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Tumor Biology |
Matsunaga-Conte Prostate Cancer Research Center, Scott Department of Urology [K. M. S., S. F. S., C. N., A. K. L., W. S.] and Department of Pathology [T. M. W.], Baylor College of Medicine, and The Methodist Hospital, Houston, Texas 77030; Departments of Urology and Biostatistics, Memorial Sloan-Kettering Cancer Center, New York, New York 10021 [M. W. K.], and the Departments of Urology, Biochemistry, and Molecular Biology, Mayo Clinic/Foundation, Rochester, Minnesota 55905 [C. Y. F. Y., D. J. T.]
We
developed a highly sensitive splice variant-specific reverse
transcriptase-PCR (RT-PCR) assay for human glandular kallikrein
(hK2) mRNA and tested its ability to detect metastatic disease in men
with clinically localized prostate cancer. An RT-PCR assay using
primers spanning intron IV and including a significant portion of the
3' untranslated region of the hKLK2 gene,
with maximum nonhomology to both hK1 and hK3, was developed. The limit
of detection of the assay was five copies of hK2 cDNA and one LNCaP
cell in 109 lymphoblasts. RT-PCR-hK2 was performed on
preoperative peripheral blood specimens from 228 consecutive radical
prostatectomy patients as well as 7 metastatic prostate cancer patients
and 14 healthy men without prostate cancer. This new RT-PCR-hK2 assay
amplifies two distinct fragments. The larger fragment (hK2-U) is
approximately 680 bp in length and corresponds to the amplified product
of a previously reported splice variant in the splice donor site of
intron IV in the hKLK2 gene. The smaller fragment
(hK2-L) is 
643 bp in length and corresponds to the amplified product
of the native hK2 mRNA. Whereas the RT-PCR-hK2-L assay was positive in
71% of our patients with metastatic prostate cancer, 14% of healthy
control men also tested positive. By univariate
(P = 0.028) and multivariate
(P = 0.0269) analysis, which controlled
for preoperative PSA, clinical stage, and biopsy Gleason score,
RT-PCR-hK2-L status added prognostic information to the prediction of
lymph node-positive disease. We have developed a new RT-PCR assay which
demonstrates a high sensitivity for detecting hK2 mRNA. Preoperative
RT-PCR-hK2-L status helps predict pathological lymph node positivity in
patients with clinically localized prostate cancer.
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