Dynamic BH3 profiling before and after menin inhibition — a one-week assay on primary cells that would choose the BH3-mimetic partner, and has never been run in lymphoid disease
also called DBP under revumenib · apoptotic priming under menin inhibition · menin inhibitor plus venetoclax mechanism test
There is a quick laboratory test that says how close a cell is to dying and which survival protein is holding it back. Several trials are already combining a menin inhibitor with a drug that targets one of those survival proteins, and nobody has run the test to check the combination makes sense.
The same thing, for a scientist
Three facts in this graph sit next to each other and have never been put together. (1) Menin inhibition induces **rapid dose-dependent cell death** in KMT2A-rearranged ALL and slow differentiation in AML [E1] — so in the lymphoid disease the drug's proximate mechanism is apoptotic. (2) KMT2A-rearranged infant ALL is the one paediatric ALL subgroup that is BCL-2-dependent rather than BCL-XL-co-dependent, and the dependence **moves to MCL-1** within hours of a targeted agent [E2]. (3) A menin inhibitor plus venetoclax is being pursued clinically, and the only study combining a menin inhibitor with BH3 profiling is myeloid and does not report a KMT2A-rearranged before-and-after priming result [E3]. Dynamic BH3 profiling — short drug exposure, then peptide panel, then cytochrome c or depolarisation readout — answers the missing question directly: does menin inhibition raise priming, lower it, or shift the dependence, and to what. The assay runs in days on primary material, is established in this exact disease, and would say whether venetoclax, an MCL-1 inhibitor, or neither is the right partner, and whether the answer differs between AML and ALL.
Design
Material. Primary KMT2A-rearranged blasts, both ALL (infant where obtainable) and AML, plus SEM and MOLM-13 as anchors, plus a KMT2A-wild-type B-ALL comparator. The lineage contrast is the point.
Exposure. Revumenib (clinical agent, so the result transfers) at pharmacologically relevant concentrations, for a short window (hours) for dynamic profiling and a longer window (days) for the differentiated or dying state. A menin-inhibitor time course matters because E1 reports the two lineages respond on different timescales.
Profiling. Standard BH3 peptide panel — BIM, BID, BAD (BCL-2/BCL-XL), HRK (BCL-XL), MS1 or NOXA (MCL-1) — reading cytochrome c release or mitochondrial depolarisation. Baseline and post-exposure, same sample.
Primary outputs.
- Change in overall priming (delta-priming) under menin inhibition, per lineage.
- Change in which anti-apoptotic protein holds the threshold.
- Whether a TP53-mutant subset behaves differently, which is predicted by the existing revumenib-resistance result.
Secondary. Repeat on the small/quiescent fraction if exp-elutriation-phase-and-size-resolved-kmt2a arm B succeeds, since the persister compartment's priming is the clinically decisive number and is currently unmeasurable.
What each outcome would mean
| Result | Interpretation | Action |
|---|---|---|
| Priming rises, dependence stays BCL-2 | menin inhibition lowers the death threshold and venetoclax is the right partner | supports current combination trials |
| Priming rises, dependence shifts to MCL-1 | the cell escapes into MCL-1 as it does under kinase inhibitors | pair with an MCL-1 inhibitor, sequentially; venetoclax would underperform |
| Priming does not change | the drug kills by a route BH3 profiling does not read, or the death is not mitochondrial | the combination rationale is not apoptotic and needs a different basis |
| Priming falls | menin inhibition de-primes surviving cells | the combination could be antagonistic, and the sequencing matters |
| AML and ALL differ | the combination cannot be chosen once for 'KMT2A-rearranged acute leukaemia' | stratify the trials by lineage |
The fourth row is the one worth the experiment on its own. A drug class given with venetoclax in open trials, which de-primes the cells it does not kill, would produce exactly the pattern of early response and early relapse this disease shows, and no current assay would detect it.
