DIVEMA derivatives

DIVEMA and some DIVEMA derivatives were synthesized and tested for toxicity and immunological behaviour. 

There are, however, many unsolved problems concerning toxicity, endocytic uptake and immunogenicity of DIVEMA. In order to learn more about the structure dependency of these properties, we sythesized some DIVEMA derivatives and investigated DIVEMA and the new derivatives with appropriate routine biological test systems. 

One disadvantage of DIVEMA is its relatively high toxicity which is mainly caused by high molecular weight fractions ( 7). In an attempt to lower the toxicity of DIVEMA, derivatives (3) and (4) were synthesized. 

Effects of DIVEMA and DIVEMA Derivatives on the Immune System. 

Are the DIVEMA derivatives mitogenic, i.e. do they stimulate lymphocyte proliferation ... 

Mitogenic Activity of DIVEMA Derivatives. A first approach to the effects of a compound on the immune system is to test its ability to stimulate lymphocyte proliteration. An appropriate test is to measure the incorporation of 3H-thymidine into DNA of dividing cells ... 

None of the substances caused significant enzyme secretion by macrophages. Only the pyrrolidone derivative (3a) caused a slight increase of LDH with increasing dose of the polymer. This finding corresponds to the higher acute toxicity of (3b) as compared to unsubstituted DIVEMA derivative (1b). 

Figure 2. Supernatant-mediated cytotoxicity caused by DIVEMA derivative (3b in Table II) (three independent runs).

Polymers (1a), (1b), (1d), (3a), (4a) and (5) were tested. Fi gure 3 shows the Endocytic Index (28) measured. The DIVEMA derivatives (1a), (3a) and (4a) Tfave no significant effect on the pinocytosis of 25j pyp (they do not influence the rate of formation of pinocytic vesicles). Polymer (1d) shows marked inhibition of pinocytosis at the highest dose.(This latter finding in vitro corresponds to in vivo results of Breslow et aTi (29)). They found that the removal of colloidal carbon from the blood of test animals is inhibited by high molecular weight DIVEMA). 

Figure 3. Stimulation of endocytosis by D1VEMA and DIVEMA derivatives. Endocytic index in the macrophage test system and ml-PVP used as standard substance.

DIVEMA (Hydrolyzed form of divinyl ether and maleic anhydride copolymer) Cyclophos- phamide Ester bond between drug derivative hydroxyl and polymer carboxyl None N/A N/T Anticancer activity of conjugate was comparable to free drug against L1210 leukemia bearing mice 58... 

The compounds tested showed mitogenic effects,i.e. they stimulated lymphocyte proliferation. Astonishingly, a DIVEMA-DNP conjugate did not induce the production of DNP-specific antibodies. Our results suggest that this is due to the formation of DNP-specific active suppressor cells. Using serum free culture media, we could show for the first time that DIVEMA and derivatives stimulate macrophages to release cytotoxic factors into the supernatant. However, no significant release of lytic enzymes was detected. 

DIVEMA and the derivatives tested failed to stimulate endocytosis by macrophages of iZ5I-PVP and 198/ u (colloidal) higher mol. wt. DIVEMA inhibited pinocytosis. We conclude that the only effect of the compounds tested on macrophages is the release of cytotoxic factors into the supernatant. 

DIVEMA samples (1a - 1d) were kindly supplied by Dr.D. Breslow, HERCULES Inc. The synthesis of the derivatives (11) will be published elsewhere. The compounds investigated are listed in Table I-... 

Table II. Acute toxicity of DIVEMA and some derivatives...

DIVEMA is known to act upon different types of immune cells (J 3) such as the antibody producing B cells (bone marrow derived lymphocytes) on T-cells (thymus processed lymphocytes) and on macrophages (cells which phagocytose foreign material). The mechanisms of interaction between polymers and the immune system are very complex. In order to investigate them, many different experimental approaches can be used. 

DIVEMA Cydophosphamide derivatives L1210-beaiing mice 110)... 

The antitumour drug cyclophosphamide requires activation by microsomal ai-zymes before it displays the alkylating activity essential for the pharmacological effect. The active metabolite of cyclophosphamide is 4-hydroxycyclophosphamide which is not very stable and therefore unsuitable for attachment to carriers. Hirano et al. attached stable mercapto derivaties of 4-hydroxycyclophosphamide to DIVEMA and later evaluated the rate of hydrolysis of the polymeric derivatives and their activity against L1210-bearing mice Unfortunately, the polymeric derivatives... 

Wang synthesized a munber of similar platinum-containing polymers derived from ring-opened DIVEMA (46). Along with the 1,2-cyclohexanediamine platinum compound, other complexes of cis-PtL2Cl2 and cis-PtL2l2 were studied. These include compounds where L = NH3, isopropylamine, aziridine, and L2 = 2,2 -bipyridine, 1,3-propanediamine, and o-phenylenediamine. 

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