The menin inhibitor in this head-to-head is at 50 nM and all three DOT1L inhibitors are at 100 nM - the comparison is dose-biased AGAINST the menin inhibitor - and compound 10's target engagement at exactly 100 nM is demonstrated inside the same series by a spike-in-normalised H3K79me2 collapse of 5.6-fold by day 3
also called GSE149183 dose control
A sceptic would ask whether the menin drug only looked better because it was given at a higher dose. It was given at half the dose of the others, and the same study separately proves the other drugs were working hard at the dose they got.
The same thing, for a scientist
Two controls that decide whether the 0/19-versus-14/19 menin result in `ana-q136-the-menin-inhibitor-clears-menin-off-every-canonical-target-locus-and-three-dot1l-inhibitors-do-not` is a dose artefact. Both come from the deposit's own sample metadata and its sibling arm, not from the paper. **1. The doses run the wrong way for a dose artefact.** Per-sample `treatment` characteristics: **VTP-50469 50 nM**; **EPZ5676 100 nM**; **compound 10 100 nM**; **compound 11 100 nM** - all at 4 d in MOLM13 [E1]. The menin inhibitor is at **half** the concentration of every DOT1L inhibitor it is being compared with, so under-dosing cannot explain why the DOT1L arms retain menin. **2. Compound 10 is demonstrably engaging DOT1L at exactly 100 nM in this series.** The H3K79me2 arm of GSE149183 treats MOLM13 with **compound 10 at 100 nM** - the same compound, same concentration, same cell line - and the depositors' own dm6 spike-in table gives a ratio-to-DMSO of **5.57 at day 3**, 17.93 at day 6 and 19.49 at day 9, with the fly read fraction rising from 1.69% (DMSO d3) to 9.43% [E2]. A 5.6-fold spike-in-normalised H3K79me2 collapse by day 3 is full target engagement one day before the factor ChIP was taken. **So the class claim rests on compound 10, and the other two ride on it.** Compound 10 is the arm with both a demonstrated on-target effect at its own dose and 14/19 menin peaks retained. **EPZ5676's engagement at 100 nM is NOT demonstrated in this deposit** - the H3K79me2 arm dosed EPZ5676 at 1 microM, ten-fold higher - so EPZ5676's 14/19 is formally compatible with under-dosing, and only compound 10's arm closes that loophole [E2]. **A metadata inconsistency to carry forward.** Every sample's `treatment_protocol` field reads *DMSO or VTP-50469 (in DMSO) were added at 1:3000 dilution, for final concentration of DMSO 0.33% and VTP-50469 100 to 330nM*, while the per-sample `treatment` characteristic for GSM4491617 and GSM4491622 reads **VTP50469 (50nM)**. The protocol text is boilerplate shared across all 31 samples and contradicts the per-sample field. **50 nM is the value used here** because it is sample-specific, and it is also the conservative choice: the protocol's 100-330 nM would make the menin arm equi- or higher-dosed and would weaken nothing, so the direction of this uncertainty does not threaten the result [E1].
Why the spike-in ruler can be used here but not on the factor ChIPs
The dm6 spike-in table is deposited for the H3K79me2 samples only. That is enough to certify target engagement - a question about the mark DOT1L writes - and it is not enough to put the MEN1 or DOT1L ChIPs on an absolute scale, because those libraries have no fly chromatin in them. The two uses are different and only the first is available; see absence-q136-the-dm6-spike-in-table-covers-only-h3k79me2-so-the-factor-chips-have-no-absolute-scale.
| script | analyses/menin-inhibitor-against-dot1l-inhibitors-in-one-deposit/run.py |
|---|---|
| status | run |
| geo | GSE149183 |
| concentration source | per-sample GEO characteristics field |
| conflicting protocol text | VTP-50469 100 to 330nM (shared boilerplate across all 31 samples) |
| dose bias direction | AGAINST the menin inhibitor |
concentrations NM — the numbers behind the chart
{
"VTP50469": 50,
"EPZ5676": 100,
"comp10": 100,
"comp11": 100
}engagement control
{
"compound_10_100nM_MOLM13_H3K79me2_dm6_ratio_to_dmso": {
"d3": 5.565127467,
"d6": 17.93472718,
"d9": 19.48662863
},
"pct_fly_reads": {
"DMSO_d3": 1.694850919,
"comp10_d3": 9.4320614
}
}engagement not demonstrated for
[ "EPZ5676 at 100 nM (H3K79me2 arm used 1 microM)", "compound 11 at any dose (no H3K79me2 arm)" ]
- A 50 nM menin-inhibitor exposure clearing menin completely from target loci makes the occupancy endpoint look easy to hit, which invites using it as a low-bar pharmacodynamic marker in trials. A marker that saturates at a low dose cannot report dose-response and is therefore useless for dose-finding.
