從遺傳性皮膚疾病看

皮膚生理的奧妙

成大醫院皮膚科 趙曉秋醫師 Genodermatoses

 Keratinization disorders

 Epidermal fragility disorders

 Multisystem disorders

 Cancer disorders

 Hair disorders

 Pigmentation disorders

 Porphyrias

Diseases Characterized by in

Diseases Keratin involved simplex K5, K14 Epidermolytic K1, K10 Epidermolytic palmoplantar K9 Nonepidermolytic K1 Focal nonepidermolytic palmoplantar K16 keratoderma bullosa of Siemens K2e White sponge K4, K13 type I K6a, K16 Pachyonychia congenita type II K6b, K17 Meesmann corneal dystrophy K3, K12 Unknown K7, K8, K18, K19, K20 Palmoplantar Keratoderma (PPK) EPPK

R162W the 1A segment of K9

Pachyonychia congenita type II (PC-2)

PC-1: K6a, K16 PC-2: K6b, K17 Dermatol Sinica 19(3):204-209, 2001 White Sponge Nevus (WSN)

*E417K in the 2B segment of K4 Br J Dermatol, 2003 Novel Mutations in the Helix Termination Motif of Keratin 3 and Keratin 12 in 2 Taiwanese Families with Meesmann Corneal Dystrophy

Ying-Ting Chen, MD,*† Sung-Huei Tseng, MD,* and Sheau-Chiou Chao, MD‡ Cornea 2005;24:928–932 Epidermolysis Bullosa (EB)

 EB is caused by defects in one or more of the that make up the . These proteins are defective because the genes that code for the proteins have mistakes in the code.

 3 major types of EB, 10 genes, which encode skin proteins

14 and 5, plectin (EBS)

VII (RDEB and DDEB)

, integrins, collagen XVII (JEB).

. Epidermolysis Bullosa * EBS-DM: R125C in the helix 1A of K14 * EBS-K: A413T in the 2B segment of K14

EBS: J Formos Med Assoc 100(6):407-411, 2000; J Formos Med Assoc 101(4):287-290, 2001 EBS

Dystrophic Epidermolysis Bullosa

 A hereditary mechanobullous disorder characterized by fragility of the skin and mucous membrane due to abnormalities of anchoring fibrils.

 Both dominant and recessive DEB have been shown to be caused by mutations in COL7A1, the encoding type VII collagen which is the major component of anchoring fibrils. Dystrophic Epidermolysis Bullosa

 In DEB, there is considerable phenotypic as well as genetic variability.

 In the majority of the cases with severe, Hallopeau-Siemens type of recessive DEB (HS-RDEB)

 mutilating scarring of the skin and complete absence of the anchoring fibrils

 premature termination codon mutations (PTC) in both COL7A1 Dystrophic Epidermolysis Bullosa

 The milder, mitis form of RDEB

 is frequently caused by a missense or an in-frame deletion in one or both alleles

 mutations which are less disruptive to the PCR Amplification and Heteroduplex Analyses

 Amplification of segments of COL7A1 spanning all 118 exons of the gene using primers synthesized on the basis of intronic sequences (GenBank L23982).

 Heteroduplex analysis using conformation- sensitive gel electrophoresis (CSGE).

 The PCR products demonstrating shifted bands were subjected to directed automated sequencing

吳xx，8 y/o boy

Mother: 3373insGG within the exon 25 Father: 7769delG within the exon 104

黃xx ，4 y/o girl ﹝借 卵﹞

Father: 4603 G>T, E1535X (exon 46) Mother: 8407+5 G>C (intron 113)

劉 x 廷

4歲 劉x順 12/yo boy 劉x廷 8 y/o boy

Father: 6182 G>A, G2061E (exon 74) Mother: 8620+26 G>A (intron 116) 劉xx 15 y/o girl

 The genetic confirmation of mutation in DEB has an impact in genetic counseling for patients.

 In recessive DEB, the risk of clinically unaffected parents producing an affected child in the subsequent is 25% and the patient having affected offspring is about as low as in the general population.  To cure EB, we must therefore introduce into the skin either enough of the correct form of the protein which is faulty (protein therapy), or the correct form of the gene that can make that protein (), or enough cells containing the correct gene ( therapy).  Protein therapy

 A Phase I clinical trial of collagen protein therapy in humans is planned for 2010

 Ex-vivo Gene Therapy

 Clinical trials of gene therapy for RDEB (type VII collagen) in humans planned for 2009/ 2010

 Hovnanian et al (France)

 Lane et al. (USA)

 Fibroblast cell therapy

 A Phase II trial of fibroblast therapy is planned for 2009/ 2010

 Bone-marrow stem-cell therapies

 -----

MalMal dede MeledaMeleda Disease,Disease SLURP-1

G86R

Pseudodominant inheritance with the G86R mutation in the ARS gene in Mal de Meleda Hailey-Hailey Disease & Darier’s Disease

HHD

DD Hailey-Hailey Disease & Darier’s Disease 2+  HHD: 3q21-24, ATP2C1, Ca -transport ATPase; sequesters calcium into Golgi

 DD: 12q23-24.1, ATP2A2 gene, encoded the sarco/endoplasmic reticulum Ca2+-ATPase type 2 isoform (SERCA2) 2+  Elevated cytoplasmic Ca might act by altering post-translation modification such as glycosylation, folding, trafficking and/or sorting of key molecules involved in cell-to-cell adhesion

 Mechanism: unclear

* HHD: Br J Dermatol 146(4):595-600, 2002; Dermatol Sinica, 2003 * DD : Br J Dermatol 146(6): 958-963, 2002

