rAAV2-mediated restoration of LEKTI in LEKTI-deficient cells from Netherton patients
Introduction
Netherton syndrome (NS, MIM 256500) is an autosomal-recessive skin disorder clinically characterized by congenital erythroderma, trichorrhexis invaginata (bamboo hair), and atopic diathesis. It accounts for up to 18% of congenital erythrodermas [1], which can be life-threatening in the neonatal period due to severe complications such as hypernatremic dehydration, hypothermia, extreme weight loss, bronchopneumonia, recurrent bacterial infections and sepsis [2].
NS is caused by genetic defects in the gene SPINK5 (serine protease inhibitor kazal-type 5) [3], [4], [5], [6]. SPINK5 is located on chromosome 5q32 and encodes serine protease inhibitor LEKTI (lympho-epithelial Kazal-type related inhibitor). The protein LEKTI is a 15-domain Kazal-type related serine protease inhibitor [7], [8]. In the epidermis, LEKTI is confined to the granular layer and stratum corneum [6], [8] where it co-localizes with kallikreins (KLK) 5 and 7 [9]. Several domains of LEKTI have shown to inhibit KLK 5 and 7 [10] that are involved in the degradation of corneodesmosomes such as desmoglein 1 (Dsg1) and desmocollin 1 (Dsc1) [11]. Normally, the interplay between the inhibitory pressure of LEKTI and the proteolytic activity of KLK 5 und 7 is well-balanced leading to a controlled desquamation process and an intact skin barrier.
In NS, mutations result in a complete or severe loss of LEKTI and subsequently a reduced inhibition of serine proteases. The enhanced activity of KLK 5 and 7 induce premature degradation of corneodesmosomal cadherins as demonstrated by a dramatically reduction of Dsg1 and Dsc1 [11], [12]. This mechanism has been suggested to result in loss of stratum corneum integrity and severe skin barrier defect.
Various therapeutic options have been used in NS to reduce clinical symptoms, among them topical steroids, calcipotriene, calcineurin-inhibitors, topical and systemic retinoids, PUVA, and intravenous immunoglobulin all with variable and only temporary success [13], [14]. Today, no causal and permanent treatment for NS does exist, although the severity and continuity of the diseases are demanding new developments. Herein, we describe the first developmental steps towards a genetic treatment approach for NS.
Section snippets
Cell culture
Human primary NS-keratinocytes were isolated from punch biopsies derived from 4 NS patients after informed consent had been given; normal human keratinocytes (NHK) came from circumcised foreskin (provided by Dr. Thomas A. Angerpointer, Pediatric Surgery Practice, Munich, Germany). The specimens were processed according to a standard protocol described previously [15], [16], [17]. Briefly, skin was cleaned of fat, cut into small piece (if necessary) and incubated overnight at 4 °C in thermolysin
Netherton patients
Keratinocytes of four patients with typical clinical presentation of NS were available for the study. The underlying genetic defects are summarized in Table 1. Mutation c.316delAG and c.1432-13 G > A had been published previously [6], [9]. All mutations resulted in premature termination of SPINK5. In addition all demonstrated reduced mRNA expression of SPINK5, and increase hydrolytic activity consistent with a deficiency of LEKTI.
Increased expression of SPINK5 in Netherton keratinocytes following gene transfer of SPINK5
In the skin, SPINK5 expression is known to be restricted to the
Discussion
Netherton syndrome is a genetic disorder with severe and chronic skin manifestations that can currently only be treated in a symptomatic way. There are two promising molecular concepts at the horizon that might offer causal therapeutic benefit – protein replacement therapy and gene therapy. In theory, protein replacement by means of topical application of recombinant LEKTI might allow LEKTI to permeate the impaired skin barrier of NS patients and become active at the granular layer and thus
Conflict of interest
The authors declare no conflict of interest.
Acknowledgements
This work was supported by grants of the Wilhelm-Sander Stiftung (2003.117.1), Munich, Germany. Work presented in this publication was performed as part of the doctoral thesis of D.R.
References (26)
- et al.
The spectrum of pathogenic mutations in SPINK5 in 19 families with Netherton syndrome: implications for mutation detection and first case of prenatal diagnosis
J Invest Dermatol
(2001) - et al.
Netherton syndrome: disease expression and spectrum of SPINK5 mutations in 21 families
J Invest Dermatol
(2002) - et al.
SPINK5 and Netherton Syndrome: novel mutations, demonstration of missing LEKTI, and differential expression of transglutaminases
J Invest Dermatol
(2004) - et al.
Serine protease activity and residual LEKTI expression determine phenotype in Netherton syndrome
J Invest Dermatol
(2006) - et al.
Corneodesmosomal cadherins are preferential targets of stratum corneum trypsin- and chymotrypsin-like hyperactivity in Netherton syndrome
J Invest Dermatol
(2006) - et al.
Comèl–Netherton syndrome defined as primary immunodeficiency
J Allergy Clin Immunol
(2009) - et al.
Elevated stratum corneum hydrolytic activity in Netherton syndrome suggests an inhibitory regulation of desquamation by SPINK5-derived peptides
J Invest Dermatol
(2002) - et al.
Neonatal and infantile erythrodermas: a retrospective study of 51 patients
Arch Dermatol
(2000) - et al.
A clinical and immunological study of Netherton's syndrome
Br J Dermatol
(1994) - et al.
Mutations in SPINK5, encoding a serine protease inhibitor, cause Netherton syndrome
Nat. Genet
(2000)
Biochemical features, molecular biology and clinical relevance of the human 15-domain serine proteinase inhibitor LEKTI
Biol Chem
LEKTI proteolytic processiong in human primary keratinozytes, tissue distribution and defective expression in Netherton syndrome
Hum Mol Genet
LEKTI fragments specifically inhibit KLK5, KLK7, and KLK14 and control desquamation through a pH-dependent interaction
Mol Biol Cell
Cited by (25)
Nucleic acid delivery into skin for the treatment of skin disease: Proofs-of-concept, potential impact, and remaining challenges
2015, Journal of Controlled ReleaseCitation Excerpt :Essentially, protease hyperactivity results in excessive and premature epidermal desquamation [79]. Netherton syndrome may be treated by down-regulating production of serine proteases in the skin, or alternatively, inducing expression of wild-type serine protease inhibitor [80]. Due to the significant reduction in skin barrier function, initial treatment with topical NAs may not require dermal enhancement.
Proteolytic activation cascade of the netherton syndrome-defective protein, LEKTI, in the epidermis: Implications for skin homeostasis
2011, Journal of Investigative DermatologyCitation Excerpt :In addition, they gain upon the design of a previously unreported and effective therapeutic intervention for NS, based on the replacement of the missing LEKTI polypeptides. The currently utilized therapies for this life-threatening and disabling disease are only aimed at reducing the clinical symptoms (Saif and Al-Khenaizan, 2007;Renneret al., 2009). Nevertheless, in spite of the encouraging results (Diet al., 2010;Roedlet al., 2011), theex-vivo gene transfer cannot be envisaged as an imminent therapeutic option.
Netherton syndrome in a Bulgarian patient: Presentation of a case and an update of therapeutic options
2023, Wiener Medizinische WochenschriftGenome Editing in Therapy of Genodermatoses
2022, Molecular BiologyMetformin inhibits melanoma cell metastasis by suppressing the miR-5100/SPINK5/STAT3 axis
2022, Cellular and Molecular Biology LettersNetherton Syndrome in Children: Management and Future Perspectives
2021, Frontiers in Pediatrics