From therapeutic antibodies to chimeric antigen receptors (CARs): making better CARs based on antigen-binding domain

Expert Opinion on Biological Therapy

Volumn 16, Issue 12, 2016, Pages 1469-1478

  Wu, Yanling ^a   Jiang, Shibo ^a   Ying, Tianlei ^a  

^a FUDAN UNIVERSITY   (China)

Author keywords

CAR; immunogenicity; mAb; scFv

Indexed keywords

CHIMERIC ANTIGEN RECEPTOR; MONOCLONAL ANTIBODY; SINGLE CHAIN FRAGMENT VARIABLE ANTIBODY; LYMPHOCYTE ANTIGEN RECEPTOR;

ANTIGEN BINDING; CANCER IMMUNOTHERAPY; CELL SPECIFICITY; CELL THERAPY; HEMATOLOGIC MALIGNANCY; HUMAN; IMMUNOGENICITY; INTRACELLULAR SIGNALING; REVIEW; T LYMPHOCYTE; ANIMAL; BINDING SITE; CLINICAL TRIAL (TOPIC); IMMUNOLOGY; METABOLISM; NEOPLASMS; PHYSIOLOGY; PROCEDURES; SIGNAL TRANSDUCTION;

ANIMALS; BINDING SITES; CLINICAL TRIALS AS TOPIC; HUMANS; NEOPLASMS; RECEPTORS, ANTIGEN, T-CELL; SIGNAL TRANSDUCTION; SINGLE-CHAIN ANTIBODIES;

EID: 84994442778     PISSN: 14712598     EISSN: 17447682     Source Type: Journal    
DOI: 10.1080/14712598.2016.1235148     Document Type: Review

References (87)

