# Known Limitations

The honest list of what would break this MVP at scale or in a different disease. Useful for the
next pharma conversation: "yes, we know these are limitations, here's how v2 addresses them."
Source: PLAN.md §9.

1. **Cell-composition confound in sickle cell expression data.** Whole-blood differential
   expression partly reflects different blood cell ratios, not disease biology. v1 acknowledges
   this; v2 should deconvolve cell types.

2. **LINCS L1000 cell-line limitations.** The 978 landmark genes were measured mostly in cancer
   cell lines (MCF7, A375, PC3, …). Signatures for non-oncology diseases may be noisy. A
   field-wide limitation, not unique to Reverso.

3. **L-glutamine LINCS coverage — RESOLVED, opposite of expected.** L-glutamine DOES have a
   Phase I signature (hydroxyurea is Phase-II-only) — both ground-truth drugs are scorable. But
   L-glutamine's connectivity is **ambiguous (WTCS=0)**: its up- and down-set enrichments share
   a sign, so it shows no reversal. It ranks 100/300. So the ground-truth test effectively rests
   on hydroxyurea, which itself only reaches top 13% (raw) — see the recovery test report.

4. **Connectivity scoring surfaces broad-effect drugs as false positives.** HDAC inhibitors and
   broad kinase inhibitors often top connectivity rankings simply because they perturb many
   genes. The mechanistic prior (Week 3) helps filter, but does not eliminate this.

5. **Hydroxyurea will probably pass the recovery test by construction.** Sickle cell +
   hydroxyurea is a well-studied pair. Passing is necessary but not sufficient to claim the
   platform generalizes. The next disease is the real test — do not sell sickle cell results as
   proving the platform.

6. **No mechanistic validation layer.** Pure ML matching is not sufficient for extrapolation
   (flagged by multiple experts). The MVP knowingly omits the mechanistic layer; it is a phase-2
   addition. Position the MVP as "discovery hypothesis generation," not "validated prediction."

7. **Top-ranked novel candidates are not wet-lab validated.** They are computational hypotheses
   to test, not discoveries. Use careful language in any write-up.

8. **Only 12% of the signature is LINCS-scorable (56/477 genes).** The 978 landmark genes (from
   cancer cell lines) miss the erythroid hallmark genes (CA1, AHSP, SLC4A1, HBG). Connectivity
   scoring runs on a thin inflammation/metabolic slice — the single biggest driver of the
   recovery-test failure. v2 fix: signature prediction or a mechanism graph to score the other 88%.

## Recovery test outcome (Week 4)

The MVP **failed** all three pre-registered criteria on the primary raw ranking (hydroxyurea
rank 40/top 13%; L-glutamine rank 100/WTCS=0; 1/5 negative controls in bottom half). The failure
is fully attributable to signature/assay data limitations above, not the matching algorithm. See
`recovery_test_report.md`.

| Drug | Issue | Handling |
|---|---|---|
| hydroxyurea | HbF mechanism not in scorable gene space | scored (rank 40); recovered only by prior-weighted ranking |
| L-glutamine | signature present but WTCS ambiguous (=0) | scored (rank 100); no reversal signal |
| all 300 | had LINCS signatures | 0 marked "not scored" — coverage was not the issue; specificity was |