Table Info

Table 1.

Preclinical studies of gene therapy for CAH to date

Vector and transgene	Model	Dose and route	Key outcome(s)	Longest benefit duration	Reference
Adenovirus CYP21A2	H-2^aw18 mice (Cyp21a1^-/-)	1 × 10⁸ pfu/adrenal Open bilateral intra-adrenal injections	CYP21A2 mRNA expressed in ~ 30% of cortex and medulla, peaking at 2–7 days and still detectable at 14 days. Normalised corticosterone production to 40 days. Improved progesterone/deoxycorticosterone ratio at 7 days but not at 40 days. Transduced adrenals still produced corticosterone ex vivo at 40 days. Improvement in adrenocortical ultrastructure at 7 days.	40 days	Tajima et al. 1999 [62]
Retrovirus Cyp21a1	H-2^aw18 mice (Cyp21a1^-/-) fibroblasts	5 × 10⁵ cells/mouse Autologous subcutaneous transplantation of ex vivo transduced fibroblasts	Four weeks post-implantation, 4 of 6 experimental mice had reduction in progesterone/deoxycorticosterone ratio.	4 weeks	Naiki et al. 2016 [64]
AAV2 Cyp21a1	H-2^aw18 mice (Cyp21a1^-/-)	1 × 10¹¹ vgc/mouse Intra-muscular injection	All mice (n = 4) had reduction in progesterone/deoxycorticosterone ratio (statistically insignificant). One mouse followed to 8 months retained “relatively low” progesterone/deoxycorticosterone ratio until 7 months.	7 months	Naiki et al. 2016 [64]
AAVRh10 CYP21A2	H-2^aw18 mice (Cyp21a1^-/-)	2 × 10¹³ vgc/kg Intravenous injection	Correction in body weight up to 15 weeks. Correction of kidney renin expression at 18 weeks. Urinary progesterone near WT levels up to 15 weeks. Correction in stress response behavioural traits. No correction in adrenocortical morphology.	18 weeks	Perdomini et al. 2017 [5]
AAVRh10 CYP21A2-HA	H-2^aw18 mice (Cyp21a1^+/+)	6.5 × 10¹¹ vgc/mouse Intravenous injection	Left adrenal showed waning CYP21A2 expression, from 65% positive cells (2 weeks) to 2.1% positive cells (32 weeks). Liver retained CYP21A2 expression at 32 weeks, while adrenal gland lost expression.	16 weeks	Markmann et al. 2018 [4]
AAVRh10 CYP21A2-HA	H-2^aw18 mice (Cyp21a1^-/-)	6.5 × 10¹¹ vgc/mouse Intravenous injection	Progesterone significantly decreased at 2 weeks, no effect by 10 weeks. ACTH decreased from 2–8 weeks, no significant effect by 32 weeks.	8 weeks	Markmann et al. 2018 [4]
AAVRh10 CYP21A2	Cynomolgus macaques	5 × 10¹², 1.5 × 10¹³, or 4.5 × 10¹³ vgc/kg Intravenous injection	Detectable vector genomes in adrenals and liver up to 24 weeks (high dose). Transgene expression increased from 4 to 12 weeks, decline evident at 24 weeks (high dose).	24 weeks	Eclov et al. 2020 [66]
AAV9 CYP11B1	Cyp11b1^-/- mice	1 × 10¹⁰ vgc/adrenal Intra-adrenal injection	All mice (n = 4) had significant decrease in serum deoxycorticosterone/corticosterone ratio at 4 weeks. Serum deoxycorticosterone/corticosterone ratio remained low in 2 mice followed to 5 months. No apparent transgene expression in adrenal glands of injected mice 11B-hydroxylase activity did not improve (data not shown).	22 weeks	Naiki et al. 2022 [61]
AAV8 CYP21A2	H-2^aw18 mice (Cyp21a1^-/-)	5 × 10¹¹ vgc/mouse Intravenous injection	All mice (n = 10) normalised serum aldosterone and renal renin expression. All mice increased serum corticosterone and reduced adrenal hyperplasia.	4 weeks	Graves et al. 2024 [67]

AAV: adeno-associated virus; ACTH: adrenocorticotropic hormone; CAH: congenital adrenal hyperplasia; pfu: plaque-forming units; vgc: vector genome copies; WT: wild-type

Declarations

Acknowledgments

Figures were created in Biorender.com.

Author contributions

EBVD: Conceptualization, Writing–original draft, Writing–review & editing. SLG and IEA: Validation, Writing–review & editing, Supervision. LEG: Conceptualization, Writing–review & editing, Supervision. All authors read and approved the submitted version.

Conflicts of interest

The authors declare that they have no conflicts of interest.

Ethical approval

Not applicable.

Consent to participate

Not applicable.

Consent to publication

Not applicable.

Availability of data and materials

Not applicable.