1. S.A.Rosenberg, P.Spiess, R.Lafreniere A new approach to the adoptive immunotherapy of cancer with tumor-infiltrating lymphocytes. Science. 1986;233(4770):1318–1321.
2. S.A.Rosenberg, M.E.Dudley. Adoptive cell therapy for the treatment of patients with metastatic melanoma. Curr Opin Immunol. 2009;21:233–240. DOI:10.1016/j.coi.2009.03.002.
3. + T cell clones for the treatment of patients with metastatic melanoma:in vivo persistence, migration, and antitumor effect of transferred T cells. Proc Natl Acad Sci U S A. 2002;99(25):16168–16173. DOI:10.1073/pnas.242600099
4. + T cells for the treatment of patients with metastatic melanoma. J Clin Oncol. 2006;24(31):5060–5069. DOI:10.1200/JCO.2006.07.1100
5. T.M.Clay, M.C.Custer, J.Sachs, et al. Efficient transfer of a tumor antigen-reactive TCR to human peripheral blood lymphocytes confers anti-tumor reactivity. J Immunol. 1999;163(1):507–513.
6. P.F.Robbins, R.A.Morgan, S.A.Feldman, et al. Tumor regression in patients with metastatic synovial cell sarcoma and melanoma using genetically engineered lymphocytes reactive with NY-ESO-1. J Clin Oncol. 2011;29:917–924. DOI:10.1200/JCO.2010.32.2537.
7. A.P.Rapoport, E.A.Stadtmauer, G.K.Binder-Scholl, et al. NY-ESO-1-specific TCR-engineered T cells mediate sustained antigen-specific antitumor effects in myeloma. Nat Med. 2015;21(8):914–921. DOI:10.1038/nm.3910
8. F.Garrido, I.Algarra. MHC antigens and tumor escape from immune surveillance. Adv Cancer Res. 2001;83:117–158.
9. F.M.Marincola, E.M.Jaffee, D.J.Hicklin, et al. Escape of human solid tumors from T-cellrecognition:molecular mechanisms and functional significance. Adv Immunol. 2011;74:181–273.
10. J.D.Stone, D.T.Harris, D.M.Kranz. TCR affinity for p/MHC formed by tumor antigens that are self-proteins:impact on efficacy and toxicity. Curr Opin Immunol. 2015;33:16–22. DOI:10.1016/j.coi.2015.01.003.
11. G.P.Linette, E.A.Stadtmauer, M.V.Maus, et al. Cardiovascular toxicity and titin cross-reactivity of affinity-enhanced T cells in myeloma and melanoma. Blood. 2013;122(6):863–871. DOI:10.1182/blood-2013-03-490565
12. B.J.Cameron, A.B.Gerry, J.Dukes, et al. Identification of a Titin-derived HLA-A1-presented peptide as a cross-reactive target for engineered MAGE A3-directed T cells. Sci Transl Med. 2013;5(197):197ra103. DOI:10.1126/scitranslmed.3006034
13. B.D.Cheson, J.P.Leonard. Monoclonal antibody therapy for B-cell non-Hodgkin’s lymphoma. New Engl J Med. 2008;359:613–626. DOI:10.1056/NEJMra0708875.
14. C.L.Vogel, M.A.Cobleigh, D.Tripathy, et al. Efficacy and safety of trastuzumab as a single agent in first-line treatment of HER2-overexpressing metastatic breast cancer. J Clin Oncol. 2002;20(3):719–726.
15. E.Van Cutsem, C.-H.Köhne, E.Hitre, et al. Cetuximab and chemotherapy as initial treatment for metastatic colorectal cancer. N Engl J Med. 2009;360(14):1408–1417. DOI:10.1056/NEJMoa0805019
16. G.Gross, T.Waks, Z.Eshhar. Expression of immunoglobulin-T-cell receptor chimeric molecules as functional receptors with antibody-type specificity. Proc Natl Acad Sci U S A. 1989;86(24):10024–10028.•• The earliest report that T cells could be genetically modified to express an antibody’s variable domains fused to TCR with the ability to effectively transmit the signal for T-cell activation and execution of its effector function.
17. J.N.Kochenderfer, W.H.Wilson, J.E.Janik, et al. Eradication of B-lineage cells and regression of lymphoma in a patient treated with autologous T cells genetically engineered to recognize CD19. Blood. 2010;116(20):4099–4102. DOI:10.1182/blood-2010-04-281931• Effective clinical treatment with anti-CD19 CAR T cells was first reported in a patient with advanced stage lymphoma.
18. R.J.Brentjens, I.Rivière, J.H.Park, et al. Safety and persistence of adoptively transferred autologous CD19-targeted T cells in patients with relapsed or chemotherapy refractory B-cell leukemias. Blood. 2011;118(18):4817–4828. DOI:10.1182/blood-2011-04-348540
