SELF-ASSEMBLING ION PAIR DRUGS

Abstract: By supplying soluble forms of positive and negative ion components of an ion pair capable of formation in situ in a targeted organism, cell or tissue, the administration of an ion pair conjugate affective to modify the target can be facilitated. In particular, ion pairs using tetrachloroplatinate as a negative ion and organic cations are disclosed.

Description: SELF-ASSEMBLING ION PAIR DRUGS Technical Field The invention relates to treatment of target conditions such as tumors and infectious agents in in vitro and in vivo environments. More specifically, the invention concerns in situ formation of ion pair conjugates which are active modifying agents for such targets. Background Art U.S. Patent 4,812,449, issued to the applicant herein and incorporated herein by reference discloses the general approach of providing active materials in the form of component precursors so as to permit self assembly in a target icroenvironment. As described in the above referenced patent, a "target" refers to an organism, tissue, cell, or other biologically responsive material which it is desired to modify. The target will occur in a general environment which may be an in vitro or an in vivo environment, and supplies its own "microenvironment". The patent discloses a large number of reaction types which may be employed to form a conjugate in situ, all of them involving covalent bond formation. As explained in this issued patent, the self- assembly of a conjugate which is intended to interact with the target in situ has a number of advantages, including selectivity provided by the individual components, delay of the effect until the assembly is completed in the microenvironment in which it is intended to take place, enhancement of rate constants and the selectivity of the components, and/or a resulting dosage enhancement as compared to administration of the preformed conjugate. The invention herein represents a particularly advantageous application of the general approach of in situ conjugate formation, namely, use of a self- assembling ion-pair in contrast to covalent bond formation. The ion pair then can modify the target after administration of the components either in vivo or in vitro. In addition, it is possible to enhance the selectivity for target by conjugating one or both of the components of the self-assembling ion pair conjugate to a target-selective ligand. Disclosure of the Invention The invention provides methods and materials useful in therapy or prophylaxis or other modification of target organisms, cells or tissues. The active agents are ion pair conjugates which are formed in situ by positive ion and negative ion components supplied separately to the environment of a target condition. The approach is particularly advantageous because the positive and negative ion components can be supplied in soluble form and differentially attracted to the target substrate, and because the formation of the ion pair may reduce toxicity of the individual components to the general environment. Accordingly, in one aspect, the invention is directed to a method to modify a target organism, cell or tissue, which method comprises supplying to the environment of said target the positive ion component and negative ion component which are members of an ion pair, wherein the ion pair is effective in modifying the target. The invention is also directed, in another aspect, to pharmaceutical compositions containing the ionic components of the ion pair. Brief Description of the Drawings Figure 1 shows the structures of positive and negative components of representative ion pair conjugates of the invention. Figure 2 shows the toxicity of rhodamine-123 cation, tetrachloroplatinate anion, and mixtures thereof. Figure 3 shows the therapeutic effect of dequalinium ion, tetrachloroplatinate ion, and mixtures thereof. Figure 4 shows the effect of the presence of tetrachloroplatinate ion on the uptake of dequalinium ion by leukemia cells. Modes of Carrying Out the Invention Complex members of an ion pair conjugate are often more tightly bound in aqueous media than simple salts of the positive and negative components. Advantage is taken of this phenomenon to permit self-assembly of these components in situ when a target condition is sought to be treated. Thus, the complex positive ion component can be administered as a simple chloride, sulfate, or acetate, the negative component as a potassium or sodium salt. In the milieu in which typical target cells, organisms or tissues reside, the more complex ions tightly associate to form an ion pair conjugate which is capable of a modifying effect. Furthermore, supplying the components as a mixture can reduce the toxicity of the individual materials. Figure 1 shows the structures of several complex ion components which are useful in the invention. Rhodamine-123 ion and dequalinium ion are useful as positive ion components; tetrachloroplatinate is useful as a negative component. Certain background information is available on these candidates. It is known that the salt of rhodamine-123 ion and tetrachloroplatinate causes DNA cleavage, although the individual ionic components do not do so (Herman et al. , Cancer Res (1988) 48:2335-2341) . It is also known that when tumor cells are treated with the salt, they take up the ionic components in the correct ratio for their ionic salt (Teicher, B.A., et al., Inorg Che (1986) 25_:3980-3983) . It is also known that the ion pair complex has significant activity against L1210 leukemia in mice, although the individual components do not (Teicher, B.A. , et al., Biochem Pharmacol (1986) 3_5:3365-3369; Weiss et al. , Proc Natl Acad Sci (1987) 84.:5444-5448; Macquet et al., JNCI (1983) 20:899-905) . The administration of the components of an ion pair in separate form has several advantages. First, the solubility of the individual components can be controlled by supplying the individual components as soluble salts with simple counterions. On the other hand, association of the two components in vivo, even outside the target environment, may reduce the toxicity of the individual components, as illustrated in Example 1 below, so that enhanced amounts of each component may be employed. The effect of the ion pair, therefore, can be focused on the target condition per se. Second, one or both components may be coupled to a target-specific ligand which permits the control of the assembly. The capacity of the ion pair eventually to home to the target may be adjusted by separately controlling the ligand coupling status of each component. Thus, the homing specificity of the components can be enhanced by coupling the component to a targeting ligand using general coupling methods known in the art, including direct coupling, but preferentially utilizing linker molecules which permit more controlled coupling reactions. Commercially available linkers, such as those from Pierce Chemical Company, Rockford, IL, may be used, for example. The targeting ligand can be, for example, an antibody or an immunoreactive fragment thereof, such as an Fab or Fab' fragment, a receptor ligand wherein the receptor is characteristic of the target cell, organism or tissue, or a substrate for active molecules present in high concentration in the microenvironment of the target. The targeting ligand can be coupled to either the positive ion or the negative ion component or to both. Coupling to both greatly enhances the selectivity of the conjugation. The formation of an ion pair conjugate is a unique application of the general approach to self assembly outlined in U.S. Patent 4,812,449, referenced above, which involved conjugates formed by covalent bonding. Ion pair formation permits additional flexibility by taking advantage of the relative strengths of ionic bonding between the components as administered and their counterions as compared to the desired ion pair. Thus, the in situ formation of the ion pair depends not only on the intrinsic affinity of the ion pair members for each other, but also on the affinity of the negative component, for example, for its positive counter ion and the affinity of the positive ion component for its counterion in the forms in which these components are administered. The following examples are intended to illustrate but not to limit the invention. Exa ple 1 Toxicity of the Components Various amounts of potassium tetrachloroplatinate and rhodamine-123 chloride were administered interperitoneally in a three-day protocol on days 1, 3 and 5 to mice and the percent survival 15 days after the last injection was recorded. A similar protocol used varying amounts of a 2:1 mixture of the rhodamine-123 chloride and potassium tetrachloro¬ platinate. The results of this experiment are shown in Figure 2. As seen, the survival rate went to 0 when only small amounts of the potassium tetrachloroplatinate and rhodamine-123 chloride were administered. However, 100% survival was obtained when 68 μmol/kg of rhodamine-123 chloride and 34 μmol/kg potassium tetrachloroplatinate were administered in two separate injections. Precipitation of the ions when mixed in vitro is known (Teicher, B.A., et al. Radiation Res (1987) 109:58-67) ; the association thus evidently occurs in vivo as well. Example 2 Enhancement of Solubility In a similar experiment, various amounts of dequalinium chloride, potassium tetrachloroplatinate, and mixtures thereof were injected to mice in a three-day protocol on days l, 3 and 5. The murine subjects had implanted L1210 tumors. Increase in lifespan was recorded as a function of dose. Figure 3 shows the results. As shown in Figure 3, neither dequalinium ion or tetrachloroplatinate ion alone had a significant effect on increase in lifespan. However, a 50% increase in lifespan was achieved at an optimal dose of a mixture of dequalinium chloride and potassium tetrachloro- platinate at a ratio of 1:4.75 w/w. The optimum concentration was between 3 and 4 mg/kg per injection of the dequalinium chloride component. Further, the presence of the chloroplatinate ion enhanced the uptake of dequalinium ion by L1210 leukemia cells. Aliquots of 10 7 L1210 cells were cultured in 5 ml RPMI supplemented with 10% fetal bovine serum in 1 mM glutamine. The cultured cells were exposed to dequalinium chloride or potassium tetrachloroplatinate or both for 3 hours at 37°C, 7% C0~. The cells were washed three times by centrifugation and resuspension, counted by hemocytometry, repelleted and sonicated in 3 ml 0.3 M HC1 and 50% aqueous ethanol. The delaquinium ion in the extracts was determined by fluorimetry. The results are shown in Figure 4. The addition of 80 μM tetrachloroplatinate ion markedly increased the dequalinium ion concentration in the cells from 1 fmol/cell when 15 μM dequalinium ion was supplied to over 2 fmol/cell.

