Peter Dukor

Immunopharmacological activities and clinical development of muramyl peptides with particular emphasis on murabutide

Certain immunopharmacological activities of muramyl peptides have been associated with inflammatory and undesirable side-effects typically observed following the administration of the prototype molecule muramyl dipeptide. This activity is now demonstrated not to be linked to a direct activation of inflammatory processes in endothelial cells. Neither MDP nor other structural derivatives were able to induce inflammatory cytokines release or E-selectin gene expression in cultured human umbilical vein endothelial cells. However, oral administration of muramyl peptides has been reported to induce certain biological effects, including the downregulation of anamnestic, antigen-specific IgE responses, which are not observed following parenteral administration. We elaborate on these findings and extend them to show the efficacy of a new muramyl peptide in suppressing polyclonally induced serum IgE levels in anti-IgD-treated mice. The comparative effects of muramyl peptides, selected for clinical development, on the induction of cytokines in human whole blood are then presented at the level of mRNA accumulation and protein secretion. Moreover, the cytokine profile induced in vitro and in vivo by the combination of the safe immunostimulant, Murabutide, with interferon-alpha is examined. This combination reveals a selective and beneficial synergistic activity and induces anti-inflammatory cytokines in the absence of synergistic toxicity. The potential and the implications for the use of a therapeutic combination of an immunostimulant with a cytokine are discussed.

Modulation of myelopoiesis by different bacterial cell-wall components: Induction of colony-stimulating activity (by pure preparations, low-molecular-weight degradation products and a synthetic low-molecular analog of bacterial cell-wall components) in vitro

Abstract Chemically pure preparations of three structurally unrelated components of the cell wall of gram-negative bacteria (BCWC), lipid A, outer-membrane lipoprotein, and murein, were tested for lymphocyte mitogenicity and the ability to induce colony-stimulating activity (CSA) in various serum-free tissue-culture systems. All three components were B-cell mitogens and induced CSA in spleen-cell cultures. However, in lymphnode-cell cultures the concentrations of these agents required for either mitogenicity or CSA induction differed markedly. Moreover, in contrast to thymidine incorporation, CSA induction was not influenced by pre-irradiation of the cells. Conversely, after removal of phagocytic cells with the iron-magnet technique, CSA was no longer inducible by BCWC, while lymphocyte proliferation was barely impaired. All three BCWC readily induced CSA release in cultures of adherent peritoneal cells without influencing the release of a cytoplasmic enzyme. BCWC-dependent CSA release from adherent peritoneal cells was not influenced by pretratment of the cultures with anti-immunoglobulin, but completely suppressed by preincubation with anti-macrophage-1.2 alloantiserum and complement. CSA induction in macrophage cultures was also achieved with a low-molecular-weight synthetic muramyldipeptide and degradation products of lipoprotein. The results suggest that the induction of CSA is not directly related to the mitogenic, immunogenic, or antigenic properties of the BCWC, but that BCWC-mediated CSA production is caused by a direct “hormone-like” interaction of the agents with mature macrophages.

Selective Stimulation of Mouse Lymphocyte Populations by Lectins

The mitogenic effect of a number of plant agglutinins was investigated in mouse lymphocyte cultures set up with different populations of T and B cells (lymph node cells and thoracic duct cells from no

Induction of tumouricidal leucocytes by the intranasal application of MTP-PE, a lipophilic muramyl peptide

SummarySingle intransal applications of MTP-PE, a lipophilic muramyl peptide, induce tumouricidal and tumouristatic leucocytes in the lungs of rats. In ex vivo assays the tumouristatic activity was detectable for 8 days after drug administration. By separation of the effector cells on Ficoll-Hypaque gradients, it was shown that both neutrophils and macrophages are responsible for this activity. Using the B16/BL6 melanoma system in mice, there was a high survival rate after repeated intranasal applications of MTP-PE

New developments in drugs enhancing the immune response: activation of lymphocytes and accessory cells by muramyl-dipeptides

Recently, stimulation of host defence mechanisms has become a major goal of pharmacotherapeutic research. Immunopotentiating compounds exert their effects in different ways. On the one hand, they may enhance non-specific effector mechanisms operative in the resistance of infectious agents and to neoplastic cells. On the other hand, they may non-specifically increase specific immune responses elicited by the recognition of antigenic determinants. Of course, the two mechanisms operative in the resistance to infectious agents and to neoplastic cells. On may be mediated by the pharmacological activation of common target cells, that is macrophages and other accessory cells, such as polymorphonuclear leucocytes.