19. C.U.Louis, B.Savoldo, G.Dotti, et al. Antitumor activity and long-term fate of chimeric antigen receptor-positive T cells in patients with neuroblastoma. Blood. 2011;118:6050–6056. DOI:10.1182/blood-2011-05-354449• Clinical data demonstrated that GD2-CAR T cells induced complete tumor responses in patients with active neuroblastoma.
20. H.M.Finney, A.D.Lawson, C.R.Bebbington, et al. Chimeric receptors providing both primary and costimulatory signaling in T cells from a single gene product. J Immunol. 1998;161(6):2791–2797.
21. C.Carpenito, M.C.Milone, R.Hassan, et al. Control of large, established tumor xenografts with genetically retargeted human T cells containing CD28 and CD137 domains. Proc Natl Acad Sci U S A. 2009;106:3360–3365. DOI:10.1073/pnas.0813101106.
22. S.Guedan, X.Chen, A.Madar, et al. ICOS-based chimeric antigen receptors program bipolar TH17/TH1 cells. Blood. 2014;124:1070–1080. DOI:10.1182/blood-2013-10-535245.
23. S.J.van der Stegen, M.Hamieh, M.Sadelain. The pharmacology of second-generation chimeric antigen receptors. Nat Rev Drug Discov. 2015;14:499–509. DOI:10.1038/nrd4597.
24. A.A.Hombach, H.Abken. Costimulation by chimeric antigen receptors revisited the T cell antitumor response benefits from combined CD28-OX40 signalling. Int J Cancer. 2011;129:2935–2944. DOI:10.1002/ijc.25960.
25. M.Jonnalagadda, A.Mardiros, R.Urak, et al. Chimeric antigen receptors with mutated IgG4 Fc spacer avoid fc receptor binding and improve T cell persistence and antitumor efficacy. Mol Ther. 2015;23(4):757–768. DOI:10.1038/mt.2014.208
26. M.Hudecek, D.Sommermeyer, P.L.Kosasih, et al. The nonsignaling extracellular spacer domain of chimeric antigen receptors is decisive for in vivo antitumor activity. Cancer Immunol Res. 2015;3(2):125–135. DOI:10.1158/2326-6066.CIR-14-0127
27. A.Chailyan, P.Marcatili, A.Tramontano. The association of heavy and light chain variable domains in antibodies:implications for antigen specificity. FEBS J. 2011;278(16):2858–2866. DOI:10.1111/j.1742-4658.2011.08207.x.
28. T.Yokota, D.E.Milenic, M.Whitlow, et al. Rapid tumor penetration of a single-chain Fv and comparison with other immunoglobulin forms. Cancer Res. 1992;52(12):3402–3408.
29. A.Bradbury. scFvs and beyond. Drug Discov Today. 2003;8(16):737–739.
30. T.C.Meeker, J.Lowder, D.G.Maloney, et al. A clinical trial of anti-idiotype therapy for B cell malignancy. Blood. 1985;65(6):1349–1363.
31. J.M.Reichert, C.J.Rosensweig, L.B.Faden, et al. Monoclonal antibody successes in the clinic. Nat Biotechnol. 2005;23(9):1073–1078. DOI:10.1038/nbt0905-1073
32. S.Miotti, S.Canevari, S.Ménard, et al. Characterization of human ovarian carcinoma-associated antigens defined by novel monoclonal antibodies with tumor-restricted specificity. Int J Cancer. 1987;39(3):297–303.•• The first CAR T cells clinically evaluated as a cancer treatment.
33. P.Hwu, G.E.Shafer, J.Treisman, et al. Lysis of ovarian cancer cells by human lymphocytes redirected with a chimeric gene composed of an antibody variable region and the Fc receptor gamma chain. J Exp Med. 1993;178(1):361–366.
34. P.Hwu, J.C.Yang, R.Cowherd, et al. In vivo antitumor activity of T cells redirected with chimeric antibody/T-cell receptor genes. Cancer Res. 1995;55(15):3369–3373.
35. M.H.Kershaw, J.A.Westwood, L.L.Parker, et al. A phase I study on adoptive immunotherapy using gene-modified T cells for ovarian cancer. Clin Cancer Res. 2006;12(20 Pt 1):6106–6115. DOI:10.1158/1078-0432.CCR-06-1183
36. J.N.Kochenderfer, S.A.Rosenberg. Treating B-cell cancer with T cells expressing anti-CD19 chimeric antigen receptors. Nat Rev Clin Oncol. 2013;10:267–276. DOI:10.1038/nrclinonc.2013.46• Provides a comprehensive description of anti-CD19 CAR in clinical trials.
37. M.Kalos, B.L.Levine, D.L.Porter, et al. T cells with chimeric antigen receptors have potent antitumor effects and can establish memory in patients with advanced leukemia. Sci Transl Med. 2011;3(95):95ra73. DOI:10.1126/scitranslmed.3002842
