Comparison of Sodium Selenite and Selenium-Enriched Spirulina Supplementation Effects After Selenium Deficiency on Growth, Tissue Selenium Concentrations, Antioxidant Activities, and Selenoprotein Expression in Rats.

Selenium contributes to physiological functions through its incorporation into selenoproteins. It is involved in oxidative stress defense. A selenium deficiency results in the onset or aggravation of pathologies. Following a deficiency, the repletion of selenium leads to a selenoprotein expression hierarchy misunderstood. Moreover, spirulina, a microalga, exhibits antioxidant properties and can be enriched in selenium.. Our objective was to determine the effects of a sodium selenite or selenium-enriched spirulina supplementation. Thirty-two female Wistar rats were fed for 12 weeks with a selenium-deficient diet. After 8 weeks, rats were divided into 4 groups and were fed with water, sodium selenite (20 µg Se/kg body weight), spirulina (3 g/kg bw), or selenium-enriched spirulina (20 µg Se/kg bw + 3 g spirulina/kg bw). Another group of 8 rats was fed with normal diet during 12 weeks. Selenium concentration and antioxidant enzyme activities were measured in plasma, urine, liver, brain, kidney, heart, and soleus. Expression of GPx (1, 3), Sel (P, S, T, W), SEPHS2, TrxR1, ApoER2, and megalin were quantified in liver, kidney, brain, and heart. We showed that a selenium deficiency leads to a growth delay, reversed by selenium supplementation despite a minor loss of weight in week 12 for SS rats. All tissues displayed a decrease in selenium concentration following deficiency. The brain seemed protected. We demonstrated a hierarchy in selenium distribution and selenoprotein expression. A supplementation of sodium selenite improved GPx activities and selenoprotein expression while a selenium-enriched spirulina was more effective to restore selenium concentration especially in the liver, kidney, and soleus.

Atypical schwannoma of the median nerve. A case report.

Schwannomas of the hand are very rare tumors and represent less than 3% of all soft tissue tumors in the hand. These tumors share clinical, epidemiological and imaging characteristics with the other soft tissue and peripheral nerve tumors; thus, it can be difficult to make a preoperative diagnosis. Here we report the case of a 48-year-old woman who presented with a schwannoma arising from the palmar branch of the median nerve. The tumor measured 54 × 41 x 52 mm and was located in the thenar eminence. The first hypothesis was a vascular tumor. After surgery and histological analysis, the final diagnosis of an atypical schwannoma was established. The presence of shared immunohistochemical characteristics with cellular histiocytoma and myoepithelial tumors forced us to adopt an aggressive follow-up protocol. As of the last follow-up at 9 years, the patient had good clinical outcomes and no recurrence. This case highlights the difficulties encountered in clinical practice to diagnose such tumors.

[Incidence of distal ulna fractures associated with distal radius fractures: Treatment options]. / Incidence des fractures de l'extrémité distale de l'ulna associées aux fractures de l'extrémité distale du radius (processus styloïde ulnaire). Nos choix thérapeutiques.

Fractures of the neck and/or head of the ulna or distal ulna fracture (DUF) other than ulnar styloid fractures can occur in combination with distal radius fractures (DRF). This combination can have a significant influence on the treatment and prognosis since it causes the entire distal forearm to be unstable. In a series of 1279 consecutive unilateral DRFs, we found an associated ulnar neck fracture in 5.9% of cases, ulnar head and neck fracture in 1.6%, and isolated ulnar head fracture in 1.4%. Overall, 9% of cases in this study had a DUF with a DRF. The frequency of extra-articular "A" (11%) and intra-articular "C" (9%) DRFs according to the AO classification was about the same. There were no cases of DUF combined with partial "B" DRF. There was a correlation between combined DUF with DRF and the patient's group in the PAF classification. DUF are more frequently associated with DRF in elderly patients. Specific distal ulnar locking plates were recently introduced and they may be a useful adjunct to distal radius locking plates when treating patients with combined DUF and DRF.

Prognostic value of tyrosinase reverse transcriptase PCR analysis in melanoma sentinel lymph nodes: long-term follow-up analysis.