Funding

EBVD was supported by the Research Training Program Scholarship, Australian Government. EBVD was additionally supported by the Children’s Medical Research Institute PhD Scholarship. LEG was supported by the Yass Memorial Scholarship, Children’s Medical Research Institute. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Copyright

References

Mallappa A, Merke DP. Management challenges and therapeutic advances in congenital adrenal hyperplasia. Nat Rev Endocrinol. 2022;18:337–52. [DOI] [PubMed] [PMC]

Speiser PW, Arlt W, Auchus RJ, Baskin LS, Conway GS, Merke DP, et al. Congenital adrenal hyperplasia due to steroid 21-hydroxylase deficiency: an endocrine society clinical practice guideline. J Clin Endocrinol Metab. 2018;103:4043–88. [DOI] [PubMed] [PMC]

Graves LE, Torpy DJ, Coates PT, Alexander IE, Bornstein SR, Clarke B. Future directions for adrenal insufficiency: cellular transplantation and genetic therapies. J Clin Endocrinol Metab. 2023;108:1273–89. [DOI] [PubMed]

Markmann S, De BP, Reid J, Jose CL, Rosenberg JB, Leopold PL, et al. Biology of the adrenal gland cortex obviates effective use of adeno-associated virus vectors to treat hereditary adrenal disorders. Hum Gene Ther. 2018;29:403–12. [DOI] [PubMed]

Perdomini M, Dos Santos C, Goumeaux C, Blouin V, Bougnères P. An AAVrh10-CAG-CYP21-HA vector allows persistent correction of 21-hydroxylase deficiency in a Cyp21^-/- mouse model. Gene Ther. 2017;24:275–81. [DOI] [PubMed]

Doudna JA, Charpentier E. Genome editing. The new frontier of genome engineering with CRISPR-Cas9. Science. 2014;346:1258096. [DOI] [PubMed]

Grabek A, Dolfi B, Klein B, Jian-Motamedi F, Chaboissier MC, Schedl A. The adult adrenal cortex undergoes rapid tissue renewal in a sex-specific manner. Cell Stem Cell. 2019;25:290–6. [DOI] [PubMed]

Fu Y, Foden JA, Khayter C, Maeder ML, Reyon D, Joung JK, et al. High-frequency off-target mutagenesis induced by CRISPR-Cas nucleases in human cells. Nat Biotechnol. 2013;31:822–6. [DOI] [PubMed] [PMC]

Buchlis G, Podsakoff GM, Radu A, Hawk SM, Flake AW, Mingozzi F, et al. Factor IX expression in skeletal muscle of a severe hemophilia B patient 10 years after AAV-mediated gene transfer. Blood. 2012;119:3038–41. [DOI] [PubMed] [PMC]

10.

Lu B, Javidi-Parsijani P, Makani V, Mehraein-Ghomi F, Sarhan WM, Sun D, et al. Delivering SaCas9 mRNA by lentivirus-like bionanoparticles for transient expression and efficient genome editing. Nucleic Acids Res. 2019;47:e44. [DOI] [PubMed] [PMC]

11.

Finn JD, Smith AR, Patel MC, Shaw L, Youniss MR, van Heteren J, et al. A single administration of CRISPR/Cas9 lipid nanoparticles achieves robust and persistent in vivo genome editing. Cell Rep. 2018;22:2227–35. [DOI] [PubMed]

12.

Liu J, Chang J, Jiang Y, Meng X, Sun T, Mao L, et al. Fast and efficient CRISPR/Cas9 genome editing in vivo enabled by bioreducible lipid and messenger RNA nanoparticles. Adv Mater. 2019;31:e1902575. [DOI] [PubMed] [PMC]

13.

Qiu M, Glass Z, Chen J, Haas M, Jin X, Zhao X, et al. Lipid nanoparticle-mediated codelivery of Cas9 mRNA and single-guide RNA achieves liver-specific in vivo genome editing of Angptl3. Proc Natl Acad Sci U S A. 2021;118:e2020401118. [DOI] [PubMed] [PMC]

14.

Musunuru K, Chadwick AC, Mizoguchi T, Garcia SP, DeNizio JE, Reiss CW, et al. In vivo CRISPR base editing of PCSK9 durably lowers cholesterol in primates. Nature. 2021;593:429–34. [DOI] [PubMed]

15.

Assessment report: Onpattro [Internet]. London: European Medicines Agency; c2018 [cited 2024 Jan 21]. Available from: https://www.ema.europa.eu/en/documents/assessment-report/onpattro-epar-public-assessment-report_en.pdf

16.

Mérian J, Boisgard R, Decleves X, Thezé B, Texier I, Tavitian B. Synthetic lipid nanoparticles targeting steroid organs. J Nucl Med. 2013;54:1996–2003. [DOI] [PubMed]

17.

