| Title of Invention | "RADIOLABELED FORM OF DRUG COMPOSITION" |
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| Abstract | Radiolabelled drug formulations particularly, radiolabelled salbutamol, salmetrol or terbutaline (or their salts) preparations, that upon mixing with different excipients can be inhaled or administered orally, trans-nasally, transdermally, and used as an effective diagnostic substance. |
| Full Text | FIELD OF THE INVENTION: The present invention relates to preparation and development of radiolabeled form of drugs acting on beta-adrenergic receptor, particularly beta-2 receptor agonists, exemplified by salbutamol sulfate. The uses of this class of radiopharmaceutical include in-vivo diagnostic imaging, including biodistribution and biokinetics studies, and invitro or invivo analytical studies of these drugs, particularly receptor-based studies and assays but without implying any limitation thereof, in all conditions and tests where these preparations can be used with benefit and advantage. BACKGROUND ART Several drugs like salbutamol sulphate interact with beta-2 adrenergic receptors and stimulate these. These specific receptors are present over the smooth muscle cells in the body. This results in a cascade or series of cellular biochemical reactions, ultimately resulting in relaxation in the tone of smooth muscle. Though the action is predominantly limited to stimulation of Beta-2 receptors, there is some agonist action on Beta-1 receptors present on the cardiac muscle also. Other drugs having similar structure and function include salmetrol and terbutalin. These drugs have several pharmacological uses. Due to their Beta-1 action, the drug finds clinical application in asthma, chronic obstructive airway diseases and related disorders, and on uterus in certain conditions of threatened abortion or impending premature delivery. These drugs do not have any diagnostic or testing uses at present, whether in-vitro or in-vivo. Beta-2 agonists, particularly Salbutamol Sulfate, are available commercially as tablets, respirable fluids (through nebulization), respirable powder (through rota-cap technology, upon inhalation) or as injections. Several new formulations or combinations are being added to the commercial market regularly for which many invitro and in-vivo test are needed for regulatory and scientific purposes including quantitative biokinetic and pharmacokinetic information. It is not possible to obtain this data directly in humans. This is usually done through chemical, optical, or other means in in-vitro samples, followed by animal studies where a big number of animals need to be sacrificed at various time intervals to collect their organs for analysis for obtaining biokinetic data after the drug/drug formulation administration, and estimating the concentration of the drug in various target organs. Using the same principles, dissolution rates of formulation and sites of dissolution are estimated by estimating serial drug concentration in blood of animals short-term and long-term toxicity is studied in animals by histopathological changes in the target organs and biochemistry alternations in blood and tissues. This animal data is then extrapolated for humans as a poor substitute because many times the animal data does not mimic human data and sometimes may even be misleading. Only after compilation of the relevant biokinetic and toxicity data in animals, human phase 1-3 clinical studies are done. A very significant amount of time and funds are spent (besides loss of animals) before efficacy of the drug/formulation can even be evaluated in humans. Thus, a number of different animal studies are to be performed in animals with new formulations at a huge cost in terms of time and money to generate information that is at best indirect and extrapolated for humans. However, the method continues to be followed diligently for each new molecule/formulation for want of a non-invasive safe method that may be able to provide this information directly in humans. A new field of pharmacoscintigraphy (Nuclear Medicine) has been established that has a significant bearing on the above algorithm or drug development protocols. Nuclear medicine or scintigraphy is the field of science where a radioactive molecule (radionuclide) is attached (usually chemically) to a chemical or a pharmaceutical that follows a particular biokinetic pathway in the body, or localizes in a particular organ/organ system of the body. The combined molecule/complex is called a radiopharmaceutical whose biokinetics and uptake in various organs can be tracked from outside using Gamma-Camera or PET camera in real time. The radionuclide attached is chosen in a manner that gives least radiation to the person injected while maximizing the information. Nuclear medicine is used routinely in scanning of various organs/organ systems, giving important medical diagnostic information through bone scan, heart