The DOT1L inhibitor removes menin from fusion promoters too, which a pure interface model does not predict
A drug aimed at a completely different enzyme - DOT1L, which writes a chemical mark on histones - also strips menin off the fusion's target genes. Menin's position on chromatin is therefore not controlled only by its handshake with the fusion.
The same thing, for a scientist
At 4 days, EPZ-5676 (pinometostat) 250 nM reduces median MENIN TSS signal at the authors' target genes to **0.467** of DMSO (n=202) against **0.744** at non-target genes (n=2,283). VTP-50469 gives 0.315 / 0.651 and the dTAG degron gives 0.437 / 1.271. So a **DOT1L catalytic inhibitor removes menin from fusion-occupied promoters about as effectively as destroying the fusion does** (0.467 vs 0.437). A model in which menin's chromatin residence at these promoters is set solely by the menin-KMT2A contact does not predict that. The parsimonious readings are (i) menin residence is partly downstream of the active-transcription state that DOT1L maintains, or (ii) DOT1L inhibition destabilises fusion occupancy, which then releases menin - and the same table supports (ii), since EPZ-5676 reduces HA at target genes to 0.362.
Why this is filed as a caution rather than a refutation
Four days is long enough for the leukaemic programme to collapse and for cells to differentiate, so menin loss under EPZ-5676 may be a consequence of the programme failing rather than a direct chromatin event. The 3 h degron arm, which is too fast for that, shows menin loss too - so the direct route exists. What this entry establishes is that the observed menin loss at fusion promoters is not diagnostic of the interface being broken, because a drug that does not touch that interface produces a similar loss.
Scale and limits
No spike-in exists anywhere in GSE173599, so every number is a within-sample share, and the treated arms are scaled by a 21-gene housekeeping panel whose own raw 3 h H3K27ac ratio averages 0.809 and is forced to 1. A housekeeping normalisation DEFINES the global change to be zero, so only the DIFFERENCE between strata is measurable; a uniform genome-wide loss of any of these proteins would be invisible here. n = 1 per arm for every channel except HA, which has two independent 3 h batches. The cells are HCB1, transformed by KMT2A::MLLT3 (MLL::AF9) - NOT the index fusion of this project, KMT2A::AFF1 - and a prior beat in this corpus found that per-gene ordering transfers between MLLT3 and AFF1 lines while magnitude does not. All 4-day arms; EPZ-5676 at 250 nM in the 1912x2 batch. DOT1L inhibition is known to act slowly because H3K79me2 turnover is slow, so a 4-day window is the shortest that shows an effect and also the window in which secondary effects appear.
| script | analyses/menin-retention-by-ha-occupancy/analyze_controls.py |
|---|---|
| status | run |
| result summary | MENIN at target genes, 4 d: EPZ-5676 0.467, VTP-50469 0.315, dTAG-VHL 0.437. At non-target genes: 0.744, 0.651, 1.271. |
| peak width ratio note | Signal instrument; no peak-width ratio applies. |
inputs
[ "GSE173599_TSS_signal_Figure4.xlsx" ]
- If a DOT1L inhibitor produces the same menin-occupancy change as a menin inhibitor, then menin occupancy cannot be used as a target-engagement biomarker for a menin inhibitor in a combination trial - and the combination is exactly what this paper argues for.
- Menin's residence being partly downstream of transcription means a menin inhibitor's chromatin readout will look best in the cells that were already responding, inflating apparent target engagement in responders.
- Does EPZ-5676 reduce menin occupancy at fusion promoters at a timepoint too early for differentiation - for instance 24 h - or only after the programme has collapsed? No such arm exists in this deposit.
- If menin residence is partly downstream of the transcriptional state, is menin occupancy a marker of active MLL-fusion transcription rather than a cause of it?
- E1moderate confidence 3 of 5: One strong study, or several smaller consistent ones.dataset
After 4 days of EPZ-5676, median MENIN TSS signal at MLL::AF9 target genes is 0.467 of DMSO, against 0.437 for the dTAG degron in the same table.
GSE173599_TSS_signal_Figure4.xlsx - per-gene HA, DOT1L and MENIN TSS signal after 4 days of EPZ-5676, VTP-50469 or dTAG-VHL (batch 1912x2) · Olsen SN et al. · NCBI GEO supplementary file 2022What this evidence cannot show
n=1 per arm; 4-day treatment with differentiation confound; the MENIN sheet holds 9,574 genes after the authors' floor, of which 202 are target genes.
Evidence that argues against this (1)
- DOT1L inhibition reduces fusion occupancy itself (HA 0.362 at target genes in the same table), so the menin loss may be entirely secondary to fusion loss rather than independent of the interface. Olsen SN et al. 2022
- E2strong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.paper
Combined DOT1L and MENIN inhibition phenocopies MLL::AF9 degradation and releases the oncoprotein from chromatin globally, whereas single agents have more modest effects.
MLL::AF9 degradation induces rapid changes in transcriptional elongation and subsequent loss of an active chromatin landscape · Olsen SN, Godfrey L, Healy JP, Choi YA, Kai Y, Hatton C, Perner F, Haarer EL, Nabet B, Yuan GC, Armstrong SA · Molecular Cell 82(6):1140-1155.e11 2022What this evidence cannot show
This is the paper's own headline conclusion; it is a 4-day drug comparison in one cell line plus MOLM13, n=1 per arm.
Evidence that argues against this (1)
- The phenocopy claim is made at 4 days in two cell lines with n=1 per arm; a combination that resembles a degron on a chromatin readout need not resemble it on the transcriptional or phenotypic readouts that matter. Olsen SN et al. 2022
- E3established confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.paper
DOT1L catalytic inhibition with EPZ-5676 has independent clinical and preclinical support as an MLL-rearranged leukaemia strategy.
MLL::AF9 degradation induces rapid changes in transcriptional elongation and subsequent loss of an active chromatin landscape · Olsen SN, Godfrey L, Healy JP, Choi YA, Kai Y, Hatton C, Perner F, Haarer EL, Nabet B, Yuan GC, Armstrong SA · Molecular Cell 82(6):1140-1155.e11 2022What this evidence cannot show
Background; the paper cites the primary DOT1L inhibitor literature rather than generating it.
Evidence that argues against this (1)
- Pinometostat single-agent clinical activity in MLL-rearranged leukaemia has been modest, so preclinical support for DOT1L inhibition has not translated, and read-across from EPZ-5676 chromatin effects to clinical benefit is not warranted. Olsen SN et al. 2022
- related to →Pinometostat (EPZ-5676) — the DOT1L inhibitor, and the most instructive failure in this field
EPZ-5676 at 4 d reduces MENIN occupancy at fusion target genes to 0.467, close to the degron's 0.437
menin occupancy at fusion promoters is not diagnostic of the menin-KMT2A interface being broken
DOT1L catalytic inhibition removes menin from fusion promoters