harvested in remission when - by definition - no malignant cells are 
detectable. It had therefore been impossible to undertake any form of quality 
control to determine if residual malignant cells have genuinely been 
eliminated. Moreover, there was no evidence that malignant cells with the 
ability to reestablish the malignancy after ABMT were physically, 
phenotypically, or biochemically identical to the cells of the "mature" tumor. 
Until recently, therefore, the justification for bone marrow purging had been 
largely a matter of faith. This was cause for considerable concern. The 
techniques of purging almost invariably damage normal progenitor cells, so 
that the engraftment of purged marrow is generally substantially slower than 
the engraftment of untreated marrow (33-35). Morbidity and mortality from 
the complications of hemopoietic and immune system failure were 
correspondingly increased (10,27). 
2.4 Use of Gene Marking to Determine the Need for Purging 
Since September 1991, we have been using the LNL6 or GIN retroviruses to 
mark one third of the marrow harvested for ABMT. These vectors confer 
Neomycin resistance which can be detected phenotypically or genetically, 
using the polymerase chain reaction. By November 1992 we had treated 9 
AML patient marrows, 5 with LNL6 and 4 with GIN. Two of these patients 
subsequently relapsed, and in both a proportion of the malignant cells at 
relapse were marker gene positive. Details of these patients are given in 
Appendix D. We therefore have unequivocal evidence that marrow harvested 
in clinical remission of AML contains cells capable of contributing to disease 
recurrence. Effective marrow purging will therefore be one essential 
precondition to improving outcome of disease treatment. 
2.5 Biology of Autologous Reconstitution 
These gene marked marrows also provided information about the mechanisms 
of marrow repopulation after BMT about which comparatively little had been 
known. We found that the cryopreserved marrow contained viable long lived 
"stem cells" which could be marked and which reconstituted the patient. 
Marked hemopoietic cells - including pluripotent GEMM progenitor cells - 
have been detected beyond one year post-BMT and T and B lymphoid 
lineages are also marked at this time (see Appendix E). Thus, the autograft 
does not simply provide temporary replenishment of committed progenitor 
cells whilst surviving host stem cells gradually repopulate the individual. 
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Recombinant DNA Research, Volume 17 