scan, renal scan etc. The same principle can be, and has been, used with advantage in drug evaluation. The equivalence of the original molecule and the radiolabeled molecule can be established in many cases by objective means. Once it is done, the information acquired from scintigraphy of the radiolabelled molecule is used to derive data/information pertaining to the original molecule / formulation in pre-clinical, animal and clinical studies. Though the radiopharmaceutical is usually injected intravenously, it can also be administered intramuscularly, inhalation, transdermally, orally, or through any other desired route to give information on the respective route of administration. The only absolute requirement of the test is that the radiolabelled product should behave predominantly like the parent molecule physically and chemically at least during the period of study. Apart from the biokinetics of the molecules, scintigraphy with the radiolabelled drug gives data relevant to the formulation containing the drug. Dissolution rate, absorption rate, bioavailability and excretion profile of the formulation can be estimated using radiolabelled drug. In many situations particularly, oral, transnasal and inhalation formulations. Data generated by nuclear medicine is considered the gold standard with respect to real-time invivo studies in human subjects and is considered an important test in evaluation a drug/formulation whose radiolabelled form exists or can be prepared. Besides human use, radiolabelled drug can have many analytic invitro uses like dissolution rate of formulation in test-tube, and filtration rate through dialyzing membrane or skin samples. Counting radioactivity makes the analytic procedures very fast, objective and cost effective. With regards to beta-2 adrenergic receptors, particularly Salbutamol Sulphate, no radiolabelled equivalent exists, particularly with Tc-99m. Therefore, no radiolabelled method exists for evaluating or studying drug/drug formulations containing Salbutamol Sulphate either invitro or invivo. Similarly there is no scintigraphy method to image the physiology or pathology related to B-2 receptors invivo or in-vitro. Also, there are no commercial RIA or IRMA or radioceptor assays to study or estimate Beta-2 receptors. Present methods of Salbutamol Sulphate assay and its Deficiencies The present bioassay methods for Salbutamol Sulphate include infrared and visible band spectroscopy, ultra-violet spectroscopy, chemical assays and chromatography and HPLC. The infrared spectrum of Salbutamol base gives major bands as follows: 3320, 3200, 3160, 1610, 1370, 1270, 1190 cm"1. Other fingerprint bands characteristics to Salbutamol (determined in KBrdisc) are: 1038, 1075, 1263, 1228 and 1333 cm'1 The UV light absorbance of salbutamol exhibits a maximum only at about 276nm. The chemical colorimetric methods include a) aminophenazone - potassium ferricyanide method that gives orange-red colour in the chloroform layer, b) FeCI3 test that gives a reddish-orange colour which does not change on the addition of sodium hydrogen carbonate solution, c) and other colour tests include the Clarke's test. The chromatographic methods include Paper chromatography (Clark's method). Thin layer chromatography methods (Clarke et al, 1975), (Evans et al, 1973) & HPLC. The deficiencies associated with one, more or all of the above methods include: a) All methods as described are unable to give visual information, or imaging of the drug distribution. b) All methods as described have comparatively low sensitivity. c) None of the above methods can assess body organ distribution except through blood concentration estimation. d) None of the above methods can give real-time dynamic information in human volunteers and subjects. e) None of the above methods can tract movement of the drug molecules in-vivo from the administered drug formulation, whether oral, inhalation, transdermal, intravenous or any other route. f) None of the above methods can be used to identify Beta-2 receptors invitro or invivo. Considering the widespread medical use of Beta-2 adrenergic receptor agonists and their formulations (oral, injectable of inhalation products) particularly of salbutamol sulphate, it was realized that radiolabeled drugs, particularly salbutamol sulphate, will have important diagnostic and analytical uses in nuclear medicine as a radiopharmaceutical as well as in generating invitro parameteric information about salbutamol. Considering the specific interaction of salbutamol, particularly its salt salbutamol sulphate with Beta-2 adrenergic receptor, it was realized that radiolabeled salbutamol will become an important tool to the study the physiology, pathology and in concentration and quantification of these receptors both invivo and invitro. In view of the severe limitations of the current testing methods of salbutamol as exposed above, and the prospective uses of the radiolabeled salbutamol, the