Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Reexamination Certificate
2001-07-17
2003-11-04
Myers, Carla J. (Department: 1655)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
C435S091200, C536S023500, C536S024310, C536S024330
Reexamination Certificate
active
06642002
ABSTRACT:
FIELD OF THE INVENTION
This invention relates generally to a method of identifying an individual having an increased susceptibility to developing Familial Primary Pulmonary Hypertension (FPPH), as well as to a method for diagnosing an individual suffering from FPPH. The invention also relates to a method of identifying an individual having an increased susceptibility to developing non-familial, or sporadic, Primary Pulmonary Hypertension (PPH), as well as to a method for diagnosing an individual suffering from sporadic PPH. The invention also relates to a method of identifying an agent capable of altering the symptoms of PPH in an individual suffering from familial or sporadic PPH, comprising contacting a test agent with Bone Morphogenic Protein Receptor II (BMPR-II) and determining whether the test agent alters BMPR-II activity, wherein an alteration in BMPR-II activity in the presence of the test agent as compared with BMPR-II activity in the absence of the test agent indicates that the test agent is capable of altering the symptoms of PPH in an individual suffering from familial or sporadic PPH.
BACKGROUND OF THE INVENTION
Primary pulmonary hypertension (PH) is characterized by sustained elevation of pulmonary artery pressure (greater than 25 mmHg at rest and greater than 30 mmHg during exercise) and with no identifiable cause, such as recurrent thromboembolism, chronic hypoxic lung disease or left-sided cardiac disease. PPH is twice as common in females than males and symptoms develop typically in the 3
rd
and 4
th
decades of life, although the disease may occur at any age. Despite advances in therapy, mortality in PPH remains high with mean survival from onset of disease only 2.5 year.
At least 6% of individuals diagnosed with PPH have a known family history of the disorder. The disease can be classified as being either familial (more than one affected relative has been identified in at least 6% of cases (familial PPH; MIM 178600) (ref. 3)) or sporadic. Familial PPH (FPPH) segregates as an autosomal dominant disorder, with markedly reduced penetrance.
There is a need to identify the genetic basis for this devastating disease in order to better diagnose and treat patients suffering from PPH.
BRIEF SUMMARY OF THE INVENTION
The invention relates to a method of identifying a subject having an increased susceptibility for developing pulmonary hypertension, comprising detecting a mutant Bone Morphogenic Protein Receptor II (BMPR-II) polypeptide or a mutated Bone Morphogenic Protein Receptor 2 (BMPR2) nucleic acid in the subject, thereby identifying a subject having an increased susceptibility for developing pulmonary hypertension. Wild-type BMPR2 nucleotide sequence is SEQ ID NO:1. Wild-type BMPR-II amino acid sequence is SEQ ID NO:2.
In one aspect, the mutated BMPR2 nucleic acid or mutant BMPR-II polypeptide has a sequence associated with pulmonary hypertension.
In another aspect, the mutated BMPR2 nucleic acid comprises a missense mutation.
In yet another aspect, the mutated BMPR2 nucleic acid comprises a nonsense mutation.
In another aspect, the mutated BMPR2 nucleic acid comprises a deletion mutation.
In another aspect, the mutated BMPR2 nucleic acid comprises an insertion mutation.
In another aspect, the mutated BMPR2 nucleic acid comprises a truncation mutation. Preferably, the mutated BMPR2 nucleic acid is truncated at a nucleotide position of the sequence set forth in SEQ ID NO:1 which is 3′ to nucleotide position 2695.
In another aspect, the subject having an increased susceptibility for developing pulmonary hypertension is identified by detecting a BMPR2 nucleic acid having a sequence associated with pulmonary hypertension.
In a preferred aspect, the pulmonary hypertension is primary pulmonary hypertension. In another aspect, the pulmonary hypertension is secondary pulmonary hypertension.
In various preferred embodiments, the mutated BMPR2 nucleic acid can include a missense mutation or a nonsense mutation.
