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Peripheral blood lymphocytes from healthy, HIV sero-negative blood donors have been in vitro immunized using penv9, a recombinant fragment of the envelope of HIV-1. This primary in vitro immunization followed by Epstein-Barr virus (EBV) transformation and somatic cell fusion subsequently gave rise to several specific anti-penv9 monoclonal antibodies (MO28, MO30 and MO43) of μ isotype. The hybridomas have been kept in culture for over 6 months and the antibody productivity for MO30 was measured to 18 μg × (24 hr × 106 cells)-1. The fine specificity of the antibodies was mapped by a peptide inhibition enzyme immunoassay, using overlapping synthetic pentadeca peptides covering the whole penv9. These human monoclonal antibodies exhibited a similar epitope specificity directed against a non-sequential determinant, including the amino acids 632–646, 677–681 and 687–691. This specificity is very rarely found in immune sera from sero-positive patients and presently not reported in human monoclonal antibodies derived from in vivo immunized individuals, indicating that different antibody specificities can be obtained by the in vitro immunization technology. These human monoclonal antibodies did not neutralize HIV. The results presented here demonstrate the feasability of generating human monoclonal antibodies against HIV by primary in vitro immunizations, thereby avoiding the use of lymphocytes derived from infected patients when human monoclonal antibodies for therapeutic purposes are to be produced. [ABSTRACT FROM AUTHOR]

HIV-2, compared to HIV-1, elicits potent and broadly neutralizing antibodies, and uses a broad range of co-receptors. However, both sensitivity to neutralization and breadth of co-receptor use varies between HIV-2 isolates, and the molecular background is still not fully understood. Thus, in the current study, we have deciphered relationships between HIV-2 neutralization sensitivity, co-receptor use and viral envelope glycoprotein (Env) molecular motifs. A panel of primary HIV-2 isolates, with predefined use of co-receptors, was assessed for neutralization sensitivity using a set of HIV-2 Env-directed monoclonal antibodies and co-receptor indicator cell lines. Neutralization sensitivity of the isolates was analysed in relation target cell co-receptor expression, in addition to amino acid motifs and predicted structures of Env regions. Results showed that HIV-2 isolates were more resistant to neutralizing antibodies when entering target cells via the alternative co-receptor GPR15, as compared to CCR5. A similar pattern was noted for isolates using the alternative co-receptor CXCR6. Sensitivity to neutralizing antibodies appeared also to be linked to specific Env motifs in V1/V2 and C3 regions. Our findings suggest that HIV-2 sensitivity to neutralization depends both on which co-receptor is used for cell entry and on specific Env motifs. This study highlights the multifactorial mechanisms behind HIV-2 neutralization sensitivity. [ABSTRACT FROM AUTHOR]

Alemtuzumab is a humanised monoclonal antibody directed against the CD52 antigen. It is approved for the treatment of B-cell chronic lymphocytic leukemia (CLL) as a monotherapy and is being investigated as combination therapy and consolidation therapy for CLL, as well as in conditioning regimens for stem cell transplantation. The pharmacokinetics (PK) of alemtuzumab is best described by a two-compartment model with large interpatient variability in all PK parameters. Analyses of small patient cohorts suggest that higher serum alemtuzumab levels are associated with better treatment responses. Several patient-specific factors, such as disease status, tumor burden and soluble CD52 levels, appear to influence serum alemtuzumab level. Future studies are needed to improve the PK model of alemtuzumab and to explore a PK-guided dosing schedule, with the goal of maximising the therapeutic benefit of this agent. [ABSTRACT FROM AUTHOR]