present invention therefore relates to the development of a new molecule/radiopharmaceutical using a new process that may be very effective in scintigraphy, receptor imaging and evaluation or testing of drug/drug formulations containing salbutamol, particularly salbutamol sulfate. OBJECTS OF THE INVENTION The most important object of the present invention is to develop a new class of radiopharmaceuticals (radiolabelled Beta-2 agonists) that can be used as a scintigraphy method, receptor imaging method and for testing or evaluating drug/drug formulations containing such drugs using Gamma Spectrometry methods. Another important object of the present invention is to make a novel radiopharmaceuticals whose biodistribution correlates with that of the parent compound upon intravenous administration and whose kinetics equate or correlate with that of the parent drug in vitro work. It is yet another object of the present invention to have radiolabelled drug formulations particularly radiolabelled salbutamol, salmetrol or terbutaline (or their salts) preparations, that upon mixing with different excipients shall result in making various formulations that can be inhaled or administered orally, trans-nasally, transdermally, or through any other route with properties same or similar to non-radiolabelled salbutamol or its salts. Still another object of the present invention is to have radiolabelled Beta-2 adrenergic receptor agonist drugs (or their salts) that can be used in place of non-radiolabelled drugs (or their salts) for the purpose of invitro evaluation of the molecule or its formulations, including receptor-based studies or assays. ADVANTAGES OF THE PRESENT INVENTION: The presently disclosed invention is a radiopharmaceutical class (radiolabelled Beta-2 adrenergic receptor agonist drugs) that constitutes a new method of diagnosis using scintigraphic of Gamma Spectroscopic techniques. The presently disclosed invention constitutes a new method of performing receptor binding assays using Gamma Spectroscopy and scintigraphy imaging with respect to Beta-2 adrenergic receptors. The presently disclosed invention constitutes a new method of performing receptor binding assays using Gamma Spectroscopy and scintigraphy imaging with respect to Beta-2 adrenergic receptors. Still further an advantage is its utility in in-vivo real time quantitative study of new drug formulations containing radiolabelled Beta-2 adrenergic receptor agonist drugs or its salts or derivatives through any route. The methodology is better than the current means of evaluation and also makes it possible to study blood biokinetics quantitatively. Yet further an advantage of the product is in vitro evaluation of drug formulations. It may be more sensitive than the current means of evaluation. Still further an advantage of using the product is increase in accuracy, reduction in time of study and cost, and ease of analysis. Further objects and advantages of the present invention will be more apparent from the following description, which has been explained with the help of a particular preparation method, which is not intended to limit the scope of the present invention. SUMMARY OF THE INVENTION Accordingly, towards achieving the stated objectives and overcoming the disadvantages of the prior art a radiolabeled form of drug composition comprising of a. drug solution in the ratio of .02-30%. b. a reducing agent in the ratio of .0002-20%. c. a stabilizer in the ratio of 0-1.8% d. phosphate buffers in the ratio of 0-8.3% e. required amount of radioactive material and f. the rest of the 100% composition being water or 0.1 N HCI or organic solvents or their combination. is herein described. In another embodiment, a process of making radiolabeled form of drug composition comprising the steps of: a) mixing drug solution in the ratio of .02-30% with a reducing agent in the ratio of .0002-20%, phosphate buffers in the ratio of 0-8.3% and water or 0.1 N HCI or organic solvents or their combination in a controlled environment of pH 2-9 and temperature of at least 20° C; b) incubating the solution with a stabilizer in the ratio of 0-1.8% for atleast 1 minute; c) mixing required amount of radioactive material with the solution of (a) and (b); d) incubating the solution for at least 3 minutes. is also herein described In yet another embodiment, a radiopharmaceutical kit comprising of a. Salbutamol Sulphate in the ratio of 0.02-1% b. Stannous Tartarate or Chloride in the ratio of 0.0002-0.2% c. Ascorbic acid uptol .8% d. Gentisic acid upto 2% e. Phosphate buffers upto 8.3% f. 0.1N HCL or water or their combination being the rest of the composition to be mixed with 10OMBq of Na99mTc04 at the time of use is herein described. DESCRIPTION OF THE INVENTION: According to this invention, radiolabeled Beta-2 adrenergic receptor agonist drugs including salbutamol, or its derivative, or their salts are prepared by the process comprising of the following