Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Reexamination Certificate
1998-09-17
2004-08-24
Navarro, Mark (Department: 1645)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
C435S007100, C435S007200, C435S007210, C435S007230, C435S021000, C435S028000, C435S069300, C435S070300, C435S410000, C436S512000, C436S513000, C436S516000, C436S517000, C436S518000
Reexamination Certificate
active
06780596
ABSTRACT:
FIELD OF THE INVENTION
The present invention is directed to a method for assessing the biological or pharmacological activity of a test material without the need to identify any of the constituents of that material, by exposing mammalian cells to the material and assessing whether structural changes, such as protein phosphorylation or protein-protein interaction, are induced in proteins present in the mammalian cells. While the present methods are useful for any test material, these methods have particular utility for testing materials which include complex mixtures of molecules where, for example, the physiological effects of the complex mixture may be a result of a synergy between two or more constituents present in the mixture. Thus, the present methods can be used to assess the biological activity of herbs, herbal extracts, plant extracts, animal extracts, natural or synthetic compounds, or combinations thereof. This method facilitates the formulation of natural products with consistent biological or pharmacological activities without the need to identify any of the chemical constituents responsible for the biological or pharmacological response.
BACKGROUND OF THE INVENTION
Modern medical and pharmaceutical sciences typically attempt to treat disease by prescription of a single, highly purified and well-characterized pharmaceutical compound whose activity has been carefully measured so that a precise dosage may be administered. The use of such highly purified drugs facilitates the manufacture of uniform dosage forms because drug concentration can simply and accurately be used to predict the appropriate pharmacological dosage.
However, use of a single highly purified drug is not always appropriate nor desirable for treating disease. In some instances, nutrients and/or pharmacologically active compounds may act together, in synergy. For example, research indicates that several constituents in saw palmetto extracts operate in synchrony to inhibit proliferation of cells in benign prostatic hyperplasia (BHP).
The most frequent reason men consult a urologist is because of an impairment in urinary flow. In men over 45 years of age, the cause of impaired urinary flow is often benign prostatic hyperplasia (BHP). The cause of BHP is an abnormal, but nonmalignant, proliferation of cells and tissues within the prostate gland. Eventually, urethral obstruction leads to urinary retention, kidney damage, and infection. In advanced stages, surgical resection is the treatment of choice.
To understand how medications may affect BPH, medical researchers seek the mechanism(s) that are believed to cause the condition. In the prostate, testosterone from the blood is converted by an enzyme to the more potent androgen, dihydrotestosterone (DHT). DHT increases the expression of proteins with resultant changes in cell metabolism and proliferation. In the process of normal growth, sex accessory organs are relatively insensitive to testosterone and DHT after puberty. However, in hyperplastic prostatic tissues the concentrations of DHT may be four to six times those of normal prostatic tissue. Thus, researchers infer that these high concentrations of DHT result in increased growth of the gland in mature males. Drugs have been developed to reduce the effects of androgens, for example, estrogens. However, while estrogens do reduce the effect of androgens they cause feminization, impotence, and cardiovascular toxicity in men—side effects which are highly undesirable.
In addition to androgenic stimulation, infiltration of the prostate by inflammatory cells is an etiologic factor in the development of BPH. These inflammatory cell types, such as polymorphonuclear neutrophils, produce chemotactic mediators and contribute to the development of the disease. Among the chemotactic factors generated by inflammatory cell types, derivatives of arachidonic acid have been extensively studied. Thus, medical research indicates that the best therapeutic regimen for BHP would address both androgenic and inflammatory mechanisms.
Several plants contain compounds with antiandrogenic and anti-inflammatory properties, for example, saw palmetto which consists of the partially dried, ripe fruit of
Serenoa repens
. Saw palmetto was recognized as a “drug” in the United States from 1906 to 1950 and was once widely used for a variety of ailments, particularly those of the urogenital tract, until losing popularity in the United States after World War II. European scientists continued to study saw palmetto and recognized that, in patients suffering form BPH, an extract of the fruit produced increased urinary flow, reduced residual urine, increased ease in commencing micturition, and decreased frequency of urination.
While extensive clinical and laboratory studies have been undertaken and reported, the mechanism of action of saw palmetto is poorly understood. Studies have shown that a liposterolic extract of the berries reduced cellular uptake of both testosterone and DHT by more than 40 percent. This mechanism is confirmed by the observation that saw palmetto extract does not induce changes in the level of testosterone, or other hormones, in the plasma. Other studies have indicated that an extract of saw palmetto reduces the conversion of less active testosterone to the more active DHT by inhibiting the enzyme 5&agr;-reductase.
In addition to their antiandrogenic properties, saw palmetto berries may also have anti-inflammatory activity. This may to be due to the inhibition of the cyclooxygenase and 5-lipoxygenase pathways, thereby preventing the biosynthesis of inflammation-producing prostaglandins and leukotrienes. Together, the antiandrogenic and anti-inflammatory effects seem to account for the beneficial role of the herb in treating BPH. Placebo-controlled, double-blind clinical studies carried out on more than 2,000 BPH patients in Germany have confirmed the effectiveness of a saw palmetto extract in such conditions.
A large number of possibly active ingredients have been isolated from saw palmetto including large amounts of beta-sitosterol-3-D-glucoside. Anthranilic acid, caffeic acid, chorogenic acid, tannin, sugars, and polysaccharides, are also present. Unfortunately, the active antiandrogenic principles remain unidentified, although they are known to reside in the acidic lipophilic fraction of the berries. The inability to identify a single, active compound indicates that a combination of ingredients may be responsible for the beneficial activities of saw palmetto. Alteration of the combination through purification of single ingredients results in a concomitant loss of the original biological activity.
However, the pharmaceutical industry generally relies upon purifying and quantifying an active ingredient in order to standardize preparation of medications.
The present invention provides a solution to this problem by providing methods for assessing the biological activity of a test material without the need for identifying and quantifying the active constituents of the test material. According to the present invention, cellular health and function can be assessed by observing the type and amount of certain proteins in the cell. Thus, the methods of the present invention expose mammalian cells to a test material and assess whether new proteins are synthesized by the cell or structural changes are induced in cellular proteins.
Cellular function is a direct result of both transcriptional and translational control processes. It has long been recognized that transcriptional control is necessary to differentiate one cell type from another. However, in addition to the variety of transcriptional controls developed by cells, cellular function is also dependent on the type and amount of post-translational modification of cellular proteins. One such post-translational modification is protein phosphorylation. Phosphorylation of serine, threonine and tyrosine residues on proteins is a fundamental post-translational regulatory process for mediating signal transduction, gene transcription, RNA splicing, cellular adhesion, apoptosis and c
Babish John G.
Lee M. Lisa
Pacioretty Linda M.
Ashni Naturaceuticals, Inc.
Hines J.
Kenyon & Kenyon
Navarro Mark
LandOfFree
Methods for determining the activity of complex mixtures does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Methods for determining the activity of complex mixtures, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Methods for determining the activity of complex mixtures will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-3358047