Preparation and anti-HIV activity of O-acylated heparin and dermatan sulfate derivatives with low anticoagulant effect.

In order to increase the ratio of anti-HIV activity to anticoagulant activity, glycosaminoglycan derivatives selectively substituted at OH and/or COOH groups were prepared. Standard heparin, heparin fragments, or dermatan sulfate were converted to their tributylammonium or tetrabutylammonium salts. Their selective O-acylation to various (controlled) degrees was carried out in a homogeneous way in N,N-dimethylformamide using carboxylic acid anhydrides and 4-(dimethylamino)pyridine as catalyst. Esterification of the COOH groups was performed by the addition of alkyl halide to an N,N-dimethylformamide solution of glycosaminoglycan tetrabutylammonium salts. The in vitro anticoagulant activity, the activity against HIV-1 and HIV-2 cytopathicity, the cytotoxicity, and the activity on the induction of giant cell formation were determined. O-acylation (O-butyrylation or O-hexanoylation) of the heparin fragments obtained by periodate depolymerization (compounds 2d and 2e), and their esters (compounds 7i and 7j), yielded products with very low anticoagulant effects in vitro, yet potent activity against both HIV-1 and HIV-2 induced cytopathicity, and low, if any, cytotoxicity. As compared to other anionic polysaccharides, these acylated derivatives are more active as inhibitors of HIV-induced giant-cell formation. Their anti-HIV activity is related to the degree of O-acylation and is mainly due to the inhibition of virus adsorption to the target cells.

O-acylated heparin derivatives with low anticoagulant activity decrease proliferation and increase alpha-smooth muscle actin expression in cultured arterial smooth muscle cells.

Selectively O-acylated derivatives of various glycosaminoglycans were prepared and tested in vitro for their anticoagulant activity and their antiproliferative effect on rat and rabbit smooth muscle cells. When O-acylation (butyrylation or hexanoylation) had been performed on periodate-depolymerized heparin fragments having very low anticoagulant activity, the antiproliferative potency was markedly increased (IC50 = 2 and 1 micrograms/ml respectively, versus 31 micrograms/ml for starting compound) without an increase in anticoagulant activity. The antiproliferative activity was related to the degree of acylation. The O-acylated derivatives of heparin fragments were also very active in reversing the de-differentiation of smooth muscle cell in culture, as estimated by the increase in the expression of alpha-smooth muscle actin and alpha-smooth muscle actin mRNA.

Antithrombotic activity, bleeding effect and pharmacodynamics of a succinyl derivative of dermatan sulphate in rabbits.

This paper compares the pharmacological properties of a new succinyl dermatan sulphate derivative (Suc-DS) to those of the natural dermatan sulphate (DS). Suc-DS was on average 2-3 times more potent than DS in catalysing the inhibition of thrombin by heparin cofactor II and in prolonging the activated partial thromboplastin time and the thrombin clotting time. After bolus injection, Suc-DS was also 2-3 times more potent than DS to prevent experimental venous thrombosis in a Wessler model. Thromboplastin or human serum were used as the thrombogenic stimulus. In contrast, the bleeding effect assessed by rat tail transection technique was comparable. After bolus intravenous injection, the pharmacodynamics of Suc-DS indicated a lower volume of distribution, which was close to the plasma volume, and a slightly lower clearance of elimination. Therefore this chemical alteration of natural DS yields a new compound with an improved antithrombotic benefit/haemorrhagic risk ratio.

[Pulmonary and muscular localization of Wegener's granulomatosis. Apropos of a case]. / Localisation pulmonaire et musculaire d'une granulomatose de Wegener. A propos d'un cas.

The authors report their observations on a patient presenting with Wegener's granulomatosis with initially isolated pulmonary involvement. Following treatment with a combination of steroid therapy and immunosuppressives muscular involvement developed with specific histological changes. The disease was only controlled by substituting at the same dose prednisone for the methylsulphobenzoate of prednisolone which was prescribed initially; this suggests in equivalent doses the superiority of one product over the other was achieved by a different pharmacodynamic behaviour.

Induction of murine macrophage tumoricidal activity and treatment of experimental pulmonary metastases by liposomes containing lipophilic muramyl dipeptide analogs.