| status | proposed |
|---|---|
| assay | dynamic BH3 profiling: short drug exposure then BH3 peptide panel, readout cytochrome c release or mitochondrial depolarisation |
| agent | revumenib, a clinical agent, so the result transfers to open trials |
| depends on | for the persister compartment, exp-elutriation-phase-and-size-resolved-kmt2a arm B |
| cost | days to weeks; established assay; no new reagents |
peptide panel
[ "BIM", "BID", "BAD (BCL-2/BCL-XL)", "HRK (BCL-XL)", "MS1 or NOXA (MCL-1)" ]
arms
[ "primary KMT2A-rearranged ALL (infant where obtainable)", "primary KMT2A-rearranged AML", "SEM (KMT2A::AFF1) and MOLM-13 (KMT2A::MLLT3)", "KMT2A-wild-type B-ALL comparator", "TP53-mutant subset, predicted to differ" ]
primary outputs
[ "delta-priming under menin inhibition, per lineage", "identity of the anti-apoptotic dependence before and after", "whether TP53 status modifies the shift" ]
decision table
{
"priming_up_bcl2_retained": "venetoclax is the right partner; supports current trials",
"priming_up_shift_to_mcl1": "sequential MCL-1 inhibition indicated; venetoclax would underperform",
"no_change": "death is not read by this assay; the combination rationale is not apoptotic",
"priming_down": "combination may be antagonistic; sequencing critical",
"aml_all_divergence": "stratify combination trials by lineage"
}atlas refs
[
{
"label": "ALL dies rapidly and AML differentiates slowly under revumenib",
"pmid": "38892207",
"title": "Distinct Responses to Menin Inhibition and Synergy with DOT1L Inhibition in KMT2A-Rearranged Acute Lymphoblastic and Myeloid Leukemia.",
"authors": "Adriaanse FRS, Schneider P, Arentsen-Peters STCJM, et al.",
"venue": "Int J Mol Sci",
"year": 2024,
"title_provenance": "Title/authors/venue backfilled from PubMed esummary on 2026-09-20 because this citation stored a bare PMID with no title, which made it unscoreable by verify-citations.py. BACKFILLING IS NOT VERIFICATION: it confirms the PMID resolves to a real paper, not that this is the right paper for the claim. That still requires reading."
},
{
"label": "MCL-1 adaptation detected by dynamic BH3 profiling in KMT2A-rearranged cells",
"pmid": "34568318",
"title": "MCL-1 Inhibition Overcomes Anti-apoptotic Adaptation to Targeted Therapies in B-Cell Precursor Acute Lymphoblastic Leukemia.",
"authors": "Manzano-Muñoz A, Alcon C, Menéndez P, et al.",
"venue": "Front Cell Dev Biol",
"year": 2021,
"title_provenance": "Title/authors/venue backfilled from PubMed esummary on 2026-09-20 because this citation stored a bare PMID with no title, which made it unscoreable by verify-citations.py. BACKFILLING IS NOT VERIFICATION: it confirms the PMID resolves to a real paper, not that this is the right paper for the claim. That still requires reading."
}
]- A null result would not clear the menin-plus-venetoclax combination; it would mean the assay cannot see the relevant mechanism, and the combination would remain unjustified rather than justified.
- If priming falls under menin inhibition, several open combination trials are running a potentially antagonistic pairing, and the finding would arrive after rather than before enrolment.
- Priming measured ex vivo is measured on cells removed from the niche, and this graph records that dormancy-associated drug resistance in ALL reverses on removal from the niche — so the assay may systematically measure the more sensitive version of the cell.
- Acting on an MCL-1 shift is constrained by tolerability, not by mechanism: MCL-1 inhibition with venetoclax caused lethal tumour lysis in most treated mice in infant KMT2A-rearranged models.
- Would the assay even register the death menin inhibition causes in KMT2A-rearranged ALL? If that death is not mitochondrial, dynamic BH3 profiling returns a null and the combination rationale needs a different instrument.
- Can enough primary infant KMT2A-rearranged material be obtained for a peptide panel at two timepoints? This is the practical constraint that has probably prevented the experiment, not the idea.
- Should the profiling be done on the surviving cells rather than the bulk? The bulk answer averages over cells that died, and the clinically relevant priming is that of the survivors.
- E1strong confidence 4 of 5: Replicated in peer-reviewed work by more than one group.paper
In vitro, KMT2A-rearranged ALL samples showed rapid dose-dependent induction of leukaemic cell death in response to revumenib while KMT2A-rearranged AML responses were much slower and promoted myeloid differentiation; acquired resistance in KMT2A-rearranged ALL arose either through MEN1 mutations or independently of them, and revumenib was significantly synergistic with the DOT1L inhibitor pinometostat in KMT2A-rearranged ALL.
Distinct Responses to Menin Inhibition and Synergy with DOT1L Inhibition in KMT2A-Rearranged Acute Lymphoblastic and Myeloid Leukemia · Adriaanse FRS, Schneider P, Arentsen-Peters STCJM, Fonseca AMND, Stutterheim J, Pieters R, et al., Stam RW · Int J Mol Sci 2024What this evidence cannot show
In vitro, with sample numbers per arm not given in the abstract and fusion partner unresolved, so lineage and partner are confounded. 'Cell death' is scored by viability assays that do not establish the death is mitochondrial apoptosis - which is precisely the gap this experiment would close, and also a reason the experiment could return a null. Read: abstract only.
