INTERNATIONAL JOURNAL OF MOLECULAR MEDICINE 3: 417-419, 1999 The role of the detection of hematogenous micrometastasis in prostate adenocarcinoma and malignant melanoma by RT-PCR IACOPOSARDI 1 , SILVIA MORETTI 2 , ROBERTO PONCHIETTI 3 , SILVIA ARRIGUCCI 1 , RENATO GUAZZELLI 1 and ENRICO MONTALI 1 Medical Genetics Unit, Department of Clinical Physiopathology, 2 2nd Dermatology Unit, S.M. Nuova Hospital, Azienda Sanitaria di Firenze, 3 2nd Urological Clinic, Florence University Medical School, Florence, Italy Received February 4, 1999; Accepted February 17, 1999 Abstract. Recent studies reported the possibility of detecting prostate adenocarcinoma and malignant melanoma cells in peripheral blood using RT-PCR of prostatic specific antigen (PSA), prostatic specific membrane antigen (PSMA) and Tyrosinase mRNAs. The PCR results showed high variability, ranging between 0% and 100% of positivity in patients with advanced disease. Our purpose was to evaluate the presence of tumor marker mRNAs in peripheral blood of prostate cancer and melanoma patients by means of RT-nested-PCR. We tested 70 and 36 peripheral blood samples from prostate carcinoma and malignant melanoma patients, respectively. The RT-PCR analysis showed the presence of PSA cDNA in 9 out of 70 (12.9%); PSMA cDNA in 14 out of 70 (20%); and Tyrosinase cDNA in 2 out of 36 (5.5%) peripheral blood samples from melanoma patients. Our study confirms the applicability of this sensitive method to monitor disease status. Although, the RT-nested-PCR of Tyrosinase is able to detect neoplastic cells in peripheral blood specimens, we suggest the necessity of a great caution in interpreting PCR results when the nested method has been used. Introduction The clinical course of patients affected by prostate adeno- carcinoma and malignant melanoma is mainly monitored by careful and instrumental examinations, but such approaches generally fail to point out the presence of micrometastases, which may be suspected when circulating neoplastic cells are Correspondence to: Dr Iacopo Sardi, Medical Genetics Unit, Department of Clinical Physiopathology, Florence University Medical School, V.le Pieraccini 6,1-50139, Florence, Italy Key words: hematogenous micrometastasis, prostate adeno- carcinoma, malignant melanoma, prostatic specific antigen, prostatic specific membrane antigen, Tyrosinase detectable in the peripheral blood. Although, the invasiveness of tumor cells is a result of cumulative genetic changes that determine a progressive metastatic capacity through various mechanisms and so the presence of hematogenous neoplastic cells would seem to imply a metastatic disease, this has not consistently been demonstrated. Indeed, several studies have showed the inefficiency of circulating neoplastic cells in forming metastatic tumor (1,2). In the past six years, the RT-nested-PCR method has been widely used for detecting the presence of Tyrosinase, PSA (prostatic specific antigen) and PSMA (prostatic specific membrane antigen) mRNAs in peripheral blood of melanoma and prostate cancer patients, respectively (3-5). This molecular method can detect a single tumor cell in over 1 million peripheral blood cells (6,7). For several neoplasms this method for detecting tumor cells is primarily limited by use of tumor- specific mRNA markers. PSA and Tyrosinase are most specific tumor markers for prostate adenocarcinoma and malignant melanoma, respectively. This technique fulfils all requirements for the identification of micrometastasis and might have the capacity to predict the course of disease both in melanoma and in prostate adenocarcinoma patients. Several studies using this highly sensitive nested-PCR method have obtained positive results in control patients (8,9) [O'Hara SM, et al, J Urol 155 (Suppl.): abs. 430A, 1996]. In 1989 Chelly et al (10) demonstrated the existence of a molecular phenomenon of basal transcription of any gene in any cell type. During DNA replication, the presence of ubiquitous transcriptional factors which bind various DNA elements give rise to a very low, but not null, level of transcription of any gene. This 'illegitimate transcription' occurs under stressed sensitivity conditions, as several PCR cycles and elevated initial amount of total RNA in the RT.-PCR reaction. Materials and methods We examined 36 melanoma and 70 prostate adenocarcinoma patients for the presence of circulating neoplastic cells using the RT-nested-PCR method. A group of controls was also analyzed. This group comprised 21 healthy young women and