The ability of three members of a new class of lipophilic muramyl dipeptide derivative to induce murine macrophage tumoricidal activity after liposomal incorporation was investigated. Liposomes containing the glycerol dipalmitate (GDP) derivatives of N-acetylmuramyl-L-alanyl-D-isoglutamine, N-acetylmuramyl-L-alanyl-D-glutamine-n-butyl ester, and N-acetylmuramyl-D-alanyl-D-isoglutamine were 5000, 2000, and greater than 10,000-fold more potent than the free muramyl dipeptides in inducing peritoneal macrophage tumoricidal activity in vitro. In situ activation of peritoneal macrophage tumoricidal activity showed that liposomal muramyl dipeptide-GDP derivatives were more potent than free hydrosoluble or sonicated muramyl dipeptide-GDP preparations. In situ induction of alveolar macrophage tumoricidal activity after i.v. treatment was observed with liposomes containing muramyl dipeptide-GDP derivatives, but not with hydrosoluble or sonicated lipophilic derivatives. Liposomes containing muramyl dipeptide-GDP derivatives were therapeutically active against experimentally induced pulmonary B16 melanoma tumors in C57BL/6 mice. These results demonstrate that when incorporated within liposomes this class of lipophilic muramyl dipeptide derivative is a potent inducer of macrophage tumoricidal activity both in vitro and in situ, and possesses antitumor activity in therapeutic treatment protocols.

A novel muramyl peptide derivative stimulates tumoricidal activity of macrophages and antibody production by B cells.

A novel analog of MDP, the 3'-iodo-4'-azido-L-phenylalanine methyl ester derivative of N-acetyl-L-alanyl-D-isoglutamine, has been prepared. This compound is capable of activating macrophages to the tumoricidal state and increasing the specific immune response of B cells. It thus appears to exhibit similar biological activities to MDP. Moreover, this compound is of potential interest for receptor photolabelling studies.

A comparative 1H-n.m.r. study of MurNAc-L-Ala-D-iGln (MDP) and its analogue murabutide: evidence for a structure involving two successive beta-turns in MDP.

The active principle, MurNAc-L-Ala-D-iGln (MDP), of complete Freund's adjuvant and its analogue, MurNAc-L-Ala-D-Gln-OnBu (murabutide), which express immunomodulatory as well as other biological properties, have been studied by 2D-1H-n.m.r. spectroscopy at 500 MHz. The results suggest the presence in MDP of two successive turns involving the MurNAc-L-Ala and L-Ala-D-iGln moieties, respectively, whereas only the former turn persists in murabutide. This turn mimics the type II beta-turn found in L-D depsipeptides, whereas the other is a typical type II beta-turn for L-D peptides.

Macrophage stimulation in vitro by an inactive muramyl dipeptide derivative after conjugation to a multi-poly(DL-alanyl)-poly(L-lysine) carrier.

It has been previously reported that N-acetyl-muramyl-L-alanyl-D-isoglutamine (MDP), which represents the minimal structure that can substitute for mycobacteria in Freund complete adjuvant, activated macrophages in vitro and in vivo. In the present study we show that, in contrast to MDP, the nonadjuvant MDP(DD) stereoisomer has no effect on cytostatic activity of thioglycolate-induced macrophages as measured by uptake of [3H]thymidine. However, surprisingly, after conjugation to an inert carrier, multi-poly(DL-alanyl)-poly(L-lysine), this compound activates macrophages in vitro and becomes at least as effective as MDP. It has also been shown in other studies that after conjugation MDP(DD) remained devoid of antigenicity and of adjuvant activity although such a conjugate could increase resistance to infection. It, therefore, appears that there exists no correlation between the structure required for adjuvant activity and the structure required for macrophage activation or for enhancement of nonspecific immunity.

Immunostimulant activities of a lipophilic muramyl dipeptide derivative and of desmuramyl peptidolipid analogs.

