Oncofusion

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Explainerestablished confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.Q118-menin-at-the-human-index-fusion

An MLL1 N-terminus ChIP sees the fusion and wild-type MLL1 in one track, and a menin ChIP has the same problem from the other side - so the standard reagents of the menin-inhibitor literature cannot attribute occupancy to the fusion, and the masking factor is now measured at 3.5 to 4.0-fold

also called the attribution problem, named by reagent · MLL1n is not fusion-specific · rule 74 applied to KMT2A

The antibody used in most of these experiments recognises the front end of the MLL1 protein, and the front end is exactly the part the cancer fusion keeps - so the antibody cannot tell the cancer protein from the normal one. Measured side by side, that ambiguity hides three quarters of the drug's effect.

The same thing, for a scientist

**Every reagent in the founding VTP-50469 occupancy dataset is a shared-epitope reagent, and the antibody names say so** [E1]: | target | reagent as deposited | what it can see | what it cannot separate | |---|---|---|---| | **MLL1n** | anti-MLL1n, Bethyl **A300-086A** lots 5 or 6 | an MLL1 **N-terminus** epitope | the fusion (which retains the N-terminus) from **wild-type MLL1** | | **MENIN** | anti-Menin, Bethyl **A300-105A** Lot 8 | menin | fusion-bound menin from **wild-type-MLL1-bound** menin and from **MLL1-independent** menin | | **DOT1L** | anti-DOT1L, CST **#77087S** clone **1W4Z** | DOT1L | fusion-recruited DOT1L from wild-type DOT1L complexes | | **H3K79me2** | anti-H3K79me2, CST **#5427** lot 4 | a histone mark | a consequence from a cause | This is rule 74 (*for an antibody readout the clone is the identifier, not the label*) applied to the family this project is about. **`MLL1n` is not a hedge in the sample title - it is the reagent's specificity, stated.** **The consequence is not theoretical and it is now quantified.** In human RS4;11 (KMT2A::AFF1), at the same 2 kb bins in the same cells, the MENIN within-sample share ratio under VTP-50469 is **0.1999-0.2560** and the MLL1-N share ratio is **0.7996-0.9069** - a **3.54 to 4.00-fold** discrepancy that depends only on which antibody was used [E2]. RS4;11 is heterozygous for t(4;11) and retains a wild-type KMT2A allele, so both species are present in every sample. **Two readings, and no reagent in this series can choose between them.** Either the fusion left with menin and wild-type MLL1 is what remains (dilution), or an MLL1-N-containing complex genuinely stays after menin departs (retention, which is this graph's `hyp-the-fusion-stays-on-chromatin-and-stops-working`). **Three things follow for how claims should be written.** 1. **Write the reagent's subject, not the intended one.** "MLL1-N-reactive material" and "menin-occupied sites", never "the fusion binds X", from these tracks. This graph already holds `concept-eviction-names-menin-not-the-fusion` for the menin half; this is the MLL1 half. 2. **An MLL1 C-TERMINUS antibody is the missing control.** The C-terminus is lost in the fusion, so a C-terminal ChIP is a wild-type-only track and the fusion's contribution could be obtained by subtraction. **No such sample exists in GSE127507 and none is known to this graph in any menin-inhibitor dataset.** 3. **A tagged fusion is the only clean route**, and Q117's numbers are attributable precisely because its fusion was FLAG-tagged. The human equivalent does not exist and is written up as `exp-q118-a-tagged-kmt2a-aff1-chip-in-rs4-11-under-vtp-50469`.