Multisystem disorders Disease Mutated gene Affected protein/function Dyskeratosis DKC1 Dyskerin congenita Incontinentia IKBKG NEMO (modulator of NF- pigmenti kappaB signaling)

Netherton syndrome SPINK5 inhibitor

Pseudoxanthoma ABCC6 MRP6, a multidrug elasticum resistance-associated protein Sjögren-Larsson FALDH Fatty aldehydrogenase syndrome

Trichorhinophalange TRPS1 TRPS1 al syndrome type I

Werner syndrome WRN DNA helicase X-linked anhidrotic EDA Ectodysplasin A NF1, NF2 Neurofibromin, Merlin Neurofibromatosis – SPINK5 –LEKT1 絲胺酸酶抑制劑

A *K754X

B

Pseudoxanthoma Elasticum (PXE)

 Progressive calcification of elastic fibers in skin, eye, and the cardiovascular system

The 27th Annual meeting of the Chinese Dermatological Society, Kaohsiung, November 17-18, 2001. PXE  ABCC6 locus at 16p13.1

 ATP-binding cassette subfamily C member 6 transporter  Multidrug resistance-associated protein 6 (MRP6)

 Physiological targets are not known

 Influence extracellular matrix environment for the correct

assembly of elastic fibers  Association in plasma lipoprotein ?  R1141X association with a sharply increased risk of CAD ?

*2477TC L826P Trichorhinophalangeal syndrome type I

.Momeni et al. (2000) positional cloned a gene, designated TRPS1, encodes a zinc finger protein .*Q705X, 1184delG, 1140CTA, A919T Dermatol Sinica 13:111-118, 1995; J Formos Med Assoc 100(9):635-638, 2001; The 3rd JSID, Sept 2000, Gifu, Japan X-linked hypohidrotic Ectodermal Dysplasia 性聯遺傳少汗性外胚層發育不良

 Ectodermal Dysplasia  150 syndromes of unknown pathogenesis  abnormal development of eccrine sweat glands, hair, and teeth  Hypohidrotic Ectodermal Dysplasia  estimated incidence 1 per 100,000 births  sparse hair, abnormal or missing teeth, lack of sweat glands  life-threatening and brain damaging episodes of hyperthermia X-linked hypohidrotic Ectodermal Dysplasia

 X-linked recessive

 X-linked anhidrotic (hypohidrotic) ectodermal dysplasia (XLHED or EDA, OMIM 305100)

 one of the most common types of EDs

 ED1, the gene for XLHED encodes Ectodysplasin A

 AR

 AD, less likely

A

B

C .Sparse eyebrows and eyelashes, and fine, light brown scalp hair .Fingernails and toenails were normal Skin was dry with widespread erythema, excoriation and lichenification Six conical-shaped teeth that had erupted at the age of 2 years

Homozygous sequence del nt 1242-1248 (CGCAGCA) Proband A

Wild-type sequence Exon 9 Father B

Heterozygous sequence Exon 9/del nt 1242-1248 Mother C The TNF-like ligand induces signals leading to cell death, proliferation or differentiation and it is important in the early epithelial-mesenchymal interaction that regulates ectodermal appendage formation Peutz-Jeghers Syndrome

 Peutz-Jeghers syndrome (PJS, OMIM 175200) is a rare autosomal dominant inherited disorder

 Characterized by melanocytic macules of the lips, buccal mucosa, and digits, multiple GI hamartomatous polyps, and an increased risk of various neoplasms (at least 10 times greater) Peutz-Jeghers Syndrome

 It occurs in approximately 1 in 8,300 to 29,000 live births

 PJS typically presents in early childhood with pigmentation or with complications of small bowel polyps – intussusception, obstruction or bleeding

Serine/Threonine Kinase STK11/LKB1 gene

 Extends over 1.3 kb and is composed of 9 exons

 Nearly 50% (18-70%) of patients with PJS are caused by germline mutations in the STK11/LKB1 gene

 LKB1/STK11 has a role in a number of pathways involved in control of cell growth Oncogene 2003; 22:4752-6. AGG (Arg)  AAG (Lys) Mutant R297K

Normal

Backward primer Congenital Atrichia (hairless gene) Nevoid basal cell carcinoma syndrome

 9q22.3

 PTCH functions as a

transmembrane receptor for the Sonic hedgehog protein (HH) and interacts with another transmembrane protein called SMO

 Gatekeeper gene

HHSP Mutations of PTCH gene in arsenic-related BCCs in Taiwan  Arsenical BD, SCC and BCC mainly occur in sun- protected sites

 Analyze the PTCH gene mutations in DNA isolated from arsenical BCCs, 75% of the BCCs (15/20) showed somatic mutation of PTCH gene

 PTC

 Cyclopamine

Molecular basis of inherited skin disease

 Clinical, biochemical, immuno- histochemical, and ultrastructural clues

 Genetic linkage and positional cloning analysis

 A better understanding of the significance of individual structural proteins and regulatory enzymes in keratinocyte adhesion and differentiation Clinical implications of molecular for patients with heritable skin diseases  Improved diagnosis and refined classification with prognostic implications

 Profound consequences for genetic counseling concerning the mode of inheritance

 DNA-based prenatal or pre-implantation genetic diagnosis in families at risk for recurrence

 Basis for future development of gene therapy and other novel treatment modalities Newer forms of therapy

 Gene manipulation

 in vivo (direct injection)

 ex vivo (grafting of transfected cultured keratinocytes) Difficult issues

 Stem cell identification

 Optimal vector design

 Delivery system

 Sustain long-term in vivo gene expression