Gao C L, Maheshwari S, Dean R C, Tatum L, Mooneyhan R, Connelly R R, McLeod D G, Srivastava S, Moul J W
Center for Prostate Disease Research, Department of Surgery, Uniformed Services University of the Health Sciences, Bethesda, Maryland 20814-4799, USA.
Urology. 1999 Apr;53(4):714-21. doi: 10.1016/s0090-4295(98)00608-6.
The reverse transcriptase-polymerase chain reaction (RT-PCR)-prostate-specific antigen (PSA) assay to detect presumed occult micrometastatic prostate cancer has been controversial, and this molecular staging has been thought to be clinically useful by some groups but not others.
We used a sensitive nested RT-PCR assay with specific primers derived from the PSA sequence and a very stringent two-step PCR protocol with denaturing temperature of 94 degrees C annealing and extension temperature of 68 degrees C. This method enabled us to detect PSA-expressing LNCaP prostate cancer (PC) cells as low as one cell of 10 million lymphocytes (1/10(7)). Ninety-six patients with PC were studied, including 85 before radical prostatectomy (RP), and 22 controls, including healthy men and women and men with benign prostatic hyperplasia.
In 85 patients undergoing RP, a minimum of two independent RT-PCR-PSA assays detected circulating prostate cells preoperatively in 27 patients (31.8%). Of 12 patients with locally advanced or advanced stage cancer, RT-PCR-PSA was positive in 5 (41.7%); of the 22 controls, no patient was RT-PCR-PSA positive. In 10 randomly selected cases, the RT-PCR product was confirmed as PSA by DNA sequencing. Of the 27 patients undergoing RP who were RT-PCR positive, 11 (40.7%) had non-organ-confined disease (pT3a or greater), and of the 58 patients who were RT-PCR negative, 32 (55.2%) had non-organ-confined disease. Patients with RT-PCR positive results also had lower margin positivity (9 of 27, 33.3%) than did patients with RT-PCR negative results (21 of 58, 36.2%). Finally, at a mean follow-up of 25.7 months, 5 (18.5%) of 27 RT-PCR positive patients had recurrence (PSA) compared with 14 (24.1%) of 58 RT-PCR negative patients.
On the basis of this blinded study, RT-PCR for PSA-expressing cells in 85 patients before RP is not related to clinical stage, age, race, grade, Gleason sum, serum PSA or prostatic acid phosphatase, tumor volume, or tumor multifocality. RT-PCR positivity did not predict pathologic stage or early PSA recurrence. A standardized RT-PCR assay needs to be developed to account for interlaboratory discrepancies.
逆转录聚合酶链反应（RT-PCR）-前列腺特异性抗原（PSA）检测用于检测假定隐匿性微转移前列腺癌一直存在争议，一些研究团队认为这种分子分期具有临床应用价值，而另一些则不然。
我们使用了一种灵敏的巢式RT-PCR检测方法，其特异性引物来源于PSA序列，并采用非常严格的两步PCR方案，变性温度为94℃，退火和延伸温度为68℃。该方法使我们能够检测到表达PSA的LNCaP前列腺癌细胞，低至每1000万个淋巴细胞中有一个细胞（1/10⁷）。对96例前列腺癌患者进行了研究，其中85例在根治性前列腺切除术（RP）前，22例为对照，包括健康男性和女性以及良性前列腺增生男性。
在85例行RP的患者中，至少两次独立的RT-PCR-PSA检测在27例患者（31.8%）术前检测到循环前列腺细胞。在12例局部晚期或晚期癌症患者中，RT-PCR-PSA阳性的有5例（41.7%）；22例对照中，无患者RT-PCR-PSA阳性。在随机选择的10例病例中，RT-PCR产物经DNA测序确认为PSA。在27例RT-PCR阳性的行RP患者中，11例（40.7%）有非器官局限性疾病（pT3a或更高分期），在58例RT-PCR阴性患者中，32例（55.2%）有非器官局限性疾病。RT-PCR阳性结果的患者切缘阳性率（27例中的9例，33.3%）也低于RT-PCR阴性结果的患者（58例中的21例，36.2%）。最后，平均随访25.7个月，27例RT-PCR阳性患者中有5例（18.5%）复发（PSA升高），而58例RT-PCR阴性患者中有14例（
