Bax is an essential mediator of mitochondria-dependent programed cell death. Bax belongs to the Bcl-2 family of proteins and its activities are regulated through interaction with other member proteins in the Bcl-2 family. To date, several apoptosis-inducing drugs activating Bax have been developed, and some of them are already in the market as therapeutics against cancer. However, at present, there are no clinically effective pharmacological Bax inhibitors protecting essential cells. Previously, we developed Bax-Inhibiting Peptides (BIPs) that belong to the peptide group of Cell-Penetrating Peptides (CPPs). CPPs have the ability to deliver cargo molecules into the cell. In this review, we will describe the mechanism of action of BIPs together with the recent applications of BIPs in disease models in vitro and in vivo. However, BIPs have several limitations in their use to treat human diseases, and other types of Bax inhibitors need to be developed for future therapeutics. Recently, several groups reported the successful development of novel small compounds inhibiting Bax. We will review these Bax inhibitors to discuss current strategies to develop pharmacological Bax inhibitors.
Impact statement
Bax induces mitochondria-dependent programed cell death. While cytotoxic drugs activating Bax have been developed for cancer treatment, clinically effective therapeutics suppressing Bax-induced cell death rescuing essential cells have not been developed. This mini-review will summarize previously reported Bax inhibitors including peptides, small compounds, and antibodies. We will discuss potential applications and the future direction of these Bax inhibitors.
KorsmeyerSJ.BCL-2 gene family and the regulation of programmed cell death. Cancer Res1999;
59:1693s–700s
2.
KaleJOsterlundEJAndrewsDW.BCL-2 family proteins: changing partners in the dance towards death. Cell Death Differ2018;
25:65–80
3.
KarchJMolkentinJD.Regulated necrotic cell death: the passive aggressive side of Bax and Bak. Circ Res2015;
116:1800–9
4.
YoshidaTTomiokaINagaharaTHolystTSawadaMHayesPGama V, Okuno M, Chen Y, Abe Y, Kanouchi T, Sasada H, Wang D, Yokota T, Sato E, Matsuyama S. Bax-inhibiting peptide derived from mouse and rat Ku70. Biochem Biophys Res Commun 2004;321:961–6
5.
GomezJAGamaVMatsuyamaS. Cell-permeable penta-peptides derived from Bax-inhibiting peptide. In: Langel U (ed) Cell penetrating peptide. 2nd ed. 2006, Boca Raton, FL: CRC Press, pp. 469–81
6.
GomezJAGamaVYoshidaTSunWHayesPLeskovK, Boothman D, Matsuyama S.
Bax-inhibiting peptides derived from Ku70 and cell-penetrating pentapeptides. Biochm Soc Trans2007;
35:797–801
7.
GomezJAChenJNgoJHajkovaDYehIJGamaVMiyagi M, Matsuyama S.
Cell-penetrating penta-peptides (CPP5s): measurement of cell entry and protein-transduction activity. Pharmaceuticals2010;
3:3594
8.
DownsJAJacksonSP.A means to a DNA end: the many roles of Ku. Nat Rev Mol Cell Biol2004;
5:367–78
9.
LiYYokotaTGamaVYoshidaTGomezJAIshikawaKSasaguri H, Cohen HY, Sinclair DA, Mizusawa H, Matsuyama S.
Bax-inhibiting peptide protects cells from polyglutamine toxicity caused by Ku70 acetylation. Cell Death Differ2007;
14:2058–67
10.
GamaVGomezJAMayoLDJacksonMWDanielpourDSongKHaas AL, Laughlin MJ, Matsuyama S.
Hdm2 is a ubiquitin ligase of Ku70-Akt promotes cell survival by inhibiting Hdm2-dependent Ku70 destabilization. Cell Death Differ2009;
16:758–69
11.
El-AndaloussiSHolmTLangelU.Cell-penetrating peptides: mechanisms and applications. Curr Pharm Des2005;
11:3597–611
12.
MurrielCLDowdySF.Influence of protein transduction domains on intracellular delivery of macromolecules. Expert Opin Drug Deliv2006;
3:739–46
13.
ZorkoMLangelU.Cell-penetrating peptides: mechanism and kinetics of cargo delivery. Adv Drug Deliv Rev2005;
57:529–45
14.
FutakiS.Membrane-permeable arginine-rich peptides and the translocation mechanisms. Adv Drug Deliv Rev2005;
57:547–58
15.
FrankelADPaboCO.Cellular uptake of the tat protein from human immunodeficiency virus.Cell1988;
55:1189–93
16.
GreenMLoewensteinPM.Autonomous functional domains of chemically synthesized human immunodeficiency virus tat trans-activator protein. Cell1988;
55:1179–88
17.
