The molecular mechanism of HIV-1's drug resistance against protease inhibitors including darunavir and the development of novel resistance-repelling protease inhibitors

• Project/Area Number: 26293239
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Partial Multi-year Fund
• Section: 一般
• Research Field: Infectious disease medicine
• Research Institution: Kumamoto University
• Principal Investigator: MITSUYA Hiroaki 熊本大学, 医学部附属病院, 特別招聘教授 (20136724)
• Project Period (FY): 2014-04-01 – 2017-03-31
• Project Status: Completed (Fiscal Year 2016)
• Budget Amount: ¥16,770,000 (Direct Cost: ¥12,900,000、Indirect Cost: ¥3,870,000); Fiscal Year 2016: ¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000); Fiscal Year 2015: ¥4,810,000 (Direct Cost: ¥3,700,000、Indirect Cost: ¥1,110,000); Fiscal Year 2014: ¥7,150,000 (Direct Cost: ¥5,500,000、Indirect Cost: ¥1,650,000)
• Keywords: HIV-1 / 新薬開発 / プロテアーゼ阻害剤 / 構造解析 / エイズ / 新規抗HIV剤 / 相互作用解析 / 結晶構造解析 / 新規抗HIV-1剤開発 / 薬剤耐性 / 薬剤開発 / 薬剤耐性メカニズム

Outline of Final Research Achievements

We identified newly generated novel protease inhibitors (GRL-058, -059, -077, -078, -079, -010 and -014), which were designed based on the structural, pharmaceutical and anti-HIV activity features of GRL-085 and KU-241 (Aoki & Mitsuya, J Virol. 90, 2180-2194, 2016). We also determined the anti-HIV activity and crystal structures of these derivatives. These compounds exerted potent activity with IC50 values of 10-11 to 10-8 molar concentrations against wild type HIV-1 (HIVWT) and darunavir (DRV)-resistant HIV-1 variants (HIVDRVRs). We also identified the chemical moieties, which confer the potent activity against HIVDRVRs on some compounds through forming strong halogen and hydrogen bonds with amino acids located at the flap region of protese. Additionally, we demonstrated that such compounds had higher genetic barrier than DRV using in vitro selection assay for inducing drug resistant HIV-1 strains.

Research Products

• [Journal Article] Probing Lipophilic Adamantyl Group as the P1-Ligand for HIV-1 Protease Inhibitors: Design, Synthesis, Protein X-ray Structural Studies, and Biological Evaluation. (2016)
  • Author(s): Ghosh AK, Osswald HL, Glauninger K, Agniswamy J, Wang YF, Hayashi H, Aoki M, Weber IT, Mitsuya H
  • Journal Title: J Med Chem. Volume: 59(14) Issue: 14 Pages: 6826-37
  • DOI: 10.1021/acs.jmedchem.6b00639
  • Peer Reviewed
• [Journal Article] Development and validation of a cell-based assay system to assess human immunodeficiency virus type 1 integrase multimerization. (2016)
  • Author(s): Nakamura T, Campbell JR, Moore AR, Otsu S, Aikawa H, Tamamura H, Mitsuya H
  • Journal Title: J Virol Methods. Volume: 236 Pages: 196-206
  • DOI: 10.1016/j.jviromet.2016.07.023
  • Peer Reviewed
• [Journal Article] Structural basis of HIV inhibition by translocation-defective RT inhibitor 4'-ethynyl-2-fluoro-2'-deoxyadenosine(EFdA). (2016)
  • Author(s): Salie ZL, Kirby KA, Michailidis E, Marchand B, Singh K, Rohan LC, Kodama EN, Mitsuya H, Parniak MA, Sarafianos SG.
