Cystic Fibrosis (CF) is the most prevalent autosomal recessive disorder in the Caucasian population (Gorelick (1991) Gastroenterology 103:681-693). Approximately 1 in 2000 live births are afflicted with CF and 5% of Caucasians in the United States are carriers of the abnormal CF gene. CF individuals rarely survive past their mid-thirties, and most mortalities are a result of recurrent pulmonary infection and, ultimately, pulmonary failure. Two other major clinical manifestations of CF are pancreatic dysfunction and male infertility.
By 1989, the CF gene had been cloned and was found to code for a chloride channel. Activation of the channel in the normal pancreas activates the chloride/bicarbonate exchanger, resulting in a net secretion of bicarbonate into the lumenal space and alkalinization of the pancreatic juice. Mutations in the chloride channel like those found in CF result in a reduced chloride conductance and a reduced ability of ductal cells to secrete bicarbonate into the lumenal space. This results in the formation of inspissated plugs within the ducts leading to obstruction of the pancreatic ducts.
In recent years, the focus in CF research has shifted towards the coupling of defective chloride channel function and membrane recycling. Recent research has demonstrated that membrane internalization at the apical plasma membrane of the pancreatic acinar cell is dependent on pH of the acinar lumen (Freedman et al., Eur. J. Cell Biol. (1998) 75:153-63), Freedman et al., (1994) Am. J. Physiol. 267:G40-G51, and Freedman et al., (1994) Eur. J. Cell Biol. 65:354-365). Since pH of the acinar lumen is reflective of ductal bicarbonate secretion from the proximal duct cells, a phenomenon regulated via the chloride channel, a coupling may exist between duct and acinar cell function, (Freedman et at., (1994) Am. J. Physiol. 267:G40-G51, and Freedman et al, (1994) Eur. J. Cell Biol. 65:354-365). Research has also confirmed the hypothesis that lack of alkalinization of the acinar lumen leads to inhibition of apical membrane internalization and defective apical endocytosis in pancreatic acinar cells from CF mice. This block in the recycling of membranes following exocytosis leads to eventual deficiency in membranes for reformation of secretory granules. Thus, pancreatic insufficiency appears to be a result of defects in membrane recycling with obstruction of the ducts occurring as a secondary event.
Although approximately 90% of CF mortalities are the result of pulmonary infection and failure, the mechanism by which CF produces lung disease remains unknown. Abnormal and viscous mucus secretions from the pulmonary epithelia often lead to lung infections. Hydration of the secretions appears to be one underlying factor. Another factor may be the inability of CF individuals to produce dipalmitoyl phosphatidylcholine, the major component of fetal-lung surfactant, which is responsible for decreasing surface tension at the alveolar air-liquid inferface. In addition, other organs, such as intestine, biliary tract and testis, become impaired due to viscous secretions. In the small intestine, this results in villous hypertrophy which compensates for decreased absorption.
Despite recent advances in our understanding of the pathogenesis of cystic fibrosis, there is no effective treatment for this life-threatening condition. As premature death is still the norm, there is a pressing need to develop novel forms of therapy to treat this disease. Although after the discovery of the gene mutation in cystic fibrosis there was great hope that gene therapy could be used as a potential treatment for this disease, these expectations were soon dismissed mainly due to (i) the inefficiency of gene transfer to epithelial cells; and (ii) the low stability over time of the transfected gene. In addition, it is not clear whether or not insertion of normal copies of the cystic fibrosis transmembrane regulator (CFTR) gene will reverse the clinical manifestations of the disease.
CF gene mutations lead to three important clinical events: 1) increased mucus secretion; 2) increased inflammatory response; and 3) decreased immune function. Previous reports have indicated that arachidonic acid (AA) levels are increased in CF patients, i.e., an increase in AA has been demonstrated in lavage fluid from the lungs of CF patients. However, this was thought to be secondary to infection and not a primary process. Our research demonstrates that there is an increase in archidonic acid (AA) while levels of docosahexaenoic acid (DHA) are significantly decreased. Thus, the ratio of DHA to AA is affected. It appears that the ratio of docosahexaenoic acid (DHA) to AA is affected. Similar findings are made in chronic inflammatory diseases.