Gidlöf S, Falhammar H, Thilén A, von Döbeln U, Ritzén M, Wedell A, et al. One hundred years of congenital adrenal hyperplasia in Sweden: a retrospective, population-based cohort study. Lancet Diabetes Endocrinol. 2013;1:35–42. [DOI] [PubMed]

18.

Claahsen-van der Grinten HL, Speiser PW, Ahmed SF, Arlt W, Auchus RJ, Falhammar H, et al. Congenital adrenal hyperplasia-current insights in pathophysiology, diagnostics, and management. Endocr Rev. 2022;43:91–159. [DOI] [PubMed] [PMC]

19.

Lai F, Srinivasan S, Wiley V. Evaluation of a two-tier Screening pathway for congenital adrenal hyperplasia in the New South Wales newborn screening programme. Int J Neonatal Screen. 2020;6:63. [DOI] [PubMed] [PMC]

20.

Krone N, Arlt W. Genetics of congenital adrenal hyperplasia. Best Pract Res Clin Endocrinol Metab. 2009;23:181–92. [DOI] [PubMed] [PMC]

21.

Turcu AF, Auchus RJ. Adrenal steroidogenesis and congenital adrenal hyperplasia. Endocrinol Metab Clin North Am. 2015;44:275–96. [DOI] [PubMed] [PMC]

22.

Falhammar H, Wedell A, Nordenström A. Biochemical and genetic diagnosis of 21-hydroxylase deficiency. Endocrine. 2015;50:306–14. [DOI] [PubMed]

23.

Bertagna X. Effects of chronic ACTH excess on human adrenal cortex. Front Endocrinol (Lausanne). 2017;8:43. [DOI] [PubMed] [PMC]

24.

El-Maouche D, Arlt W, Merke DP. Congenital adrenal hyperplasia. Lancet. 2017;390:2194–210. [DOI] [PubMed]

25.

Krone N, Braun A, Roscher AA, Knorr D, Schwarz HP. Predicting phenotype in steroid 21-hydroxylase deficiency? Comprehensive genotyping in 155 unrelated, well defined patients from southern Germany. J Clin Endocrinol Metab. 2000;85:1059–65. [DOI] [PubMed]

26.

Finkielstain GP, Chen W, Mehta SP, Fujimura FK, Hanna RM, Van Ryzin C, et al. Comprehensive genetic analysis of 182 unrelated families with congenital adrenal hyperplasia due to 21-hydroxylase deficiency. J Clin Endocrinol Metab. 2011;96:E161–72. [DOI] [PubMed] [PMC]

27.

Yau M, Gujral J, New MI. Congenital adrenal hyperplasia: diagnosis and emergency treatment. In: Feingold KR, Anawalt B, Blackman MR, Boyce A, Chrousos G, Corpas E, et al., editors. Endotext. South Dartmouth (MA): MDText.com, Inc.; 2000. [PubMed]

28.

New MI. Extensive clinical experience: nonclassical 21-hydroxylase deficiency. J Clin Endocrinol Metab. 2006;91:4205–14. [DOI] [PubMed]

29.

Falhammar H, Nordenström A. Nonclassic congenital adrenal hyperplasia due to 21-hydroxylase deficiency: clinical presentation, diagnosis, treatment, and outcome. Endocrine. 2015;50:32–50. [DOI] [PubMed]

30.

Tusie-Luna MT, Traktman P, White PC. Determination of functional effects of mutations in the steroid 21-hydroxylase gene (CYP21) using recombinant vaccinia virus. J Biol Chem. 1990;265:20916–22. [DOI] [PubMed]

31.

Rushworth RL, Torpy DJ, Falhammar H. Adrenal crisis. N Engl J Med. 2019;381:852–61. [DOI] [PubMed]

32.

Reisch N, Willige M, Kohn D, Schwarz HP, Allolio B, Reincke M, et al. Frequency and causes of adrenal crises over lifetime in patients with 21-hydroxylase deficiency. Eur J Endocrinol. 2012;167:35–42. [DOI] [PubMed]

33.

Falhammar H, Frisén L, Norrby C, Hirschberg AL, Almqvist C, Nordenskjöld A, et al. Increased mortality in patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency. J Clin Endocrinol Metab. 2014;99:E2715–21. [DOI] [PubMed]

34.

Keely E, Malcolm J. Congenital adrenal hyperplasia in pregnancy: approach depends on who is the ‘patient’. Obstet Med. 2012;5:154–60. [DOI] [PubMed] [PMC]

35.

Muthusamy K, Elamin MB, Smushkin G, Murad MH, Lampropulos JF, Elamin KB, et al. Clinical review: adult height in patients with congenital adrenal hyperplasia: a systematic review and metaanalysis. J Clin Endocrinol Metab. 2010;95:4161–72. [DOI] [PubMed]

36.

