TWI656876B

TWI656876B - 治療ｐａｈ之抗增生劑

Info

Publication number: TWI656876B
Application number: TW106135422A
Authority: TW
Inventors: 史蒂芬伊凡斯
Original assignee: 美商輝瑞股份有限公司
Priority date: 2016-10-20
Filing date: 2017-10-17
Publication date: 2019-04-21
Also published as: EP3804724B1; US20190240226A1; MX2019004602A; SG11201902523UA; ES2934846T3; US20210186974A1; BR112019005526A2; AU2017345367A1; US11439646B2; EP3804724A1; RU2019111887A; JP2018076290A; EP3528812B1; CN109843297A; KR20190071763A; US10849903B2; RU2019111887A3; WO2018073687A1; TW201827054A; IL266026A

Abstract

肺高血壓和相關疾病(如肺動脈高血壓)可藉由投予有效劑量之CDK抑制劑治療，該CDK抑制劑包括帕博西尼(palbociclib)，6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2基]胺基}吡啶並[2,3-d]嘧啶-7(8H)-酮或其醫藥上可接受之鹽。

Description

治療ＰＡＨ之抗增生劑

[0001]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 肺高血壓和相關疾病(如肺動脈高血壓)可藉由投予有效劑量之CDK抑制劑治療，該CDK抑制劑包括帕博西尼(palbociclib)，6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2基]胺基}吡啶並[2,3-d]嘧啶-7(8H )-酮。

[0002]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . Pulmonary hypertension (PH) has been previously classified as primary (idiopathic) or secondary.肺高血壓(PH)先前已被分類為原發性(特發性)或繼發性。最近，世界衛生組織(WHO)已將肺高血壓分為5組：第1組：肺動脈高血壓(PAH)；第2組：具有左心臟疾病之PH；第3組：具有肺病及/或低氧血症(hypoxemia)之PH；第4組：由於慢性血栓性(thrombotic)及/或栓塞性疾病所造成之PH；和第5組：雜項病症(例如類肉瘤病(sarcoidosis)、組織細胞增生症(histiocytosis)X，淋巴管瘤病(lymphangiomatosis)及肺血管壓迫性病變)。　　[0003]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . Pulmonary arterial hypertension (PAH) is a serious, complex , progressive and life-threatening disease of the pulmonary vasculature, characterized by profound vasoconstriction and an abnormal proliferation of smooth muscle cells in the walls of the pulmonary arteries.肺動脈高血壓(PAH)是一種嚴重、複雜、進行性且威脅生命的肺血管疾病，其特徵在於強烈的血管收縮和肺動脈壁中平滑肌細胞異常增生。肺部Severe narrowing血管嚴重狹窄導致肺動脈壓非常高。這些高血壓使心臟難以泵送血液通過欲充氧的肺。PAH之患者由於心臟努力掙扎以克服這些高血壓來泵送血液，因而苦於呼吸極度短促。Patients with PAH typically develop significant increases in pulmonary vascular resistance (PVR) and sustained elevations in pulmonary artery pressure (PAP), which ultimately lead to right ventricular failure and death.PAH患者通常發展出顯著增加之肺血管阻力(PVR)和持續升高之肺動脈壓(PAP)，最終導致右心室衰竭和死亡。Patients diagnosed with PAH have a poor prognosis and equally compromised quality of life, with a mean life expectancy of 2 to 5 years from the time of diagnosis if untreated.被診斷為PAH之患者的預後差且生活品質同樣受損，如果未治療的話，從診斷時間起平均之預期壽命為2至5年。　　[0004]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Current methods of treating PAH focus on reducing symptoms and prolonging patient lifespan and enhancing quality of life. 目前治療PAH之方法著重於減輕症狀及延長患者壽命並提高生活品質。該等治療方法包括投予：血管擴張劑，諸如前列環素(prostacyclin)、依前列醇(epoprostenol)和西地那非(sildenafil)；內皮素受體拮抗劑，諸如波生坦(bosentan)；鈣通道阻斷劑，諸如胺氯地平(amlodipine)、地爾硫卓(diltiazem)和硝苯地平(nifedipine)；抗凝劑，諸如華法林(warfarin)；補充氧療和利尿劑。當醫療失敗時，最終之治療選擇為肺及/或心肺移植。然而，這些方法的每一種都存在一或多種缺點，例如缺乏有效性、副作用嚴重、患者的依從性低及無法預防PAH。因此，需要更好的治療PAH之方法。本發明解決這些需求。

1C1　　The present invention provides a method of treating PAH[0005]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 本發明提供治療PAH和其他相關疾病和病症之方法，該方法係藉由投予有效劑量之CDK抑制劑(包括帕博西尼(palbociclib)and other related diseases and conditions by administering a therapeutically effective d ose of a CDK inhibitor, including palbociclib .)。　　Two k ey cell types involved in progression towards PAH are Pulmonary Arterial[0006]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 涉及朝PAH進展的二種關鍵細胞類型為肺動脈平滑肌細胞(PASMC)和肺動脈外膜纖維母細胞(PAAF)。本發明係部分基於發現帕博西尼顯著影響人和大鼠PASMC及人PAAF之抗增生作用。帕博西尼亦顯著影響PAH模型中之血流動力學(hemodynamics)、肺血管細胞增生、肺動脈形態和右心室肥大的變化。　　[0007]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Palbociclib is a potent and selective inhibitor of CDK4 and CDK6, having the structure: 帕博西尼為CDK4和CDK6之有效和選擇性抑制劑，其具有下列結構：。　　[0008]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Palbociclib is a potent and selective inhibitor of CDK4 and CDK6, having the structure: 帕Palbociclib is described inWHO Drug Information , Vol.博西尼係描述於WHO Drug Information , Vol. 27, No. 2,第172頁(2013) 中。帕博西尼及其醫藥上可接受之鹽和調製劑揭示於下列文獻中：國際刊物第WO 2003/062236號和美國專利案編號6,936,612、7,208,489和7,456,168；國際刊物第WO 2005/005426號和美國專利案第7,345,171和7,863,278號；國際刊物第WO 2008/032157號和美國專利案第7,781,583號；國際刊物第WO 2014/ 128588號；和國際申請案第PCT/IB2016/053040號。前述各參考文獻之內容以引用其整體之方式併入本文。　　Palbociclib is approved in the United States for the treatment of hormone receptor (HR)-positive, human epidermal growth factor 2 (HER2)-negative advanced or metastatic breast cancer in combination with letrozole as initial endocrine therapy in postmenopausal women or in combination with fulvestrant following disease progression on endocrine therapy.[0009]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Current methods of treating PAH focus on reducing symptoms and prolonging patient lifespan and enhancing quality of life. Palbociclib is a potent and selective inhibitor of CDK4 and CDK6, having the structure:帕博西尼在美國被核准與來曲唑(letrozole)組合以作為停經後婦女之初始內分泌療法或在內分泌療法後疾病進展上與氟維司群(fulvestrant)組合，以用於治療激素受體(HR)陽性，人表皮生長因子2(HER2)陰性之末期或轉移性乳癌。　　In another embodiment of the invention, palboci c lib is administered to a subject diagnosed with or at risk of developing pulmonary hypertension or PAH includ ing , but are not limited to, idiopathic PAH, hereditary or familial PAH, and secondary pulmonary hypertension (eg hypertension resulting from pulmonary emboli, emphysema, pulmonary fibrosis, and congenital heart disease).