steps: Drug solution, preferably in aqueous medium is mixed with a small quantity of a reducing agent, preferably a stannous salt, particularly its chloride or tartrate in a controlled environment of pH (2.5-8.5) and temperature (preferably more than 20 C). The molar ratio is kept at 1:1.1 or more, preferably more than 1:20.0. It is incubated, preferably in presence of a stabilizer like ascorbic or gentisic acid, for more than one minute. If storage is not required, the require amount of radioactivity (preferably Tc-99m pertechnetate with high specific activity with molar ratio 2:1 or more) is mixed to the solution and further incubated for 5 minute or more before use. If storage is required, the solution is freeze dried before adding radioactivity, or kept continuously at subzero centigrade temperature continuously, and radioactivity solution is mixed just before use and injected/ administered after incubation period (preferably more than one minute). After incubation, sufficient radiolabeling occurs with Tc-99m to give radiolabeled form of the drug for various invitro and invivo uses mentioned in the above sections but not implying any limitation thereof. Embedded within the preparation methodology are a number of quality control measures including thin layer paper chromatography to check and confirm high labelling efficiency (should be more than 85% always and more than 95% routinely), sustenance of labelling invitro and invivo and the quantitative estimation of other radiochemical species. For human use particularly, all the ingredients should be of chemical or medical grade, laboratory-ware should be sterile and Good Laboratory Practice principles should be adhered to. Radiochemical, chemical and other impurities should be below the prescribed levels. Sterility & pyrogencity-free certification needs to be obtained before human/animal diagnostic use. The essential instrumentation needed for invivo imaging use (of humans or animals) is a Gamma Camera or a SPECT system and Gamma Spectrometer (or an equivalent instrument to count radioactivity in samples) for invitro use. When radiolabeling is done with Tc-99m, the preferred embodiment of the invention, the spectrophotometer peak is centered at 140 key or their about. In accordance to the preferred embodiment of the present invention, the sequence of predetermined steps and methodology of mixing may be changed in a manner not detrimental to the stability and efficacy of the medical product. The working of the present invention will be more apparent from the following experiments, which are not intended to limit the scope of the present invention. EXAMPLE 1 0.1N hydrochloric acid was used as a solvent for 100 microgram/ml solution of SBM. 1ml of this solution was added to 180 microgram of stannous tartarate. 20 millicurie of freshly eluted Na99mTc04 was added and incubated for 15 minutes. Radiochemical purity of the radio-complex was determined using ITLC that was 98% after 1 hr of making the preparation and 97% after 45 hours of making the preparation. 10 mCi of the preparation was injected intravenously into a human being slowly through an antecubital vein after estimating the radioactivity count rate. Whole body and spot view (scintigraphic imaging) were taken at various intervals of time till 24 hours on a Gamma Camera (Fig. 1). Organ-wise distribution was estimated using computer software to provide drug distribution profile inside the human body at various time intervals and data compared with the known behavior of the salt. It was found that the radiolabeled drug was excreted through the reneal and hepatobiliary route, did not cross blood brain barrier and was preferentially showing uptake in lung parenchyma which have a high density of beta-2 adrenergic receptors. The biodistribution profile and invivo behavior of the radiolabeled drug was similar to that known with non-labeled salbutamol. EXAMPLE 2 0.1N hydrochloric acid was used as a solvent for 10 microgram/ml solution of SBM. 1ml of this solution was added to 20 microgram of stannous tartarate. 1.5 millicurie of freshly eluted Na99mTc04 was added and incubated for 20 minutes. Radiochemical purity of the radio-complex was determined using ITLC was 97% after 1 hr of making the preparation and 97% after 4 hours and 92% after 24 hrs of making the preparation. 1 mCi or less of the preparation was injected intravenously into groups of mice and rabbits for estimating biodistribution and biodistribution of the radiolabeled molecule. Mice were sacrificed at different time intervals and their organs cleared and pulverised Radioactivity content in different organs was found using Gamma Spectrophotometer peaked at 140 Kev. Blood samples of group of rabbits was taken at different time intervals to estimate protein binding, blood clearance half-life, volume of distribution and excretion rates after counting radioactivity in blood samples on a Gamma Spectrophotometer. Whole body and spot view (scintigraphic imaging) were taken at various intervals of time till 24 hours on a Gamma Camera. Organ-wise distribution and other parameters obtained as discussed above were compared with the known behavior of the salt. It was found that the data were compatible signified similar behavior of the radiolabeled salt with respect to non-radiolabeled salt. EXAMPLE 3 Radiolabeled drug was manufactured as disclosed above. 