Mendes-Dos-Santos CT, Martins DL, Guerra-Júnior G, Baptista MTM, de-Mello MP, de Oliveira LC, et al. Prevalence of testicular adrenal rest tumor and factors associated with its development in congenital adrenal hyperplasia. Horm Res Paediatr. 2018;90:161–8. [DOI] [PubMed]

37.

Jenkins-Jones S, Parviainen L, Porter J, Withe M, Whitaker MJ, Holden SE, et al. Poor compliance and increased mortality, depression and healthcare costs in patients with congenital adrenal hyperplasia. Eur J Endocrinol. 2018;178:309–20. [DOI] [PubMed]

38.

Aso K, Izawa M, Higuchi A, Kotoh S, Hasegawa Y. Stress doses of glucocorticoids cannot prevent progression of all adrenal crises. Clin Pediatr Endocrinol. 2009;18:23–7. [DOI] [PubMed] [PMC]

39.

Bartter FC, Albright F, Forbes AP, Leaf A, Dempsey E, Carroll E. The effects of adrenocorticotropic hormone and cortisone in the adrenogenital syndrome associated with congenital adrenal hyperplasia: an attempt to explain and correct its disordered hormonal pattern. J Clin Invest. 1951;30:237–51. [DOI] [PubMed] [PMC]

40.

Wilkins L, Lewis RA, Klein R, Gardner LI, Crigler JF Jr, Rosemberg E, et al. Treatment of congenital adrenal hyperplasia with cortisone. J Clin Endocrinol Metab. 1951;11:1–25. [DOI] [PubMed]

41.

Wilkins L, Lewis RA, Klein R, Rosemberg E. The suppression of androgen secretion by cortisone in a case of congenital adrenal hyperplasia. Bull Johns Hopkins Hosp. 1950;86:249–52. [PubMed]

42.

Schröder MAM, Claahsen-van der Grinten HL. Novel treatments for congenital adrenal hyperplasia. Rev Endocr Metab Disord. 2022;23:631–45. [DOI] [PubMed] [PMC]

43.

Weitzman ED, Fukushima D, Nogeire C, Roffwarg H, Gallagher TF, Hellman L. Twenty-four hour pattern of the episodic secretion of cortisol in normal subjects. J Clin Endocrinol Metab. 1971;33:14–22. [DOI] [PubMed]

44.

Mallappa A, Sinaii N, Kumar P, Whitaker MJ, Daley LA, Digweed D, et al. A phase 2 study of Chronocort, a modified-release formulation of hydrocortisone, in the treatment of adults with classic congenital adrenal hyperplasia. J Clin Endocrinol Metab. 2015;100:1137–45. [DOI] [PubMed] [PMC]

45.

Nella AA, Mallappa A, Perritt AF, Gounden V, Kumar P, Sinaii N, et al. A phase 2 study of continuous subcutaneous hydrocortisone infusion in adults with congenital adrenal hyperplasia. J Clin Endocrinol Metab. 2016;101:4690–8. [DOI] [PubMed] [PMC]

46.

Wright C, O’Day P, Alyamani M, Sharifi N, Auchus RJ. Abiraterone acetate treatment lowers 11-oxygenated androgens. Eur J Endocrinol. 2020;182:413–21. [DOI] [PubMed] [PMC]

47.

Merke DP, Mallappa A, Arlt W, Brac de la Perriere A, Lindén Hirschberg A, Juul A, et al. Modified-release hydrocortisone in congenital adrenal hyperplasia. J Clin Endocrinol Metab. 2021;106:e2063–77. [DOI] [PubMed] [PMC]

48.

Mallappa A, Nella AA, Sinaii N, Rao H, Gounden V, Perritt AF, et al. Long-term use of continuous subcutaneous hydrocortisone infusion therapy in patients with congenital adrenal hyperplasia. Clin Endocrinol (Oxf). 2018;89:399–407. [DOI] [PubMed] [PMC]

49.

Pang S, Hotchkiss J, Drash AL, Levine LS, New MI. Microfilter paper method for 17 alpha-hydroxyprogesterone radioimmunoassay: its application for rapid screening for congenital adrenal hyperplasia. J Clin Endocrinol Metab. 1977;45:1003–8. [DOI] [PubMed]

50.

Gene, cell, & RNA therapy landscape Q2 2021 quarterly data report [Internet]. Waukesha: American Society of Gene + Cell Therapy; [cited 2024 Jan 21]. Available from: https://www.asgct.org/global/documents/asgct-pharma-intelligence-quarterly-report-july-20.aspx

51.

Atchison RW, Casto BC, Hammon WM. Adenovirus-associated defective virus particles. Science. 1965;149:754–6. [DOI] [PubMed]

52.

Li C, Samulski RJ. Engineering adeno-associated virus vectors for gene therapy. Nat Rev Genet. 2020;21:255–72. [DOI] [PubMed]

53.

