Acute Leukemias - Republican Scientific Medical Library

a 20.2 · Methods for Detection of MRD 249
Table 20.1 (continued)
Flow cytometric immunophenotyping
Disadvantages Instability of antigenic expression
on leukemic cells
(lineage switch, loss of antigens)
during or after the
treatment course
(Immunophenotypic shifts)
This chapter will focus on current strategies for monitoring
MRD in ALL and will attempt to address these
questions by summarizing some of the recent studies
of MRD monitoring in ALL.
20.2 Methods for Detection of MRD
MRD detection techniques rely on the ability to identify
a unique marker on the leukemia cells. The two methods
that typically have been employed for MRD detection
and monitoring include polymerase chain reaction
(PCR) methods and flow cytometry (FC). For PCR techniques,
monitoring of a leukemia-specific fusion gene
(e.g., BCR-ABL) or a clone-specific rearrangement of
the immunoglobulin heavy chain (IgH) or T-cell receptor
(TCR) genes have been used. For flow cytometric
MRD monitoring, an aberrant immunophenotype present
on the cell surface of the leukemic blasts can be
identified at diagnosis and used for MRD monitoring.
These techniques have far greater sensitivity than standard
cytogenetic analysis and may detect anywhere
from one in ten thousand to one leukemia cell in a background
of one million normal cells. General characteristics
of each of these techniques are described below
and summarized in Table 20.1.
20.2.1 Flow Cytometric Detection of MRD
Multiparameter flow cytometry is a widely applicable
and reliable approach for monitoring MRD in ALL
PCR analysis of chromosome
aberration
Lack of reproducibility of
results when small numbers
of transcripts are present
Presence of oligoclonal
populations that can cause
both false-negative and
false-positive results
Difficult quantification of
MRD
PCR analysis of IgH/TCR genes
Risk of RNA degradation and inefficiency
during conversion of mRNA to
cDNA (which may reduce the sensitivity
of RT-PCR monitoring)
Lack of reproducibility of results
when small numbers of transcripts
are present
Presence of oligoclonal populations
that can cause both false-negative
and false-positive results
due to the presence of aberrant or unusual immunophenotypes
expressed on the cell surface of the lymphoblast.
These aberrant immunophenotypes can be the result
of cross-lineage expression (e.g., presence of myeloid
antigens on a lymphoid progenitor cell), asynchronous
expression of lymphoid maturation antigen (e.g.,
when two or more antigens not normally present at
the same stage of normal hematopoietic differentiation
are coexpressed on the lymphoblast), antigen overexpression,
absence of normal maturation antigens, and/
or ectopic antigen expression [4–6].
FC detection of MRD can be utilized in the majority
of cases of both B- and T-lineage ALL and is rapid,
relatively sensitive, and quantitative, with the ability
to detect one leukemia cell in a background of 10 3 –10 4
normal cells. Disadvantages to this technique include
a lack of standardization across laboratories, with significant
variation depending on the expertise of the operator,
difficulty in distinguishing between normal
regenerating bone marrow progenitors and residual
leukemic blasts, and the instability of the antigenic expression
of the leukemic clone with resultant immunophenotypic
shifts during treatment that can result in
false-negative MRD results [7, 8]. Conversely, the selection
of inappropriate antigens to distinguish leukemic
cells from normal cells may result in false-positive
MRD results.