Red blood cell phenotyping is a sensitive technique for monitoring chronic myeloid leukaemia patients after T‐cell‐depleted bone marrow transplantation and after donor leucocyte infusion

Abstract

Summary. Fifteen consecutive patients with Philadelphia chromosome (Ph)-positive chronic myeloid leukaemia (CML) who relapsed from T-cell-depleted bone marrow transplantation (BMT) were successfully treated with donor leucocyte infusions (DLIs). Chimaerism was analysed using red blood cell phenotyping (RCP), and the results were compared with cytogenetic analysis and outcome of qualitative and quantitative polymerase chain reaction (PCR) for breakpoint molecules. In all patients, an increase in autologous erythrocytes and/or a decrease in donor red cells indicated relapse. Donor erythrocytes started to increase from 4 to 20 (median 12) weeks after DLI. At 6 and 12 months after DLI, complete donor chimaerism was found in 11 and 15 patients, respectively, and all patients were in cytogenic remission. A high percentage of autologous red cells at the time of DLI predicted pancytopenia. During relapse and after DLI, the percentage of autologous red cells was strongly correlated with the reappearance and disappearance of Phpositive metaphases (r 0?90; P < 0?001 and r 0?96; P < 0?001 respectively). The same was true for the correlation between the percentage of autologous red cells and positivity/negativity in PCR for Bcr-Abl breakpoint molecules (r 0?94; P < 0?001). A normalized Bcr-Abl dose of greater than 10 À3 in real-time quantitative PCR correlated well with relapse and the presence of autologous red blood cells (r 0?77; P < 0?001). We conclude that RCP is a sensitive, easy to perform and fast technique for the prediction of pending relapse after BMT for CML. RCP also predicts the response to DLI and the occurrence of bone marrow aplasia after DLI.

How and when should we monitor chimerism after allogeneic stem cell transplantation?

“…These methods include, for example, cytogenetics, 20,21 red cell phenotyping, 22,23 restriction fragment length polymorphism analysis (RFLP) [24][25][26][27] and fluorescence in situ hybridization of sex chromosomes. [28][29][30][31][32][33] A major limitation of these different techniques was that they were time consuming and did not offer the possibility to study all patients (for reviews, see Thiede et al 17 and Thomas et al 34 ).…”

Significance of chimerism in hematopoietic stem cell transplantation: new variations on an old theme

“…36 Due to this, patients transplanted for CML are monitored frequently by RCP. An increasing percentage of autologous red blood cells and/or a decreasing number of erythrocytes of donor origin always correlates with relapse.…”

“…In sex-mismatched HSCTs, Ychromosome-specific markers, 31 X-chromosome-specific markers 32 and amelogenin loci 25 have been frequently used, and recently dual-color fluorescent in situ hybridization (FISH) has been introduced as the technique of choice in cases of sex-mismatched HSCTs. [33][34][35] Also used in routine analysis to establish chimerism status are cytogenetic techniques, erythrocyte phenotyping 36 and restriction fragment length polymorphisms (RFLPs). Some of the studies have also used PCR-based allele-specific amplification techniques like the amplified refractory mutation system (ARMS) 37 or restriction endonuclease in situ digestion (REISDs) 38 to detect the mixed chimerism.…”

Erythrocyte antigen and reticulocyte engraftment after allogeneic hematopoietic stem cell transplantation

“…RFLP is a highly sensitive method; however, there is a need for qualified staff and specialized equipment and it is time consuming. 9,13,14 Using EA to demonstrate chimerism provides an opportunity to follow the process of engraftment using red cells alone, [15][16][17] and offers the advantages of simplicity and speed without requirement for either special equipment or staff.…”