[0010]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . Current methods of treating PAH focus on reducing symptoms and prolonging patient lifespan and enhancing quality of life.於本發明之另一實施態樣中係對被診斷患有或處於發展出肺高血壓或PAH之風險中的個體投予帕博西尼，該肺高血壓或PAH包括，但不限於特發性PAH、遺傳性或家族性PAH和繼發性肺動脈高血壓(例如由肺栓塞、肺氣腫、肺纖維化和先天性心臟病引起)。In one embodiment, the subject is diagnosed with idiopathic PAH or hereditary PAH.於一實施態樣中，該個體被診斷出患有特發性PAH或遺傳性PAH。於一些實施態樣中，該處於發展出PAH之風險中的個體在編碼骨形態發生之第2型蛋白受體的基因中具有突變。　　[0011]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .The present invention also includes a kit for the treatment or prevention of pulmonary arterial hypertension. 本發明亦包括用於治療或預防肺動脈高血壓之套組。In one embodiment, the kit comprises palbociclib and an administration device.於一實施態樣中，該套組包含帕博西尼和投予裝置。該投予裝置可經過設計以用於肺部遞送，諸如吸入器、噴霧器、吹入器、滴管和霧化器。於其他實施態樣中，該投予裝置可經過設計以用於靜脈內或動脈內遞送，諸如導管。帕博西尼可視需要地配製成供儲存在投予裝置中。該套組可進一步包含用於對個體(例如人)投予帕博西尼的說明書以治療或預防本文所討論之PAH或其他疾病。In one embodiment, the instructions elaborate and qualify the mode of administration, for example, by specifying the days of administration for palbociclib during a 28 day cycle.於一實施態樣中，該說明書闡述並使投予模式合格，例如藉由具體指定在28天週期之期間內投予帕博西尼的日子。

[0017]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .DETAILED DESCRIPTION OF THE INVENTIONPulmonary hypertension, like systemic hypertension, is not a single disease but a group of diseases, which share the defining element of a mean pulmonary arterial pressure ＞ 25 mm Hg. 肺動脈高血壓(如全身性高血壓)並非單一疾病，而是一組疾病，此組疾病共享平均肺動脈壓力為≥ 25mmHg的界定要素。PHPH has been classified and divided into 5 groups (Galie et al. ERJ (2009) Dec.; 34(6);1219-63).已被分類並分成5組(Galie et al. ERJ (2009) Dec.; 34(6);1219-63)。This invention generally relates to , but is not limited to, treating the World Health Organization (WHO) group 1, or PAH group of, diseases characterized by elevated pulmonary arterial pressure and elevated blood flow resistance due to a precapillary pulmonary microangiopathy.本發明大致上關於，但不限於治療世界衛生組織(WHO)第1組或PAH組的疾病，該第1組或PAH組疾病之特徵為由於微血管前肺微血管病所導致之肺動脈壓升高和血流阻力增加。　　A key feature in the PAH lung is abnormal cellular proliferation, which leads to progressive obliteration of the lumina of the pulmonary vasculature, resulting in pathological increases in vascular resistance.[0018]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .DETAILED DESCRIPTION OF THE INVENTIONPulmonary hypertension, like systemic hypertension, is not a single disease but a group of diseases, which share the defining element of a mean pulmonary arterial pressure ＞ 25 mm Hg. PAH肺中之一種關鍵特徵為異常的細胞增生，此導致肺脈管系統之腔管進展性閉塞，造成病理性血管阻力增加。肺血管重塑為主動的結構改變過程，其本質上與細胞生長、細胞死亡、細胞遷移、細胞分化和細胞外基質合成或降解之修飾關聯。Schermuly, R.T., et al. Mechanisms of disease: pulmonary arterial hypertension.Nature reviews. Cardiology 8, 443-455 (2011)。 While a number of pharmacological classes already exist as approved therapies in clinical PAH, they primarily induce vasorelaxation and thereby reduce pulmonary vascular resistance.雖然已有許多藥理學類別被核准作為臨床PAH療法，但它們主要誘導血管舒張，從而降低肺血管阻力。再者，鑑於患者之個別差異，單一療法可能不適當。Humbert, M. & Ghofrani, HA The molecular targets of approved treatments for pulmonary arterial hypertension.Humbert, M. & Ghofrani, H.A. The molecular targets of approved treatments for pulmonary arterial hypertension.Thorax 71, 73-83 (2016)。因此，這些療法不能解決血管重塑的複雜性。　　C hanges in the pulmonary vasculature involve[0019]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . DETAILED DESCRIPTION OF THE INVENTIONPulmonary hypertension, like systemic hypertension, is not a single disease but a group of diseases, which share the defining element of a mean pulmonary arterial pressure ＞ 25 mm Hg.肺脈管系統中之變化涉及PASMC、PAAF和肺動脈內皮細胞(PAEC)。Guignabert, C, et al.Guignabert, C, et al.Pathogenesis of pulmonary arterial hypertension: lessons from cancer. European respiratory review:an official journal of the European Respiratory Society 22, 543-551 (2013)。新近對PAH之研究的回顧討論增生信號傳導“轉發站(hub)”以靶向信號轉導的多個途徑。Pullamsetti, SS, et al.American journal of respiratory and critical care medicine (2016)，於2016年9月14日首次在線上以DOI：10.1164/rccm.201606 -1226PP發表。另一篇回顧考慮肺部血流中之變化與肺血管壁內皮細胞中之癌樣變化之間的關係。Heppe, C.M., et al.Vascular Pharmacology 83 (2016) 17-25。　　N ew approaches have focused on targeting the pro-proliferative phenotype that underpins the pulmonary vascular remodeling in PAH patients, with the aim of achieving improved efficacy and enhanced survival.[0020]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . DETAILED DESCRIPTION OF THE INVENTIONPulmonary hypertension, like systemic hypertension, is not a single disease but a group of diseases, which share the defining element of a mean pulmonary arterial pressure ＞ 25 mm Hg.新方法已著重在靶向該支持PAH患者之肺血管重塑的促增生表型，目的是達成提升效能和增進存活。在臨床前研究中，已經有幾個實例其中顯示出腫瘤藥物能有效減緩PAH疾病進展。參見Savai, R., et al. Pro-proliferative and inflammatory signaling converge on FoxO1 transcription factor in pulmonary hypertension.Nature medicine 20, 1289-1300(2014)；Ciuclan, L., et al. Imatinib attenuates hypoxia-induced pulmonary arterial hypertension pathology via reduction in 5-hydroxytryptamine through inhibition of tryptophan hydroxylase 1 expression.American journal of respiratory and critical care medicine 187, 78-89 (2013)；和Schermuly, R.T., et al. Reversal of experimental pulmonary hypertension by PDGF inhibition.The Journal of clinical investigation 115, 2811-2821 (2005)。　　