Another condition associated with abrogated levels of DHA is respiratory distress syndrome (RDS) in premature born babies. Every year, in the United States alone, 400,000 babies are born prematurely from an estimated total of four million pregnancies. Respiratory distress syndrome due to surfactant deficiency is one of the main complications in the premature newborn. Of those babies who survive, some remain compromised with lifelong disabilities including mental retardation, cerebral palsy, deafness, and blindness.
The introduction in 1971 of the lecithin/sphingomyelin ratio in amniotic fluid as a test to predict the risk of RDS provided the obstetrician with a valuable diagnostic tool to improve the management of pregnancies at risk for premature delivery (Gluck et al., (1971) Am. J. Obstct Gynecol. 109:440-445). Thus, in situations where the fetal lungs are immature, the obstetrician can delay the delivery to allow the lungs to mature. However, despite the significant improvement in the accuracy of tests to predict the risk of RDS, knowledge of the maturity status of the fetal lung does not significantly change the management of these premature babies in situations where preterm deliver is imminent (e.g., preterm labor). Although corticosteriod therapy has been used to induce fetal lung maturation in pregnancies at risk for premature delivery, its use remains highly controversial and its efficacy very limited (Crowley et al., (1995) Am. J. Obstct. Gynecol. 173:322-335). The recent introduction of exogenous surfactant as a routine therapy to treat babies born prematurely has been a major development in efforts to decrease the severity of RDS (Cooke et al., (1995) J. Obstct. Gynecol. 102:679-681). However, despite the use of surfactant therapy, RDS continues to be one of the main causes of morbidity and mortality in the premature newborn. DHA treatment of the mother or newborn to prevent or treat RDS has not been used.
Thus, a need still exists to develop novel methods for treating disorders, such as cystic fibrosis and respiratory distress syndrome due to surfactant deficiency, in which DHA levels are affected.
Accordingly, an object of the invention is to provide a method for treating a disorder in which the serum, tissue or membrane levels and/or ratios of DHA and AA are affected.
Another object of the invention is to provide a method of treating cystic fibrosis in a subject by administering to the subject a therapeutically effective amount of an omega 3 fatty acid comprising 22-24 carbon atoms and 5 or more double bonds.
A still other object of the invention is to provide a method of treating a chronic inflammatory disease, e.g., ulcerative colitis, Crohn""s disease, chronic pancreatitis, asthma, rheumatoid arthritis or chronic gastritis.
A further object of the invention is to provide a method for treating hypertrophy of small intestine in a subject suffering from a disorder in which DHA levels are affected.
An additional object of the invention is to provide a method of restoring normal morphology to a cell or tissue affected with a disorder in which DHA levels are affected.
A still further object of the invention is to provide a method for diagnosing in a subject a disorder in which DHA levels are affected.
Another object of the invention is to provide a method of ameliorating affects of cystic fibrosis in a newborn.
A still additional object of the invention is to provide a method for increasing surfactant levels in a fetus.
These and other objects and features of the invention will be apparent from the following description and from the claims.
The invention features a novel method of treating a subject suffering from a disorder in which DHA levels are affected. The method includes administering to the subject a therapeutically effective amount of an omega 3 fatty acid comprising 22-24 carbon atoms and 5 or more double bonds, (e.g., DHA and DHA precursors, e.g., docosapentaenoic acid, tetracosapentaenoic acid and tetracosahexaenoic acid). The dosage may be of the fatty acid itself or in a form more easily incorporated into the tissue such as a derivative or a structured lipid. This method is particularly useful in treating subjects suffering from a disorder characterized by a defect in the CF gene, e.g., cystic fibrosis; or a chronic inflammatory disorder, e.g., ulcerative colitis, Crohn""s disease, chronic pancreatitis, asthma, rheumatoid arthritis or chronic gastritis. The method is also useful in treating a disorder characterized by reduced levels of fetal surfactant, e.g., by abrogation of fetal surfactant biosynthesis.
Proper levels of treatment can be determined in a variety of ways, e.g., measurement of the DHA levels in plasma which are raised to at least about 170 xcexcg/ml. Preferably, the DHA levels are raised to the range of about 200 xcexcg/ml to about 500 xcexcg/ml.