OBJECTIVE: To determine the prognostic value of detecting tyrosinase transcripts in melanoma sentinel lymph nodes (SLNs). METHODS: Reverse transcription (RT) PCR for tyrosinase mRNA was performed on negative SLNs of 76 patients with melanoma. RESULTS: Tyrosinase mRNA was found in 39 patients (51.3%). After a median follow-up period of 51 months, significant differences were found in overall survival (OS) but not in disease-free survival (DFS). The 5-year OS and DFS rates were 97.2% and 80%, respectively, for RT-PCR tyrosinase-negative (TN) patients vs. 78.67% and 66.24% for RT-PCR tyrosinase-positive (TP) patients (P = 0.019 and P = 0.38, respectively). Of four progressing patients in the TN group, three relapsed with subcutaneous, soft-tissue or lymph-node metastases, while seven out of nine progressing patients in the TP group relapsed at visceral sites. CONCLUSIONS: No significant differences in DFS were found by RT-PCR tyrosinase expression analysis at melanoma SLNs. Significant differences in OS could be related to a different pattern of relapse and must be confirmed after a longer follow-up time.

Melanoma inhibiting activity protein (MIA), beta-2 microglobulin and lactate dehydrogenase (LDH) in metastatic melanoma.

BACKGROUND: Serum levels of melanoma markers may have a role in monitoring disease evolution in metastatic melanoma. PATIENTS AND METHODS: Serial measurements of melanoma inhibiting activity protein (MIA), lactate dehydrogenase (LDH), S-100 and beta2-microglubulin were obtained from 42 metastatic melanoma patients during their biochemotherapy treatment. RESULTS: High pre-treatment serum levels of S-100, LDH, MIA and P2-microglobulin were detected in 50%, 57%, 50% and 24% of the patients, respectively. Only S-100 had prognostic significance for both disease-free (p=0.011) and overall survival (p=0.021). In patients who responded to treatment, S-100 levels decreased significantly from pre-treatment to the time of response (p = 0.050). When patients progressed, levels of MIA and P2-microglobulin increased significantly (p =0.028 and p =0.030, respectively). CONCLUSION: Correlation with disease evolution was found for S-100, MIA and P2-microglobulin levels. Despite the small sample size of the study, S-100 was a significant prognostic marker for overall survival and disease-free survival.

S-100beta and MIA in advanced melanoma in relation to prognostic factors.

We compared the sensitivity and specificity of S-100 and MIA in advanced melanoma, in 96 patients with no evidence of disease (NED) and 86 patients with metastatic melanoma. Abnormal S100 (>0.2 microg/l) and MIA (>14 ng/ml) results were found in 1.1% and 3.2% of NED patients and in 59.3% and 54.6% of the patients with active melanoma (p<0.001). Using both tumor markers simultaneously, the sensitivity increased up to 69.8% with the same specificity 96.8%. S100 serum levels were not related to growth patterns. By contrast, MIA levels seemed to be related to the growth pattern, with higher levels in nodular melanoma (60.6+/-87.1 ng/ml) compared with acral-lentigous melanoma (11.9+/-5.4 ng/ml) (p=0.02). Likewise, S100 was related to the metastases site with significantly higher sensitivity and mean concentrations in patients with brain metastases (p=0.01) with the lowest in those with lung MI. MIA was related to the same metastases locations but without statistical significance. In summary, both S100 and ML4 are useful markers related to prognostic factors, being more effective when used in combination.

Serum protein s-100 predicts clinical outcome in patients with melanoma treated with adjuvant interferon--comparison with tyrosinase rt-PCR.

OBJECTIVE: To study the clinical value of the determination of serum S-100 protein as a single tumor marker or in combination with tyrosinase RT-PCR in patients with melanoma receiving adjuvant interferon. PATIENTS AND METHODS: Patients were tested for serum S-100 protein luminoimmunometric assay and for blood tyrosinase mRNA (RT-PCR), before starting interferon and every 2-3 months thereafter. RESULTS: One hundred and six patients (stage IIA, 27; IIB, 19; III, 49; and IV, 11) were included in the study. Median follow-up was 51 months (range 2-76). In the univariate analysis, under treatment S-100 > or =0.15 microg/l and a positive RT-PCR correlated with a lower disease-free survival and overall survival (OS). In the multivariate analysis, clinical stage, under therapy positive RT-PCR and S-100 levels > or =0.15 mug/ml, were independent prognostic factors for OS. The hazard ratio for OS was 3.9 (95% CI, 1.67-9.15; p = 0.004) and 2.2 (95% CI, 1.05-4.6; p = 0.016) for S-100 > or =0.15 microg/l and positive RT-PCR, respectively. When both techniques where combined, a positive RT-PCR indicated a poorer clinical outcome only in patients with S-100 <0.15 microg/l. CONCLUSIONS: S-100 > or =0.15 microg/l and a positive RT-PCR during adjuvant interferon therapy indicate a high risk of death in resected melanoma patients. S-100 determination has a higher positive predictive value than RT-PCR, while tyrosinase RT-PCR adds prognostic information in patients with S-100 <0.15 microg/l.