[0021]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Cyclin dependent kinases and related serine/threonine protein kinases are important cellular enzymes that perform essential functions in regulating cell division and proliferation. 週期蛋白(cyclin)依賴性激酶和相關之絲胺酸/蘇胺酸蛋白激酶為重要的細胞酶，其在調節細胞分裂和增生中執行重要的功能。週期蛋白依賴性激酶催化單位被稱為週期蛋白之調節次單位激活。現已鑑定出至少十六種哺乳動物週期蛋白(Johnson DG, Walker CL. Cyclins and Cell Cycle Checkpoints. Annu. Rev. Pharmacol. Toxicol. (1999) 39:295 312)。週期蛋白B/CDK1、週期蛋白A/CDK2、週期蛋白E/CDK2、週期蛋白D/CDK4、週期蛋白D/CDK6及其他可能之異頻驅使(heterodyne)(包括CDK3和CDK7)為細胞週期進程的重要調節劑。週期蛋白/ CDK異頻驅使之其他功能包括調節轉錄、DNA修復、分化和凋亡(Morgan DO. Cyclin dependent kinases: engines, clocks, and microprocessors. Annu. Rev. Cell. Dev. Biol. (1997) 13:261 291)。　　T he complexity of pulmonary vascular remodeling provides many pathways for investigation such as targeting cell cycle of proliferation[0022]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 肺血管重塑之複雜性提供許多調查途徑，諸如靶向涉及稱為週期蛋白依賴性激酶(CDK)之蛋白質的細胞增生週期。這些CDK與特定之週期蛋白聯結以形成活性所必要之全酶(holoenzyme)複合物。對增生週期的每個步驟而言，需要不同的週期蛋白，且各週期蛋白之相對表現增加及/或減少以周期性激活其特異性CDK。Charron, T., et al. The cell cycle: a critical therapeutic target to prevent vascular proliferative disease.The Canadian journal of cardiology 22 Suppl B, 41B-55B (2006)。　　[0023]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .The present invention is based, in part, on the discovery that CDK inhibitors (CDKi's) 本發明係部分基於發現CDK抑制劑(CDKi)抑制人和大鼠PASMC及人PAAF中之有絲分裂原的增生作用並在PAH模型中減少血流動力學in hemodynamics, pulmonary vascular cell proliferation, pulmonary artery morphology, and right ventricle hypertrophy in PAH models.、肺血管細胞增生、肺動脈形態和右心室肥大的升高。　　The present invention is further based, in part, on the discovery that palbociclib significantly affected the anti-proliferative effects in human and rat[0024]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . The present invention is based, in part, on the discovery that CDK inhibitors (CDKi's)本發明進一步部分基於以下發現：帕博西尼顯著影響人和大鼠PASMC及人PAAF中之抗增生作用，並在PAH模型中顯著影響血流動力學in hemodynamics, pulmonary vascular cell proliferation, pulmonary artery morphology, and right ventricle hypertrophy in PAH models.、肺血管細胞增生、肺動脈形態和右心室肥大的變化。　　A n o ther embodiment of the invention includes a method o f treat ing[0025]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 本發明之另一實施態樣包括治療PAH(肺動脈高血壓)之方法，該方法包含對有需要治療PAH之個體投予有效量之6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2基]胺基}吡啶並[2,3-d]嘧啶-7(8H)-酮或其醫藥上可接受之鹽。　　[0026]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . A nother embodiment of the invention includes a method o f treat ing本發明之另一實施態樣包括治療PAH(肺動脈高血壓)之方法，該方法包含對有需要治療PAH之個體投予有效量之6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2基]胺基}吡啶並[2,3-d]嘧啶-7(8H)-酮。　　[0027]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Another embodiment of the invention includes a method of treating PAH comprising administering to a subject in need thereof an effective amount of a CDK inhibitor. 本發明之另一實施態樣包括治療PAH之方法，該方法包含對有需要治療PAH之個體投予有效量之CDK抑制劑。　　Another embodiment of the invention includes a method of treating PAH comprising administering to a subject in need thereof an effective amount of a CDK i nhibitor selected from any of the following compounds, or pharmaceutically acceptable salts thereof :[0028]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Another embodiment of the invention includes a method of treating PAH comprising administering to a subject in need thereof an effective amount of a CDK inhibitor. 本發明之另一實施態樣包括治療PAH之方法，該方法包含對有需要治療PAH之個體投予有效量之CDK抑制劑，該CDK抑制劑係選自下列任一化合物或其醫藥上可接受之鹽：　　 trilaciclib (G1T28), G1T38 , alvocidib曲來西尼(trilaciclib) (G1T28)、G1T38、歐弗西地(alvocidib)SEL-24 , milciclib , AGM-130 , AT 7519 , BAY 1143572 / 1112054 , BCD-115 , CYC065 , FLX 925 , SHR 6390 , TG-02 , A-1467729, ABC 1183, AZD 5576 , BPI 1178, senexin , ICEC0942, CCT-68127, CCT-251921 , ON 123300 , voruciclib , SEL120-34, SRX-3177, TP 1287, SY1365 , UD-017 , or VS2-370 . SEL-24、密西尼(milciclib)、AGM-130、AT 7519、BAY 1143572 /1112054、BCD-115、CYC065、FLX 925、SHR 6390、TG-02、A-1467729、ABC 1183、AZD 5576、BPI 1178、塞內辛(senexin)、ICEC0942、CCT-68127、CCT-251921、ON 123300、渥魯西尼(voruciclib)、SEL120-34、SRX-3177、TP 1287、SY1365、UD-017或VS2-370。　　[0029]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . Another embodiment of the invention includes a method of treating PAH comprising administering to a subject in need thereof an effective amount of a CDK inhibitor.本發明之另一實施態樣包括治療PAH之方法，該方法包含對有需要治療PAH之個體投予有效量之CDK抑制劑，該CDK抑制劑係選自：　　1. Ribociclib (7-cyclopentyl-N,N-dimethyl-2-((5-(piperazin-1-yl)pyridin-2-yl)amino)-7H-pyrrolo[2,3-d]pyrimidine-6-carboxamide), or pharmaceutically acceptable salts thereof: A CDKi targeting cyclin D1/CDK4 and cyclin D3/CDK6 cell cycle pathway, with potential antineoplastic activity.1. 利博西尼(ribociclib) (7-環戊基-N,N-二甲基-2-((5-(六氫吡嗪-1-基)吡啶-2-基)胺基)-7H-吡咯並[2,3-d]嘧啶-6-羧醯胺)或其醫藥上可接受之鹽：一種CDKi，其靶向週期蛋白D1/CDK4和週期蛋白D3/CDK6細胞週期途徑，具有潛在的抗贅瘤活性。利博西尼特異性抑制CDK4和6；　　2.2. 艾貝馬西尼(abemaciclib) (2-嘧啶胺 N-(5-((4-乙基-1-六氫吡嗪基)甲基)-2-吡啶基)-5-氟-4-(4-氟-2-甲基-1-(1-甲基乙基)-1H-苯並咪唑-6-基))或其醫藥上可接受之鹽：一種CDKi，其靶向CDK4和CDK6細胞週期途徑；或　　3.3.地納西尼(dinaciclib) (2-[(2S)-1-[3-乙基-7-[(1-氧離子基吡啶-1-鎓-3-基)甲胺基]吡唑並[1,5-a]嘧啶-5-基]六氫吡啶-2-基]乙醇) pyrazolo[1,5-a]pyrimidin-5-yl]piperidin-2-yl]ethanol), or pharmaceutically acceptable salts thereof: A CDKi that selectively targets CDK1, CDK2, CDK5, and CDK9.或其醫藥上可接受之鹽：一種CDKi，其選擇性地靶向CDK1、CDK2、CDK5和CDK9。　　