CaiSRXuGBecker-HapakMMaMDowdySFMcLeodHL.The kinetics and tissue distribution of protein transduction in mice. Eur J Pharm Sci2006;
27:311–9
18.
SchwarzeSRHoAVocero-AkbaniADowdySF.In vivo protein transduction: delivery of a biologically active protein into the mouse. Science1999;
285:1569–72
19.
WadiaJSStanRVDowdySF.Transducible TAT-HA fusogenic peptide enhances escape of TAT-fusion proteins after lipid raft macropinocytosis. Nat Med2004;
10:310–5
20.
JoliotAPernelleCDeagostini-BazinHProchiantzA.Antennapedia homeobox peptide regulates neural morphogenesis. Proc Natl Acad Sci U S A1991;
88:1864–8
21.
DerossiDJoliotAHChassaingGProchiantzA.The third helix of the Antennapedia homeodomain translocates through biological membranes. J Biol Chem1994;
269:10444–50
22.
FutakiSSuzukiTOhashiWYagamiTTanakaSUedaKSugiura Y.
Arginine-rich peptides. An abundant source of membrane-permeable peptides having potential as carriers for intracellular protein delivery. J Biol Chem2001;
276:5836–40
OehlkeJSchellerAWiesnerBKrauseEBeyermannMKlauschenzEMelzig M, Bienert M.
Cellular uptake of an alpha-helical amphipathic model peptide with the potential to deliver polar compounds into the cell interior non-endocytically. Biochimica et Biophysica Acta1998;
1414:127–39
25.
PoogaMHallbrinkMZorkoMLangelU.Cell penetration by transportan. FASEB J1998;
12:67–77
26.
PoogaMKutCKihlmarkMHallbrinkMFernaeusSRaidRLand T, Hallberg E, Bartfai T, Langel U.
Cellular translocation of proteins by transportan. FASEB J2001;
15:1451–3
27.
FoergCMerkleHP.On the biomedical promise of cell penetrating peptides: limits versus prospects. J Pharm Sci2008;
97:144–62
28.
JoliotAProchiantzA.Transduction peptides: from technology to physiology. Nat Cell Biol2004;
6:189–96
NakaseITadokoroAKawabataNTakeuchiTKatohHHiramotoKNegishi M, Nomizu M, Sugiura Y, Futaki S.
Interaction of arginine-rich peptides with membrane-associated proteoglycans is crucial for induction of actin organization and macropinocytosis. Biochemistry2007;
46:492–501
31.
PerezGIJurisicovaAWiseLLipinaTKanisekMBechardATakai Y, Hunt P, Roder J, Grynpas M, Tilly JL.
Absence of the proapoptotic Bax protein extends fertility and alleviates age-related health complications in female mice. Proc Natl Acad Sci U S A2007;
104:5229–34
32.
KnudsonCMTungKSTourtellotteWGBrownGAKorsmeyerSJ.Bax-deficient mice with lymphoid hyperplasia and male germ cell death. Science1995;
270:96–9
33.
NgoJMatsuyamaMKimCPoventud-FuentesIBatesASiedlakSLLee HG, Doughman YQ, Watanabe M, Liner A, Hoit B, Voelkel N, Gerson S, Hasty P, Matsuyama S.
Bax deficiency extends the survival of Ku70 knockout mice that develop lung and heart diseases. Cell Death Dis2015;
6:e1706–25811803
34.
D'OrsiBMateykaJPrehnJHM.Control of mitochondrial physiology and cell death by the Bcl-2 family proteins Bax and Bok. Neurochem Int2017;
109:162–70
35.
GalluzziLBlomgrenKKroemerG.Mitochondrial membrane permeabilization in neuronal injury. Nat Rev Neurosci2009;
10:481–94
36.
GibsonMEHanBHChoiJKnudsonCMKorsmeyerSJParsadanianMHoltzman DM.
BAX contributes to apoptotic-like death following neonatal hypoxia-ischemia: evidence for distinct apoptosis pathways. Mol Med2001;
7:644–55
37.
WangXHanWDuXZhuCCarlssonYMallardCJacotot E, Hagberg H.
Neuroprotective effect of Bax-inhibiting peptide on neonatal brain injury. Stroke2010;
41:2050–5
38.
SelkoeDJHardyJ.The amyloid hypothesis of Alzheimer's disease at 25 years. EMBO Mol Med2016;
8:595–608
39.
KudoWLeeHPSmithMAZhuXMatsuyamaSLeeHG.Inhibition of Bax protects neuronal cells from oligomeric Abeta neurotoxicity. Cell Death Dis2012;
3:e309–22592316
40.
