and anti-AIDS virus activity. When administered intravenously to normal rhesus monkeys, hu5A8 bound to all target CD4+ cells without depletion and showed a significantly longer plasma half-life than mu5A8. Nevertheless, an anti-hu5A8 response directed predominantly against V region determinants did eventually appear within 2 to 4 weeks in most animals. However, when hu5A8 was administered to rhesus monkeys chronically infected with the simian immunodeficiency virus of macaques, anti-hu5A8 antibodies were not detected. Repeated administration of hu5A8 in these animals resulted in sustained plasma levels and CD4+ cell coating with humanized antibody for 6 weeks. These studies demonstrate the feasibility of chronic administration of CD4-specific MAb as a potential means of treating or preventing HIV-1 infection.     

Assessment of regional sympathetic nerve activity in vasospastic angina: analysis of iodine 123-labeled metaiodobenzylguanidine scintigraphy

Injury of the normal central nervous system is a major concern in the radiotherapy of brain tumors, but the pathogenesis of injury remains poorly understood. Modulation of the production of growth factors is associated with ischemia and traumatic injury in the central nervous system. Ionizing radiation has been shown to induce basic fibroblast growth factor in endothelial cells and in cells of a human breast carcinoma cell line. The inducibility of basic fibroblast growth factor after irradiation and its potential role in the recovery response of the central nervous system led us to investigate the effects of radiation on the expression of this growth factor in primary cultures of normal rat type 1 astrocytes. Astrocyte monolayers were exposed to ionizing radiation (1 to 10 Gy). Northern blot analysis revealed that doses of 2 to 10 Gy markedly reduced the expression of basic fibroblast growth factor as early as 1 h after irradiation, and that it remained below levels in unirradiated cells for at least 24 h. The effect was not associated with astrocyte cytotoxicity, and it appears to have some specificity for basic fibroblast growth factor since the levels of mRNA coding for ciliary neurotrophic factor and glial fibrillary acidic protein were not affected.     

FcalphaRI (CD89) as a novel trigger molecule for bispecific antibody therapy

Promising results from clinical trials with unconjugated antibodies stimulated renewed interest in immune effector mechanisms of monoclonal antibodies (MoAbs). We investigated the potential of IgA as antibody isotype for cell- or complement-mediated tumor cell lysis and assessed the potential of its myeloid Fc receptor, FcalphaRI (CD89), as trigger molecule for bispecific antibody (BsAb)-mediated immunotherapy. Comparing hapten-directed antibodies of human IgA2 with IgG1 or IgG3 isotypes, we found all three to mediate effective killing of sensitized tumor target cells in whole blood assays. Analysis of effector mechanisms showed IgG-mediated lysis to be predominantly complement-dependent, whereas IgA-dependent killing was primarily effector cell-mediated. A comparison of effector cell populations in antibody-dependent cell-mediated cytotoxicity (ADCC) showed neutrophils to be most important for IgA-dependent tumor cell killing, involving FcalphaRI as shown with Fc receptor blocking antibodies. Reverse ADCC experiments against target cells sensitized with Fc receptor antibodies, or assays with FcalphaRI-directed bispecific antibodies confirmed FcalphaRI as effective trigger molecule in polymorphonuclear neutrophil (PMN)-mediated lysis. During granulocyte colony-stimulating factor (G-CSF ) therapy, (FcalphaRI x HER-2/neu) bispecific antibodies induced enhanced killing of HER-2/neu positive SK-BR-3 breast cancer cells in whole blood assays. This enhanced cytotoxicity was paralleled by increased PMN counts, which lead to higher effector to target cell ratios in G-CSF-primed blood. Furthermore, bispecific antibodies, directed to FcalphaRI and Candida albicans, enhanced neutrophils' phagocytosis of fungi. In summary, these results identify IgA as an effective antibody isotype for immunotherapy, working primarily via FcalphaRI on neutrophils. They suggest FcalphaRI-directed bispecific antibodies and G-CSF to be an attractive combination for malignant or infectious diseases.     