Another embodiment of the invention includes a method of treating PAH comprising administering to a subject in need thereof an effective amount of a CDK inhibitor, or pharmaceutically acceptable salts thereof, as the CDK inhibitor is presented in any one or combination in the non-exclusive list including: WO03/062236, WO07140222 , WO10075074, WO10020675, WO16015598, WO16015597, WO16015605, WO16015604, WO2012/066508, WO04004632, WO11026911, WO11026904, WO 2011/101409, WO2006/074985, WO2012/061156 , WO1000913, WO10009155, and WO0183469.[0030]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . Another embodiment of the invention includes a method of treating PAH comprising administering to a subject in need thereof an effective amount of a CDK inhibitor.本發明之另一實施態樣包括治療PAH之方法，其包含對有需要治療PAH之個體投予有效量之CDK抑制劑或其醫藥上可接受之鹽，該CDK抑制劑為呈現於下列非排他性列表中之任何一CDK抑制劑或組合，包括： WO03/062236、WO07140222、WO10075074、WO10020675、WO16015598、WO16015597、WO16015605、WO16015604、WO2012/066508、WO04004632、WO11026911、WO11026904、WO2011/101409、WO2006/074985、WO2012/061156、WO1000913、WO10009155和WO0183469。　　Related diseases to be treated include any one or more or combination of the following: pulmonary capillary hemangiomatosis (PCH) (Group 1), pulmonary hypertension due to left heart disease (Group 2 PH), pulmonary hypertension due to lung disease and/or hypoxia (Group 3), chronic thromboembolic pulmonary hypertension (Group 4), pulmonary hypertension with unclear or multifactorial etiologies (Group 5), idiopathic forms of pulmonary vascular disease, or a lung disorder that including any form of acute and chronic lung injury and inflammation (ARDS, ILD, pneumonia, COPD, asthma), or primary lung vascular disorders (idiopathic, collagen vascular-associated, liver disease-associated, drug-associated, HIV-associated, blood clot-induced pulmonary hypertension).[0031]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 欲治療之相關疾病包括下列疾病之任何一或多種或其組合：肺微血管血管瘤病(PCH)(第1組)、由於左心臟疾病造成之肺高血壓(第2組PH)、由於肺病及/或缺氧造成之肺高血壓(第3組)、慢性血栓性肺高血壓(第4組)、具有不明或多因素病因之肺高血壓(第5組)、特發型肺血管疾病或包括任何形式之急性和慢性肺損傷和炎症的肺病(ARDS、ILD、肺炎、COPD、哮喘)、或原發性肺血管病(特發性、膠原蛋白血管相關性、肝臟疾病相關性、藥物相關性、HIV相關性、血凝塊誘發之肺高血壓)。本發明之另一實施態樣包括治療本文所討論之相關疾病discussed herein comprising administering to a subject in need thereof an effective amount of 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d]pyrimidin-7(8H)-one, or pharmaceutically acceptable salt thereof or no such salt.的方法，其包含對有需要治療之個體投予有效量之6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2-基]胺基}吡啶並[2,3-d]嘧啶-7(8H)-酮或其醫藥上可接受之鹽或無該等鹽。　　P albociclib was examined for its activity in PDGF- BB or serum-induced proliferation of human and rat PASMCs.[0032]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 在由PDGF-BB或血清誘導之人和大鼠PASMC的增生中檢查帕博西尼(Palbociclib)之活性。參見實施例1。帕博西尼有效地抑制由PDGF- BB誘導之人初代PASMC增生，藉由BrdU所得之IC₅₀ 為7.4nM，為3組之平均值(第1a圖)，並以相對螢光單位(rFU)表示。由5%胎牛血清(FBS)誘導之人類和大鼠初代PASMC二者的增生類似被帕博西尼處理所抑制(IC₅₀ 分別為7.4nM和27.4nM，各為3組之平均值)(第1b圖和第1d圖)。Palbociclib was also very potent against a nother cell type involved in vascular remodeling, the adventitial fibroblast (PAAF) .帕博西尼在對抗涉及血管重塑之另一細胞類型，外膜纖維母細胞(PAAF)亦非常有效。由血清誘導之人初代PAAF的增生受抑制之IC₅₀ =21.6nM (為3組之平均值)(第1c圖)。　　[0033]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .To demonstrate the anti-proliferative effects of CDK inhibitor s on PASMCs and PAAF were achieved via the pharmacological inhibition of CDK activity, three additional compounds ( a bemaciclib, dinaciclib and r ibociclib) were profiled in parallel to p albociclib . 為了證明CDK抑制劑對PASMC和PAAF之抗增生作用係經由藥學上抑制CDK之活性來達成，與帕博西尼並行找出另外三種化合物(艾貝馬西尼、地納西尼和利博西尼)的概況。表1提供如第1e、1f 和1g圖中所呈現之基於%增生作用的IC₅₀ FIG.值。　　It should be noted that due to experimental logistics , the human PASMC donor cells associa ted with FIG .[0034]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . To demonstrate the anti-proliferative effects of CDK inhibitor s on PASMCs and PAAF were achieved via the pharmacological inhibition of CDK activity, three additional compounds ( a bemaciclib, dinaciclib and r ibociclib) were profiled in parallel to p albociclib .應注意的是，由於實驗組織工作，與第1a和1b圖，及第2a和2b圖關聯之人類PASMC供體細胞係使用來自一位供體之細胞產生，而與第1e和2c圖關聯之數據係使用來自不同供體之細胞產生。同樣地，支持第1c和1f圖之人類PAAF亦衍自不同供體的細胞。最後，第1d和1g圖之大鼠PASMC製劑係從衍自不同動物之細胞製劑產生。如實施例1中所討論之各化驗法之間亦有差異。P albociclib was then examined for its effects on the phosphorylation of Rb in human PASMCs using in-cell western analysis .[0035]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 然後使用細胞中西方分析來檢查帕博西尼對人PASMC之Rb磷酸化的作用。參見實施例1。帕博西尼有效抑制由PDGF-BB或血清誘導之Rb磷酸化的IC₅₀ FIG.分別為5.2nM(第2b圖)和14.9nM(第2a圖)。因此，帕博西尼非常有效地阻斷PAH中涉及病理性血管重塑之關鍵細胞類型增生。　　In additional studies , where palbociclib was run in parallel with a bemaciclib, d inaciclib and r ibociclib ,[0036]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 於其中帕博西尼與艾貝馬西尼、地納西尼和利博西並行運作之另外的研究中，使用10%FBS誘導增生，而所有CDK抑制劑均顯示出抑制該經誘導之Rb磷酸化。參見表2，其係基於如第2c圖中呈現之數據2c, where the percent effect is calculated similarly to the percent effect for proliferation.，其中該百分比作用之計算類似於增生之百分比作用所用者。 Table 2: I nhibition of P hosphorylation of R b in h PASMCsMCT has been shown to induce pulmonary vascular injury primarily after hepatic generation of the toxic metabolite, monocrotaline pyrrole.[0037]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . MCT已顯示主要在肝臟產生毒性代謝物野百合鹼吡咯後誘導肺血管損傷。The MCT model offers the advantage of several key aspects of human PAH, including vascular remodeling, proliferation of smooth muscle cells, endothelial dysfunction, upregulation of inflammatory cytokines, and right ventricle ( RV ) failure .MCT模型之優點為提供人類PAH之幾個關鍵態樣，包括血管重塑、平滑肌細胞增生、內皮功能障礙、炎性細胞因子上調及右心室(RV)衰竭。