The immunostimulant properties of a new muramyl dipeptide (MDP) derivative bearing a lipophilic moiety on the C-terminal end of the peptide chain are described. It is shown, in particular, that 1,O-(acetylmuramyl-L-alanyl-D-isoglutamine-L-alanyl)-glycerol-3-mycolate had increased immunostimulant activity in comparison with MDP. It induced hypersensitivity even when administered with an antigen in saline, and it gave higher protection against bacterial infections than did MDP. A quite unexpected finding was obtained with the corresponding desmuramyl compound 1,O-(L-alanyl-D-isoglutamine-L-alanyl)-glycerol-3-mycolate, which had no activity in producing humoral antibodies but was just as active as the muramic acid-containing compound in stimulating nonspecific resistance to bacterial infections. It was not pyrogenic. Modifications of the peptide moiety or the lipid moiety of this peptidolipid led to decrease, or even loss, of activity. These results show the importance of the N-acetylmuramyl moiety in MDP for humoral antibody production. The peptidolipid 1,O-(L-alanyl-D-isoglutamine-L-alanyl)-glycerol-3-mycolate is the first member of a new category of nonspecific immunostimulants.

[Bacterial counts in sputum. The value and interpretation in lower respiratory tract infections (author's transl)]. / La numération bactérienne des expectorations. Sa valeur et son interprétation dans les infections des voies respiratoires inférieures.

A convenient and effective microtechnic of bacteria numeration in sputum has been used and its results are presented, considering the accompanying clinical data. One has therefore been able to appreciate the signification of the number and prevalance of bacteria isolated in lower respiratory tract infections: 1) The concentration of 10(7) bacteria/ml is a useful critical value, but it does not mean either virulence or lower limit of pathogenicity. 2) The importance of the predominance of a bacterial strain is an essential parameter in quantitative analysis. 3) The interpretation of the bacteriological results can only be made after thoroughly examining the cytology, the clinical infection symptoms, the anamnestic data, and considering the possibility of an antibiotic therapy preceding the test.

[Pulmonary fibrosis during a case of dermatomyositis (author's transl)]. / Fibrose pulmonaire au cours d'une dermatomyosite.

The authors describe the case of a pulmonary interstitial fibrosis, the discovery of which preceeded the skin and muscular manifestations of dermatomyositis which was confirmed by a muscular biopsy. The aetiopathogenesis of the pulmonary disorder is not well known, nevertheless an immunological mechanism demonstrated in the experimental myositis is suggested but has yet to be proved.

[Mucoepidermoid tumors in the bronchi. About three cases (author's transl)]. / Les tumeurs muco-épidermoïdes des bronches. A propos de trois cas.

The little frequent mucoepidermoid tumors in the bronchi can be distinguished by their histological characteristics associating epidermoid and mucilage-like cells identified through alcian blue and PAS colour agents. Although these tumors are normally considered as "benign", some of them may have a capacity to develop in very malign way. We enclose three new findings which all developped unfavorably in a short period.

Synthesis of N-acetyl-muramyl-L-alanyl-D-glutamic-alpha-amide(MDP) or -alpha-methyl ester derivatives, bearing a lipophilic group at the C-terminal peptide end.

We report the synthesis of nine lipophilic derivatives of N-acetyl-muramyl-L-alanyl-D-glutamic-alpha-amide (MDP) or -alpha-methyl ester in which the gamma-carboxyl function of the D-glutamyl residue is either esterified by a medium chain alcohol or substituted by an L-alanyl residue esterified by a medium or long chain alcohol. A new method is described which easily allows one to obtain derivatives of MDP, bearing a free or substituted amino-acyl or peptidyl residue on the gamma-carboxyl function.

[A critical study of immuno-enzymatic reactions coupled with the precipitation tests on cellulose acetate membranes]. / Etude critique des réactions immuno-enzymatiques couplées aux tests de précipitation sur membranes d'acétate de cellulose.

Precipitating tests carried out on cellulose acetate membrane can be increased by treating the immune complexes with enzyme linked anti-immunoglobulin antibodies. From our trials in parastic diseases (Amoebiasis, schistosomiasis, fasciolasis, filariasis, hydatidosis, trichinosis) and mycosis (Aspergillosis, Candidiasis), it seems that the immuno-enzymatic labelling of the "active" precipitating reactions should only be taken in consideration. We must insit on the importance of ELIEDA (enzyme-linked-immuno-electrodiffusion-assay) and ELIDEPA (enzyme-linked-immuno-double-electro-phoresis-assay). Both of these analytical assays are particularly sensitive. The sequence of appearance of the multiple precipitating systems of the complex parasitic mosaic can easily be watched. The different classes of immunoglobulins and their kinetics are determined by the use of monospecific antibodies linked to different enzymes which give a polychromic specific staining.