A three-cell mosaic culture: In vitro immune response by a combination of pure B- and T-cells with peritoneal macrophages

Abstract Lymph node cell suspensions from BD 2 F 1 and nu/+ mice contained very low numbers of macrophages and failed to produce PFC upon exposure to sheep erythrocytes if cultured alone. Addition of 99%-pure, attached mononuclear peritoneal phagocytes resulted in the development of PFC comparable to that seen with spleen cells. Spleen cells from congenitally athymic nu/nu mice were lacking θ-positive cells and responded to sheep erythrocytes only if cortisone-resistant nu/+ thymus cells were added. Lymph node cells from nu/nu mice (devoid of both θ-positive cells and phagocytes) responded in culture provided both cortisone-resistant thymus cells and purified macrophages from nu/+ donors were present. None of the three components displayed any activity if cultured alone. Omission of only one component of the three-cell system also greatly diminished the response. Full reactivity of the composite cultures could be achieved by the addition of a relatively small proportion of cortisone-resistant thymus cells. The results are discussed in terms of nonspecific T-cell activation and of a three-cell model of the in vitro immune response to sheep red cells.

Oral administration of muramyl dipeptide into mice modulates cell proliferation, immunoglobulin synthesis and cytokine mRNA levels in gut associated lymphoid tissues

Muramyl peptides (MDPs) are synthetic immunostimulants capable of potentiating a multitude of immune functions following parenteral administration into a host. The parent molecule MDP was also found to exert certain activities when applied via the oral route. Thus, we have studied the effect of oral treatment of mice with MDP on the lymphoproliferative responses, immunoglobulin secretion and cytokine induction in gut-associated lymphoid tissues (GALT) employing lymphocyte transformation test, ELISA and RT-PCR, respectively. Cells derived from Peyers patches (PP) of mice orally primed with MDP were found to have enhanced proliferative responses to different mitogens and to secrete significantly more IgG, IgM and IgA immunoglobulins than cells from unprimed mice. These effects were not observed with cells derived from mesenteric lymph nodes (MLN) or spleens of MDP-primed mice. However, oral administration of MDP resulted in the up-regulation of interleukin-6 (IL-6) mRNA in MLN and down-regulation of IL-4 and tumour necrosis factor-alpha (TNF-alpha) mRNAs in MLN, spleens and PP. These studies suggest that selective modulations of GALT responses by orally administered MDP are achievable and imply that these agents may be useful in the therapy and prophylaxis of allergic diseases.

Suppression of in vivo IgE and tissue IL-4 mRNA induction by SDZ 280.636, a synthetic muramyl dipeptide derivative

Modulation of IgE isotype expression on B cells is one of the numerous effects of muramyl peptides on the regulation of the immune system. A non toxic diacyl glycerol derivative of muramyl dipeptide (MDP), in which the L-alanine is replaced by L-threonine (MDP-Threo-GDP; SDZ 280.636), is currently under investigation as lead compound for the development of an anti-allergic drug. In this report, the modulatory effect of orally administered SDZ 280.636 in a murine model on polyclonally induced IgE levels is described. In this model, mice are injected i.v. with goal anti mouse IgD (GAMD) and challenged three to four weeks later with goal IgG (GIG). Both the primary and secondary immune responses lead to an increase of serum IgE levels. We demonstrate the efficacy of this muramyl dipeptide derivative in selectively inhibiting a polyclonal IgE response in GAMD-primed, GIG challenged mice without affecting the levels of other immunoglobulin classes. It is further shown that the induction of interleukin 4 (IL-4) gene transcript levels in lymphoid organs, which is observed as a consequence of boosting GAMD pretreated mice with GIG, is selectively suppressed in gut associated lymphoid tissues (GALT) and mesenteric lymph nodes but not in spleen. In contrast, interleukin 13 (IL-13) mRNA levels are not affected by SDZ 280.636. The findings that SDZ 280.636 inhibits polyclonal IgE responses and suppresses IL-4, but not IL-13 mRNA expression point towards differences in the regulatory pathways of IL-4 and IL-13 gene transcription in lymphoid organs. Thus the mechanism of action appears to involve a specific suppression of IL-4 gene transcription in cells occurring in Peyers patches and mesenteric lymph nodes which are among the first constituents of the immune system encountered by an orally administered drug.

Functional and Morphologic Characteristics of Complement Receptor Lymphocytes in Mice

Complement receptor lymphocytes (CRL) (1) are bone marrow-derived (2) lymphoid cells capable of binding antigen-antibody-complement complexes through a membrane receptor for modified C3. CRL are located in follicular areas of peripheral lymphoid tissues (3) and have been shown to comprise the precursors of antibody forming cells (4).