Pilot study of treatment of biochemotherapy-refractory stage IV melanoma patients with autologous dendritic cells pulsed with a heterologous melanoma cell line lysate.

Eleven AJCC stage IV melanoma patients with progressive disease after treatment with biochemotherapy were treated with autologous dendritic cells pulsed with heterologous tumor cell lysates. The vaccine used mature DCs (CD1a+++, CD40++, CD80++, CD83+, and CD86+++) generated from peripheral blood monocytes in the presence of GM-CSF and IL-4. After 7 days, DCs were matured with a defined cocktail of cytokines (IL-1+IL-6+TNF-alpha+PGE2) and simultaneously pulsed with lysates of heterologous melanoma cell lines, for 2 days. A total of 4 x 10(6) DCs was injected monthly under ultrasound control in an inguinal lymph node of normal appearance. The study was closed when all patients died as a consequence of tumor progression. No sign of toxicity was observed during the study. One patient experienced a partial response lasting 5 months, and two patients showed a mixed response which lasted 3 months. The median survival of the whole group was 7.3 months (range 3-14 months). This vaccination program had specific antitumoral activity in highly pretreated and large tumor burden stage IV melanoma patients and was well tolerated. The clinical responses and the median survival of the group of patients, together with the low toxicity of our DC vaccine, suggest that this approach could be applied to earlier AJCC stage IV melanoma patients.

Gamma-detecting probe used intraoperatively to locate the sentinel lymph node in patients with malignant melanoma.

OBJECTIVE: To assess the usefulness of lymphoscintigraphy and intraoperative gamma probe in the detection of sentinel lymph nodes. DESIGN: Prospective open study. SETTING: University hospital, Spain. SUBJECTS: 40 patients with malignant melanoma (24 stage I/II, 16 stage III). INTERVENTION: The day before operation a lymphoscintigram with 99mTc-nanocolloid was taken and the first lymph node identified was considered to be the sentinel node. A hand-held gamma probe was used for intraoperative mapping. MAIN OUTCOME MEASURE: Identification of the sentinel node. RESULTS: Sentinel nodes were identified in 39/40 patients (98%). In 24 patients with stage I/II disease, 34 sentinel nodes were found (6 invaded and 28 clear of melanoma). A total number of 161 regional lymph nodes were harvested, none of them invaded by melanoma. In 16 patients with stage III disease, 22 sentinel nodes were located (14 invaded and 8 clear of melanoma). A total of 89 regional lymph nodes were excised in patients with invaded sentinel nodes (44 of which were invaded and 45 clear of disease). 41 lymph nodes were excised from patients with clear sentinel nodes, and all were also clear of melanoma. CONCLUSIONS: We conclude that this is a useful technique for the selection of patients with melanoma who may require lymphadenectomy.

Large deletions of chromosome 9p in cutaneous malignant melanoma identify patients with a high risk of developing metastases. Hospital Clinic Malignant Melanoma Group, University of Barcelona.

Cutaneous malignant melanoma (CMM) is an aggressive tumour with a high metastatic potential. Deletions of chromosome 9p have been detected in CMM, some of which involve the CDKN2A/p14ARF genes. Loss of heterozygosity (LOH) of 16 microsatellite markers on 9p and mutations in the CDKN2A/p14ARF genes had been previously studied in 32 melanoma patients by our group. 9p deletions were detected in 15 primary tumours (45.5%) and are here correlated with the clinical outcome over 5 years and compared with classical prognostic factors. Eight of the 32 patients developed metastases (25%). The metastases were all detected within 768 days of the initial diagnosis. The patients without metastases were last monitored at least 1621 days after diagnosis. None of the 21 patients with more than eight microsatellites conserved developed metastases, whereas all of the eight patients who developed metastases had eight or more markers deleted. The sensitivity of this analysis to predict metastases was 100% (specificity 84%), whereas the sensitivity for the same sample using a Breslow thickness > 3 mm was 62.5% (specificity 68%). LOH of eight or more of the 9p microsatellite markers is therefore a useful prognostic factor to predict the development of metastases in the first 4.4-6.3 years (1621-2294 days).