Westhaus A, Cabanes-Creus M, Rybicki A, Baltazar G, Navarro RG, Zhu E, et al. High-Throughput in vitro, ex vivo, and in vivo screen of adeno-associated virus vectors based on physical and functional transduction. Hum Gene Ther. 2020;31:575–89. [DOI] [PubMed] [PMC]

54.

Lee EJ, Guenther CM, Suh J. Adeno-associated virus (AAV) vectors: rational design strategies for capsid engineering. Curr Opin Biomed Eng. 2018;7:58–63. [DOI] [PubMed] [PMC]

55.

Alexander IE, Cunningham SC, Logan GJ, Christodoulou J. Potential of AAV vectors in the treatment of metabolic disease. Gene Ther. 2008;15:831–9. [DOI] [PubMed]

56.

Srivastava A. In vivo tissue-tropism of adeno-associated viral vectors. Curr Opin Virol. 2016;21:75–80. [DOI] [PubMed] [PMC]

57.

Cunningham SC, Dane AP, Spinoulas A, Alexander IE. Gene delivery to the juvenile mouse liver using AAV2/8 Vectors. Mol Ther. 2008;16:1081–8. [DOI] [PubMed]

58.

Penaud-Budloo M, Le Guiner C, Nowrouzi A, Toromanoff A, Chérel Y, Chenuaud P, et al. Adeno-associated virus vector genomes persist as episomal chromatin in primate muscle. J Virol. 2008;82:7875–85. [DOI] [PubMed] [PMC]

59.

Riepe FG, Tatzel S, Sippell WG, Pleiss J, Krone N. Congenital adrenal hyperplasia: the molecular basis of 21-hydroxylase deficiency in H-2(aw18) mice. Endocrinology. 2005;146:2563–74. [DOI] [PubMed]

60.

Gotoh H, Sagai T, Hata J, Shiroishi T, Moriwaki K. Steroid 21-hydroxylase deficiency in mice. Endocrinology. 1988;123:1923–7. [DOI] [PubMed]

61.

Naiki Y, Miyado M, Shindo M, Horikawa R, Hasegawa Y, Katsumata N, et al. Adeno-associated virus-mediated gene therapy for patients’ fibroblasts, induced pluripotent stem cells, and a mouse model of congenital adrenal hyperplasia. Hum Gene Ther. 2022;33:801–9. [DOI] [PubMed]

62.

Tajima T, Okada T, Ma XM, Ramsey W, Bornstein S, Aguilera G. Restoration of adrenal steroidogenesis by adenovirus-mediated transfer of human cytochromeP450 21-hydroxylase into the adrenal gland of 21-hydroxylase-deficient mice. Gene Ther. 1999;6:1898–903. [DOI] [PubMed]

63.

Macapagal MC, Slowinska BS, Nimkarn S, De BP, Licholai T, Marshall I, et al. Gene therapy of 21-hydroxylase deficient mice utilizing an adeno-associated virus vector. ENDO 2002: the Endocrine Society’s 84th annual meeting; 2002 June 19–22; San Francisco, US.

64.

Naiki Y, Miyado M, Horikawa R, Katsumata N, Onodera M, Pang S, et al. Extra-adrenal induction of Cyp21a1 ameliorates systemic steroid metabolism in a mouse model of congenital adrenal hyperplasia. Endocr J. 2016;63:897–904. [DOI] [PubMed]

65.

Merke DP, Auchus RJ, Sarafoglou K, Geffner ME, Kim MS, Escandon RD, et al. Design of a phase 1/2 open-label, dose-escalation study of the safety and efficacy of gene therapy in adults with classic congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency through administration of an adeno-associated virus (AAV) serotype 5-based recombinant vector encoding the human CYP21A2 gene. J Endocr Soc. 2021;5:A82. [DOI]

66.

Eclov RJ, Lewis TEW, Kapadia M, Scott DW, McCoy DD, Rouse JL, et al. OR25-01 durable CYP21A2 gene therapy in non-human primates for treatment of congenital adrenal hyperplasia. J Endocr Soc. 2020;4:OR25–01. [DOI]

67.

Graves LE, van Dijk EB, Zhu E, Koyyalamudi S, Wotton T, Sung D, et al. AAV-delivered hepato-adrenal cooperativity in steroidogenesis: implications for gene therapy for congenital adrenal hyperplasia. Mol Ther Methods Clin Dev. 2024;32:101232. [DOI] [PubMed] [PMC]

68.

CAH letter to community Q1 2023 [Internet]. Raleigh: Adrenas Therapeutics; [cited 2024 Jan 21]. Available from: https://adrenastx.com/wp-content/uploads/CAH-Letter-to-Community-Q1-2023-Update-FINAL.pdf

69.