38. S.Gill, C.H.June. Going viral:chimeric antigen receptor T-cell therapy for hematological malignancies. Immunol Rev. 2015;263(1):68–89. DOI:10.1111/imr.12243.
39. I.C.Nicholson, K.A.Lenton, D.J.Little, et al. Construction and characterisation of a functional CD19 specific single chain Fv fragment for immunotherapy of B lineage leukaemia and lymphoma. Mol Immunol. 1997;34(16–17):1157–1165.
40. D.L.Porter, B.L.Levine, M.Kalos, et al. Chimeric antigen receptor-modified T cells in chronic lymphoid leukemia. N Engl J Med. 2011;365:725–733. DOI:10.1056/NEJMoa1103849.
41. S.L.Maude, N.Frey, P.A.Shaw, et al. Chimeric antigen receptor T cells for sustained remissions in leukemia. N Engl J Med. 2014;371(16):1507–1517. DOI:10.1056/NEJMoa1407222
42. B.E.Bejcek, D.Wang, E.Berven, et al. Development and characterization of three recombinant single chain antibody fragments (scFvs) directed against the CD19 antigen. Cancer Res. 1995;55(11):2346–2351.
43. R.J.Brentjens, J.-B.Latouche, E.Santos, et al. Eradication of systemic B-cell tumors by genetically targeted human T lymphocytes co-stimulated by CD80 and interleukin-15. Nat Med. 2003;9(3):279–286. DOI:10.1038/nm827
44. R.J.Brentjens, E.Santos, Y.Nikhamin, et al. Genetically targeted T cells eradicate systemic acute lymphoblastic leukemia xenografts. Clin Cancer Res. 2007;13(18 Pt 1):5426–5435. DOI:10.1158/1078-0432.CCR-07-0674
45. R.J.Brentjens, M.L.Davila, I.Riviere, et al. CD19-targeted T cells rapidly induce molecular remissions in adults with chemotherapy-refractory acute lymphoblastic leukemia. Sci Transl Med. 2013;5:177ra138. DOI:10.1126/scitranslmed.3005930• Clinical data demonstrated the marked antitumor efficacy of 19-28z CAR-modified T cells in patients with relapsed/refractory B-ALL.
46. U.Testa, R.Riccioni, S.Militi, et al. Elevated expression of IL-3Ralpha in acute myelogenous leukemia is associated with enhanced blast proliferation, increased cellularity, and poor prognosis. Blood. 2002;100:2980–2988. DOI:10.1182/blood-2002-03-0852.
47. Q.Sun, J.M.Woodcock, A.Rapoport, et al. Monoclonal antibody 7G3 recognizes the N-terminal domain of the human interleukin-3 (IL-3) receptor alpha-chain and functions as a specific IL-3 receptor antagonist. Blood. 1996;87(1):83–92.
48. S.Tettamanti, V.Marin, I.Pizzitola, et al. Targeting of acute myeloid leukaemia by cytokine-induced killer cells redirected with a novel CD123-specific chimeric antigen receptor. Br J Haematol. 2013;161:389–401. DOI:10.1111/bjh.12282.
49. I.Pastan, R.Hassan. Discovery of mesothelin and exploiting it as a target for immunotherapy. Cancer Res. 2014;74(11):2907–2912. DOI:10.1158/0008-5472.CAN-14-0337.
50. P.S.Chowdhury, J.L.Viner, R.Beers, et al. Isolation of a high-affinity stable single-chain Fv specific for mesothelin from DNA-immunized mice by phage display and construction of a recombinant immunotoxin with anti-tumor activity. Proc Natl Acad Sci U S A. 1998;95:669–674.
51. Q.Li, C.F.Verschraegen, J.Mendoza, et al. Cytotoxic activity of the recombinant anti-mesothelin immunotoxin, SS1(dsFv)PE38, towards tumor cell lines established from ascites of patients with peritoneal mesotheliomas. Anticancer Res. 2004;24:1327–1335.
52. M.V.Maus, A.R.Haas, G.L.Beatty, et al. T cells expressing chimeric antigen receptors can cause anaphylaxis in humans. Cancer Immunol Res. 2013;1:26–31. DOI:10.1158/2326-6066.CIR-13-0006• First description that clinical anaphylaxis results from mesothelin-targeting CAR-modified T cells.
53. C.H.Lamers, R.Willemsen, P.van Elzakker, et al. Immune responses to transgene and retroviral vector in patients treated with ex vivo-engineered T cells. Blood. 2011;117(1):72–82. DOI:10.1182/blood-2010-07-294520
54. S.H.Petersdorf, K.J.Kopecky, M.Slovak, et al. A phase 3 study of gemtuzumabozogamicin during induction and postconsolidation therapy in younger patients with acute myeloid leukemia. Blood. 2013;121(24):4854–4860. DOI:10.1182/blood-2013-01-466706