Facts and figures
definitionAn MLL1 N-terminus antibody detects both the KMT2A fusion (which retains the N-terminus) and wild-type MLL1; a menin antibody detects menin bound to either, plus MLL1-independent menin. Neither can attribute chromatin occupancy to the fusion.
measured masking factor rs4113.54x to 4.00x on the within-sample share ratio at genome scale
missing controlan MLL1 C-TERMINUS ChIP, which would be wild-type-only because the C-terminus is lost in the fusion; no such sample exists in GSE127507
shown for fusionthe reagents are the ones used for KMT2A::AFF1 (RS4;11), KMT2A::MLLT3 (MOLM13) and KMT2A::AFDN (ML-2), and in none of those is the fusion distinguished from wild-type MLL1
absence classnever-measured
absence class noteThe gap this token describes is the MLL1 C-TERMINUS ChIP under a menin inhibitor: no instrument has been pointed at it in any deposited series known to this graph. It does NOT describe the tagged-fusion experiment, which has been done in mouse (GSE260742, FLAG-MLLAF9) and not in a human KMT2A::AFF1 line.
antibodies
{
  "MLL1n": "Bethyl A300-086A, lots 5 or 6 - MLL1 N-terminus",
  "MENIN": "Bethyl A300-105A Lot 8",
  "DOT1L": "Cell Signaling #77087S clone 1W4Z lot 0",
  "H3K79me2": "Cell Signaling #5427 lot 4"
}
How this could bite us back
  • Every pharmacodynamic biomarker in this field built on an MLL1 ChIP is measuring a quantity whose relationship to fusion occupancy is uncertain by a measured factor of four. A compound triaged out for failing to move that readout may have been active.
  • The reverse error is worse: a claim that the drug does NOT displace the fusion, made from an MLL1-N track, would be repeated as a mechanistic fact. This graph holds such a claim, and it is correct about MLL1-N-reactive material and silent about the fusion.
Still open
  1. Where exactly is the A300-086A epitope? If it is mapped, the fraction of clinically observed KMT2A breakpoints that lie C-terminal to it can be computed, and any fusion variant breaking N-terminal to the epitope would be invisible to every experiment in this literature.
  2. Does any deposited dataset anywhere carry an MLL1 C-terminal ChIP in a KMT2A-rearranged line with and without a menin inhibitor? That single dataset would convert every MLL1-N measurement in this field into a fusion measurement by subtraction, and no archive search for it was run in this beat.
  3. How many of this graph's occupancy claims about the fusion rest on an MLL1-N or MLL1-C antibody rather than a tagged construct? The corpus already records that 0 of 24 sampled load-bearing experiments rest on a dedicated human KMT2A::AFF1 experiment, but no reagent-level audit of the occupancy claims specifically has been done.
Every source behind this entry (2)
  1. E1established confidence 5 of 5: Multiple independent lines of evidence; treated as settled in the field.dataset

    GSE127507's four ChIP targets are anti-MLL1 N-terminus (Bethyl A300-086A lots 5 or 6), anti-Menin (Bethyl A300-105A Lot 8), anti-DOT1L (Cell Signaling #77087S clone 1W4Z lot 0) and anti-H3K79me2 (Cell Signaling #5427 lot 4), as recorded in the !Sample_characteristics_ch1 antibody field of all 26 sample records, and none of them is fusion-specific.

    What this evidence cannot show

    The catalogue numbers and lots are transcribed from the deposited records and were not checked against the vendors' current specifications. That the N-terminus is retained in KMT2A::AFF1 is a property of the fusion architecture recorded elsewhere in this graph and is not re-derived here. Whether A300-086A's epitope lies N-terminal to every clinically observed KMT2A breakpoint was not verified; it is described by the vendor as an N-terminal antibody and the sample title calls it MLL1n.

    Evidence that argues against this (1)
    • If the A300-086A epitope happened to lie between the most common KMT2A breakpoints and the N-terminus in a way that excluded some fusion variants, the attribution problem would be different rather than absent - the antibody would then be partly fusion-blind rather than fusion-ambiguous. The exact epitope coordinates were not established here and that is a real gap in this entry. Krivtsov AV et al. 2019
  2. E2strong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.dataset

    In RS4;11 the MENIN and MLL1-N within-sample share ratios at the same 2 kb bins under VTP-50469 differ by 3.54-fold at the MLL1-N held set and 4.00-fold at the MENIN held set, and by 2.68 to 5.12-fold on the fraction-retained-in-own-top statistic across RS4;11, MOLM13 and ML-2.

    What this evidence cannot show

    NEITHER ANTIBODY IN THIS SERIES IS FUSION-SPECIFIC AND THE ANTIBODY NAME SAYS SO. MLL1n is anti-MLL1 N-terminus (Bethyl A300-086A, lots 5 or 6); the N-terminus is retained in KMT2A::AFF1, so that track is the sum of the fusion and wild-type MLL1 in unknown proportion. MENIN is Bethyl A300-105A Lot 8; menin binds the fusion, wild-type MLL1 and non-KMT2A partners. Q117 could attribute its numbers because its fusion was FLAG-tagged; nothing here can. Every occupancy statement in this beat is about MLL1-N-reactive material or menin-occupied sites, never about the fusion. No scale factor exists for any sample. The depositors state that a Drosophila spike-in was run on one batch, that fly/human ratios were 'found to be small', and that NO spike-in normalisation was applied; no number for that measurement is reported and none is deposited. Every cross-condition figure here is therefore a WITHIN-SAMPLE share or a within-sample rank, which cancel an unknown per-sample scale factor exactly, and never a fold change. n = 1 per antibody per condition per line - there is no replicate anywhere in GSE127507 - so no number here carries a spread and none is precise. The MENIN and MLL1n ChIPs are separate experiments with different efficiencies, so the ratio of their share ratios is not a direct measurement of how much wild-type MLL1 there is. The MOLM13 figure partly reflects a 4-day MENIN exposure against a 3-day MLL1n one.

    Evidence that argues against this (1)
    • The discrepancy is exactly what would be seen if the fusion were not evicted and menin simply dissociated from an intact chromatin-bound complex - in which case nothing is masked and the MLL1-N number is the right one. The entry states both readings and cannot choose. Iqbal Lab oncofusion knowledge graph 2026
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