CD34+ subset and tumor cell quantitation by flow cytometry--step toward quality assessment of autografts in B cell malignancies

Abstract

To day it is possible to predict the probability of fast engraftment based on a very simple flow cytometry standard analysis of CD34+ cells as documented by the 28 laboratories within the NSCL-G. However, the risk for delayed platelet engraftment still needs to be predicted in clinical practice for patients receiving less than 10 x 10(6) CD34+ cells/kg. Here we present data from our center supporting that identification by double staining of uncommitted (CD34+/CD38-) and lineage specific (CD34+/CD61+) progenitors may allow us to predict patients at high risk for prolonged platelet recovery. Following high dose therapy more than 30% of patients with haematological malignancies do suffer from disease recurrence within the first 3-6 months following high dose therapy. Today there are strong indications that such patients may have been transplanted with an autograft contaminated with a high number of potentially malignant B cells. Here we present a novel methodology for quantitation of blood circulating tumor cells by combining flow cytometry, cell sorting, limiting dilution and single cell RT-PCR. Such methodology has documented mobilization of clonal B cells following priming of the peripheral blood stem cell harvest and it can be used to identify minor populations and predict the efficacy of patient specific purging strategy. Consequently, quality assessment of autografts may include techniques which can predict fast three lineage engraftment as well as the risk for prolonged platelet recovery and can identify the group of patients/autografts with a strong contamination of potential tumor cells with a risk of early relapse. The future supportive cell therapy may depend upon improvements of such technologies and strategies including the selective administration of lineage specific growth factors e.g., trombopoietin as well as patient specific controlled purging strategies.