Stenmark, K.R., et al. Animal models of pulmonary arterial hypertension:the hope for etiological discovery and pharmacological cure.American journal of physiology. Lung cellular and molecular physiology 297, L1013-1032 (2009)。在第3和4圖方面，Palbo為帕博西尼，Sild為西地那非，Rio為利奧西呱(riociguat)，而Veh為載劑。在實施例3所描述之MCT PAH模型describ ed in Example 3 , rats exposed to MCT consistently developed pulmonary hypertension, with an increase of mean (MPAP), systolic (SPAP), and diastolic (DPAP) pulmonary arterial pressures.中，暴露於MCT之大鼠持續發展出肺高血壓，其平均肺動脈壓(MPAP)、肺動脈收縮壓(SPAP)和肺動脈舒張壓(DPAP)增加。暴露於MCT後富爾頓(Fulton)指數亦增加。富爾頓指數為基於下列重量之比例： right ventricle [RV] / (left v entricle [LV]+ septum [S])右心室[RV]/(左心室[LV]+心室隔膜[S]) 　　For these studies, a n agent was considered to be administered[0038]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 在這些研究方面，當在第0天首次投予該藥劑時，該藥劑被認為是預防性投予，同一天亦投予第二藥劑(例如MCT)以引起該模型中發展出PAH症狀。However, an agent was considered to be administered therapeutically when the agent was administered after the animal had developed symptoms of PAH , eg, the agent was first administered on day 14 of the study .然而，當動物已發展出PAH症狀後才投予藥劑時，該藥劑被認為是治療性投予，例如該藥劑是在研究的第14天首次投予。　　As expected, s ildenafil (50 mg/kg/dose; PO BID), when dosed prophylactically (day 0-28), significantly reduced PAH-related increases in MPAP (FIG. 3b) , SPAP (FIG. 3h) , DPAP (FIG. 3i) , Fulton Index ( FIG. 3c )[0039]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 如所預期者，當預防性地給藥(第0-28天)時，西地那非(50mg/kg/劑量；PO，BID)顯著減少MPAP(第3b圖)、SPAP(第3h圖)、DPAP(第3i圖)、富爾頓指數(第3c圖)和心臟重量(第3j圖)的PAH-相關增加。然而，當治療性投予時(第14-28天)；西地那非(已被核准之用於PAH的療法)僅顯示出小幅且不明顯地降低本研究中之MPAP、SPAP、DPAP、富爾頓指數和心臟重量。　　[0040]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 當治療性投予(第14-28天)時，Palbociclib (100 mg/kg/day; PO; QD),帕博西尼(100 mg/kg/天；PO；QD)遠超過預期。第3a圖顯示研究之藥物的投予頻率。當預防性給藥時，帕博西尼不僅符合，且超過西地那非之正面血流動力學和生理作用。　　Histomorphometrically, p albociclib, but not s ildenafil, dramatically improved and in some cases fully normalized structural changes in the pulmonary vasculature (FIG. 3f) .[0041]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 帕博西尼組織形態計量學上顯著地改善(且在一些情況下完全正常化)肺脈管系統之結構變化，但西地那非則不(第3f圖)。與經投予載劑/MCT的組別相比較，經投予帕博西尼的組別(第14-28天)之肺泡內肺動脈(≤50μm外徑)和前肺泡肺動脈(51-100μm外徑)的平均血管壁厚度(彈性蛋白染色)已顯著降低(降至0.58x MCT)，超出西地那非的有利效果(第0-28天)(0.76x MCT)，而西地那非組(第14-28天)對這些測量值的作用很小(第3d圖)。相較於西地那非，帕博西尼(第14-28天)亦使完全肌化之肺泡內血管的密度降低更多，西地那非僅在第0至28天給藥時有效(αSMA染色；第3e圖)。此外，與經投予載劑/MCT之組別(範圍1-4；平均3)相比較，經投予帕博西尼之組別(第14-28天)的疾病嚴重性評分亦顯著較低(範圍0-2；平均1)，再次優於經投予西地那非之組別(第0-28天；範圍1-4；平均2.5)(第3g圖3g ).)。　　Chronic hypoxia plus[0042]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 慢性缺氧加vascular endothelial growth factor (VEGF) receptor blockade (Semaxinib/Su5416) cause severe PAH in rats.血管內皮生長因子(VEGF)受體阻斷(塞美西尼(Semaxinib)/Su5416)在大鼠中引起嚴重的PAH。Stenmark, K.R.，見下文。如實施例4中所描述者In the Su5416/Hx PAH model , as describ ed in Example 4 , p albociclib (100 mg/kg/day; PO; QD) was evaluated against two drugs common ly used to treat PAH patients: s ildenafil (30 mg/kg/dose; PO; BID) and r iociguat (10 mg/kg/day, PO; QD).，在Su5416/Hx PAH模型中，相對於常用來治療PAH患者的二種藥物評估帕博西尼(100mg/kg/天；PO；QD)：西地那非(30mg/kg/劑量；PO；BID)和利奥西呱(riociguat) (10 mg/kg/天，PO；QD)。Lang, M., et al. The soluble guanylate cyclase stimulator riociguat ameliorates pulmonary hypertension induced by hypoxia and SU5416 in rats.PloS one 7, e43433 (2012)。所有三種藥物均係在第7至28天投予(第4a圖)。雖然一如預期，以西地那非預防性治療時顯著降低MPAP(第4b圖)、SPAP(第4h圖)和DPAP(第4i圖)，以利奥西呱預防性治療時顯著降低MPAP和DPAP，與載劑/Su 5416/Hx組相比較，本研究中使用任一此二藥劑進行治療性治療時僅導致富爾頓指數略為降低，肺或心臟重量無變化。然而，帕博西尼取代西地那非和利奥西呱二者而將血流動力學終點恢復至接近非患病之對照組(第4b圖比較MPAP結果，且第4c圖比較富爾頓指數值)。當檢視治療性治療時，僅帕博西尼組顯示出富爾頓指數(第4c圖)和心臟重量顯著降低(第4j圖)。　　Histomorphometrically, p albociclib, but not s ildenafil or riociguat , dramatically improved and in some cases fully normalized structural changes in the pulmonary vasculature (FIG. 4f).[0043]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 帕博西尼組織形態計量學上顯著改善(且在一些情況下完全正常化)肺血管中之結構變化，但西地那非或利奥西呱則不(第4f圖)。組織形態計量學分析顯示出帕博西尼顯著降低肺泡內血管和前肺泡血管二者之平均血管壁厚度，但利奥西呱或西地那非則不(第4d圖)。The p albociclib-dosed group also had significantly lower (0.68x Su 5416/ Hx) muscularized intra-acinar vessel density compared with vehicle/Su 5416/ Hx group.與載劑/Su5416/Hx組相比較，經投予帕博西尼的組別亦顯著降低肌化之肺泡內血管密度(0.68x Su5416/Hx)。經投予Sildenafil- and Riociguat-dosed group means were not statistically significantly differ ent from vehicle/SuHx group (FIG . 4 e ).西地那非或利奥西呱之組別的平均值與載劑/SuHx組的差別並非統計上有意義的(第4e圖)。此外，與載劑/Su5416/Hx組(範圍3-4；平均3.65)相比較，該經投予帕博西尼之組別之疾病嚴重性評分幾乎完全正常化(範圍0-1.5；平均0.25)，優於經投予西地那非和利奥西呱之組別(第4g圖)。令人驚訝地，在大鼠SU5416/Hx PAH模型中，帕博西尼治療導致血流動力學改變幾乎完全被抑制及肺脈管系統的結構顯著改善，優於西地那非和利奥西呱。　　IHC analysis of dual αSMA/Ki67- and dual CD31/phosphoR b-stained rat lung sections (FIG . 5 a and FIG. 5 b ) from thein vivo studies showed that MCT caused a significant increase of vessel-associated Ki67⁺ proliferating cells (FIG. 5c) and a trend of increase in numbers of vessel-associated phoshoRb⁺ cells (FIG. 5d) .[0044]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . IHC分析來自活體內研究之經雙重αSMA/Ki67染色和雙重CD31/磷酸Rb染色的大鼠肺切片(第5a和5b圖)顯示出MCT引起血管相關之Ki67⁺ 增生細胞顯著增加(第5c圖)且血管相關之磷酸Rb⁺ 細胞數有增加的趨勢(第5d圖)。See Example 5. These target-specific endpoints were dramatically decreased to control levels by p albociclib administration (FIG . 5 c ).參見實施例5。經由投予帕博西尼，這些靶特異性終點顯著降低至對照水準(第5c圖)。類似地，Su5416/治療與缺氧之組合可誘導肺脈管系統中之血管相關之Ki67⁺ 和pRb⁺ 細胞顯著增加。帕博西尼充分逆轉這些增加；whereas ,然而，西地那非或利奥西呱顯示對這些生物標記物沒有明顯的有利作用。