Page 302 - Haematologica Vol. 109 - July 2024

LETTER TO THE EDITOR
Selection of dormant cells during treatment of T-lineage lymphoblastic leukemia and CREB as a therapeutic target
The slower clearance of acute lymphoblastic leukemia (ALL) cells during induction therapy is associated with an increased risk of relapse. Insight into the mechanism(s) of the multidrug resistance, which these cells clearly dis- play, may lead to more specific, minimal residual disease (MRD)-directed therapies. We have recently reported on the use of single-cell, high dimensional mass cytometry to functionally characterize B-lineage ALL cells in both drug-naïve cells at disease presentation and in those persisting during induction.1 The large panel of more than 30 antibodies allowed for the discrimination of leukemia cells from normal cells, based on the identification of a leukemia-associated immunophenotype (LAIP), and provid- ed information on cell cycle, apoptosis and the activation state of key signaling pathways, known to be recurrently activated in ALL. We demonstrated the presence of a strongly activated CREB pathway across a broad spectrum of cytogenetic groups and, using high dimensional anal-
Table 1. Details of patients used in the study.
yses, showed that activated CREB was prominent in all viable cell clusters at both presentation and in MRD. We also showed that minor subpopulations with hyperactive CREB visible at presentation had a selective advantage during induction therapy. Furthermore, using a specific inhibitor of CREB transcriptional function, we validated CREB as a therapeutic target in ALL cells. Here, we expand on our previous study using a mass cytometric antibody panel specific for T-lineage ALL (T-ALL) and similarly show the presence of activated CREB in almost all viable cell clusters during therapy, their sensitivity to CREB inhibition and importantly show that MRD is made up of non-cycling cells that appear to be selected for during therapy. These data suggest that MRD-directed therapies should be cell cycle-independent and that the CREB pathway is a valid therapeutic target in T-ALL and, as we have previously shown, B-lineage ALL.
Bone marrow samples were accessed through the New-
Sample ID
Sex
Flow MRD ALL %
ALL cell numbers analyzed by CyTOF
Age in years at presentation
Presentation WBC x109/L
Survival status
LK970 Presentation
Male
74.95
209,263
11
316
Alive
LK14 Presentation
Female
86.7
69,828
11
450
Alive
LK134 Presentation
UN
77.9
46,967
UN
UN
UN
LK145 Presentation
Male
2.26*
2,239
2
50
Alive
LK189 Presentation
Male
74.98
78,564
18
92
Alive
LK203 Presentation
Male
76.81
54,573
4
760
Alive
LK214 Presentation
Male
64.9
50,094
11
43
Alive
LK214 MRD day 8
12.95
14,318
LK223 Presentation
Female
90
118,289
4
294
Alive
LK223 MRD day 8
15.6
565
LK223 MRD day 29
0.3
104
LK287 Presentation
Male
69.69
111,012
15
183
Alive
LK287 MRD day 21
37.03
22,883
LK287 MRD day 29
4
821
LK290 Presentation
Male
89.95
68,762
16
76
Alive
LK296 Presentation
Male
95.72
74,729
13
20
Alive
LK304 Presentation
Male
5.60**
98,906
5
11
Alive
LK342 Presentation
Male
80.1
105,999
8
91
Alive
LK342 MRD day 29
Positive by PCR**
189
LK350 Presentation
Male
80.54
103,067
1
578
Died from disease
LK350 MRD day 8
56.7
171,494
LK350 MRD day 29
13.33
11,810
Minimal residual disease (MRD) was assessed using flow cytometry following the principles of our standardized method for B-lineage acute lymphoblastic leukemia (ALL) and involved 5 panels of 6 antibody combinations, along with Syto 41 to exclude red cell contamination.10 *Sam- ple was from a mediastinal mass. **There was inadequate sample for Flow MRD but polymerase chain reaction (PCR) MRD was reported as >0.01%. ***Patient had been exposed to 2 days of dexamethasone. UN: unknown; CyTOF: cytometry time of flight; WBC: white blood cell count.
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