  • Journal Title: Proc Natl Acad Sci U S A. Volume: 113 Issue: 33 Pages: 9274-9276
  • DOI: 10.1073/pnas.1605223113
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] A Modified P1 Moiety Enhances In Vitro Antiviral Activity against Various Multidrug-Resistant HIV-1 Variants and In Vitro Central Nervous System Penetration Properties of a Novel Nonpeptidic Protease Inhibitor, GRL-10413. (2016)
  • Author(s): Amano M, Salcedo-Gomez PM, Zhao R, Yedidi RS, Das D, Bulut H, Delino NS, Sheri VR, Ghosh AK, Mitsuya H
  • Journal Title: Antimicrob Agents Chemother. Volume: 60(12) Pages: 7046-7059
  • Peer Reviewed
• [Journal Article] C-5-Modified Tetrahydropyrano-Tetrahydofuran-Derived Protease Inhibitors (PIs) Exert Potent Inhibition of the Replication of HIV-1 Variants Highly Resistant to Various PIs, including Darunavir (2015)
  • Author(s): Aoki, M. Hayashi, H. Yedidi, R. S. Martyr, C. D. Takamatsu, Y. Aoki-Ogata, H. Nakamura, T. Nakata, H. Das, D. Yamagata, Y. Ghosh, A. K. Mitsuya, H.
  • Journal Title: Journal of Virology Volume: 90 Issue: 5 Pages: 2180-94
  • DOI: 10.1128/jvi.01829-15
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Design of HIV-1 Protease Inhibitors with Amino-bis-tetrahydrofuran Derivatives as P2-Ligands to Enhance Backbone-Binding Interactions: Synthesis, Biological Evaluation, and Protein–Ligand X-ray Studies. (2015)
  • Author(s): Ghosh, AK., Martyr, CD., Osswald, HL., Sheri, VR., Kassekert, LA., Chen, S., Agniswamy, J., Wang, YF., Hayashi, H., Aoki, M., Weber, IT., and Mitsuya, H.
  • Journal Title: J Med Chem Volume: 58 Issue: 17 Pages: 6994-7006
  • DOI: 10.1021/acs.jmedchem.5b00900
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] A NOVEL TRICYCLIC LIGAND-CONTAINING NONPEPTIDIC HIV-1 PROTEASE INHIBITOR, GRL-0739, EFFECTIVELY INHIBITS THE REPLICATION OF MULTI-DRUG-RESISTANT HIV-1 VARIANTS AND HAS A DESIRABLE CENTRAL NERVOUS SYSTEM (CNS) PENETRATION PROPERTY IN VITRO (2015)
  • Author(s): Masayuki Amano, Yasushi Tojo, Pedro Miguel Salcedo-Gómez, Garth L. Parham, Prasanth R. Nyalapatla, Debananda Das, Arun K. Ghosh and Hiroaki Mitsuya
  • Journal Title: Antimicrobial Agents and Chemotherapy Volume: 59 Issue: 5 Pages: 2625-2635
  • DOI: 10.1128/aac.04757-14
  • Peer Reviewed / Open Access
• [Journal Article] Structure-based design, synthesis, X-ray studies, and biological evaluation of novel HIV-1 protease inhibitors containing isophthalamide-derived P2-ligands. (2015)
  • Author(s): Arun K. Ghosh, Jun Takayama, Luke A. Kassekert, Jean-Rene Ella-Menye, Sofiya Yashchuk, Johnson Agniswamy, Yuan-Fang Wang, Manabu Aoki, Masayuki Amano, Irene T. Weber, Hiroaki Mitsuya.
  • Journal Title: Bioorg Med Chem Lett. Volume: 25 Issue: 21 Pages: 4903-4909
  • DOI: 10.1016/j.bmcl.2015.05.052
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Design of gem-Difluoro-bis-Tetrahydrofuran as P2 Ligand for HIV-1 Protease Inhibitors to Improve Brain Penetration: Synthesis, X-ray Studies, and Biological Evaluation. (2015)
  • Author(s): Arun K. Ghosh, Sofiya Yashchuk, Akira Mizuno, Nilanjana Chakraborty, Johnson Agniswamy, Yuan-Fang Wang, Manabu Aoki, Pedro Miguel Salcedo Gomez, Masayuki Amano, Irene T. Weber, Hiroaki Mitsuya.