In another aspect, the invention features a method of identifying a mutant BMPR-II polypeptide or a mutated BMPR2 nucleic acid, including detecting, in a patient with PPH, a BMPR-II polypeptide that is not present in normal subjects or a BMPR2 nucleic acid that is not present in normal subjects, thereby identifying a mutant BMPR-II polypeptide or a mutated BMPR2 nucleic acid.
In another aspect, the invention features a method of increasing BMPR-II biological activity.
In another aspect, the invention features a method of decreasing BMPR-II biological activity.
In another aspect, the invention features a method of identifying a compound that modulates the biological activity of a BMPR-II polypeptide, including: a) contacting a sample including a BMPR-II polypeptide or a BMPR2 nucleic acid with the compound; and b) measuring BMPR-II biological activity in the sample, whereby an increase or decrease in BMPR-II biological activity, compared to BMPR-II biological activity in an identical sample not contacted with the compound, identifies a compound that modulates the biological activity of the BMPR-II polypeptide.
In various embodiments of this aspect of the invention, BMPR-II biological activity is increased or decreased; the BMPR-II polypeptide is a wild-type BMPR-II polypeptide or the BMPR2 nucleic acid is a wild-type BMPR2 nucleic acid; the BMPR-II polypeptide is a polymorphic variant of a BMPR-II polypeptide or the BMPR2 nucleic acid is a polymorphic variant of a BMPR2 nucleic acid; or the BMPR-II polypeptide is a mutant BMPR-II polypeptide or the BMPR2 nucleic acid is a mutated BMPR2 nucleic acid.
In another aspect, the invention features a non-human mammal having a deleted, mutated, or polymorphic variant BMPR2 gene. In various aspects of the twelfth aspect of the invention, the non-human mammal is a mouse; and/or the non-human mammal is homozygous for the deleted, mutated, or polymorphic variant BMPR2 gene.
Additional advantages of the invention will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. The advantages of the invention will be realized and attained by means of the elements and combinations particularly pointed out in the appended claims. It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the invention, as claimed.
REFERENCES:
patent: 3610795 (1971-10-01), Antoine
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patent: WO 02/04684 (2002-01-01), None
patent: WO 02/16398 (2002-02-01), None
Deng et al. American Journal of Human Genetics. Sep. 2000. 67: 737-744.*
Machado et al. American Journal of Human Genetics. Jan. 2001. 68: 92-101.*
Deng et al., Familial Primary Pulmonary Hypertension (Gene PPH1) Is Caused by Mutations in the Bone Morphogenetic Protein Receptor-II Gene,Am. J. Hum. Genet. 67:737-744 (2000).
Lane et al., Heterozygous germline mutations in BMPR2, encoding a TGF-&bgr; receptor, cause familial primary pulmonary hypertension,Nature Genetics, vol. 26:81-84 (Sep. 2000). XP002907136.
Machado et al., BMPR2 Haploinsufficiency as the Inherited Molecular Mechanism for primary Pulmonary Hypertension,American Journal of Human Genetics, 68:92-102 (2001). XP002909517.
Thomson et al., Sporadic primary pulmonary hypertension is associated with germline mutations of the gene encoding BMPR-II, a receptor member of the TGF-&bgr; family,J. Med. Genet. 37:741-745 (2000). XP001118640.
Abenhaim et al. Appetite-Suppressant Drugs and the Risk of Pulmonary Hypertension.N. Eng. J. MEd. 335(9):609-616 (1996).
Agrawal et al. Cell-Cycle kinetics and VSV-G pseudotyped retrovirus-mediated gene transfer in blood-derived CD34+cells.Exper. Hematol. 24:738-747 (1996).
Alvarez et al. A Phase I Study of Recombinant Adenovirus Vector-Mediated Intraperitoneal Delivery of Herpes Simplex Virus Thymidine Kinase (HSV-TK) Gene and Intravenous Genciclovir for Previously Treated Ovarian and Extraovarian Cancer Patients.Hum. Gene Ther. 8:597-613 (1997).
Foroud Tatiana
Lane Kirk B.
Loyd James E.
Machado Rajiv D.
Nichols William C.
Myers Carla J.
Needle & Rosenberg P.C.
Vanderbilt University
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