Clinical relevance of DNA ploidy and proliferative activity in childhood rhabdomyosarcoma: a retrospective analysis of patients enrolled onto the Italian Cooperative Rhabdomyosarcoma Study RMS88.

PURPOSE: Evaluation of the possible clinical relevance of DNA ploidy and proliferative activity assessed as S-phase fraction (SPF) in childhood rhabdomyosarcoma (RMS). PATIENTS AND METHODS: We conducted a retrospective study on 59 RMS patients enrolled onto the ICS-RMS88 protocol (seven botryoid, 35 embryonal, and 17 alveolar RMS), for which formalin-fixed paraffin-embedded (FFPE) tissue was available. Nuclear suspensions for cytometric investigation were obtained using a mechanical disaggregation. Tumors were distinguished according to their DNA index (DI) value as follows: diploid (0.9 < DI < 1.1), hyperdiploid (1.1 < or = DI < 1.8 or DI > or = 2.2), and tetraploid (1.8 < or = DI < 2.2); for analysis of SPF, a cutoff value of 14% was used. RESULTS: DNA histograms were diploid in 19 (33%) cases, hyperdiploid in 29 (49%), and tetraploid in 10 (32%). One patient showed both a hyperdiploid and a tetraploid peak. The 5-year overall survival (OS) rate by ploidy status was 73% in hyperdiploid patients as compared with 33% and 25% in diploid and tetraploid patients, respectively (P = .0012). A striking difference emerged when the 5-year OS for the combined diploid and tetraploid RMS groups was compared with survival of the hyperdiploid RMS group: 30% versus 73%, respectively (P = .0006). In addition, the SPF was prognostically relevant: 5-year OS by SPF less than or greater than 14% was 70% and 36%, respectively (P = .009). Multivariate analysis confirmed the importance of DNA content (P = .0006) and SPF (P = .034) in predicting survival. CONCLUSION: These findings confirm that ploidy and SPF are important new prognostic factors that are able to identify selected groups of patients at high risk of treatment failure, even if the tumor's presentation is favorable according to standard criteria.

Prognostic value of rhodamine-efflux and MDR-1/P-170 expression in childhood acute leukemia.

The evidence that mechanisms other than P-170 expression may influence its "pump" and the retention/efflux of chemotherapeutic agents, prompted us to investigate the value of a functional multidrug resistance (MDR) assay in a series of childhood acute leukemia samples. Forty acute leukemia cases, mainly of lymphoid origin (ALL), were evaluated for MDR expression using a functional test based on rhodamine-123 efflux (Rhd-E). This was correlated with the quantification of P-170 external epitopes based on the positivity with the 4E3.16 and MRK16 monoclonal antibodies (MAbs). When compared with the status of the disease and response to treatment, the mean (m) Rhd-E value was significantly lower in patients at diagnosis (m = 7.1% versus m = 22.4% at relapse) and in patients who achieved a complete remission (m = 8.81% versus 31.5% in resistant cases). In the 22 samples analyzed, an overall correlation was found between the functional assay and the P-170 expression (r = 0.6), despite the much lower level of MDR positivity recognized by the immunocytometric method (m = 0.78% and 0.9% in cases at diagnosis versus m = 3.7% and 4.1% at relapse, with the 4E3.16 and MRK16 MoAbs). These data suggest that the assessment of the clinical impact of MDR expression in pediatric ALL should be based on methodological approaches capable of providing information extended to the P-170 pump function, rather then only on its gene and protein expression.