Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Cyclopentanohydrophenanthrene ring system doai
Reexamination Certificate
1999-06-17
2001-11-27
Lankford, Jr., Leon B. (Department: 1651)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Cyclopentanohydrophenanthrene ring system doai
C514S04400A, C514S051000, C514S170000, C514S557000, C514S581000, C514S851000
Reexamination Certificate
active
06323191
ABSTRACT:
The present invention relates to a novel method of treating cystic fibrosis. More particularly, the present invention relates to the use of artificial chloride channels or transporters as a therapeutic for cystic fibrosis. The invention also relates to a method of increasing cell membrane halide permeability.
Cellular lipid bilayers are highly impermeable to charged molecules. However, ionophores can increase the permeability of cell membranes to specific inorganic ions. For example, the antibiotic valinomycin complexes with potassium ions (K
+
) and readily passes through the cell membrane. In the absence of valinomycin, K
+
passes through the cell membrane very slowly. Ionophores, which may be small hydrophobic molecules that dissolve in lipid bilayers, enable ions to be transported across the cellular bilayer because they form lipid-soluble complexes with specific ions.
The two classes of ionophores, mobile ion carriers and channel formers, operate by shielding the charge of the transported ion enabling the charged molecule to penetrate the hydrophobic interior of the cellular wall. Valinomycin, a mobile ion carrier, picks up K
+
on one side of the membrane, diffuses across the bilayer, and releases K
+
, on the other side. Gramicidin A, a channel-forming ionophore, forms a transmembrane channel across the bilayer, which selectively allows ions to flow through the channel and across the bilayer.
Many ionophores are useful as antibiotics because in addition to transporting ions, the ionophore disrupts the cell membrane which leads to leakage of vital cellular constituents and cell destruction. There has recently been a great deal of interest in the design and synthesis of ionophores and other membrane disrupting compounds in the search for novel antibiotic agents. A well studied example is Amphotericin B (Amp B) (Nagawa et al.,
J. Am. Chem. Soc.,
113, pp. 7237-7240 (1991)). Amp B is an ionophore that generates transmembrane pores. These pores, which allow leakage of vital cellular constituents and trigger cell destruction, make Amp B an effective antibiotic. Subsequently, the structural elements of Amp B have become a starting point for designing compounds with similar functional characteristics. Many of these compounds, such as 5-Androstene-3B,17B-bis[(oxycarbonyl) hexaethylene Glycol], also possess interesting ionophoric characteristics (Stadler, et al,
J. Am. Chem. Soc.,
116, pp. 6677-6682 (1994)).
The antibiotic squalamine is a novel sterol-spermidine conjugate that has recently been isolated from tissues of the dogfish shark, Squalus acanthias (Moore et al.,
Proc. Natl. Acad. Sci. USA,
90, pp. 1354-1358 (1993)). This steroid, which is an adduct between spermidine and an anionic bile salt intermediate, has demonstrated potent antibacterial activity against both gram-negative and gram-positive bacteria. Unfortunately, squalamine is only found in limited quantities in nature.
In the search for compounds functionally equivalent to Amp B, several mimics of squalamine have been synthesized (Sadownik, et al.,
J. Am. Chem. Soc.,
117, pp. 6138-6139 (1995)). The sterol-spermine conjugates that have been made are both structurally similar to squalamine and demonstrate extraordinary antibiotic properties. The compounds' ability to exhibit potent activity against a broad spectrum of microorganisms are of particular interest. European patent application nos. WO 9638464, WO 9632404, and WO 9004401 describe the utility of this class of compounds as an antibiotic, the disclosures of which are hereby incorporated by reference.
In addition to their broad antibiotic activity, the synthetic mimics of squalamine possess the unique ionophoric activity of cell membrane and transport anion selectivity (Deng, et al.,
J. Am. Chem. Soc.,
118, pp.8975-8976 (1996)). The transport of ions across negatively charged bilayers is favored over transport across neutral ones; and no Na
+
transport activity is observed while effective Cl
−
transport activity is observed.
