The European Myeloma Network (EMN) organized two flow cytometry workshops. The first aimed to identify specific indications for flow cytometry in patients with monoclonal gammopathies, and consensus technical approaches through a questionnaire-based review of current practice in participating laboratories. The second aimed to resolve outstanding technical issues and develop a consensus approach to analysis of plasma cells. The primary clinical applications identified were: differential diagnosis of neoplastic plasma cell disorders from reactive plasmacytosis; identifying risk of progression in patients with MGUS and detecting minimal residual disease. A range of technical recommendations were identified, including: 1) CD38, CD138 and CD45 should all be included in at least one tube for plasma cell identification and enumeration. The primary gate should be based on CD38 vs. CD138 expression; 2) after treatment, clonality assessment is only likely to be informative when combined with immunophenotype to detect abnormal cells. Flow cytometry is suitable for demonstrating a stringent complete remission; 3) for detection of abnormal plasma cells, a minimal panel should include CD19 and CD56. A preferred panel would also include CD20, CD117, CD28 and CD27; 4) discrepancies between the percentage of plasma cells detected by flow cytometry and morphology are primarily related to sample quality and it is, therefore, important to determine that marrow elements are present in follow-up samples, particularly normal plasma cells in MRD negative cases.

A B S T R A C T PurposeHalf the patients with acute myeloid leukemia (AML) who achieve complete remission (CR), ultimately relapse. Residual treatment-surviving leukemia is considered responsible for the outgrowth of AML. In many retrospective studies, detection of minimal residual disease (MRD) has been shown to enable identification of these poor-outcome patients by showing its independent prognostic impact. Most studies focus on molecular markers or analyze data in retrospect. This study establishes the value of immunophenotypically assessed MRD in the context of a multicenter clinical trial in adult AML with sample collection and analysis performed in a few specialized centers. Patients and MethodsIn adults (younger than age 60 years) with AML enrolled onto the Dutch-Belgian Hemato-Oncology Cooperative Group/Swiss Group for Clinical Cancer Research Acute Myeloid Leukemia 42A study, MRD was evaluated in bone marrow samples in CR (164 after induction cycle 1, 183 after cycle 2, 124 after consolidation therapy). ResultsAfter all courses of therapy, low MRD values distinguished patients with relatively favorable outcome from those with high relapse rate and adverse relapse-free and overall survival. In the whole patient group and in the subgroup with intermediate-risk cytogenetics, MRD was an independent prognostic factor. Multivariate analysis after cycle 2, when decisions about consolidation treatment have to be made, confirmed that high MRD values (Ͼ 0.1% of WBC) were associated with a higher risk of relapse after adjustment for consolidation treatment timedependent covariate risk score and early or later CR. ConclusionIn future treatment studies, risk stratification should be based not only on risk estimation assessed at diagnosis but also on MRD as a therapy-dependent prognostic factor.

Cytomegalovirus (CMV) infection is an

We conclude that perforin expression can be severely reduced in sJIA. This finding may implicate defective cytotoxicity and haemophagocytosis and could thus explain why sJIA may be complicated by macrophage activation syndrome. ASCT leads to a reconstitution of the (T cell) immune system with a normal expression of perforin.

We studied the recovery of CMV-specific CD4 + and CD8 + T-cell immunity in 52 recipients of allogeneic stem cell transplantation (SCT). The proportions of IFN-c-producing CD4+ and CD8 + T cells upon in vitro activation using peptide pools representing the CMV pp65 and IE-1 proteins were assessed at multiple time points post SCT, and correlated with the occurrence of CMV reactivation. In a retrospective analysis, recurrent CMV reactivations occurred in 9 patients and were associated with low pp65-specific CD4 + T-cell and low IE-1-specific CD8 + T-cell reactivities, whereas patients without detectable CMV reactivation (n = 30) or a single reactivation (n = 13) showed a better recovery of these immune responses. CD4 + T-cell responses to IE-1 were infrequent in most patients, whereas CD8 + T-cell responses to pp65 occurred frequently, but did not correlate with protection against (recurrent) reactivation. Prospectively, CMV-specific T-cell responses could be studied prior to 14 reactivation episodes in 8 patients. CD4 + T-cell responses to IE-1 and pp65 were positive in only 1 and 2 episodes, respectively. CD8 + T-cell responses against IE-1 were positive in 4, but against pp65 in 12 episodes, again showing that CD8 + T-cell reactivity against pp65 did not prevent CMV reactivation. Thus, monitoring of particular CMV-specific CD4 + and CD8 + T-cell responses after allogeneic SCT may identify patients at risk for recurrent CMV reactivations. q