帕博西尼在PAH疾病模型中抑制肺脈管系統中之細胞增生。　　The invention includes a method of treating PAH, or a disorder disclosed herein, where the method includes using palbociclib as monotherapy or also in a combination therapy in which a patient in need of treatment is administered palbociclib in combination with one or more drugs (referred to also as active agent) approved for the treatment of PAH, for the treatment of a PAH associated condition, or for the treatment of a disorder disclosed herein, or as combination thereof.[0045]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 本發明包括治療PAH或本文所揭示之病症的方法，其中該方法包括使用帕博西尼作為單一療法或者亦將帕博西尼用於組合療法中，該組合療法中係投予有需要治療之患者帕博西尼組合上一或多種藥物(亦稱為活性劑)，該一或多種藥物係經核准用於治療PAH、用於治療與PAH相關之病症、或用於治療本文所揭示之病症、或彼等之組合。For example, an additional active agent may include but is not limited to a prostaglandin (eg, epoprostenol, treprostinil, iloprost, selexipag), an endothelin receptor antagonist (eg, bosentan, ambrisentan, macitentan), a guanylate cyclase inhibitor (eg, riociguat), vasodilators (eg, prostacyclin and sildenafil), calcium channel blockers (eg, amlodipine, diltiazem, and nifedipine);例如，另外之活性劑可包括，但不限於前列腺素(例如依前列醇(epoprostenol)、曲前列素(treprostinil)、伊洛前列素(iloprost)、西利前列素(selexipag))、內皮素受體拮抗劑(例如波生坦、安立生坦(ambrisentan)、馬西替坦(macitentan))、鳥苷酸環化酶抑制劑(例如利奥西呱)、血管擴張劑(例如前列環素和西地那非)、鈣通道阻斷劑(例如胺氯地平、地爾硫卓和硝苯地平)；抗凝劑(例如華法林)和利尿劑。提及例如西地那非之藥物時係包括西地那非和所有醫藥上可接受之鹽，例如西地那非檸檬酸鹽。　　In another aspect, palbociclib is used in the manufacture of a medicament for the treatment or prevention of PAH.[0046]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 於另一態樣中，帕博西尼係用於製造用於治療或預防PAH之藥物。再於另一態樣中，帕博西尼係用於製造治療或預防如本文所討論之相關疾病的藥物。於各種實施態樣中，該藥物係經調配成用於經由口服投予，包括立即釋出和持續釋出之藥物調製劑。於其他實施態樣中，該藥物係經調配成用於經由吸入投予。在所有這些實施態樣中，本發明提供該藥物之單位劑量形式。　　The present invention also includes isotopically labelled compounds, which are identical to palbociclib , but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature.[0047]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 本發明亦包括經同位素標記之化合物，該經同位素標記之化合物與帕博西尼相同，除了有一或多個原子被具有與自然界中常見之原子質量或質量數不同的原子質量或質量數之原子所替代。可被併入本發明化合物之同位素的實例包括氫、碳、氮、氧、磷、硫、氟和氯之同位素，諸如分別為² H、³ H、¹³ C、¹¹ C、¹⁴ C、¹⁵ N、¹⁸ O、¹⁷ O、³¹ P、³² P、³⁵ S、¹⁸ F和³⁶ Cl。含有上述同位素及/或其他原子之其他同位素的本發明化合物、其前藥及該化合物或該前藥之醫藥上可接受之鹽係在本發明之範圍內。本發明之某些經同位素標記之化合物，例如那些被併入放射性同位素(諸如³ H 和¹⁴ C)的化合物乃有用於藥物及/或受質組織分佈分析。氚化(即³ H)和碳-14(即¹⁴ C)同位素為特佳者，因其容易製備和檢測。此外，以較重之同位素的取代，諸如氘(即² H)可因代謝穩定性較高而提供某些治療優勢，例如體內半衰期增加或劑量需求降低，因此，在某些情況下為較佳者。經同位素標記之帕博西尼及其前藥通常係藉由進行下文中所揭示之方案及/或實施例和製備方法中之程序來製備，但以易於取得之經同位素標記的試劑來取代非經同位素標記之試劑。　　[0048]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 當本文中提及帕博西尼時，其包括6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2基]胺基}吡啶並[2,3-d]嘧啶-7(8H)-酮或形成包含鹽之醫藥上可接受之調配劑之其醫藥上可接受之鹽，該包含鹽之醫藥上可接受之調配劑包括，但不限於帕博西尼之酸加成鹽、溶劑化物和N-氧化物。參見 WO 03/062236 (其以引用方式併入本文)。　　[0049]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 如本文所使用之術語“個體”係指人、靈長類動物、伴侶動物(包括貓或狗)。術語“個體”包括患者。　　[0050]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 如本文所使用之術語“治療” 係指逆轉、減輕、抑制進展或預防該術語適用之失調或病症、或預防該等病症或失調的一或多種症狀。如本文所使用之術語“治療”係指治療的行為，如上句所定義之“治療”。　　[0051]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 帕博西尼可調配成廣泛多種口服和腸胃道外劑型並投予(包括經皮和直腸投予)。本技藝之技術熟習人士將理解下列劑型可包含帕博西尼或帕博西尼之對應的醫藥上可接受之鹽或溶劑化物作為活性成分。　　[0052]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 本發明亦包含醫藥調製劑，該醫藥調製劑包含治療有效量之帕博西尼與用醫藥上可接受之載體、稀釋劑或賦形劑。為了製備具有本發明化合物之醫藥組成物，醫藥上可接受之載體可為固體或液體。固體形式之製劑包括粉劑、片劑、丸劑、膠囊、扁囊劑、栓劑和可分散顆粒劑。固體載體可為亦可作為稀釋劑、調味劑、黏合劑、防腐劑、片劑崩解劑、或包封材料之一或多種物質。在片劑劑型方面，根據劑量，帕博西尼可構成該劑型之1wt%至80wt%，通常為該劑型之5wt%至60wt%，更典型為約10wt%至約35wt%，或再更典型為約15wt%至約25wt%。於具體之實施態樣中，帕博西尼包含該劑型(按重量計)之約20wt%。　　[0053]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 於本發明之固體劑型中，該載體可包含各種醫藥上可接受之賦形劑，包括，例如稀釋劑、崩解劑、黏合劑、潤滑劑、助流劑和表面活性劑。調製劑亦可包括賦形劑，諸如防腐劑、抗氧化劑、調味劑和著色劑，及本技藝已知之其他賦形劑。　　[0054]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 固體劑型，諸如片劑，通常含有稀釋劑，例如乳糖(單水合物、噴霧乾燥之單水合物、無水物，等)、甘露醇、木糖醇、右旋糖、蔗糖、山梨醇、可壓縮之糖、微晶型纖維素、粉狀纖維素、澱粉、預膠化之澱粉、葡萄糖結合劑(dextrate)、葡聚醣、糊精、右旋糖、麥芽糖糊精、碳酸鈣、磷酸氫鈣、磷酸鈣、硫酸鈣、碳酸鎂、氧化鎂、泊洛沙姆、聚環氧乙烷、羥丙基甲基纖維素及彼等之混合物。不同類型之微晶型纖維素可能適合用於本文所描述之調製劑中。微晶型纖維素之實例包括Avicel®類型：PH101、PH102、PH103、PH105、PH112、PH113、PH200、PH301和其他類型之微晶型纖維素，諸如矽化之微晶型纖維素(SMCC)。於一些實施態樣中，該稀釋劑係選自由下列所組成之群組：微晶型纖維素、乳糖一水合物、甘露醇、山梨醇、木糖醇、碳酸鎂、磷酸氫鈣、磷酸鈣、或彼等之混合物。於某些實施態樣中，該稀釋劑包含微晶型纖維素。於一些實施態樣中，該稀釋劑包含一或多種類型之微晶型纖維素，例如Avicel®PH105、Avicel®PH200或彼等之混合物。於一些該等實施態樣中，該稀釋劑不包括乳糖一水合物。於其他該等實施態樣中，該稀釋劑包含微晶型纖維素，且進一步包含乳糖一水合物。稀釋劑常佔該固體劑型之約25wt%至約75wt%，且較佳地，該劑型之約50wt%至約75wt%。　　[0055]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 固體劑型常含有崩解劑。崩解劑之實例包括澱粉羥乙酸鈉、羧甲基纖維素鈉、羧甲基纖維素鈣、交聯羧甲基纖維素鈉、交聯聚維酮、聚乙烯吡咯啶酮、甲基纖維素、微晶型纖維素、低級烷基取代之羥丙基纖維素、澱粉、預膠化之澱粉和藻酸鈉。於一些實施態樣中，該崩解劑為交聯聚維酮。任何等級之交聯聚維酮均可使用；例如CL、CL-SF和XL等級之交聯聚維酮適合用於本文所描述之調製劑中。具體實例包括Kollidon、Kollidon CL®、Kollidon CL-M®、Polyplasdone XL®、Polyplasdone XL-10®和Polyplasdone INF-10®。於一些實施態樣中，該載體包含至少一種選自由下列所組成之群組的崩解劑：交聯聚維酮、交聯羧甲基纖維素鈉和澱粉羥乙酸鈉。於特定之實施態樣中，該崩解劑為交聯聚維酮。崩解劑常佔該劑型之約1wt%至約25wt%，較佳為約5wt%至約20wt%，更佳為約5wt%至約10wt%。　　[0056]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 黏合劑可用於賦予片劑調製劑黏合(cohesive)性質。合適之黏合劑包括微晶型纖維素、明膠、糖、聚乙二醇、天然和合成樹膠、聚乙烯吡咯啶酮、預膠化之澱粉、羥丙基纖維素及羥丙基甲基纖維素。於一些實施態樣中，該黏合劑係選自由下列所組成之群組：微晶型纖維素、羥丙基纖維素和羥丙基甲基纖維素。於特定之實施態樣中，該黏合劑為微晶型纖維素，例如Avicel®PH105。當存在時，黏合劑可包含該劑型之約0wt%至約15wt%、或約0.2wt%至約10wt%。於一些實施態樣中，該黏合劑包含該劑型之約5wt%至約10wt%。於特定之實施態樣中，該黏合劑包含該劑型之約10wt%。　　[0057]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 固體劑型常含有一或多種潤滑劑。潤滑劑之實例包括硬脂酸鎂、硬脂酸鈣、硬脂酸鋅、硬脂富馬酸鈉、硬脂酸鎂與十二烷基硫酸鈉之混合物、或二或更多種該等物質之混合物。於一些實施態樣中，該潤滑劑為硬脂酸鎂及/或硬脂富馬酸鈉。於一些實施態樣中，該潤滑劑為硬脂酸鎂。於一些該等實施態樣中，該固體劑型為包含顆粒內和顆粒外硬脂酸鎂之片劑。