55. U.S. Food and Drug Administration News Release. Pfizer voluntarily withdraws cancer treatment Mylotarg from U.S. market. 2010. Available from::http://www.fda.gov/NewsEvents/Newsroom/PressAnnouncements/2010/ucm216448.htm
56. V.Marin, I.Pizzitola, V.Agostoni, et al. Cytokine-induced killer cells for cell therapy of acute myeloid leukemia:improvement of their immune activity by expression of CD33-specific chimeric receptors. Haematologica. 2010;95:2144–2152. DOI:10.3324/haematol.2010.026310.
57. A.Dutour, V.Marin, I.Pizzitola, et al. In vitro and in vivo antitumor effect of anti-CD33 chimeric receptor-expressing EBV-CTL against CD33 acute myeloid leukemia. Adv Hematol. 2012;2012:683065. DOI:10.1155/2012/683065.
58. S.S.Kenderian, M.Ruella, O.Shestova, et al. CD33-specific chimeric antigen receptor T cells exhibit potent preclinical activity against human acute myeloid leukemia. Leukemia. 2015;29(8):1637–1647. DOI:10.1038/leu.2015.52
59. Q.-S.Wang, Y.Wang, H.-Y.Lv, et al. Treatment of CD33-directed chimeric antigen receptor-modified T cells in one patient with relapsed and refractory acute myeloid leukemia. Mol Ther. 2015;23(1):184–191. DOI:10.1038/mt.2014.164
60. F.Révillion, J.Bonneterre, J.P.Peyrat. ERBB2 oncogene in human breast cancer and its clinical significance. Eur J Cancer. 1998;34:791–808.
61. D.J.Slamon, B.Leyland-Jones, S.Shak, et al. Use of chemotherapy plus a monoclonal antibody against HER2 for metastatic breast cancer that overexpresses HER2. N Engl J Med. 2001;344:783–792. DOI:10.1056/NEJM200103153441101.
62. E.Tan-Chiu, G.Yothers, E.Romond, et al. Assessment of cardiac dysfunction in a randomized trial comparing doxorubicin and cyclophosphamide followed by paclitaxel, with or without Herceptin as adjuvant therapy in node-positive, human epidermal growth factor receptor 2-overexpressing breast cancer:NSABP B-31. J Clin Oncol. 2005;23:7811–7819. DOI:10.1200/JCO.2005.02.4091.
63. Y.Zhao, Q.J.Wang, S.Yang, et al. A herceptin-based chimeric antigen receptor with modified signaling domains leads to enhanced survival of transduced T lymphocytes and antitumor activity. J Immunol. 2009;183(9):5563–5574. DOI:10.4049/jimmunol.0900447
64. R.A.Morgan, J.C.Yang, M.Kitano, et al. Case report of a serious adverse event following the administration of T cells transduced with a chimeric antigen receptor recognizing ERBB2. Mol Ther. 2010;18(4):843–851. DOI:10.1038/mt.2010.24
65. N.Ahmed, V.S.Brawley, M.Hegde, et al. Human epidermal growth factor receptor 2 (HER2) -specific chimeric antigen receptor-modified T cells for the immunotherapy of HER2-positive sarcoma. J Clin Oncol. 2015;33(15):1688–1696. DOI:10.1200/JCO.2014.58.0225
66. C.J.Wikstrand, L.P.Hale, S.K.Batra, et al. Monoclonal antibodies against EGFRvIII are tumor specific and react with breast and lung carcinomas and malignant gliomas. Cancer Res. 1995;55(14):3140–3148.
67. P.Gupta, S.-Y.Han, M.Holgado-Madruga, et al. Development of an EGFRvIII specific recombinant antibody. BMC Biotechnol. 2010;10:72. DOI:10.1186/1472-6750-10-72.
68. C.-J.Shen, Y.-X.Yang, E.Q.Han, et al. Chimeric antigen receptor containing ICOS signaling domain mediates specific and efficient antitumor effect of T cells against EGFRvIII expressing glioma. J Hematol Oncol. 2013;6:33. DOI:10.1186/1756-8722-6-33.
69. L.A.Johnson, J.Scholler, T.Ohkuri, et al. Rational development and characterization of humanized anti-EGFR variant III chimeric antigen receptor T cells for glioblastoma. Sci Transl Med. 2015;7(275):275ra22. DOI:10.1126/scitranslmed.aad3106
70. Y.Feng, X.Xiao, Z.Zhu, et al. A novel human monoclonal antibody that binds with high affinity to mesothelin-expressing cells and kills them by antibody-dependent cell-mediated cytotoxicity. Mol Cancer Ther. 2009;8:1113–1118. DOI:10.1158/1535-7163.MCT-08-0945.