Claims: CLAIMS 1. A method to modify a target organism, cell or tissue contained within an environment with a conjugate comprised of at least one positive ion and at least one negative ion and wherein said conjugate is bonded by ion pair bonding, which method comprises providing to said environment said positive ion component and said negative ion component wherein said ion pair bonding occurs in the microenvironment of the target organisms, cell or tissue. 2. The method of claim 1 wherein the positive ion component is supplied as a water-soluble salt. 3. The method of claim 1 wherein the negative ion component is a water-soluble salt. 4. The method of claim 1 wherein the positive ion component is dequalinium ion. 5. The method of claim 1 wherein the positive ion component is rhodamine-123 ion. 6. The method of claim 1 wherein the negative ion component is tetrachloroplatinate. 7. The method of claim 1 wherein at least one of said components is further coupled to a targeting ligand specific for said target organism, cell or tissue. 8. The method of claim 7 wherein said targeting ligand is a receptor ligand, an antibody, or an immunologically reactive fragment thereof. 9. The method of claim 7 wherein both positive and negative ion components are further coupled to a targeting ligand specific for said target organism, cell or tissue. 10. The method of claim 1 wherein the positive ion component and negative ion component are administered simultaneously. 11. A pharmaceutical composition useful in modifying a target condition in an environment, which composition comprises a positive ion component and a negative ion component capable of ion pair formation in the microenvironment of the target condition. 12. The composition of claim 11 wherein the positive ion component is supplied as a water-soluble salt. 13. The composition of claim 11 wherein the negative ion component is a water-soluble salt. 14. The composition of claim 11 wherein the positive ion component is dequalinium ion. 15. The composition of claim 11 wherein the positive ion component is rhodamine-123 ion. 16. The composition of claim 11 wherein the negative ion component is tetrachloroplatinate. 17. The composition of claim 11 wherein at least one of said components is further coupled to a targeting ligand specific for said target organism, cell or tissue. 18. The composition of claim 17 wherein said targeting ligand is a receptor ligand, an antibody, or an immunologically reactive fragment thereof. 19. The composition of claim 17 wherein both positive and negative ion components are further coupled to a targeting ligand specific for said target organism, cell or tissue.

Inventors: RIDEOUT DARRYL C, RIDEOUT, DARRYL, C.
Applicants: SCRIPPS RESEARCH INST, THE SCRIPPS RESEARCH INSTITUTE
Application year: 1991
Main classification: A61K31
IPC codes: A61K31, A61K33, A61K38, A61K39, A61K45, A61P31, A61P35