Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Reexamination Certificate
2000-03-08
2001-08-28
Arthur, Lisa B. (Department: 1655)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
C536S023100, C536S024300, C536S024330
Reexamination Certificate
active
06280951
ABSTRACT:
TECHNICAL FIELD
The present invention relates to a method for predicting reactivity to parathyroid hormone in humans by analysis of genetic polymorphism. The results obtained by the analyis of genetic polymorphism can be used in predicting the severity of secondary hyperparathyroidism in hemodialyzed patients, and the severity of primary hyperparathyroidism, and in predicting the effects of the administration of parathyroid hormone on treatment of regressive osteoporosis.
BACKGROUND OF THE INVENTION
Parathyroid hormone (hereinafter referred to as PTH) is a polypeptide hormone which is produced in the parathyroid gland and secreted therefrom. The main function of this hormone is to keep the calcium level in body fluids constant. PTH promotes bone resorption and calcium resorption in the kidney, thereby increasing the calcium level in blood, and PTH also promotes excretion of phosphate, thereby decreasing the phosphate level in blood. A decrease in the calcium level in blood promotes PTH secretion, while an increase in the calcium level in blood suppresses the secretion of PTH.
Hyperparathyroidism, which is characterized by excess secretion of PTH, includes primary and secondary hyperparathyroidism. In primary hyperparathyroidism, excess secretion of PTH due to tumefaction of the parathyroid gland, etc. results in dysbolism of calcium, thereby causing disorders such as hypercalcemia, hypophosphatemia, osteitis fibrosa, nephrolithiasis, and hypertension. Secondary hyperparathyroidism is caused when a low serum calcium level continues for a long time, which condition results from decreased renal function, deficiency of activated vitamin D, decreased reactivity of bones to PTH, etc. Secondary hyperparathyroidism results in an increase in PTH secretion. In some cases, this excess secretion of PTH continues even when the serum calcium level becomes normal or elevated, and may result in hyperparathyroidism becoming serious. For example, in a patient who is subjected to hemodialysis because of chronic renal failure, hyperparathyroidism tends to be serious. One of the important factors affecting this tendency is development of resistance to PTH, which is considered to be associated with reactivity to PTH. At present, secondary hyperparathyroidism is treated by the administration of activated vitamin D
3
.
The severity of secondary hyperparathyroidism in hemodialyzed patients varies among individuals, and therefore genomic factors have become of interest. As an example of studies on genomic factors, there is a report which shows the relationship in Japanese between polymorphism of the vitamin D receptor (VDR) gene by restriction enzyme Bsm I and the serum PTH level in hemodialyzed patients (Y. Tsukamoto et al., Nature Med. vol.2, 1996, p. 1162). Polymorphism of the VDR gene is correlated with occurrence of secondary hyperparathyroidism and response to treatment with vitamin D. Also, severity of primary hyperparathyroidism is thought to be associated with the same genomic factors in some cases.
With respect to the mechanism of occurrence of hyperparathyroidism, genetic differences in reactivity to PTH have been thought to be a stronger determining factor. However, a methodology has not been established for examining genetic polymorphism to predict reactivity to PTH.
Polymorphism of the parathyroid hormone receptor gene was reported by Frank G. Hustmyer et al. in Human Mol. Gent. vol. 2, p. 1330 (Bsm I polymorphism) and by E. Schipani et al in Human Mol. Gent. vol. 3, p. 1210 (polymorphism in exon M7). However, there is no report demonstrating the polymorphism associated with PTH reactivity.
As mentioned above, a method for predicting PTH reactivity by analysis of genetic polymorphism has not been found, PTH reactivity being thought to be associated with severity of hyperparathyroidism, and predicting the severity thereof was difficult. The prediction of the severity of hyperparathyroidism is advantageous. For example, if a tendency for secondary hyperparathyroidism observed in hemodialyzed patients to become serious is predicted, a suitable method for treatment can be selected so as to prevent it from becoming serious. Alternatively, if the severity of symptoms accompanied by primary hyperparathyroidism such as hypercalcemia, bone lesions, urinary calculus, etc. is predicted, it may be possible to determine whether an operation for removal of the parathyroid gland should be performed and to determine how urgent the operation is. Furthermore, the prediction of PTH reactivity is useful for predicting the effects of PTH administration in the treatment of regressive osteoporosis.
SUMMARY OF THE INVENTION
Accordingly, an object of the present invention is to provide a method for predicting PTH reactivity by analysis of genetic polymorphism.
The present inventors have found the effects of genetic polymorphism of the parathyroid hormone receptor gene on the severity of hyperparathyroidism, and the relationship between this genetic polymorphism and PTH reaction, and have thereby completed the present invention.
The present invention provides a method for predicting reactivity to parathyroid hormone by determining a genotype with respect to genetic polymorphism of the parathyroid hormone receptor gene.
The determination of genotype may be performed by detection for the presence or absence of digestion with a restriction enzyme. In a preferred embodiment, genetic polymorphism is analyzed by treating a DNA fragment obtained by amplifying the first intron region of the parathyroid hormone receptor gene with restriction enzyme Van91 I. Preferably, amplification is performed using a primer having the sequence of SEQ ID NO:1 and a primer having the sequence of SEQ ID NO:2.
The present invention also provides a primer set for analyzing genetic polymorphism of the parathyroid hormone receptor gene, comprising a primer having the sequence of SEQ ID NO:1 and a primer having the sequence of SEQ ID NO:2, and furthermore provides a kit for prediction of reactivity to parathyroid hormone in a humany, comprising the above primer set and restriction enzyme Van91 I.
Furthermore, the present invention relates to a method for predicting the tendency for a patient being subjected to hemodialysis to suffer from severe secondary hyperparathyroidism according to the above-mentioned method, and a method for predicting the tendency for a human to suffer from severe primary hyperparathyroidism according to the analysis of polymorphism as mentioned above. Also, the present invention relates to a method for predicting the reactivity to parathyroid hormone to be administered for treatment of regressive osteoporosis according to the above-mentioned method.
REFERENCES:
Schipani et al “Polymorphism in exon M7 of the PTHR gene” Human Molecular Genetics, vol. 3, No. 7, p. 1210, 1994.*
Hustmyer et al “Bsml polymorphism at the parathyroid hormone receptor locus (PTHR) in three populations” Human Molecular Genetics, vol. 2, No. 8, p. 1330, 1993.*
Schipani et al “Pseudohypoparathroidism Type lb is not caused by mutations in the coding exons of the human PTH/PTH-related peptide receptor gene” J. of Clinical Endocrinology and Metabolism, vol. 80, No. 5, p. 1611-1621, 1995.*
Heishi et al “A novel Nan91 ! polymorphism in the 1st intron of the parathyroid hormone” Biol. Pharm, Bill, vol. 23, No. 4, p. 386-389, Apr. 2000.*
Y. Tsukamoto et al,Nature Medicine, 2(11):1162 (1996).
F. Hustmyer et al.,Human Molecular Genetics, 2(8):1330 (1993).
E. Schipani et al.,Human Molecular Genetics, 3(7):1210 (1994).
Heishi Masayuki
Ishida Osamu
Tazawa Hiromitsu
Tsukamoto Yusuke
Arthur Lisa B.
Burns Doane , Swecker, Mathis LLP
Goldberg Jeanine
Tsukamoto Yusuke
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