  • Journal Title: ChemMedChem Volume: 10 Issue: 1 Pages: 107-115
  • DOI: 10.1002/cmdc.201402358
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Dimerization of HIV-1 protease occurs through two steps relating to the mechanism of protease dimerization inhibition by darunavir. (2014)
  • Author(s): Hayashi H, Takamune N, Nirasawa T, Aoki M, Morishita Y, Das D, Koh Y, Ghosh AK, Misumi S, Mitsuya H.
  • Journal Title: Proc. Natl. Acad. Sci. U S A. Volume: 111 Issue: 33 Pages: 12234-9
  • DOI: 10.1073/pnas.1400027111
  • Peer Reviewed / Open Access
• [Journal Article] Conserved hydrogen-bonding network of P2 bis-tetrahydrofuran containing HIV-1 protease inhibitors (PI) with protease active site amino acid-backbone aid in their activity against PI-resistant HIV. (2014)
  • Author(s): Yedidi RS, Garimella H, Aoki M, Aoki H, Desai DV, Chang SB, Davis DA, Fyvie WS, Kaufman JD, Smith DW, Das D, Wingfield PT, Maeda K, Ghosh AK and Mitsuya H.
  • Journal Title: Antimicrob Agents Chemother Volume: 58 Issue: 7 Pages: 3679-88
  • DOI: 10.1128/aac.00107-14
  • Peer Reviewed
• [Presentation] 4’-Modified NRTIs’ Potent Anti-HIV Activity Stems from Strong RT Active Site Binding. (2017)
  • Author(s): Takamatsu Y, Das D, Kohgo S, Hattori S, Hayashi H, Matsuda K, Stefan G. Sarafianos SG, Mitsuya H, and Maeda K.
  • Organizer: 2017 Conference on Retroviruses and Opportunistic Infections (CROI)
  • Place of Presentation: シアトル、アメリカ
  • Year and Date: 2017-02-12
• [Presentation] KU-241が前例を見ない程強力な抗HIV活性を発揮する機序の解析 (2016)
  • Author(s): 林 宏典, 青木 学, Debananda Das, 服部 真一朗, 青木 宏美, 鍬田 伸好, 高松 悠樹, Yedidi Ravikiran, 長谷川 和也, Arun K Ghosh and 満屋 裕明
  • Organizer: 第30回日本エイズ学会学術集会・総会
  • Place of Presentation: 鹿児島
  • Year and Date: 2016-11-24
• [Presentation] 表面プラズモン共鳴法を用いた新規HIV-1侵入阻害剤活性評価法の樹立 (2016)
  • Author(s): 松田 幸樹, 林 宏典, 原田 恵嘉, 服部 真一朗, 鳴海 哲夫, 玉村 啓和, 吉村 和久, 満屋 裕明, 前田 賢次
  • Organizer: 第30回日本エイズ学会学術集会・総会
  • Place of Presentation: 鹿児島
  • Year and Date: 2016-11-24
• [Presentation] DRV耐性株を含む種々の多剤耐性変異株にも有効な新規抗HIV薬候補化合物KU-241の体内動態・脳組織内濃度・2週間反復投与毒性 (2016)
  • Author(s): 鍬田 伸好, 青木 学, 服部 真一朗, 青木 宏美, 林 宏典, 前田 賢次, Arun K. Ghosh, 満屋 裕明
  • Organizer: 第30回日本エイズ学会学術集会・総会
  • Place of Presentation: 鹿児島
  • Year and Date: 2016-11-24
• [Presentation] HIV-1プロテアーゼのflap領域と相互作用する化合物はdarunavir高度耐性HIV-1を強力に阻害する (2016)
  • Author(s): 林 宏典, 青木 学, Yedidi Ravikiran, 中田 浩智, Debananda Das, 中村 照也, 長谷川 和也, 山縣 ゆり子, Arun K Ghosh, 満屋 裕明