AllikmetsRShroyerNFSinghNSeddonJMLewisRABernsteinPSPeiffer A, Zabriskie NA, Li Y, Hutchinson A, Dean M, Lupski JR, Leppert M.
Mutation of the Stargardt disease gene (ABCR) in age-related macular degeneration. Science1997;
277:1805–7
41.
AllikmetsR.Further evidence for an association of ABCR alleles with age-related macular degeneration. The International ABCR Screening Consortium. Am J Hum Genet2000;
67:487–91
42.
MaedaAMaedaTGolczakMChouSDesaiAHoppelCLMatsuyama S, Palczewski K.
Involvement of all-trans-retinal in acute light-induced retinopathy of mice. J Biol Chem2009;
284:15173–83
43.
SawadaOPerusekLKohnoHHowellSJMaedaAMatsuyamaSMaeda T.
All-trans-retinal induces Bax activation via DNA damage to mediate retinal cell apoptosis. Exp Eye Res2014;
123:27–36
44.
QinQPatilKSharmaSC.The role of Bax-inhibiting peptide in retinal ganglion cell apoptosis after optic nerve transection. Neurosci Lett2004;
372:17–21
45.
ChenYNYamadaHMaoWMatsuyamaSAiharaMAraieM.Hypoxia-induced retinal ganglion cell death and the neuroprotective effects of beta-adrenergic antagonists. Brain Res2007;
1148:28–37
46.
KungGKonstantinidisKKitsisRN.Programmed necrosis, not apoptosis, in the heart. Circ Res2011;
108:1017–36
47.
WhelanRSKonstantinidisKWeiACChenYReynaDEJhaSCalvert JW, Lindsten T, Thompson CB, Crow MT, Gavathiotis E, Dorn GW, O'Rourke B, Kitsis RN.
Bax regulates primary necrosis through mitochondrial dynamics. Proc Natl Acad Sci U S A2012;
109:6566–71
48.
KarchJKwongJQBurrARSargentMAElrodJWPeixotoPMMartinez-Caballero S, Osinska H, Cheng EH, Robbins J, Kinnally KW, Molkentin JD.
Bax and Bak function as the outer membrane component of the mitochondrial permeability pore in regulating necrotic cell death in mice. eLife2013;
2:e00772
49.
SuzukiKYanagiharaTYokoyamaTMaeyamaTOgata-SuetsuguSArimura-OmoriMMikumo H, Hamada N, Harada E, Kuwano K, Harada T, Nakanishi Y.
Bax-inhibiting peptide attenuates bleomycin-induced lung injury in mice. Biology Open2017;
6:1869–75
50.
LiuTDe Los SantosFGPhanSH.The bleomycin model of pulmonary fibrosis. Methods Mol Biol2017;
1627:27–42
51.
MatsuyamaSPalmerJBatesAPoventud-FuentesIWongKNgoJMatsuyama M.
Bax-induced apoptosis shortens the life span of DNA repair defect Ku70-knockout mice by inducing emphysema. Exp Biol Med2016;
241:1265–71
52.
GaoMMonianPQuadriNRamasamyRJiangX.Glutaminolysis and transferrin regulate ferroptosis. Mol Cell2015;
59:298–308
53.
TanakaKKobayashiNGutierrezASRivas-CarrilloJDNavarro-AlvarezNChenYNarushima M, Miki A, Okitsu T, Noguchi H, Tanaka N.
Prolonged survival of mice with acute liver failure with transplantation of monkey hepatocytes cultured with an antiapoptotic pentapeptide V5. Transplantation2006;
81:427–37
54.
SotoPSmithLC.BH4 peptide derived from Bcl-xL and Bax-inhibitor peptide suppresses apoptotic mitochondrial changes in heat stressed bovine oocytes. Mol Reprod Dev2009;
76:637–46
55.
KellerAAMussbachFBreitlingRHemmerichPSchaeferBLorkowskiSReissmann S.
Relationships between cargo, cell penetrating peptides and cell type for uptake of non-covalent complexes into live cells. Pharmaceuticals2013;
6:184–203
56.
MussbachFFrankeMZochASchaeferBReissmannS.Transduction of peptides and proteins into live cells by cell penetrating peptides. J Cell Biochem2011;
112:3824–33
57.
KellerAABreitlingRHemmerichPKappeKBraunMWittigBSchaefer B, Lorkowski S, Reissmann S.
Transduction of proteins into leishmania tarentolae by formation of non-covalent complexes with cell-penetrating peptides. J Cell Biochem2014;
115:243–52
58.
GomezJMatsuyamaS.Cell-penetrating penta-peptides and Bax-inhibiting peptides: protocol for their application.Methods Mol Biol2011;
683:465–71
59.