於其他實施態樣中，該固體劑型為包含顆粒內硬脂酸鎂和顆粒外硬脂富馬酸鈉之片劑。當存在時，潤滑劑常包含該劑型之約0.25wt%至約10wt%，較佳為約0.5wt%至約6wt%。　　[0058]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 片劑亦可能包含助流劑，例如二氧化矽、膠態二氧化矽、矽酸鎂、三矽酸鎂、滑石及其他形式之二氧化矽，諸如聚集之矽酸鹽和水合二氧化矽。於一些實施態樣中，該助流劑為二氧化矽。當存在時，助流劑可佔該片劑之約0wt%至約10wt%，較佳為約0.2wt%至約5wt%，或約0.5wt%至約2wt%。　　[0059]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 片劑可視需要地包含表面活性劑，諸如月桂基硫酸鈉和聚山梨醇酯80。當存在時，表面活性劑可佔該片劑之0wt%至10wt%，或較佳為0.2wt%至5wt%。　　[0060]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 一般而言，本發明之固體劑型係根據製藥化學中常用之方法製備。選定之賦形劑可與該活性藥物成分一起被併入顆粒外或顆粒內隔室其中一者或二者。　　[0061]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 帕博西尼之治療有效劑量將在每天約0.01mg/kg體重至約100mg/kg體重之間變化。典型之成人劑量將為每天約0.1 mg至約3000 mg。通常，該帕博西尼之治療有效的成人劑量為每天75mg、100mg或125mg。根據該特定應用，該單位劑量製劑中之活性組分的量可在約0.1mg至約500mg，較佳為約25mg至約125mg之間變化或調整。若需要時，該組成物亦可含有其他相容之治療劑。需要以帕博西尼治療之個體的投予劑量為每天約0.6至約500mg，且較佳為每天約25mg至125mg，該劑量或是在單一劑量中投予或是在24小時之期間內在數個劑量中投予。該等治療可在連續的間隔內重複進行，只要有需要。化驗法實施例實 Example 1 施例 1 [0062]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .細胞增生化驗法：將來自正常人類個體之初代PASMC和PAAF(ScienCell研究實驗室，美國加州聖地牙哥)和大鼠之初代PASMC(Cell Applications，美國加州聖地牙哥)平皿接種在CellBIND^® 96孔盤中之含有5%胎牛血清(FBS)之Dulbecco氏修飾之Eagle培養基(DMEM)中，及在含有0.1%FBS之DMEM中血清飢餓24小時。首先將細胞與不同濃度之帕博西尼預培育30分鐘(一式三份)，然後加入30 ng/ml PDGF-BB或5%FBS以刺激細胞增生。將細胞保持48小時再使用細胞增生ELISA BrdU(Cell Proliferation ELISA BrdU)(溴脫氧尿苷)發光套組(Roche Diagnostics公司，德國曼海姆)評估增生速率。在培養基中加入BrdU，16小時後再定量併入之BrdU。使用此程序來產生如至少呈現在第1a、1b、1c和1d圖中之數據。藉由測定抑制50%反應時需要施加至該系統之化合物的濃度來計算任何指定之抑制劑的IC₅₀ 值。IC₅₀ 值係從擬合至該化合物濃度反應數據的四參數邏輯計算方程式導出。　　[0063]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Current methods of treating PAH focus on reducing symptoms and prolonging patient lifespan and enhancing quality of life. 或者，不使用BrdU，可使用EdU(5-乙炔基-2'-脫氧尿苷)。首先將細胞與不同濃度之CDK抑制劑預溫育30分鐘，然後加入5%FBS以刺激細胞增生。將細胞保持24小時再使用EDU Click-iT EdU Alexa Fluor^® 647 HCS化驗法(Thermofisher Scientific，美國)評估增生速率。在培養基中加入EdU，16小時後再定量併入之EdU。使用配備適當之螢光顯微過濾器的自動化高含量成像平台來掃描96孔盤以定量增生，該螢光顯微過濾器係用於作為Click-iT化驗法之一部分的DAPI(4',6-二脒基-2-苯基吲哚)或Hoechst染色和Cyanine5染料。定量從Cyanine5檢測之增生的EdU陽性細胞並對每孔之總細胞數標準化，其中該每孔之總細胞數係藉由DAPI或Hoechst 信號鑑定。然後計算%作用：[0064]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 每一CDK抑制劑濃度之平均(avg)增生細胞量(amt)為有FBS下。所使用之CDK抑制劑為帕博西尼、艾貝馬西尼、地納西尼和利奥西呱。　　[0065]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 最小增生量(min amt of prolif'n)係基於其中未添加FBS且無CDK抑制劑存在之孔中的細胞數。　　[0066]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 最大增生量(maxn amt of prolif'n)係基於其中添加FBS且無CDK抑制劑存在之孔中的細胞數。　　[0067]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 此程序係用來產生如至少呈現在第1e、1f和1g圖中之數據。　　[0068]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .細胞內西方分析 (In-cell western analysis) ：依上述培養來自正常人個體之初代PASMC及令其血清飢餓。首先將細胞與不同濃度之帕博西尼預溫育60分鐘，然後加入5% FBS或10% FBS或30ng/ml PDGF-BB以刺激細胞增生和Rb磷酸化。24小時後，根據製造商之說明書(Li-Cor Biosciences公司，美國內布拉斯加州林肯市)，使用細胞內西方分析評估帕博西尼對Rb磷酸化之效果。使用兔子單株抗體抗磷酸-Rb(anti-phospho-Rb) (Ser807/811) (Cell Signaling Technology公司，美國麻薩諸塞州丹佛市)測量磷酸化Rb，接著以經IRDye 800CW紅外螢光染料標記之山羊抗兔子二級抗體檢測之。使用小鼠抗肌動蛋白單株抗體(pan Ab-5)(Fisher Scientific公司，美國賓州匹茲堡；Sigma Aldrich公司，美國密蘇里州聖路易斯)測量肌動蛋白之量，隨後以經IRDye 680RD標記之山羊抗小鼠二級抗體檢測之。參見其中使用5% FBS之第2a圖、其中使用PDGF-BB之第2b圖及其中使用10% FBS之第2c圖。實施例2 　　[0069]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3- d ]pyrimidin-7(8 H )-one . MCT 誘導之大鼠 PAH 模型：在第0天，雄性Sprague-Dawley大鼠(Charles River實驗室)(260-280g；載劑對照組，n=6；所有其他組，n=15)接受溶解於載劑(33.3% 1N HCl，14.8% 1N NaOH，52.2% diH₂ O)中之MCT(50mg/kg，SC；1ml/kg BW)或單獨之載劑。或是在第0-28天預防性地投予(5ml/kg；PO)西地那非或是在第14-28天治療性地投予西地那非(50mg/kg/劑量；BID)。治療性地投予帕博西尼(100mg/kg；QD)(第14-28天)。將西地那非和帕博西尼二者再懸浮於載劑(0.5%甲基纖維素)中並經由口服投予(5ml/kg)。第28天，經由異氟烷麻醉動物，評估血流動力學參數並在安樂死後收集組織。實施例3 　　[0070]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3- d ]pyrimidin-7(8 H )-one . 由 MCT 誘導之大鼠 PAH 模型：在第0天，雄性Sprague-Dawley大鼠(Charles River實驗室)(260-280g；載劑對照組，n=6；所有其他組，n=15)接受溶解於載劑(33.3% 1N HCl，14.8% 1N NaOH，52.2% diH₂ O)中之MCT(50mg/kg，SC；1ml/kg BW)或單獨之載劑。在第0-28天預防性地投予(5ml/kg；PO)西地那非或是在第14-28天治療性地投予西地那非(50mg/kg/劑量；BID)。治療性地投予帕博西尼(100mg/kg；QD)(第14-28天)。將西地那非和帕博西尼二者再懸浮於載劑(0.5%甲基纖維素)中並經由口服投予(5ml/kg)。第28天，經由異氟烷麻醉動物，評估血流動力學參數並在安樂死後收集組織。　　[0071]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3- d ]pyrimidin-7(8 H )-one . 血流動力學測量 - 野百合鹼 (MCT) 研究：第28天，在正壓通氣機中經由2%異氟烷將動物麻醉。使用經預校準之壓力傳感器測量在異氟烷正壓通氣中之動物的肺血流動力。使用經由右頸動脈引至主動脈弓高度之肺動脈幹的Millar Solid State Mikro尖端導管(Millar儀器，德州休斯頓)測量穩態肺動脈壓和全身動脈壓。藉由生理數據採集系統，ADI Chart(ADI儀器，科羅拉多州科羅拉多斯普林斯)(柏拉圖生物製藥) 自動計算血流動力學數值。實施例4 　　[0072]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .由 SuHx 誘導之肺動脈高血壓 ：自Charles River實驗室取得Sprague-Dawley大鼠(200-300g；載劑對照組，n=5；所有其他組，n=10)。在第0天，大鼠接受溶解在DMSO中之塞美西尼，接著被飼養在低氧(13%)下，直到第28天。在第7-28天投予在載劑(0.5%甲基纖維素)中之西地那非(30mg/kg/劑量；BID)、利奥西呱(10mg/kg，QD)或帕博西尼(100mg/kg；QD)。在第28天，依類似於MCT研究之方式進行終端血流動力學評估和屍檢。　　[0073]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .血流動力學測量 Su/Hx 研究：在第28天，經由肌肉內注射氯胺酮/賽拉嗪(80/10 mg/kg)將動物麻醉。將Millar導管插入肺動脈並依Stringeret al ., (1981) Catheterization of the pulmonary artery in the closed-chest rat.J. Appl. Physiol. 51, 1047-1050之描述測量肺壓力和血流動力學。使用NOTOCORD-Hem軟體(NOTOCORD公司，法國)(CorDynamics)計算血流動力學值。　　[0074]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .測量 RV 肥大：在屍檢時採集心臟後，移出心房和大血管。將RV與LV和S分開。測量RV和LV+S之濕重以計算富爾頓指數。實施例5 　　[0075]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .組織製備方法：在麻醉下，將動物放血並以氧化(來自實施例3之動物[MCT])或肝素化(來自實施例4之動物[Su5416/Hx])鹽水沖洗肺循環。從胸腔完好取出氣管、肺和心臟並放置在冰冷的鹽水中。移出右肺葉，稱重並立即在液態氮中快速冷凍。使用充填固定劑(10%NBF)之具有附接之鈍端針(20g)的10mL注射器將左肺葉充氣並將氣道打結以保持肺葉持續擴張。然後，將左肺浸入10%NBF中固定≤48小時，再轉移到70%乙醇中≤5天。　　