The use erythrocyte glutathione as a predictive marker for malignant melanoma.

BACKGROUND: Glutathione (GSH) may provide defense against reactive oxygen species (ROS) generated by ultraviolet radiation. Furthermore, some authors have demonstrated a relationship between the GSH of peripheral blood erythrocytes (GSHe) and resistance to chemotherapy. PATIENTS & METHODS: To observe the influence of GSH on the genesis and evolution of Malignant Melanoma (MM), we assessed the concentration of GSH in erythrocytes (GSHe) in MM patients (n = 566) and controls (n = 164) by the method of Beutler (1963). RESULTS: No differences were found between the two groups (5.94 +/- 1.61 cases vs 6.08 +/- 1.49 mmol/gr Hb, controls; p > 0.05). Fifty seven patients with poor evolution (disease-free survival < 2 years) had higher GSH levels than the remaining patients (6.35 +/- 1.83 vs 5.83 +/- 1.62 mmol/g Hb; p < 0.01). GSHe increased significantly after antineoplastic therapy (4.75 +/- 1.26 vs 7.73 +/- 1.39 mmoVg Hb; p < 0.001), thus indicating a possible role in chemoresistance. 2 CONCLUSIONS: GSHe is not related to the risk of developing MM. GSHe may be related to the evolution of MM, being higher in patients who suffer relapse or metastasis. GSHe increases significantly during cytostatic treatment.

Prognostic significance of the detection of circulating malignant cells by reverse transcriptase-polymerase chain reaction in long-term clinically disease-free melanoma patients.

The purpose of this study was to assess the prognostic significance of the detection of circulating melanoma cells by reverse transcriptase-PCR in long-term clinically disease-free melanoma patients. Patients with melanoma who were free of clinical relapse for at least 6 months after primary tumor diagnosis were included and prospectively followed. Tyrosinase mRNA in peripheral blood from these patients was assayed by reverse transcriptase-PCR at the time of their inclusion in the study. One hundred six blood samples from 57 melanoma patients were analyzed. The median time between melanoma diagnosis and inclusion in the study was 24 months (range, 7-51 months). The median follow-up time calculated from the time of inclusion in the study was 27 months (range, 11-36 months). Tyrosinase mRNA in blood was detected in 10 (17.5%) of 57 patients: 2 (18%) of 11 stage I patients, 6 (19%) of 33 stage II patients, and 2 (15%) of 13 stage III patients. Actuarial 2-year DFS was 89% for the tyrosinase-negative patients versus 30% for the positive patients (P = 0.003). Actuarial 2-year OS was 97% for the tyrosinase-negative patients versus 72% for the positive patients (P = 0.001). Tyrosinase mRNA could be detected in the blood of a proportion of long-term disease-free melanoma patients, regardless of their initial clinical stage. The presence of late circulating melanoma cells in this selected group of clinically disease-free patients was significantly associated with a subsequent high risk of relapse and death.

[Sentinel lymph node detection with lymphoscintigraphy and intraoperative probe in malignant melanoma patients]. / Detección del ganglio centinela mediante linfogammagrafía y sonda detectora intraoperatoria en pacientes con melanoma maligno.

BACKGROUND: The sentinel lymph node is the first node in a lymphatic basin to receive lymphatic drainage from a tumor site. If this node is free of tumor, then radical lymphadenectomy may be avoided. The goal of this study was to assess the usefulness of lymphoscintigraphy and intraoperative gamma probe in the sentinel node detection in patients with malignant melanoma. METHOD: We prospectively studied 40 patients with malignant melanoma (24 in I/II stages and 16 in III stage). The day before surgery a lymphoscintigraphy with 99mTc-nanocolloid was performed and the first lymph node identified was considered as sentinel node. For intra-operative mapping a hand-held gamma probe was used. RESULTS: Sentinel nodes were identified in 39/40 (97.5%) patients. In 24 patients with I/II stages 34 sentinel nodes were demonstrated (six positive and 28 negative for malignant melanoma). A total amount of 161 regional lymph nodes was harvested, all of them being negative for malignant melanoma. In 16 patients with III stage, 22 sentinel nodes were located (14 positive and eight negative for malignant melanoma). A total of 89 regional lymph nodes were excised in sentinel nodes positive patients (44 positive and 45 negative for malignant melanoma) and 36 lymph nodes in sentinel node negative, all of them negative for malignant melanoma. CONCLUSIONS: In patients with malignant melanoma, lymphoscintigraphy with 99Tc-nanocolloid is useful for the detection of sentinel lymph node. Biopsy of this node is useful for the selection of patients to undertake a lymphadenectomy.