ADventure trial update [Internet]. Raleigh: Adrenas Therapeutics; [cited 2024 Jan 21]. Available from: https://cahgenetherapy.com/news/jan24

70.

White PC. Emerging treatment for congenital adrenal hyperplasia. Curr Opin Endocrinol Diabetes Obes. 2022;29:271–6. [DOI] [PubMed] [PMC]

71.

Finco I, Mohan DR, Hammer GD, Lerario AM. Regulation of stem and progenitor cells in the adrenal cortex. Curr Opin Endocr Metab Res. 2019;8:66–71. [DOI] [PubMed] [PMC]

72.

Chang SP, Morrison HD, Nilsson F, Kenyon CJ, West JD, Morley SD. Cell proliferation, movement and differentiation during maintenance of the adult mouse adrenal cortex. PLoS One. 2013;8:e81865. [DOI] [PubMed] [PMC]

73.

Ingle DJ, Higgins GM. Autotransplantation and regeneration of the adrenal gland. Endocrinology. 1938;22:458–64. [DOI]

74.

King P, Paul A, Laufer E. Shh signaling regulates adrenocortical development and identifies progenitors of steroidogenic lineages. Proc Natl Acad Sci U S A. 2009;106:21185–90. [DOI] [PubMed] [PMC]

75.

Iannaccone P, Morley S, Skimina T, Mullins J, Landini G. Cord-like mosaic patches in the adrenal cortex are fractal: implications for growth and development. FASEB J. 2003;17:41–3. [DOI] [PubMed]

76.

Salmon TN, Zwemer RL. A study of the life history of cortico‐adrenal gland cells of the rat by means of trypan blue injections. Anat. Rec. 1941;80:421–9. [DOI]

77.

Zwemer RL. A study of adrenal cortex morphology. Am J Pathol. 1936;12:107–14. [DOI] [PubMed] [PMC]

78.

Freedman BD, Kempna PB, Carlone DL, Shah M, Guagliardo NA, Barrett PQ, et al. Adrenocortical zonation results from lineage conversion of differentiated zona glomerulosa cells. Dev Cell. 2013;26:666–73. [DOI] [PubMed] [PMC]

79.

Wang D, Zhang F, Gao G. CRISPR-based therapeutic genome editing: strategies and in vivo delivery by AAV vectors. Cell. 2020;181:136–50. [DOI] [PubMed] [PMC]

80.

Stenson PD, Mort M, Ball EV, Chapman M, Evans K, Azevedo L, et al. The human gene mutation database (HGMD^®): optimizing its use in a clinical diagnostic or research setting. Hum Genet. 2020;139:1197–207. [DOI] [PubMed] [PMC]

81.

Jinek M, Chylinski K, Fonfara I, Hauer M, Doudna JA, Charpentier E. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science. 2012;337:816–21. [DOI] [PubMed] [PMC]

82.

Pickar-Oliver A, Gersbach CA. The next generation of CRISPR-Cas technologies and applications. Nat Rev Mol Cell Biol. 2019;20:490–507. [DOI] [PubMed] [PMC]

83.

Higashi Y, Yoshioka H, Yamane M, Gotoh O, Fujii-Kuriyama Y. Complete nucleotide sequence of two steroid 21-hydroxylase genes tandemly arranged in human chromosome: a pseudogene and a genuine gene. Proc Natl Acad Sci U S A. 1986;83:2841–5. [DOI] [PubMed] [PMC]

84.

Pignatelli D, Carvalho BL, Palmeiro A, Barros A, Guerreiro SG, Macut D. The complexities in genotyping of congenital adrenal hyperplasia: 21-hydroxylase deficiency. Front Endocrinol (Lausanne). 2019;10:432. [DOI] [PubMed] [PMC]

85.

Rodrigues NR, Dunham I, Yu CY, Carroll MC, Porter RR, Campbell RD. Molecular characterization of the HLA-linked steroid 21-hydroxylase B gene from an individual with congenital adrenal hyperplasia. EMBO J. 1987;6:1653–61. [DOI] [PubMed] [PMC]

86.

White PC, New MI, Dupont B. Structure of human steroid 21-hydroxylase genes. Proc Natl Acad Sci U S A. 1986;83:5111–5. [DOI] [PubMed] [PMC]

87.

White PC, Speiser PW. Congenital adrenal hyperplasia due to 21-hydroxylase deficiency. Endocr Rev. 2000;21:245–91. [DOI] [PubMed]

88.

Wang C, Pallan PS, Zhang W, Lei L, Yoshimoto FK, Waterman MR, et al. Functional analysis of human cytochrome P450 21A2 variants involved in congenital adrenal hyperplasia. J Biol Chem. 2017;292:10767–78. [DOI] [PubMed] [PMC]

89.