Evidence that argues against this (1)
- A third outcome exists that a viability assay cannot see and that a priming assay would also miss: lineage switch. In a patient sample that switched during revumenib treatment, oncoprotein and menin fell to undetectable levels while ENL persisted on oncoprotein-target loci including a GMP-like programme. Cells taking that route neither differentiate nor die, so an experiment framed entirely around the death axis would characterise two of three exits and miss the one that causes treatment failure with the drug fully on target. Read: abstract only. Janssens DH et al. 2024
- E2moderate confidence 3 of 5: One strong study, or several smaller consistent ones.paper
Dynamic BH3 profiling of B-cell precursor ALL showed that trametinib and sunitinib sharply increased apoptotic priming in an NRAS-mutant line and in a KMT2A-rearranged FLT3-high line respectively, that profiling after treatment identified anti-apoptotic adaptation in the BCL-2 family particularly involving MCL-1, that metronomic sequential addition of the MCL-1 inhibitor S63845 was synergistic and more effective than single agents, and that KMT2A-rearranged patient-derived xenograft cells responded to sunitinib with a dynamic BH3 profile analogous to the SEM cell line.
MCL-1 Inhibition Overcomes Anti-apoptotic Adaptation to Targeted Therapies in B-Cell Precursor Acute Lymphoblastic Leukemia · Manzano-Munoz A, Alcon C, Menendez P, Ramirez M, Seyfried F, Debatin KM, et al., Montero J · Front Cell Dev Biol 2021What this evidence cannot show
The KMT2A-rearranged arm is one cell line plus PDX cells, with KMT2A::AFF1 only; the drugs used are a MEK inhibitor and a multi-target TKI, neither of which is a menin inhibitor, so the MCL-1 adaptation is established for a different perturbation class. Dynamic BH3 profiling measures priming over hours and its correspondence to durable clinical benefit is inferred. Read: abstract only.
Evidence that argues against this (1)
- The in vivo record for acting on an MCL-1 adaptation in this disease is much worse than the in vitro record. In infant KMT2A-rearranged PDX models, an MCL-1 inhibitor gave no significant survival benefit as monotherapy at any dose, did not beat venetoclax alone when combined with it, and at higher doses with venetoclax caused lethal tumour lysis syndrome in the majority of treated mice; only the triple combination with dexamethasone produced durable responses. A priming result showing an MCL-1 shift would therefore not license an MCL-1 inhibitor as the partner without the in vivo tolerability problem being solved first. Read: the numerical detail is recorded in this graph's target-mcl1-with-bcl2-and-dexamethasone-infant-kmt2a-all entry from that beat's reading of the full text; the identifiers here were re-fetched from PubMed, whose record for this article carries no abstract. Karsa M et al. 2026
- E3moderate confidence 3 of 5: One strong study, or several smaller consistent ones.paper
The menin inhibitor DS-1594b combined with venetoclax was tested in AML cell lines with MLL rearrangements or NPM1 mutations and in primary AML patient samples using proliferation, viability, apoptosis and differentiation assays, western blotting and BH3 profiling, showing significant synergy with enhanced differentiation and inhibited proliferation, with menin inhibition the primary driver of transcriptional change and affecting expression of anti-apoptotic regulators, offered as the mechanistic explanation for the synergy.
Menin inhibitor DS-1594b drives differentiation and induces synergistic lethality in combination with venetoclax in acute myeloid leukemia cells with rearranged mixed-lineage leukemia and mutated nucleophosmin-1 · Ciaurro V, Sharlandjieva V, Skwarska A, Chahrour C, Baran N, Zeng Z, et al., Konopleva M · Haematologica 2026What this evidence cannot show
This is the closest existing approach to the proposed experiment and it is myeloid only, uses a non-approved menin inhibitor, and the abstract does not report a before-and-after priming result for KMT2A-rearranged cells specifically; the in vivo arm is an NPM1-mutated xenograft. So it establishes that the assay has been applied in this drug class, not that the question has been answered. Read: abstract only.
Evidence that argues against this (1)
- If a menin inhibitor already kills KMT2A-rearranged ALL rapidly on its own, a combination rationale based on lowering the apoptotic threshold may be redundant there, and a priming shift measured in myeloid cells that differentiate slowly cannot be transferred to lymphoid cells that die quickly. The experiment could therefore return a large effect in AML and a null in ALL, and the null would be the more consequential result because infants are the population in question. Read: abstract only. Adriaanse FRS et al. 2024
measures whether menin inhibition raises, lowers or redirects apoptotic priming, and to which anti-apoptotic protein
tests whether the rapid death seen in KMT2A-rearranged ALL is mitochondrial apoptosis, which decides whether a BH3-mimetic partner is rational
would say whether venetoclax, an MCL-1 inhibitor or neither is the right partner for a menin inhibitor, per lineage
a lineage-stratified priming measurement is a direct test of the hypothesis on the apoptotic axis