  • Organizer: 第26回抗ウイルス療法学会総会
  • Place of Presentation: 名古屋
  • Year and Date: 2016-05-13
• [Presentation] 前例を見ない程強力な活性を有する新規HIV-1プロテアーゼ阻害剤KU-241の同定 (2016)
  • Author(s): 服部 真一朗, 青木 学, 鍬田 伸好, 青木 宏美, 林 宏典, Arun K. Ghosh, 前田 賢次, 満屋 裕明
  • Organizer: 第26回抗ウイルス療法学会総会
  • Place of Presentation: 名古屋
  • Year and Date: 2016-05-13
• [Presentation] 本邦におけるHIV感染者・AIDS発症者数の動向とARTの医療経済的解析 (2016)
  • Author(s): 尾又 一実, 菊池 嘉, 岡 慎一, 満屋 裕明. 本邦におけるHIV感染者・AIDS発症者数の動向とARTの医療経済的解析
  • Organizer: 第26回抗ウイルス療法学会総会
  • Place of Presentation: 名古屋
  • Year and Date: 2016-05-13
• [Presentation] Novel protease inhibitors, which bind to protease’s flap region, strongly inhibit multi-drug resistant HIV-1 variants including darunavir-resistant variants (2015)
  • Author(s): Hironori Hayashi、Manabu Aoki、Ravikiran S. Yedidi、Hirotomo Nakata、Debananda Das、Teruya NakamuraKazuya Hasegawa、Yuriko Yamagata、Arun K. Ghosh、Hiroaki Mitsuya
  • Organizer: 第29回日本エイズ学会学術集会・総会
  • Place of Presentation: 東京ドームホテル（東京都）
  • Year and Date: 2015-11-30
• [Presentation] In vitro evaluation of the genotoxic potential of the novel anti-HIV protease inhibitor, KU-241, which is potent against multi-PI-Resistant HIV-1 variants (2015)
  • Author(s): Nobuyo Higashi-Kuwata、Manabu Aoki1、Shinichiro Hattori、Hiromi Ogata-Aoki、Hironori Hayashi、Kenji Maeda、Arun K. Ghosh、Hiroaki Mitsuya
  • Organizer: 第29回日本エイズ学会学術集会・総会
  • Place of Presentation: 東京ドームホテル（東京都）
  • Year and Date: 2015-11-30
• [Presentation] 新規HIV-1 プロテアーゼ阻害剤KU-241 の同定と抗HIV 活性の検討 (2015)
  • Author(s): 服部 真一朗、青木 学、鍬田 伸好、青木 宏美、林 宏典、Arun K. Ghosh、前田 賢次、満屋 裕明
  • Organizer: 第29回日本エイズ学会学術集会・総会
  • Place of Presentation: 東京ドームホテル（東京都）
  • Year and Date: 2015-11-30
• [Presentation] Novel protease inhibitors, which bind to the flap region of HIV-1 protease, strongly inhibit multi-drug resistant HIV-1 variants including darunavir- resistant variants. (2015)
  • Author(s): Hironori Hayashi, Manabu Aoki, Ravikiran S. Yedidi, Hirotomo Nakata, Debananda Das, Teruya Nakamura, Kazuya Hasegawa, Yuriko Yamagata, Arun K. Ghosh and Hiroaki Mitsuya
  • Organizer: 第16回熊本エイズセミナー
  • Place of Presentation: 阿蘇ホテルグランビリオ（熊本県阿蘇市）
  • Year and Date: 2015-10-07
• [Presentation] Modified P1-Moiety Enhances Antiviral Activity against Drug-Resistant HIV-1 Variants and Central Nervous System (CNS) Penetration Effectiveness of Newly-Generated HIV-1 Protease Inhibitors In Vitro. (2015)