MuroskiMEMorganTJJrLevensonCWStrouseGF.A gold nanoparticle pentapeptide: gene fusion to induce therapeutic gene expression in mesenchymal stem cells. J Am Chem Soc2014;
136:14763–71
60.
WolfeJMFadzenCMChooZNHoldenRLYaoMHansonGJPentelute BL.
Machine learning to predict cell-penetrating peptides for antisense delivery. ACS Cent Sci2018;
4:512–20
61.
LindstenTRossAJKingAZongWXRathmellJCShielsHAUlrich E, Waymire KG, Mahar P, Frauwirth K, Chen Y, Wei M, Eng VM, Adelman DM, Simon MC, Ma A, Golden JA, Evan G, Korsmeyer SJ, MacGregor GR, Thompson CB.
The combined functions of proapoptotic Bcl-2 family members bak and bax are essential for normal development of multiple tissues. Mol Cell2000;
6:1389–99
62.
NiuXBrahmbhattHMergenthalerPZhangZSangJDaudeMet al.
A small-molecule inhibitor of Bax and Bak oligomerization prevents genotoxic cell death and promotes neuroprotection. Cell Chem Biol2017;
24:493–506 e5
63.
HetzCVittePABombrunARostovtsevaTKMontessuitSHiverASchwarz MK, Church DJ, Korsmeyer SJ, Martinou JC, Antonsson B.
Bax channel inhibitors prevent mitochondrion-mediated apoptosis and protect neurons in a model of global brain ischemia. J Biol Chem2005;
280:42960–70
PeixotoPMTeijidoOMirzalievaODejeanLMPavlovEVAntonssonBKinnally KW.
MAC inhibitors antagonize the pro-apoptotic effects of tBid and disassemble Bax/Bak oligomers. J Bioenerg Biomembr2017;
49:65–74
66.
PieperAAXieSCapotaEEstillSJZhongJLongJMBecker L, Huntington P, Goldman SE, Shen CH, Capota M, Britt JK, Kotti T, Ure K, Brat DJ, Williams NS, MacMillan KS, Naidoo J, Melito L, Hsieh J, De Brabander J, Ready JM, McKnight SL.
Discovery of a proneurogenic, neuroprotective chemical. Cell2010;
142:939–51
67.
De Jesus-CortesHXuPDrawbridgeJEstillSJHuntingtonPTranSBritt J, Tesla R, Morlock L, Naidoo J, Melito LM, Wang G, Williams NS, Ready JM, McKnight SL, Pieper AA.
Neuroprotective efficacy of aminopropyl carbazoles in a mouse model of Parkinson disease. Proc Natl Acad Sci U S A2012;
109:17010–5
68.
TeslaRWolfHPXuPDrawbridgeJEstillSJHuntingtonPMcDaniel L, Knobbe W, Burket A, Tran S, Starwalt R, Morlock L, Naidoo J, Williams NS, Ready JM, McKnight SL, Pieper AA.
Neuroprotective efficacy of aminopropyl carbazoles in a mouse model of amyotrophic lateral sclerosis. Proc Natl Acad Sci U S A2012;
109:1617016–21
69.
MacMillanKSNaidooJLiangJMelitoLWilliamsNSMorlockLHuntington PJ, Estill SJ, Longgood J, Becker GL, McKnight SL, Pieper AA, De Brabander JK, Ready JM.
Development of proneurogenic, neuroprotective small molecules. J Am Chem Soc2011;
133:1428–37
70.
UchimeODaiZBirisNLeeDSidhuSSLiSLai JR, Gavathiotis E.
Synthetic antibodies inhibit Bcl-2-associated X Protein (BAX) through blockade of the N-terminal activation site. J Biol Chem2016;
291:89–102
71.
RobinAYIyerSBirkinshawRWSandowJWardakALuoCSShi M, Webb AI, Czabotar PE, Kluck RM, Colman PM.
Ensemble properties of Bax determine its function. Structure2018;
26:1346–59 e5
72.
FancelliDAbateAAmiciRBernardiPBallariniMCappaACarenzi G, Colombo A, Contursi C, Di Lisa F, Dondio G, Gagliardi S, Milanesi E, Minucci S, Pain, G P. Pelicci PG, Saccani A, Storto M, Thaler F, Varasi M, Villa M, Plyte S.
Cinnamic anilides as new mitochondrial permeability transition pore inhibitors endowed with ischemia-reperfusion injury protective effect in vivo. J Med Chem2014;
57:5333–47
73.
SiemenDZiemerM.What is the nature of the mitochondrial permeability transition pore and what is it not?IUBMB Life2013;
65:255–62
