Journal Reading

醫學系七年級 (Intern): 陳姿璇 (B8701059)指導教授 : 鄭建睿 醫師

Journal Reading

The American Journal of Surgical Pathology 27: 750-761, 2003

Aberrant nuclear immunoreactivity for TFE3 (Transcription Factor E3) in neoplasms with TFE3 gene fusions

A sensitive and specific immunohistochemical (IHC)assay

The department of pathology , Johns Hopkins Medical Institutions, Baltimore,Maryland and the department of pathology , Memorial Sloan-Kettering Cancer Center,NewYork

Journal Reading

Recurrent chromosomal translocations Gene fusions Novel chimeric proteins Tumor-specific diagnostic

immunohistochemistry (IHC) assays

Falini and Mason

Immunohistochemistry (IHC) assays

Antibody-fusion protein breakpoint; Difficulty in raising specific antibodies Robust antibodies-a portion of one of

the proteins; Successful, overexpression ALK1, anaplastic large cell lymphoma;

inflammatory myofibroblastic tumor The C-terminus of WT1desmoplastic small

round cell tumor

Breakpoints

Alveolar Soft Part Sarcoma (ASPS)

t(X;17),(p11.2;q25) TFE3 (Transcription Factor E3),

chromosome X p 11.2 Renal cell carcinomas, children and

young adults ASPL (Alveolar Soft Part Sarcoma Locus)

gene, chromosome 17q25 unknown

Renal cell carcinomas

t (X;1)(p11.2;q21) TFE3-PRCC(Papillary Renal Cell

Carcinoma) gene fusion t (X;17)(p11.2;q25) TFE3-ASPL (Alveolar Soft Part Sarcoma

Locus) gene fusion Xp11.2, TFE3 gene, 3’ end IHC, useful diagnostic marker

Material and Methods Positive control cases: Formalin-fixed, paraffin-embedded tissue sections Xp11.2,TFE3 ASPS, three types of renal carcinoma ASPS-positive control group: 19 Renal carcinoma-positive control group:21 t(X;17)(p11.2;q25)/

ASPL (Alveolar Soft Part Sarcoma Locus) -TFE3: 7

t(X;1)(p11.2;q21)/ PRCC (Papillary Renal Cell Carcinoma) -TFE3: 11

t(X;1)(p11.2;p34)/ PSF(PTB-associated splicing factor) -TFE3: 3

Material and Methods

Screening cases: Organ-specific tissue microarray (TMAs) Memorial Sloan Kettering Cancer Center: High-density TMA: 7-270 cores, ranging from 3 to 6

cores per tumor, with a core diameter of 0.4-1.0mm Low-density TMA: 27-35 cores, single core per tumor , 3mm in diameter The Johns Hopkins Hospital: TMA: 99 spots,2mm in diameter Positive/weak positive-donor blocks- whole sections -immunostained for TFE3

Material and Methods

Material and Methods

Test cases: Archival pediatric renal carcinoma cases

Lack of frozen tissue, molecular analysis

TFE3 antibody: P-16 polyclonal antibody

Material and Methods

Material and Methods IHC method: 4um sections mounted onto positively charged slides Xylene- deparaffinized-30 minutes Ethanol-rehydrated Steamed for 30 minutes at 98-99C in EDTA buffer in

a vegetable steamer Quenching: Hydrogen peroidase Biotin blocking: Avidin Incubation overnight:1:600dilution polyclonal

antibody in phosphate-buffered saline Detection: biotinylated secondary antibody,

horseradish peroxidase-conjugated streptavidin, 3’-3’-diaminobenzidine (Chromogen)

Material and Methods

Scoring of TFE3 nuclear immunoreactivity:

0-3+, Nucleus 1+: weak/equivocal (negative) 2+: moderate 3+ strong readily apparent at low-power

magnification (4X objective)

Material and Methods

Results

Normal tissues: lung, thyroid, lymph node, breast,colon,liver,gallbladder,pancreas,uterus,ovary,bone,kidney,bladder,adrenal, prostate and skin

No detectable TFE3 nuclear protein 1/18 chronic pancreatitis,weak(1+)

Results

Positive control cases: tumors with knownXp11.2 translocationsorTFE3 gene fusions:

39/40: moderate or strong 19 ASPS: 18 strong ;1 moderate 7 t(X;17)(p11.2;q25) (ASPL-TFE3):

2 strong; 5 moderate 3 t(X;1)(p11.2;p34)(PSF-TFE3): strong 11 t(X;1)(p11.2;q21)(PRCC-TFE3):

1 weak (negative); 2 strong; 8 moderate

Results

Results

Screening cases: Table 1 6/1476 TMA: negative

Results

Results

Results

Test cases: relationship between TFE3 immunoreactivity and morphology in pediatric renal carcinoma:

11 cases 8 TFE3: 5 ASPL-TFE3; 3PRCC-TFE3 7/8: TFE3 staining (5ASPL-FE; 2 PRCC-

TFE3)

Results

Discussion

TFE3(Transcription Factor E3):a basic helix-loop-helix DNA binding domain and a leucine zipper dimerization domain

Nuclear localization signal Native TFE3 protein: not detectable Half-life short, tightly regulated Strong nuclear immunoreactivity: highly sensitive,

highly specific Sensitivity:97.5% (39/40)---100% Bouin’s acidic fixative: render specific antigen less

detecbale by IHC Specificity: 99.6% (6/1476)---99.1%

TFE3(Transcription Factor E3)

Discussion

Discussion

3 ASPS blocks: >20 years of age, strong

time 1 PSF-TFE3 block: unstained over one

year, stored at room temperature oxidation Immunoreactivity: edges> center complete fixation

Discussion

False-positive: upregulated native TFE3 protein artifacts of fixation ASPS: PAS (Periodic acid-Schiff) stain: needle-

shaped cytoplasmic deposits Clear cell sarcoma of kidney (CCSK):

fine chromatin of CCSK nuclei , chemotherapy: doxorubicin

Xp11.2-related renal cell carcinoma: interferon

Discussion

The prevalence of these tumors in adults and children:

One 68-year-old woman, one 38-year-old man, one 39-year-old woman: ASPL-TFE3

Adult: rare

Reference

Atlas of Genetics and Cytogenetics in Oncology and Haematology

http://www.infobiogen.fr/services/chromcancer/Genes/TFE3ID86.html