  • Author(s): Masayuki Amano and Hiroaki Mitsuya
  • Organizer: 第16回熊本エイズセミナー
  • Place of Presentation: 阿蘇ホテルグランビリオ（熊本県阿蘇市）
  • Year and Date: 2015-10-07
• [Presentation] 新規HIV-1プロテアーゼ阻害剤開発と構造解析 (2015)
  • Author(s): 林 宏典、青木 学、Yedidi Ravikiran、中田 浩智、Debananda Das、中村 照也、長谷川 和也、山縣 ゆり子、Arun K Ghosh、満屋 裕明
  • Organizer: 第17回白馬シンポジウム in 米子
  • Place of Presentation: 鳥取大学医学部（鳥取県米子市）
  • Year and Date: 2015-06-19
• [Presentation] HIV-1プロテアーゼ （PR） の二量体化は2ステップで起こりdarunavirのPR二量体化阻害は最初のステップで起こる (2014)
  • Author(s): 林 宏典、高宗 暢暁、韮澤 崇、青木 学、森下 宣彦、Debanannda Das、閌 康博、三隅 将吾、満屋 裕明
  • Organizer: 第28回エイズ学会総会
  • Place of Presentation: 大阪国際会議場（大阪市北区）
  • Year and Date: 2014-12-03 – 2014-12-05
• [Presentation] Dimerization of HIV-1Protease Occurs through Two Steps, Which Relates to the Mechanism of Protease to the Mechanism of Protease Dimerization Inhibition by Darunavir (2014)
  • Author(s): 林 宏典、満屋 裕明
  • Organizer: 15th Kumamoto AIDS Seminar
  • Place of Presentation: 阿蘇グランヴィリオホテル（熊本県阿蘇市）
  • Year and Date: 2014-10-01 – 2014-10-03
• [Presentation] HIV-1プロテアーゼ (PR) の二量体化は2ステップで起こりダルナビルのPR二量体化阻害は最初のステップで起こる (2014)
  • Author(s): 林 宏典、高宗 暢暁、韮澤 崇、青木 学、森下 宣彦、Debanannda Das、閌 康博、三隅 将吾、満屋 裕明
  • Organizer: 第16回白馬シンポジウムin熊本
  • Place of Presentation: 熊本大学（熊本市中央区）
  • Year and Date: 2014-06-13 – 2014-06-14
• [Presentation] HIV-1の高度darunavir耐性発現にはdarunavir高感受性を単独で付与するV32Iの獲得が重要である (2014)
  • Author(s): 青木 学、満屋 裕明
  • Organizer: 第16回白馬シンポジウムin熊本
  • Place of Presentation: 熊本大学（熊本市中央区）
  • Year and Date: 2014-06-13 – 2014-06-14
• [Presentation] 多剤耐性HIV-1変異体に強力な活性を発揮する新規プロテアーゼ阻害剤 (2014)
  • Author(s): 青木 学、林 宏典、Ravikiran S. Yedidi、Debananda Das、青木 宏美、Cuthbert D. Martyr、Arun K. Ghosh、満屋 裕明
  • Organizer: 第24回抗ウイルス療法研究会総会
  • Place of Presentation: ハイランドリゾートホテル（山梨県富士吉田市）
  • Year and Date: 2014-05-07 – 2014-05-08
• [Presentation] 高い中枢神経系 (CNS) 移行性と強力な抗HIV-1活性を有する新規HIV関連神経認知障害 (HAND) 予防/治療薬の開発 (2014)
  • Author(s): 天野 将之、Pedro Miguel Salcedo-Gomez、Arun K. Ghosh、満屋 裕明
  • Organizer: 第24回抗ウイルス療法研究会総会
  • Place of Presentation: ハイランドリゾートホテル（山梨県富士吉田市）
  • Year and Date: 2014-05-07 – 2014-05-08
• [Patent(Industrial Property Rights)] HIV-1 PROTEASE INHIBITORS AND USES THEREOF (2014)
  • Inventor(s): GHOSH A, RAO V, MITSUYA H
  • Industrial Property Rights Holder: Purdue U. & Kumamoto U.
  • Industrial Property Rights Type: 特許
  • Filing Date: 2014-05-16
  • Acquisition Date: 2015-05-15
  • Overseas