The natural history and outcome of salvage treatment for patients with fludarabine-refractory chronic lymphocytic leukemia who are either refractory to alemtuzumab ("double-refractory") or ineligible for alemtuzumab due to bulky lymphadenopathy ("bulky fludarabine-refractory") have not been described. We present the outcomes of 99 such patients (double-refractory n = 58, bulky fludarabine-refractory n = 41) undergoing their first salvage treatment at our center. Patients received a variety of salvage regimens including monoclonal antibodies (n = 15), single-agent cytotoxic drugs (n = 14), purine analogue combination regimens (n = 21), intensive combination chemotherapy (n = 36), allogeneic stem cell transplantation (SCT; n = 4), or other therapies (n = 9). Overall response to first salvage therapy other than SCT was 23%, with no complete responses. All four patients who underwent SCT as first salvage achieved complete remission. Early death (within 8 weeks of commencing first salvage) occurred in 13% of patients, and 54% of patients experienced a major infection during therapy. Overall survival was 9 months, with hemoglobin < 11 g/dL (hazard ratio 2.3), hepatomegaly (hazard ratio 2.4), and performance status ≥ 2 (hazard ratio 1.9) being significant independent predictors of inferior survival. [ABSTRACT FROM AUTHOR]

Alemtuzumab, the monoclonal anti-CD52 antibody, has clinical activity in B-cell and T-cell malignancies at the dose of 30 mg three times weekly for 9 - 12 weeks. This standard regimen induced responses usually shorter than 6 months. To prolong time to progression, we initialized a phase II study with an identical initial scheme until partial response, followed by a maintenance therapy lasting at least 4 months. Eleven heavily pretreated patients (8 with B-chronic lymhocytic leukemia (B-CLL) and 3 with small lymphoctyic lymphoma (SLL)) have been treated with this maintenance regimen (MR patients) and were retrospectively compared to 5 patients (3 B-CLL and 2 SLL) treated with the standard regimen (SR patients). Patients characteristics before treatment were identical in both groups. Objective response was reached by 9 (82%) MR patients and 3 (60%) SR patients (p NS). After the treatment, 8 (73%) MR patients and all SR patients progressed with a median time at 12.2 months and 3 months respectively. Survival time from alemtuzumab was significatively different ( P  < 0.005). None of the patients died in the MR group with a median follow-up at 16 months. In the SR group, the median survival from alemtuzumab was 5.9 months. We did not observe any differences in terms of hematological toxicites and infections between the two groups. In conclusion, maintenance alemtuzumab therapy seems to increase the time to progression and the survival, without adding hematological toxicities and infectious complications. More patients are needed to confirm this observation. [ABSTRACT FROM AUTHOR]

In this review B cell responses in HIV-infected individuals are summarized together with the techniques used to date to produce human monoclonals to HIV and the properties of these antibodies. Profound disturbances in B cell responses are apparent both in vivo and in vitro. While there is evidence in vivo of marked polyclonal B cell activation, primary and secondary antibody responses are impaired. Similarly these cells exhibit spontaneous immunoglobulin secretion upon in vitro culture but do not readily respond to B cell mitogens and recall antigens including HIV. Furthermore, certain of these defects can be reproduced in normal B cells in vitro by incubation with HIV or HIV coded peptides. Individuals infected with HIV develop antibodies to HIV structural proteins (e.g. p17, p24, gp41 and gp120) and regulatory proteins (e.g. vif, nef, RT). Autoantibodies against a number of immunologically important molecules are also frequently observed. The anti- HIV antibodies are predominantly of the IgO1 isotype and exhibit a variety of effects on the virus in vitro. To date, using conventional immortalization strategies, an appreciable number of human monoclonals to HIV have been developed. These have been specific for gp41, gp120 and gag with antibodies of the former specificity predominating. The majority of these antibodies have been of the IgG I isotype. Only a small number of the antibodies neutralize virus in vitro and most of these react with gp 120. The neutralizing antibodies recognize conformational and carbohydrate epitopes or epitopes in amino acid positions 306-322. The predominant epitopes recognized by the anti-gp41 antibodies were in amino acid positions 579-620 and 644-662. A high percentage (≏ 25%) of these antibodies enhance viral growth in vitro. The problems relating to the production of human monoclonals to HIV are discussed together with strategies that could be used in the future. [ABSTRACT FROM AUTHOR]