CDKN2A mutations in Spanish cutaneous malignant melanoma families and patients with multiple melanomas and other neoplasia.

The CDKN2A gene has been implicated in cutaneous malignant melanoma (CMM) in about 40% of families with linkage to chromosome 9p21, while a small proportion of families have mutations in the CDK4 gene. In order to estimate the importance of these genes in the predisposition to CMM in Spanish families and patients we have analysed, by SSCA, a total of 56 subjects belonging to 34 CMM families, and nine patients with multiple CMM and other neoplasia. We have detected germline CDKN2A mutations in six out of the 34 families (17%). A frameshift mutation (358delG) and four missense mutations (G59V, G101W (two cases), D84Y, and R87W) were identified. Five CMM patients from different families (14%) carried the A148T variant, which is known not to affect p16 activity. No mutations were detected in the patients with multiple CMM or other neoplasms. We have not found mutations either in exon 1 beta of the CDKN2A gene or in exon 2A of CDK4. Linkage analysis of the 9p21 region showed exclusion for one of the families for CMM and for four families for CMM/dysplastic naevi. This study indicates a small role for CDKN2A in Spanish CMM families and suggests that other genes are also responsible for CMM predisposition.

[Detection of sentinel lymph nodes by lymphatic gammagraphy and intraoperative gamma-ray probe in patients with malignant melanoma. Initial results]. / Detección del ganglio centinela mediante linfogammagrafía y sonda de rayos gamma intraoperatoria en pacientes con melanoma maligno. Primeros resultados.

In the last years there has been an arising concern in the sentinel lymph node identification, the first lymph node to receive direct draining from the primary tumour, specially in malignant melanoma (MM). We studied 20 patients with MM: 10 with palpable regional lymph nodes and 10 without palpable LN by performing a lymphoscintigraphy using 99mTc-nanocolloid and a gamma-ray detecting probe during the surgery to locate the sentinel lymph node. In patients with palpable LN, 13 sentinel lymph nodes were identified. Ten of them were MM involved. Furthermore, 82 LN were harvested from involved lymph basins and 30 of them were positive for MM. In patients without palpable LN, 14 sentinel lymph nodes were identified (3 positives and 11 negatives for MM) and other 76 LN were resected (all of them negative). There were not <> in any patient. These preliminary results support the utility of the technique for the diagnosis and lymphadenectomy selection in patients without palpable LN but which could be involved by micrometastases.

Mid-arm sentinel lymph nodes showing surprising drainage from a malignant melanoma in the forearm.

A 51-year-old man with a malignant melanoma in his left forearm was studied to detect the sentinel lymph node and to assess the possibility of micrometastases in regional lymph nodes. Lymphoscintigraphy demonstrated two sentinel lymph nodes in the midarm. Two other nodes in the same location as well as in the left axilla were also observed. The exact location of the sentinel lymph nodes was identified with a gamma-ray detector. At the time of surgery, blue dye was injected around the primary lesion and the two sentinel lymph nodes on the inner side of the left arm were resected. Both lymph nodes were pigmented black. The histopathologic study demonstrated metastases from malignant melanoma in both nodes. This case reflects the main role of lymphoscintigraphy in identifying draining lymph nodes in unusual locations as observed in this patient.

Retention of the CDKN2A locus and low frequency of point mutations in primary and metastatic cutaneous malignant melanoma.

CDKN2A has been found mutated in melanoma families which show linkage to chromosome 9p21. In contrast, a low mutation rate has been found in melanomas, suggesting that CDKN2A might not be the first target for mutation in the development of this type of tumour. To elucidate the role of the CDKN2A gene and its alternative transcript p19ARF in the development of cutaneous malignant melanoma (CMM) we have analyzed 48 primary and metastasic CMM tumours for mutations and for loss of heterozygosity (LOH). Only one point mutation was detected (2%), while hemizygous deletions were identified in 20% of these tumours. Retention of the CDKN2A locus was found in 10 (47%) tumours with deletions at one or both sides of CDKN2A, suggesting that loss of this gene is not involved in CMM-tumour initiation and that another tumour-suppressor gene involved in melanoma is located at 9p21.