Chapman JR, Taylor MR, Boulton SJ. Playing the end game: DNA double-strand break repair pathway choice. Mol Cell. 2012;47:497–510. [DOI] [PubMed]

90.

Lieber MR. The mechanism of double-strand DNA break repair by the nonhomologous DNA end-joining pathway. Annu Rev Biochem. 2010;79:181–211. [DOI] [PubMed] [PMC]

91.

Iyer S, Suresh S, Guo D, Daman K, Chen JCJ, Liu P, et al. Precise therapeutic gene correction by a simple nuclease-induced double-stranded break. Nature. 2019;568:561–5. [DOI] [PubMed] [PMC]

92.

Shen MW, Arbab M, Hsu JY, Worstell D, Culbertson SJ, Krabbe O, et al. Predictable and precise template-free CRISPR editing of pathogenic variants. Nature. 2018;563:646–51. [DOI] [PubMed] [PMC]

93.

Moynahan ME, Jasin M. Mitotic homologous recombination maintains genomic stability and suppresses tumorigenesis. Nat Rev Mol Cell Biol. 2010;11:196–207. [DOI] [PubMed] [PMC]

94.

Riesenberg S, Kanis P, Macak D, Wollny D, Düsterhöft D, Kowalewski J, et al. Efficient high-precision homology-directed repair-dependent genome editing by HDRobust. Nat Methods. 2023;20:1388–99. [DOI] [PubMed] [PMC]

95.

Liu M, Rehman S, Tang X, Gu K, Fan Q, Chen D, et al. Methodologies for improving HDR efficiency. Front Genet. 2019;9:691. [DOI] [PubMed] [PMC]

96.

Ginn SL, Christina S, Alexander IE. Genome editing in the human liver: progress and translational considerations. Prog Mol Biol Transl Sci. 2021;182:257–88. [DOI] [PubMed]

97.

Suzuki K, Tsunekawa Y, Hernandez-Benitez R, Wu J, Zhu J, Kim EJ, et al. In vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integration. Nature. 2016;540:144–9. [DOI] [PubMed] [PMC]

98.

McCarty DM, Young SM Jr, Samulski RJ. Integration of adeno-associated virus (AAV) and recombinant AAV vectors. Annu Rev Genet. 2004;38:819–45. [DOI] [PubMed]

99.

Zuris JA, Thompson DB, Shu Y, Guilinger JP, Bessen JL, Hu JH, et al. Cationic lipid-mediated delivery of proteins enables efficient protein-based genome editing in vitro and in vivo. Nat Biotechnol. 2015;33:73–80. [DOI] [PubMed] [PMC]

100.

Aguirre AJ, Meyers RM, Weir BA, Vazquez F, Zhang CZ, Ben-David U, et al. Genomic copy number dictates a gene-independent cell response to CRISPR/Cas9 targeting. Cancer Discov. 2016;6:914–29. [DOI] [PubMed] [PMC]

101.

Ranzani M, Annunziato S, Adams DJ, Montini E. Cancer gene discovery: exploiting insertional mutagenesis. Mol Cancer Res. 2013;11:1141–58. [DOI] [PubMed] [PMC]

102.

Mullard A. Gene therapy community grapples with toxicity issues, as pipeline matures. Nat Rev Drug Discov. 2021;20:804–5. [DOI] [PubMed]

103.

Paulk N. Gene therapy: it is time to talk about high-dose AAV. Genet Eng Biotechnol News. 2020;40:14–6. [DOI]

104.

Mukai H, Ogawa K, Kato N, Kawakami S. Recent advances in lipid nanoparticles for delivery of nucleic acid, mRNA, and gene editing-based therapeutics. Drug Metab Pharmacokinet. 2022;44:100450. [DOI] [PubMed] [PMC]

105.

Akinc A, Maier MA, Manoharan M, Fitzgerald K, Jayaraman M, Barros S, et al. The Onpattro story and the clinical translation of nanomedicines containing nucleic acid-based drugs. Nat Nanotechnol. 2019;14:1084–7. [DOI] [PubMed]

106.

Mulligan MJ, Lyke KE, Kitchin N, Absalon J, Gurtman A, Lockhart S, et al. Phase I/II study of COVID-19 RNA vaccine BNT162b1 in adults. Nature. 2020;586:589–93. [DOI] [PubMed]

107.

Reichmuth AM, Oberli MA, Jaklenec A, Langer R, Blankschtein D. mRNA vaccine delivery using lipid nanoparticles. Ther Deliv. 2016;7:319–34. [DOI] [PubMed] [PMC]

108.

Xia Y, Tian J, Chen X. Effect of surface properties on liposomal siRNA delivery. Biomaterials. 2016;79:56–68. [DOI] [PubMed] [PMC]

109.

Zadory M, Lopez E, Babity S, Gravel SP, Brambilla D. Current knowledge on the tissue distribution of mRNA nanocarriers for therapeutic protein expression. Biomater Sci. 2022;10:6077–115. [DOI] [PubMed]

110.