- What is the intracellular exposure of VTP-50469 at 50 nM relative to compound 10 at 100 nM? Nominal medium concentration is the weakest form of dose matching and it is all this deposit provides.
- Would EPZ5676 at 1 microM - the dose at which this series demonstrates its engagement - still leave menin on 14/19 target peaks? That single extra ChIP would remove the only remaining under-dosing loophole in the whole comparison.
- E1established confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.dataset
The per-sample GEO `treatment` characteristics give VTP-50469 at 50 nM (GSM4491617, GSM4491622) and EPZ5676, compound 10 and compound 11 each at 100 nM (GSM4491618-GSM4491620, GSM4491623-GSM4491625), all at 4 d in MOLM13; the shared boilerplate treatment_protocol field instead states VTP-50469 100 to 330 nM.
NCBI GEO GSE149183 - MEN1, DOT1L and H3K79me2 ChIP-seq in MOLM13 and RS4;11 under DMSO, VTP-50469, EPZ5676, compound 10 and compound 11; 31 samples, Homo sapiens; SubSeries of GSE149185, BioProject PRJNA627612, SRA SRP258055 · NCBI Gene Expression Omnibus 2020What this evidence cannot show
The two fields disagree about the menin inhibitor's concentration and the deposit gives no way to adjudicate. The per-sample field is used because it is sample-specific; the alternative reading only strengthens the conclusion, so the ambiguity is not load-bearing.
Evidence that argues against this (1)
- Concentration is not exposure. A 50 nM VTP-50469 and a 100 nM compound 10 are not equipotent by construction, and neither compound's intracellular free fraction is measured here. The argument is only that the menin arm is not favoured by dose. NCBI Gene Expression Omnibus 2020
- E2established confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.dataset
In the same series, MOLM13 treated with compound 10 at 100 nM shows a dm6 spike-in ratio-to-DMSO of 5.57 at day 3, 17.93 at day 6 and 19.49 at day 9, with the fly read fraction rising from 1.69% under DMSO to 9.43% - the depositors' own numbers, from GSE149183_MOLM13_H3K79me2_TSS-ChIPseq_signal_+_dm6_spike-in_1000u_3000d_res100_sitepro.txt.gz.
NCBI GEO GSE149183 - MEN1, DOT1L and H3K79me2 ChIP-seq in MOLM13 and RS4;11 under DMSO, VTP-50469, EPZ5676, compound 10 and compound 11; 31 samples, Homo sapiens; SubSeries of GSE149185, BioProject PRJNA627612, SRA SRP258055 · NCBI Gene Expression Omnibus 2020What this evidence cannot show
These are the depositors' computed ratios, not a reanalysis. They are taken at days 3, 6 and 9 while the factor ChIPs are at day 4, so the engagement figure bracketing day 4 is the day-3 value of 5.57. A rising fly FRACTION is not by itself a falling human signal, but the human counts are reported beside it and fall from 30.20 M to 25.83 M.
Evidence that argues against this (1)
- EPZ5676 is dosed at 1 microM in the H3K79me2 arm and 100 nM in the factor arm, so this engagement control does NOT cover EPZ5676 at the dose used for its MEN1 ChIP. Compound 11's engagement is not measured at any dose in this series - there is no compound 11 H3K79me2 arm at all. NCBI Gene Expression Omnibus 2020
the menin inhibitor is at 50 nM against the DOT1L inhibitors' 100 nM, so dose bias runs AGAINST the headline result
compound 10 at 100 nM gives a dm6 spike-in H3K79me2 ratio of 5.57 by day 3 - the target-engagement control for the whole DOT1L arm
VTP-50469 dosed at 50 nM in the factor-ChIP arm, with a conflicting 100-330 nM in the deposit's shared protocol text