DermPath Update

Curtis T. Thompson, M.D.

Associated Adjunct Professor

Department of Dermatology

Oregon Health Sciences University

Portland, Oregon

Thanks. Last lecture before adjourning to beautiful weather. Dont want to batter with slides but give you a fairly concise update of advances in dermpath. Mostly molecular but there are some that are relevant to clinical dermatologists, especially those reading the so-called bread and butter themselves.

Melanocytic Neoplasms

Nevus vs MelanomaStagingSentinel lymph nodeDesmoplastic MM

Nevus versus Melanoma

H&EProtein (Immunohistochemistry)DNA (Karyotypes)DNA probes (FISH)DNA probes fancy (CGH)Protein (Immunohistochemistry)H&E

Nevus versus Melanoma

DNA AlterationTagged DNA probesFluoresence in-situ hybridization (FISH)Comparative Genomic Hybridization (CGH)

Fluorescence in-situ hybridization (FISH)

FISH

Comparative Genomic Hybridization (CGH)

Comparative Genomic Hybridization (CGH)

Array CGH

Array CGH

CGHNevus vs Melanoma

Spitzs nevus vs Melanoma

Spitzs nevus vs Melanoma

Spitz--Chromosome arm 11p gain or normalMelanomaMore alterations

Spitzs nevus vs Melanoma--CGH

Clinical Problem--CGH takes too long

Nevus versus Melanoma

FISH Nevus vs Melanoma

Four areas of interestChromosome 6p256 centromere6q2311q13

Protein (Antigen/antibody IHC)

Overexpression of Protein in MM

Kashani-Sabet et al.5 Markers identified on gene expression dataSpitz, MM in nevus, dysplastic nevus, misdiagnosed nevi.Specificity 95%Sensitivity 91%JURY IS OUT BUT HAS POTENTIAL TO REPLACE FISH WHICH IS EXPENSIVE AND TECHNICALLY DIFFICULT.

Nevus versus Melanoma

H&EProtein (Immunohistochemistry)DNA (Karyotypes)DNA probes (FISH)DNA probes fancy (CGH)Protein (Immunohistochemistry)H&E

H&E?

Melanoma Staging

Good bye to Clarks levelsHello to mitotic figuresBalch et al. Final Version of 2009 AJCC Melanoma Staging and Classification. J Clin Oncol 23:4799v1, 2009.

Mitotic Figures

5X 40x fields is 1mm2Count the most active areasCount only dermal melanocytesBe careful of counting lymphocytes and other cells.Antiphosphohistone HC (Immunohistochemistry)Casper D et al. Use of Anti-phosphohistone H3 Immunohistochemistry to Determine Mitotic Rate in Thin Melanoma, Am J Dermatopathol, In press, 2010.

Mitotic Figures

Antiphosphohistone HC (Immunohistochemistry)Casper D et al. Use of Anti-phosphohistone H3 Immunohistochemistry to Determine Mitotic Rate in Thin Melanoma, Am J Dermatopathol, In press, 2010.

Antiphosphohistone--Mitotic Figures

Sentinel Lymph Node

Jury still out on utilityInterferon or not0.75? 1.00mm? Stage 1b?Kanzler MH, Sentinel node biopsy and standard of care for melanoma: A re-evaluation of the evidence. JAAD 62(5):880-882, 2010.

Desmoplastic MM

Not as bad of an actorDiscovered later (>Breslow)Soares de Almeida L et al. Desmoplastic malignant melanoma: a clinicopathologic analysis of 113 cases. Am J Dermatopathol 30(3):207-215, 2008.

Proliferative RateKi-67 IHC

Proliferative RateKi-67 IHC

Seborrheic keratosis versus SCCISNevus vs MelanomaTrichoepithelioma/trichoblastoma vs BCCTadashi T. Pigmented Bowen disease arising in pigmented reticulated seborrheic keratosis. Int J Clin Oncology, In press, 2010.

Alopecia

Cytokeratin 15 (Stem cell in bulge)decreased in cicatricial processesPozdnyakova O. Involvement of the bulge region in primary scarring alopecia, J Cutaneous Pathology 35:922-925. 2008.

Cytokeratin 15

Stem cell

Alopecia

Will loss of Cytokeratin 15 in cicatricial (primary scarring) alopecia have prognostic significance?

Nail

OnychomatricomaCentrifuging nail specimenOrienting specimen

Nail

Onychomatricoma

Nail matrix?tumor or reactiveEpithelial and stromal componentsDistinguish from fibroma/angiofibroma

Onychomatricoma

Nail

Centrifuging nail specimen

Takiguchi R et al. Centrifuging nail specimen debris increased sensitivity of fungus detection. Abstract. American Society of Dermatopathology, Chicago, IL October 1, 2009.