Immunohistochemical study of alpha, mu and pi class glutathione S transferase expression in malignant melanoma. MMM Group. Multidisciplinary Malignant Melanoma Group.

Human pi, mu and alpha class glutathione S transferases (GST) have been localized immunohistologically in normal skin, naevi and melanoma. Pi GSTs were found principally in the stratum basalis and, to a lesser extent, in the superficial layers. Normal melanocytes showed strong nuclear and cytoplasmatic staining. Distribution of GST mu in the epidermis showed that only the stratum basale, where melanocytes are located, stained well but with weak nuclear staining. Normal melanocytes were also well stained. The alpha GSTs were relatively abundant in the upper strata and to a lesser extent, in the basal layers. The absence of nuclear staining gives these cells a target appearance. Normal melanocytes showed strong cytoplasmatic staining. The pi GSTs seem to be most persistently and strongly expressed in malignant melanoma (MM), but mu GSTs are also found, whereas the alpha GSTs were only occasionally present. The finding of the GST mu in the melanocytes of the basal layer raises new questions regarding the role of GST mu in these cells because of the inherent risk of MM in individuals with a congenital deficiency of this isoenzyme. The role of GSTs in the resistance of cells to chemotherapy is also discussed.

Inherited susceptibility to several cancers but absence of linkage between dysplastic nevus syndrome and CDKN2A in a melanoma family with a mutation in the CDKN2A (P16INK4A) gene.

Genetic predisposition plays an important role in the development of nearly 10% of cases of cutaneous malignant melanoma (CMM). The CDKN2A gene has been described as responsible for melanoma susceptibility in a proportion of families with CMM linked to 9p. CDKN2A encodes a cyclin-dependent kinase inhibitor also implicated in the carcinogenesis of several sporadic tumors. Even though the incidence of other cancers is higher in CMM families, pancreatic adenocarcinoma is the only other well demonstrated cancer associated with CDKN2A mutations in some CMM pedigrees. We describe a family with four cases of CMM, eight patients affected by other cancers, and nine patients affected by dysplastic nevus (DN) syndrome. A CDKN2A frameshift mutation (358delG) was present in all the CMM patients, in at least three of the patients with other cancers (CDKN2A status is unknown in four patients), and in only two of the DN patients (CDKN2A status is unknown in one patient). An absence of linkage between chromosome 9p markers and the 358delG CDKN2A mutation and DN was detected, indicating genetic heterogeneity for DN and CMM in this family. The study strongly suggests that CDKN2A mutations are involved not only in the predisposition to CMM but also to several other types of cancer.

Detection of circulating neoplastic cells by reverse-transcriptase polymerase chain reaction in malignant melanoma: association with clinical stage and prognosis.

PURPOSE: Circulating melanoma cells can be detected in peripheral blood by means of tyrosinase mRNA amplification by reverse-transcriptase polymerase chain reaction (RT-PCR). We conducted a prospective study to evaluate the clinical significance of the presence of circulating neoplastic cells in the blood of patients with malignant melanoma (MM). METHODS: A sensitive RT-PCR assay was used to detect tyrosinase mRNA in the peripheral blood of patients with stages I to IV melanoma. Healthy subjects or patients with other malignancies were used as negative controls. RESULTS: Ninety-one assessable patients were included in the study. There was a statistically significant association between RT-PCR positivity and clinical stage. Circulating melanoma cells were detected in 36% of patients with localized disease (stages I and II), in 45% of patients with regional nodal involvement (stage III), and in 94% of patients with metastatic disease (stage IV) (P < .001). In stage II-III patients who were RT-PCR-positive for mRNA tyrosinase in blood, the recurrence rate and disease-free survival were significantly worse than patients who were RT-PCR-negative. In multivariate analysis, RT-PCR was an independent prognostic factor for recurrence in patients with nonmetastatic disease (P = .002). CONCLUSION: The detection of circulating melanoma cells in peripheral blood by RT-PCR correlated with the clinical stage of patients with melanoma and was an independent prognostic factor for recurrence. Further studies are warranted to better assess the significance of this test in the evaluation of prognosis, early detection of relapse, and in monitoring the effectiveness of systemic therapy.