Cystic fibrosis (CF) is a complex disease affecting many organs with epithelial cell linings. The lethal genetic disorder, caused by the presence of mutations in the gene that encodes for a protein known as cystic fibrosis transmembrane conductance regulator (CFTR), affects the permeability of the epithelial cell linings to Cl
−
ions. (Welsh et al.,
Neuron,
8, 821-829 (1992)). CFTR regulates the passage of Cl
−
ions through the cell membrane epithelial cells. (Quinton,
FASEB J.,
4, 2709-2717 (1990); Jiang et al.,
Science,
262, 424-427 (1993); Smith et al.,
J. Clini. Invest.,
91, 1590-1597 (1994)). Through the regulation of ions across the cell membrane of epithelial cells, CFTR regulates the flow of fluid. In CF, the mutations of the CFTR gene cause defective transepithelial Cl
−
transport and therefore defective fluid transport.
The genetic mutations causing abnormal ion transport lead to abnormal mucous secretion, inflammation, infection and tissue damage. It is believed that CFTR regulates active ion transport-mediated fluid transport in a variety of epithelial cells including sweat glands, pancreas, intestine, genital tract, and airways. In airway epithelia, for example, it is believed that defective electrolyte and fluid transport causes impairment of airway clearance and defective bactericidal activity of salt-sensitive defensins, which subsequently results in recurrent infections and destruction of lungs in CF patients. (Jiang et al.,
Science,
262, 424-427 (1993); Smith et al.,
Cell,
85, 229-236 (1996); Goldman et al.,
Cell,
88, 553-560 (1997)). Patients suffering from CF are prone to recurrent lung infections and airway blockage, small bowel obstruction, pancreatic insufficiency, cirrhosis of the liver due to biliary tract obstruction, infertility in males, and eventually death.
Several therapeutic approaches are being developed concurrently for the treatment of CF. These include 1) use of agents that improve the anti-bacterial activity and viscosity of the mucous fluid lining the airways (Smith et al.,
Cell,
85, 229-236 (1996); Goldman et al.,
Cell,
88, 553-560 (1997)), 2) use of agents that activate alternative Cl
−
channels to compensate the CFTR Cl
−
channel defect, 3) protein and gene augmentation therapy. (Welsh et al.,
Cell
73, 1251-11254, 1993.), and 4) use of agents that reverse the mutant phenotype. There is currently no effective treatment for the disease.
Accordingly, the present invention is directed to the novel use of ionophores as artificial Cl
−
transport pathways in CF epithelia to treat the defective Cl
−
and fluid transport. Ionophores and in particular non-peptide ionophores, and in particular, small molecule ionophores, represent a potential novel means of treating CF. Preferred small molecule ionophores have a molecular weight less than or equal to 2000, or preferably less than or equal to 1750, or more preferably less than or equal to 1500, or even more preferably, less than or equal to 1000. To achieve these and other advantages and in accordance with the purpose of the invention, as embodied and broadly described, the invention treats CF by administering an effective amount of an ionophore to a patient.
In another aspect, the invention includes using an ionophore to generate chloride secretion on intact monolayers of airway epithelia cells and other epithelia cells by administering an ionophore to a mammal. Defective Cl
−
secretion in airway epithelia in vitro and in vivo can be corrected.
In a further aspect, the invention includes using an ionophore to increase cell membrane halide and anion permeability of epithelia cells by administering an ionophore to a mammal.
The present invention also relates to novel ionophores that may be useful as artificial Cl− transport pathways; that may generate chloride secretion on intact monolayers of epithelia cells; and may increase cell membrane halide permeability.
The invention also provides for pharmaceutical compositions of io
Cheng Seng H.
Harris David J.
Jiang Canwen
Lane Mathieu
Lee Edward R.
Finnegan Henderson Farabow Garrett & Dunner L.L.P.
Flood Michele C.
Genzyme Corporation
Lankford , Jr. Leon B.
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