Nail Bed/Matrix Biopsy Processing

Nail Bed/Matrix Biopsy Processing

R

L

IdeaOrientation of Tissue

+

Tissue

Nail Bed/Matrix Biopsy Processing

Nail Bed/Matrix Biopsy Processing

Tissue on

Paper guide

Nail Bed/Matrix Biopsy Processing

Nail Bed/Matrix Biopsy Processing

Nail Bed/Matrix Biopsy Processing

Cassette

Tissue on

Paper Digit

Nail Bed/Matrix Biopsy Processing

Cover with sponge
to hold in place

Cassette

Tissue on

Paper Digit

Sponge

Medial

Lateral

Nail Bed/Matrix Biopsy Processing

Submit in an excess of

10% fresh formaldehyde

Medial

Lateral

Onychomycosis Diagnostics

Sampling is an issueSubungal debris is better than nail plate for sampling.

Chang A et al. A modified approach to the histologic diagnosis of onychomycosis. JAAD 57:849-53, 2007.

Nail Fungus Diagnostics
Centrifuging Debris

Sample is reduced in volume in a large volume centrifuge.Pellet moved to Cytospin (Fisher HealthCare) centrifuge.Debris is centrifuged directly onto a glass slide.

Cytospin

Cytospin

Cytospin

Study204 Cases

132 cases positive on initial nail PAS52 cases centrifuged after negative nail PAS18 cases positive on centrifugeTotal 150 positive casesnail and centrifuge

Study204 Cases

Sensitivity increased 65% to 74% with nail centrifuge.20 cases (10%) centrifuge not performed, usually because nail was submitted dry.

ConclusionCentrifuge of Nail Debris

Positive cases--increased 9% from 73% to 82% with centrifuge.Negative centrifuge cases often associated with bacteria (45%)Clinicians should consider submitting nail debris in a cytoprep solution rather than nail plate.

AFX vs other

p63AFX--CD10, p63 and CD99

Kanner W et al. CD10, p63 and CD99 expression in the differential diagnosis of atypical fibroxanthoma, spindle cell squamous cell carcinoma and desmoplastic melanoma. J Cutaneous Pathol 37:744-750, 2010.

Merkel

PolyomavirusNeurofilament dot

Merkel

Merkel

Merkel

Merkel--Polyomavirus

Viral large T antigen gene (replication)Can send clinical samples for analysis (Memorial Sloan Kettering)

Merkel Diagnostics

Neurofilament dotMcCalmont T. Paranuclear dots of neurofilament reliably identify Merkel cell carcinoma J Cutaneous Pathol 37:821, 2010.

Merkel Diagnostics

Cytokeratin 20+ (CK7-)Synaptophysin+Chromogranin+

Merkel Diagnostics

Neurofilament dotUseful in cases in which cytokeratin is nonspecific (no perinuclear dot)

Infectious

HPV SubtypingUse probes from cervical studiesUsually patient requestsSubtypingHigh-riskLow-riskWide Screen

Infectious

SyphilisImmunohistochemistryReplaces Warthin-Starry

Autoimmune

Autoimmune

C3d Antigen/Antibody--ImmunohistochemistryBullous pemphigoid Pemphigus vulgaris.

C3d AntibodyBullous Pemphigoid

C3d AntibodyPemphigus Vulgaris

C3d Antibody

Formalin-fixed TissueNo Michels!

Pfaltz K et al. C3d immunohistochemistry on formalin-fixed tissue is a valuable tool in the diagnosis of bullous pemphigoid of the skin. J Cutaneous Pathol 37:654-8, 2009.

Lymphoma

ClassificationClonalityIn-situ Hybridization for kappa/lambda

Classification

WHO-EORTC Classification. GET INFO FROM TEACHING CASESubcutaneousBadGood

Flow Cytometry

Can be a great diagnostic tool for cellular infiltratesCan test more antigens/antibodies.Wu et al. Contribution of flow cytometry in the diagnosis of cutaneous lymphoid lesions. Journal of Investigative Dermatology 121: 1522-30, 2003.

Lymphoma

ClonalityGene rearrangement on two remote anatomic siteslook for same clone.Helps with MF patient who have rash vs MF.Thurber S et al. T-cell clonality analysis in biopsy specimens from two different skin sites shows high specificity in the diagnosis of patients with suggested mycosis fungoides. JAAD 57:782-90, 2007.

Kappa/lambda Light In-situ Hybridization

ClonalityB-cells/plasma cells

Thank you!

Brandon StokesRodd TakiguchiPhoebe RichJanet Roberts