Precise microdissection of human bladder carcinomas reveals divergent tumor subclones in the same tumor.
BACKGROUND
Human bladder carcinoma is thought to arise from a field change that affects the entire urothelium. Whether independently transformed urothelial cell populations exist in the same patient is uncertain.


METHODS
We studied the clonality of urinary bladder carcinoma in 18 female patients who underwent cystectomy for urothelial carcinoma. None had multiple tumors. Tumor samples were obtained from different areas of the same tumor. Sixty-seven tumor samples were analyzed. Tumor genomic DNA was microdissected and extracted from formalin-fixed, paraffin-embedded slides. The clonality of urothelial tumors was evaluated on the basis of a polymorphism of the X chromosome-linked human androgen receptor gene (HUMARA) locus. The technique is dependent on digestion of DNA with the methylation-sensitive restriction enzyme HhaI, polymerase chain reaction (PCR) amplification of HUMARA locus, and detection of methylation of this locus. With this method, only the methylated HUMARA allele is selectively amplified by PCR.


RESULTS
Eleven of 18 patients were informative. Nonrandom inactivation of the X chromosome was found in 9 of the 11 informative patients (82%). Seven patients showed different patterns of nonrandom X chromosome inactivation for tumor samples obtained from different regions of the same tumor. Two patients showed the same pattern of nonrandom X chromosome inactivation in all samples.


CONCLUSIONS
Some muscle-invasive urothelial carcinomas may arise from independently transformed progenitor urothelial cells, supporting the "field effect" theory for bladder carcinogenesis.
Cheng
Liang
L
Department of Pathology, Indiana University School of Medicine, Indianapolis, IN 46202, USA. lcheng@iupiu.edu
Gu
Jian
J
Ulbright
Thomas M
TM
MacLennan
Gregory T
GT
Sweeney
Christopher J
CJ
Zhang
Shaobo
S
Sanchez
Katya
K
Koch
Michael O
MO
Eble
John N
JN
D019175:DNA Methylation; D004273:DNA, Neoplasm / Q000032:analysis*; D005260:Female; D006801:Humans; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D011944:Receptors, Androgen / Q000235:genetics*; D001749:Urinary Bladder Neoplasms / Q000235:genetics* / Q000473:pathology* / Q000648:ultrastructure; D014960:X Chromosome / Q000235:genetics*
C410798:AR protein, human; D004273:DNA, Neoplasm; D011944:Receptors, Androgen