Leptospirosis: an ignored cause of acute renal failure in Taiwan

Leptospirosis, caused by a spirochete, is the most common zoonosis in domestic or wild animals. Animals excrete infected urine in soil or water and may cause human infections through abrased wound, mucosa, conjunctiva, or by swallowing contaminated water. Clinical presentations of leptospirosis are mostly subclinical. Five to ten percent of leptospirosis are fatal, causing fever, hemorrhage, jaundice, and acute renal failure (Weil's syndrome). Leptospirosis has been ignored as a cause of acute renal failure in Taiwan. We report two patients with leptospirosis who presented with high fever, abdominal pain, jaundice, and acute renal failure. Patient 1 died on day 12 of admission of multiple organ failure associated with pancytopenia, hypogammaglobulinemia, and reactive hemophagocytosis. Leptospirosis was recognized after death. Patient 2 was admitted with similar presentations 2 weeks later. Penicillin and doxycycline were given early in the course, and azotemia, jaundice, respiratory failure, and aseptic meningitis gradually improved. Renal biopsy showed interstitial nephritis. Several tubular clearance tests showed proximal tubular defect with severe bicarbonate wasting (FeHCO3- 20.9%) and incomplete type II renal tubular acidosis without affecting the distal nephron. After 80 days of treatment, this patient was discharged with recovery of conscious level and renal function. This is the first leptospirosis patient with detailed tubular functional and morphological studies of the kidney. Diagnosis of leptospirosis was made by microscopic agglutination test (MAT) for antibody to leptospira and by polymerase chain reaction (PCR) for leptospira DNA in blood and urine (interrogans serogroup australis in case 1 and Leptospira borgpetersenii serogroup ballum in case 2). Because active surveillance has resulted in 13 cases diagnosed as leptospirosis islandwide thereafter, underestimation and ignorance of leptospirosis as a cause of acute renal failure may occur in Taiwan. Therefore, an area with a low leptospirosis incidence may actually have a very high incidence. Leptospirosis should be suspected in febrile patients with jaundice and renal failure when pathogens cannot be identified by traditional culture for microorganisms.     

TNF mediates lung leak, but not neutrophil accumulation, in lungs of rats given IL-1 intratracheally

Interleukin-1 (IL-1) is increased in lung lavages obtained from patients with acute respiratory distress syndrome, and administering IL-1 intratracheally to rats causes an acute, neutrophil-dependent, oxidative lung leak. We found that rats given IL-1 intratracheally had increased lung lavage fluid tumor necrosis factor (TNF) levels, and that rats treated with TNF binding protein (TNFbp) intravenously did not develop the increased lung leak that occurs after administration of IL-1 intratracheally. In contrast, rats given IL-1 intratracheally and TNFbp intravenously had the same elevations in lung lavage neutrophil accumulation and lung lavage cytokine-induced neutrophil chemoattractant levels as rats given IL-1 intratracheally. Our results show that TNFbp decreases neutrophil-mediated lung leak, but not lung neutrophil accumulation, after administration of IL-1 intratracheally in rats.     

Small changes in essential amino acid concentrations alter diet selection in amino acid-deficient rats

A new plate designed specifically to address complex wrist pathology was used for the internal fixation of 22 complex fractures of the distal radius in 22 patients in a prospective multicenter trial. The majority of fractures were group C2- and C3-type fractures according to the Comprehensive Classification of Fractures. No plate failures, loss of reduction, nonunions, or infections occurred. Within the average follow-up time of 14 months, the functional results (including an average motion of 76% and an average grip strength of 56% of the contralateral side) were comparable to those reported for similar fractures in previous investigations. Five patients had irritation of the tendons in the second dorsal compartment. This trial serves both as a verification of the safety and efficacy of this distal radius plate as well as a demonstration of its utility in the treatment of complex fractures of the distal radius.     

Digital quantification of the enzyme-linked immunospot (ELISPOT)

Enzyme-linked immunospot (ELISPOT) is a sensitive technique for the detection of cytokines released by immune cells. The technique is well established and correlates closely with the enzyme-linked immunosorbent assay (ELISA) technique. Here, we introduce an integrated imaging-analysis system to quantitate the spots formed by the ELISPOT assay. The spots can be easily counted when the background is clear and the spots are few. However, when the number of spots increases to > 100, this task becomes challenging and time-consuming. To minimize time and standardize the counting procedure, we have used a digital camera linked to a computer system for reading the plates. In general, the computer is able to count more spots and has a smaller standard deviation when compared with the manual microscopic count. This integrated system is commercially available, and we believe that this method is objective, time-saving and consistent in routine ELISPOT assay counting, especially when numerous spots are present.