500 microCurie of the radiolabeled drug was mixed thoroughly with non-labeled drug solution containing 1 mg each of the active component and formulated into 1% salbutamol sulphate skin ointment and 1 mg salbutamol slow release (SR) tablets for oral use. After appropriate application procedures on humans (a) Salbutamol ointment rubbed over dorsal surface on hand and b) oral administration of the radioactive SR tablet), sequential Gamma Camera imaging was carried out. Aim was to a) determine rate of transdermal transport of the drug ointment and b) exact location of the tablet in Gastrointestinal tract of the human volunteer & rate of dissolution of the tablet and its blood absorption kinetics. The images and the data was analyzed to get this important pharmacological information that otherwise would have been very difficult and almost impossible to get using other means. The present invention has been explained with the help of above experiment, which is not intended to limit the scope of the present invention. We Claim 1. Radiolabeled form of drug composition comprising of a) drug solution in the ratio of .02-30%. b) a reducing agent in the ratio of .0002-20%. c) a stabilizer in the ratio of 0-1.8% d) phosphate buffers in the ratio of 0-8.3% e) required amount of radioactive material and f) the rest of the 100% composition being water or 0.1 N HCI or organic solvents or their combination. 2. A composition as claimed in claim 1 wherein the said drug solution consists of Salbutamol Sulphate, Sametrol or Tebutalin sulphate. 3. A composition as claimed in any of the preceding claims wherein the said reducing agent is selected from stannous tartarate or stannous chloride. 4. A composition as claimed in any of the preceding claims wherein the said stabilizer includes ascorbic acid, gentistic acid or the like. 5. A composition as claimed in any of the preceding claims wherein the said radioactive material comprises of Tc-99m pertechnetate. 6. A process of making radiolabeled form of drug composition comprising the steps of: a) mixing drug solution in the ratio of .02-30% with a reducing agent in the ratio of .0002-20%, phosphate buffers in the ratio of 0-8.3% and water or 0.1 N HCI or organic solvents or their combination in a controlled environment of pH 2-9 and temperature of at least 20° C; b) incubating the solution with a stabilizer in the ratio of 0-1.8% for atleast 1 minute; c) mixing required amount of radioactive material with the solution of (a) and (b); d) incubating the solution for at least 3 minutes. 7. A process as claimed in claim 6 wherein the said drug solution consists of Salbutamol Sulphate, Sametrol or Tebutalin sulphate. 8. A process as claimed in claims 6, 7 or 8 wherein the said reducing agent comprises of stannous tartarate or stannous chloride. 9. A process as claimed in claims 6, 7, 8 or 9 wherein the said stabilizer comprises of ascorbic acid or gentistic acid. 10. A composition as claimed in claims 6, 7, 8 or 9 wherein the said radioactive material comprises of Tc-99m pertechnetate. 11. A radiopharmaceutical kit comprising of a. Salbutamol Sulphate in the ratio of 0.02-1 % b. Stannous Tartarate or Chloride in the ratio of 0.0002-0.2% c. Ascorbic acid upto 1.8% d. Gentisic acid upto 2% e. Phosphate buffers upto 8.3% f. 0.1N HCL or water or their combination being the rest of the composition to be mixed with 100MBq of Na99mTcO4 at the time of use. 12. A composition as herein described in the description and accompanying drawings. 13. A composition as herein described in the description and accompanying drawings. |
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| Patent Number | 268316 | ||||||||||||||||||
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| Indian Patent Application Number | 1443/DEL/2005 | ||||||||||||||||||
| PG Journal Number | 35/2015 | ||||||||||||||||||
| Publication Date | 28-Aug-2015 | ||||||||||||||||||
| Grant Date | 26-Aug-2015 | ||||||||||||||||||
| Date of Filing | 03-Jun-2005 | ||||||||||||||||||
| Name of Patentee | DIRECTOR GENERAL, DEFENCE RESEARCH & DEVELOPMENT ORGANISATION | ||||||||||||||||||
| Applicant Address | MINISTRY OF DEFENCE, GOVERNMENT OF INDIA, WEST BLOCK-VIII, WING-I, SECTOR-1, R. K. PURAM, NEW DELHI-110066, INDIA. | ||||||||||||||||||
Inventors:
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| PCT International Classification Number | A61K 51/04 | ||||||||||||||||||
| PCT International Application Number | N/A | ||||||||||||||||||
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