Clonality of cell populations in refractory anaemia using combined approach of gene loss and X‐linked restriction fragment length polymorphism‐methylation analyses

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Abstract
We have used X‐linked restriction fragment length polymorphism (RFLP)‐methylation and gene deletion analyses to investigate the nature of the progenitor cell of origin in the myelodysplastic syndromes (MDS). Gene deletion studies were performed on the granulocyte and T‐lymphocyte fractions of six women with refractory anaemia (RA) and either a partial deletion of the long arm of chromosome 5 (5q‐) or monosomy 7. All six showed gene loss in the granulocyte but not the T‐lymphocyte fractions, indicating monoclonality of the granulocytes but not the T‐lymphocytes. In order to further investigate this finding, we subsequently performed X‐RFLP‐methylation studies using the probe M27β, and also a probe for the phosphoglycerate kinase (PGK) gene. These studies have confirmed the monoclonality of the granulocytes and the polyclonality of the T‐lymphocytes in these cases. Our findings suggest that in this group of patients with MDS the T‐lymphocytes were not involved in the disorder, and furthermore, in the one case where B‐lymphocytes were also available, that the progenitor cell of origin was restricted to the myeloid lineage.This work was supported by the Leukaemia Research Fund of the United Kingdom. We thank Dr I. Craig and Professor A. Jeffreys for the probes M27β and pγ3, respectively. The probe for the FMS gene was kindly donated by Dr N. Spurr. The probes for the HPRT and PGK gene regions were used with the permission of Dr B. Vogelstein. We thank Dr D. Y. Mason for use of the monoclonal antibodies CD22 and CD68.