Dilliard SA, Siegwart DJ. Passive, active and endogenous organ-targeted lipid and polymer nanoparticles for delivery of genetic drugs. Nat Rev Mater. 2023;8:282–300. [DOI] [PubMed] [PMC]

111.

Durymanov M, Reineke J. Non-viral delivery of nucleic acids: insight into mechanisms of overcoming intracellular barriers. Front Pharmacol. 2018;9:971. [DOI] [PubMed] [PMC]

112.

Taha EA, Lee J, Hotta A. Delivery of CRISPR-Cas tools for in vivo genome editing therapy: trends and challenges. J Control Release. 2022;342:345–61. [DOI] [PubMed]

113.

Li S, Hu Y, Li A, Lin J, Hsieh K, Schneiderman Z, et al. Payload distribution and capacity of mRNA lipid nanoparticles. Nat Commun. 2022;13:5561. [DOI] [PubMed] [PMC]

114.

Bolous NS, Chen Y, Wang H, Davidoff AM, Devidas M, Jacobs TW, et al. The cost-effectiveness of gene therapy for severe hemophilia B: a microsimulation study from the United States perspective. Blood. 2021;138:1677–90. [DOI] [PubMed]

115.

Kis Z, Kontoravdi C, Shattock R, Shah N. Resources, production scales and time required for producing RNA vaccines for the global pandemic demand. Vaccines (Basel). 2020;9:3. [DOI] [PubMed] [PMC]

116.

Pardi N, Tuyishime S, Muramatsu H, Kariko K, Mui BL, Tam YK, et al. Expression kinetics of nucleoside-modified mRNA delivered in lipid nanoparticles to mice by various routes. J Control Release. 2015;217:345–51. [DOI] [PubMed] [PMC]

117.

Kenjo E, Hozumi H, Makita Y, Iwabuchi KA, Fujimoto N, Matsumoto S, et al. Low immunogenicity of LNP allows repeated administrations of CRISPR-Cas9 mRNA into skeletal muscle in mice. Nat Commun. 2021;12:7101. [DOI] [PubMed] [PMC]

118.

Scallan CD, Jiang H, Liu T, Patarroyo-White S, Sommer JM, Zhou S, et al. Human immunoglobulin inhibits liver transduction by AAV vectors at low AAV2 neutralizing titers in SCID mice. Blood. 2006;107:1810–7. [DOI] [PubMed]

119.

Kraemer FB. Adrenal cholesterol utilization. Mol Cell Endocrinol. 2007;265–266:42–5. [DOI] [PubMed]

120.

Uhlén M, Fagerberg L, Hallström BM, Lindskog C, Oksvold P, Mardinoglu A, et al. Proteomics. Tissue-based map of the human proteome. Science. 2015;347:1260419. [DOI] [PubMed]

121.

Rosol TJ, Yarrington JT, Latendresse J, Capen CC. Adrenal gland: structure, function, and mechanisms of toxicity. Toxicol Pathol. 2001;29:41–8. [DOI] [PubMed]

122.

Burkhardt WA, Guscetti F, Boretti FS, Ivos Todesco A, Aldajarov N, Lutz TA, et al. Adrenocorticotropic hormone, but not trilostane, causes severe adrenal hemorrhage, vacuolization, and apoptosis in rats. Domest Anim Endocrinol. 2011;40:155–64. [DOI] [PubMed]

123.

Ryan US, Ryan JW, Smith DS, Winkler H. Fenestrated endothelium of the adrenal gland: freeze-fracture studies. Tissue Cell. 1975;7:181–90. [DOI] [PubMed]

124.

Zhang YN, Poon W, Tavares AJ, McGilvray ID, Chan WCW. Nanoparticle-liver interactions: cellular uptake and hepatobiliary elimination. J Control Release. 2016;240:332–48. [DOI] [PubMed]

125.

Eipel C, Abshagen K, Vollmar B. Regulation of hepatic blood flow: the hepatic arterial buffer response revisited. World J Gastroenterol. 2010;16:6046–57. [DOI] [PubMed] [PMC]

126.

Gomez-Sanchez CE. Regulation of adrenal arterial tone by adrenocorticotropin: the plot thickens. Endocrinology. 2007;148:3566–8. [DOI] [PubMed]

127.

Sapirstein LA, Goldman H. Adrenal blood flow in the albino rat. Am J Physiol. 1959;196:159–62. [DOI] [PubMed]

128.

Akinc A, Querbes W, De S, Qin J, Frank-Kamenetsky M, Jayaprakash KN, et al. Targeted delivery of RNAi therapeutics with endogenous and exogenous ligand-based mechanisms. Mol Ther. 2010;18:1357–64. [DOI] [PubMed] [PMC]