[0076]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .組織形態計量分析：使用三個3-5μm厚之左肺葉橫截面進行組織形態計量學分析。由委員會認證之獸醫病理學家藉由光學顯微鏡評估經蘇木精和伊紅(H＆E)染色之肺切片來確定整體疾病嚴重性評分。評分範圍係從0(無)、1(最小)、2(輕度)、3(中度)、4(明顯)至5(重度)並根據受PAH模型之組織變化(炎性、退行性、血管及/或增生)特徵影響之組織的近似百分比分配。使用以經修飾之Verhoeff-van Gieson彈性蛋白染色製備之肺切片評估平均血管壁厚度。利用隨機場域採樣(Visiopharm newCAST)加觀察者輔助識別肺泡內和前肺泡血管來採樣10%之組織面積進行圖像分析並應用核探頭測量血管壁之厚度。使用經雙αSMA/Ki67 IHC染色之切片及觀察者輔助計算≥70%肌化(即，SMA⁺ )、橫向切片、≤50μm外徑、具有可識別腔之肺泡內血管，以類似方式評估充分肌化之肺泡內血管密度。使用隨機場域採樣之10%組織面積及觀察者輔助識別與外徑≤100μm之血管壁(即，在內膜或中層)相關之Ki67⁺ 或pRb⁺ 細胞分別計算在經αSMA/Ki67 IHC和CD31/pRb IHC染色之肺切片中之血管相關的Ki67⁺ 和pRb⁺ 細胞。　　[0077]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .免疫組織化學，雙 α-SMA/Ki67 核抗原 ：在Leica Bond Rx自動免疫組織化學染色器上，在3μm厚，經福馬林固定之石蠟包埋的(FFPE)切片上進行所有IHC步驟。該儀器經過編程以烘烤並將玻片脫蠟。將該初級抗體，抗人αSMA(兔子多株抗體ab5694，Abcam公司，麻薩諸塞州劍橋)在Bond^TM 初級抗體稀釋劑(PV6123)中稀釋200倍(1:200)，施加在玻片上，培育30分鐘並使用Bond^TM Polymer Refine AP-Red檢測套組(DS9390，Leica Biosystems公司，伊利諾州Buffalo Grove)檢測。此步驟後接著使用Bond^TM Epitope Retrieval 1(ER1)溶液(AR9961，Leica Biosystems)進行20分鐘熱誘導之抗原決定部位修復。施用二級抗體，抗Ki67(小鼠單株抗體Ki67 [MM1]核抗原RTU，PA0118 [Leica])15分鐘，再以Bond^TM Polymer Refine HRP DAB檢測系統(DS9800)檢測。將玻片複染、脫水並蓋上蓋玻片。　　[0078]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .雙 CD31/ 磷酸 Rb ：該初始步驟與上述相同。以Bond^TM Epitope Retrieval 2(ER2)溶液(AR9640)進行20分鐘之熱誘導的抗原決定部位修復。將該初級抗體，抗磷酸-Rb(兔子多株磷酸-Rb[ser807/811]，9308，Cell Signaling Technology公司，麻薩諸塞州丹佛斯)在Bond^TM 初級抗體稀釋劑中稀釋600倍(1:600)，施加在玻片上並培育30分鐘。接著使用針對初級抗體之Bond^TM Polymer HRP-DAB Refine檢測套組。此步驟之後，使用Bond^TM Epitope Retrieval 1(ER1)溶液(AR9961，Leica Biosystems)進行10分鐘熱誘導之抗原決定部位修復。施加二級抗體，抗CD31(兔子單株抗體[EPR3094]，ab76533，Abcam公司，麻薩諸塞州劍橋)45分鐘，然後以Bond^TM Polymer Refine AP-Red檢測套組(DS9390，Leica Biosystems)檢測。將玻片複染、脫水並蓋上蓋玻片。Statistical Analysis 統計分析 [0079]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one . 以平均值±SEM呈現數據。先使用單因方差分析(ANOVA)，再使用Tukey多重比較檢驗(GraphPad Prism 6，GraphPad軟體，美國加州聖地牙哥)進行組間的統計分析。設定之統計顯著性水準為經多重調節之P 值為0.05 (標記為或*)或0.01(標記為或**)。

[0012]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Current methods of treating PAH focus on reducing symptoms and prolonging patient lifespan and enhancing quality of life.The present invention also includes a kit for the treatment or prevention of pulmonary arterial hypertension.FIG . 第 1 圖顯示1 ………..細胞增生：以經系列稀釋之帕博西尼處理細胞後，進行BrdU併入分析以測量細胞DNA合成率和增生。該劑量-反應曲線顯示帕博西尼有效抑制人PASMC(第1a和1b圖)、人PAAF(第1c圖)、大鼠PASMC(第1d圖)之增生。以經系列稀釋之帕博西尼或另一已知之CDK抑制劑處理細胞後，進行EdU併入分析以測量細胞DNA合成率和增生。該劑量-反應曲線顯示人PASMC(第1e圖)、人PAAF(第1f圖)和大鼠PASMC(第1g圖)之增生受抑制。( FIG. 1e), human PAAF (FIG. 1f ), and rat PASMCs (FIG. 1 g ) . 　　FIG .[0013]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Current methods of treating PAH focus on reducing symptoms and prolonging patient lifespan and enhancing quality of life.The present invention also includes a kit for the treatment or prevention of pulmonary arterial hypertension. 第 2 圖 2 .顯示Rb磷酸化：帕博西尼對視網膜母細胞瘤基因產物磷酸化之作用。帕博西尼有效地阻斷人初代PASMC中由5%FBS(第2a圖)或PDGF-BB(( FIG. 2 b ) in human primary PASMCs, while it had little effect on total Rb levels.((((((99(((((第2b圖)誘導之在絲胺酸807和811的Rb磷酸化(磷酸-Rb)，同時其對總Rb量幾乎沒有作用。已知CDK抑制劑抑制由10%FBS誘導之人初代PASMC中之磷酸-Rb(第2c圖)。　　FIG .[0014]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .Current methods of treating PAH focus on reducing symptoms and prolonging patient lifespan and enhancing quality of life.第 3 圖顯示顯顯顯示H emodynamic and structural changes associated with PAH pathology in th e rat monocrotaline (MCT) model .大鼠野百合鹼(monocrotaline) (MCT)模型中與PAH病理學相關之血流動力學和結構變化。第3a圖顯示研究藥物之投予頻率。實驗結果係以平均肺動脈壓(MPAP；第3b圖)、肺動脈收縮壓(SPAP；第3h圖)3h ), and diastolic pulmonary arterial pressure和肺動脈舒張壓(DPAP ; FIG. 3i ) ,(DPAP；第3i圖)、富爾頓指數(Fulton's index)(RV/(LV+S)；第3c圖)、血管壁厚度(第3d圖)、肌化之肺泡內血管密度(Muscularized intra-acinar vessel density)(第3e圖)、組織病理學(第3f圖)、疾病嚴重性評分(第3g圖)和心臟重量(第3j圖)顯示。　　FIG .[0015]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .第 4 圖顯示顯顯顯H emodynamic and structural changes associated with PAH pathology in th e rat monocrotaline (MCT) model .大鼠SuHx PAH模型中之肺血管之血流動力學變化和明顯的結構性改善。第4a圖顯示研究之藥物的投予頻率(第4a圖)。實驗結果係以平均肺動脈壓(MPAP；第4b圖)、SPAP(第4h圖)3h ), and diastolic pulmonary arterial pressure、DPAP(第4i圖)、富爾頓指數(RV/(LV + S)；第4c圖)、血管壁厚度(第4d圖)、肌化之肺泡內血管密度(第4e圖)、組織病理學(第4f圖)、疾病嚴重性評分(第4g圖)和心臟重量(第4j圖)顯示。　　[0016]Pulmonary hypertension and related diseases , like pulmonary arterial hypertension, can be trea ted by administering an effective dose of a CDK inhibitor, including p albociclib, 6-acetyl-8-cyclopentyl-5-methyl-2-{[5-(piperazin-1-yl)pyridin-2yl]amino}pyrido[2,3-d ]pyrimidin-7(8H )-one .第 5 圖顯示顯示顯顯在FIG .肺脈管系統(pulmonary vasculature)中評估之細胞增生。大鼠肺切片中之IHC of dual stained雙重染色之αSMA蛋白/ Ki67(F IG. 5a) and CD31/ phosphoRb (F IG. 5b) in rat lung sections . (第5a圖)和CD31/磷酸Rb(第5b圖)的IHC。實驗結果ppRb；以Ki67(第5c圖)和血管相關之視網膜母細胞瘤蛋白磷酸化(第5d圖)顯示。

Claims

一種使用6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2基]胺基}吡啶並[2,3-d]嘧啶-7(8H)-酮或其醫藥上可接受之鹽於製造治療肺動脈高血壓(PAH)之藥物的用途。
如申請專利範圍第1項之用途，其中係在24小時之期間內單次地或是在多個劑量中投予劑量為每天約0.6至約500mg之6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2基]胺基}吡啶並[2,3-d]嘧啶-7(8H)-酮或其醫藥上可接受之鹽。
如申請專利範圍第1項之用途，其中係在24小時之期間內單次地或是在多個劑量中投予劑量為每天約25至125mg之6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2基]胺基}吡啶並[2,3-d]嘧啶-7(8H)-酮或其醫藥上可接受之鹽。
如申請專利範圍第1至3項中任一項之用途，其中該藥物除了包括6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2基]胺基}吡啶並[2,3-d]嘧啶-7(8H)-酮或其醫藥上可接受之鹽還包括西地那非(sildenafil)。
如申請專利範圍第4項之用途，其中係將6-乙醯基-8-環戊基-5-甲基-2-{[5-(六氫吡嗪-1-基)吡啶-2基]胺基}吡啶並[2,3-d]嘧啶-7(8H)-酮使用於製造治療肺動脈高血壓(PAH)之藥物。