Omega 3 fatty acids comprising 22-24 carbon atoms and 5 or more double bonds may be administered in different modes, e.g., as a part of a nutritional additive or total formulation. If a total formulation is used, it can be as a part of an enteral nutrition solution, an emulsion or as a solid formulation. The therapeutically effective amount of an omega 3 fatty acid comprising 22-24 carbon atoms and 5 or more double bonds, e.g., DHA, is about 0.3-5% of the total caloric intake, preferably 0.5-3% of total caloric intake. The therapeutically effective amount of DHA should produce a pancreatic AA/DHA ratio between 0.2-1.5, a lung AA/DHA ratio between 0.1-1.6 and a blood AA/DHA ratio between 0.1-1.6. As noted, a preferred method of administration is as a structured lipid having at least one omega 3 fatty acid comprising 22-24 carbon atoms and 5 or more double bonds, most preferably with the DHA located at R2 position of the structured lipid. Preferred structured lipids also include medium chain or short chain fatty acids. In some embodiments, the structured lipid has DHA (or the 22-24 carbon, 5 or more double bond omega 3 fatty acid) at two or even all three positions.
The invention also features a method of increasing the level of DHA in one or more tissues of a subject having a disorder in which DHA levels are affected. The method includes administering to the subject a therapeutic composition comprising an omega 3 fatty acid comprising 22-24 carbon atoms and 5 or more double bonds in an amount effective to raise the levels of DHA in the subject""s tissue. Preferably, the therapeutic composition comprises an omega 3 fatty acid comprising 22-24 carbon atoms and 5 double bonds in the range of about 0.5-3% of the total caloric intake. Preferably, the omega 3 fatty acid comprising 22-24 carbon atoms and 5 double bonds is DHA or a structed lipid containing DHA.
Preferably, the DHA levels are increased in affected tissue, e.g., plasma, pancreatic or lung tissue, most preferably, the DHA levels are increased in plasma to at least about 170 xcexcg/ml.
In another preferred embodiment, the disease in which DHA levels are affected is selected for the group consisting of a disorder characterized by an defect in the CF gene, e.g., cysitc fibrosis, a chronic inflammatory disorder e.g., ulcerative colitis, Crohn""s disease, chronic pancreatitis, asthma, rheumatoid arthritis, chronic gastritis, or lowered fetal surfactant levels.
The invention also pertains to a method of ameliorating the effects of cystic fibrosis in a newborn. The method includes administering a therapeutic composition comprising an omega 3 fatty acid comprising 22-24 carbon atoms and 5 or more double bonds to a mother during pregnancy such that the effects of cystic fibrosis in the newborn are ameliorated.
The invention further pertains to a method of increasing surfactant levels in a fetus. The method includes administering a therapeutic composition comprising an omega 3 fatty acid comprising 22-24 carbon atoms and 5 or more double bonds to a pregnant woman who may or may not be at risk for premature delivery such that the surfactant levels in the fetus are increased.
In another aspect, the invention features a method of treating hypertrophy of small intestine in a subject suffering from a disorder in which DHA levels are affected. The method includes administering to the subject a therapeutic composition comprising an omega 3 fatty acid comprising 22-24 carbon atoms and 5 or more double bonds in an amount effective to decrease hypertrophy of small intestine by at least about 25 percent.
In yet another aspect, the invention features a method of restoring normal morphology of a cell or tissue which exhibits a disease morphology associated with disorder in which DHA levels are affected. The method includes contacting a cell, e.g., a pancreatic acinar cell or a lung cell, or a tissue, e.g., in vitro or in vivo, with a therapeutically effective amount of an omega 3 fatty acid comprising 22-24 carbon atoms and 5 or more double bonds such that the normal morphology of the cell or tissue is restored.
In still another aspect, the invention also features a method of diagnosing a disorder in which DHA levels are affected. The method includes: a) obtaining a tissue biopsy from a subject; b) determining the ratio of AA to DHA in the tissue biopsy; and c) comparing the ratio in the tissue biopsy to a standard AA to DHA ratio. If the ratio in the tissue biopsy is at least about 20% higher than the standard AA to DHA ratio, this indicates that the subject is suffering from the disorder in which DHA levels are affected.
The following description and non-limiting examples further elucidate the invention.