Human monoclonal antibodies specific for human cytomegalovirus (CMV) antigens have been established using peripheral blood lymphocytes from a seropositive donor. Immortalization of antigen-specific B cells was achieved by Epstein-Barr virus transformation followed by somatic cell fusion of antigen-specific lymphoblastoid cells. Four clones producing high-affinity antibodies (0.2-7 × 109 M-1) specific for the viral matrix protein pp65 have been further characterized with respect to epitope specificity of secreted antibodies. The studied antigen represents a major protein produced by in vitro-cultivated virus, and is important in the scrodiagnosis of CMV infection. The human monoclonal antibodies recognized different epitopes. some of which proved to be overlapping. The fine specificity of these antibodies was evaluated using synthetic peptides covering the sequence of pp65. The antibody MO58 recognized a linear epitope (residues 283-288) whereas antibody MO53 recognized a discontinuous epitope involving residues 208-216 and 280-285. Despite the close proximity of these epitopes, the antibodies did not compete with each other for the same binding site on intact antigen. [ABSTRACT FROM AUTHOR]

Human-mouse hybridoma cell lines producing human monoclonal antibodies against the cardiac glycoside digoxin were established after in vitro immunization or direct immortalization of human peripheral blood lymphocytes with digoxin. Three antibodies, designated MO6, LH92 and LH114, displayed different patterns of fine specificity against digoxin and several digoxin analogues, as elucidated by inhibition ELISA. All three monoclonal antibodies had μ heavy chains, two of them (MO6 and LH114) had κ light chains and one (LH92) λ light chains. DNA encoding the variable regions of both heavy and light chains of the three antibodies were amplified from cDNA using the polymerase chain reaction (PCR). The nucleotide sequences of the amplified DNA were determined after subcloning of PCR fragments in MI3 vectors. The deduced amino acid sequences revealed considerable sequence differences in the complementarity determining regions between the three antibodies. [ABSTRACT FROM AUTHOR]

The article presents information on author index of the December 1, 1986, issue of the periodical "Immunology." Some of the articles as their authors are, "The Ig Class Distribution of Anti-Phosphoryl Choline Responses in Mice Infected With Parasitic Nematodes," by N.M. Almond and R.M.F. Parkhouse, "Passively Acquired Antibodies to Respiratory Syncytial Virus Impair the Secondary Cytotoxic T-cell Response in the Neonatal Mouse," by C.R.M. Bangham, "Production and Characterization of Pepsin Fragments of Human IgA I to Determine Domain-Specificity of Monoclonal Anti-4gA Antibodies," by J. Biewenga, A. Faber, J.C. Pronk, J.J. Haauman, and others.

Human B cells are polyclonally activated in vitro by T cells stimulated with immobilized anti-CD3 monoclonal antibodies. We have analysed the effect of CD3 ligation on the production of antigen-specific antibodies, using peripheral blood lymphocytes from tetanus toxoid vaccinated blood donors. High levels of antigen-specific antibodies were obtained after stimulation with anti-CD3 antibodies for 7 days. Addition of a soluble recall antigen did not affect the total amount of Ig produced, but dramatically decreased the antigen-specific response. The addition of IL-2, IL-4, anti-CD40 or anti-CD28 antibodies or the removal of antigen did not restore the B cell response. Analysis using limiting dilution of B cells showed that the frequency of antigen-specific memory B cells decreased significantly in cultures stimulated with antigen. The antigen-specific B cell response could be completely restored only if the soluble antigen was cross-linked on the surface of the B cells. These results suggested that peripheral memory B cells were eliminated or anergized in the presence of soluble antigen. [ABSTRACT FROM AUTHOR]

The article provides evidence that subcutaneous Rituximab, an anti-CD20 monoclonal antibody that is routinely used as part of the management of the majority of B cell malignancies, can be safely administered without the need for any pre-medication. Topics discussed include percentage of cases were severe infusion-related reactions are seen including bronchospasm and hypotension, and two changes that are required to administer Rituximab by the subcutaneous route.

The author reflects on the role of monoclonal antibody alemtuzumab in the treatment of chronic lymphocytic leukemia (CLL). According to the author, alemtuzumab is effective against malignant B cells in the blood in CLL. In fact, he points out that alemtuzumab has been used for treatment of relapsed/refractory CLL after alkylating agents and purine analogues.