71. P.S.Adusumilli, L.Cherkassky, J.Villena-Vargas, et al. Regional delivery of mesothelin-targeted CAR T cell therapy generates potent and long-lasting CD4-dependent tumor immunity. Sci Transl Med. 2014;6(261):261ra151. DOI:10.1126/scitranslmed.3010162
72. J.H.Sampson, B.D.Choi, L.Sanchez-Perez, et al. EGFRvIIImCAR modified T-cell therapy cures mice with established intracerebral glioma and generates host immunity against tumor-antigen loss. Clin Cancer Res. 2014;20:972–984. DOI:10.1158/1078-0432.CCR-13-0709.
73. D.-G.Song, Q.Ye, M.Poussin, et al. A fully human chimeric antigen receptor with potent activity against cancer cells but reduced risk for off-tumor toxicity. Oncotarget. 2015;6(25):21533–21546. DOI:10.18632/oncotarget.4071
74. X.Xiao, M.Ho, Z.Zhu, et al. Identification and characterization of fully human anti-CD22 monoclonal antibodies. MAbs. 2009;1(3):297–303.
75. W.Haso, D.W.Lee, N.N.Shah, et al. Anti-CD22-chimeric antigen receptors targeting B-cell precursor acute lymphoblastic leukemia. Blood. 2013;121(7):1165–1174. DOI:10.1182/blood-2012-06-438002
76. P.J.Bugelski, R.Achuthanandam, R.J.Capocasale, et al. Monoclonal antibody-induced cytokine-release syndrome. Expert Rev Clin Immunol. 2009;5(5):499–521. DOI:10.1586/eci.09.31
77. D.T.Teachey, S.R.Rheingold, S.L.Maude, et al. Cytokine release syndrome after blinatumomab treatment related to abnormal macrophage activation and ameliorated with cytokine-directed therapy. Blood. 2013;121(26):5154–5157. DOI:10.1182/blood-2013-02-485623• Important article describing the effects of tocilizumab oncytokine release syndrome in T-cell-activating therapies.
78. M.Hudecek, M.-T.Lupo-Stanghellini, P.L.Kosasih, et al. Receptor affinity and extracellular domain modifications affect tumor recognition by ROR1-specific chimeric antigen receptor T cells. Clin Cancer Res. 2013;19(12):3153–3164. DOI:10.1158/1078-0432.CCR-13-0330• This study demonstrated that the functionality of CARs could be improved by increasing scFv affinity.
79. R.C.Lynn, Y.Feng, K.Schutsky, et al. High-affinity FRβ-specific CAR T cells eradicate AML and normal myeloid lineage without HSC toxicity. Leukemia. 2016;30:1355–1364. DOI:10.1038/leu.2016.35
80. M.Chmielewski, A.Hombach, C.Heuser, et al. T cell activation by antibody-like immunoreceptors:increase in affinity of the single-chain fragment domain above threshold does not increase T cell activation against antigen-positive target cells but decreases selectivity. J Immunol. 2004;173(12):7647–7653.
81. X.Liu, S.Jiang, C.Fang, et al. Affinity-tuned ErbB2 or EGFR chimeric antigen receptor T cells exhibit an increased therapeutic index against tumors in mice. Cancer Res. 2015;75(17):3596–3607. DOI:10.1158/0008-5472.CAN-15-0159
82. A.A.Hombach, V.Schildgen, C.Heuser, et al. T cell activation by antibody-like immunoreceptors:the position of the binding epitope within the target molecule determines the efficiency of activation of redirected T cells. J Immunol. 2007;178(7):4650–4657.
83. C.Monrat, S.Elias, E.Lamis, et al. Novel GD2-specific chimeric antigen receptor-modified T cells targeting osteosarcoma. In:Proceedings of the 106th Annual Meeting of the American Association for Cancer Research. Philadelphia (PA):AACR. Cancer Res. 2015;75(15Suppl):Abstract nr 3157. DOI:10.1158/0008-5472.CAN-14-3569
84. P.Holliger, P.J.Hudson. Engineered antibody fragments and the rise of single domains. Nat Biotechnol. 2005;23(9):1126–1136. DOI:10.1038/nbt1142.
85. A.L.Nelson, J.M.Reichert. Development trends for therapeutic antibody fragments. Nat Biotechnol. 2009;27(4):331–337. DOI:10.1038/nbt0409-331.
86. A.L.Nelson. Antibody fragments:hope and hype. MAbs. 2010;2(1):77–83.
87. F.R.Jamnani, F.Rahbarizadeh, M.A.Shokrgozar, et al. T cells expressing VHH-directed oligoclonal chimeric HER2 antigen receptors:towards tumor-directed oligoclonal T cell therapy. Biochim Biophys Acta. 2014;1840(1):378–386. DOI:10.1016/j.bbagen.2013.09.029