KPV Entry Through GSDMD Pores vs. a PepT1 Baseline

Decision

YELLOW — A2 unresolved.

COMP-042 supports rapid passive KPV entry within its declared pore model, but it does not establish intracellular pharmacology or physiological selectivity.

• A1, exposure-proxy diagnostic: intra-articular GREEN, subcutaneous YELLOW, oral RED under the declared route-concentration design spaces. The comparator is the lowest extracellular KPV concentration reported effective in a PepT1-positive cell assay, not an intracellular IC50, target-engagement threshold, or efficacy bar.
• A2, pore-versus-healthy heuristic: unresolved. Favorable ≥3× corners occur in the full sensitivity grid, including some intra-articular moderate- and high-PepT1 cases. The healthy-cell equation and PepT1 scenarios are unvalidated, and concurrent PepT1 transport in the pyroptotic cell is not modeled.
• Platform scope: this KPV result does not resolve the broader hypothesis that a transporter-orphan, membrane-impermeant payload could exploit GSDMD pores.

Model

The passive pore contribution uses a short cylindrical pore with two-sided access resistance:

p_pore = H · D · π · r² / (L + π·r/2)
τ_eq   = V_cell / (N_pores · p_pore)
C_pore = C_ext · (1 − exp(−t/τ_eq))

C_pore is capped at the extracellular boundary concentration. It is the modeled passive pore contribution, not total KPV in a pyroptotic cell.

The A2 response surface compares that contribution with a heuristic healthy-cell PepT1 baseline:

C_healthy = AR_lin · Km · C_ext/(Km + C_ext)
S_model   = C_pore/C_healthy

AR_lin has four unweighted scenarios: absent, low, moderate, and high. The equation omits measured synovial-macrophage Vmax, efflux, turnover, degradation, membrane potential, and proton coupling. S_model is therefore an equation-response diagnostic, not physiological selectivity or a probability.

A1 result: passive pore contribution vs. extracellular assay proxy

Route	Central extracellular design input	Central ratio to 10 nM proxy	Unweighted design-space fraction ≥ proxy	A1 state
Intra-articular	292 µM	29,200×	1.000	GREEN
Subcutaneous	0.030 µM	3.0×	0.679	YELLOW
Oral	0.001 µM	0.1×	0.036	RED

These route concentrations are not established synovial exposures. The intra-articular range is arithmetic from unsourced dose and compartment-volume assumptions; the subcutaneous and oral ranges are named pharmacokinetic design spaces.

At the central pore parameters, τ_eq is 2.17 seconds and the modeled passive contribution reaches essentially the extracellular boundary during the 300-second lifetime. Across the unweighted A1 sampling space, the equilibration fraction has a median of 1.000 and a fifth percentile of 0.975.

The deterministic lifetime × pore-count grid includes a one-pore stress case outside the main 10–10,000-pore design range. At 10 pores and 60 seconds, the modeled equilibration fraction is 0.749, so “complete equilibration at every modeled ≥10-pore condition” is not supported. Pore lifetime is low-sensitivity in much of the declared space, not universally irrelevant.

All results in this section are Mechanistic Extrapolation (computational). Dalmasso et al. provide the In Vitro extracellular assay observation used as the engineering proxy; they do not provide the modeled intracellular threshold.

A2 result: full route concentration × Km sensitivity

Each route crosses three extracellular-concentration bounds, three Km bounds, and four PepT1 scenarios: 36 cases per route, 108 total.

Route	PepT1 absent	PepT1 low	PepT1 moderate	PepT1 high
Intra-articular	9/9 ≥3×	9/9 ≥3×	2/9 ≥3×	1/9 ≥3×
Subcutaneous	9/9 ≥3×	9/9 ≥3×	0/9 ≥3×	0/9 ≥3×
Oral	9/9 ≥3×	9/9 ≥3×	0/9 ≥3×	0/9 ≥3×

The full grid contradicts the old central-only claim that favorable selectivity occurs only when PepT1 is absent or low. It does not establish the opposite claim. The crossings show what the heuristic equation permits; no scenario is known to represent synovial macrophages, and no route qualifies without a matched empirical baseline.

When the modeled healthy-cell PepT1 baseline is zero, strict JSON stores selectivity_ratio: null with selectivity_ratio_state: positive_infinity_zero_healthy_baseline. That value means mathematical positive infinity, not missing data. A future 0/0 case has the distinct state undefined_zero_over_zero.

Interpretation boundaries

1. PepT1 remains the empirical gate. Dalmasso et al. demonstrated PepT1-mediated KPV uptake in epithelial and Jurkat models (In Vitro). Functional PepT1 and KPV accumulation in synovial macrophages remain unmeasured.
2. The A2 numerator is pore-only. Concurrent PepT1 transport in the pyroptotic cell is excluded, so the model does not estimate total-cell pyroptotic-versus-intact selectivity.
3. Pharmacodynamic timing is unresolved. KPV is framed as acting upstream of GSDMD pore formation. That ordering makes therapeutic-timing sufficiency uncertain, but the transport model does not establish that all relevant activity is over or that KPV arriving through a pore cannot matter.
4. Intracellular stability is unresolved. Extracellular or serum stability cannot be transferred to intracellular retention without evidence.
5. A KPV-specific result is not a platform verdict. A transporter-orphan, membrane-impermeant, downstream-acting payload remains a cleaner probe of pore-specific delivery.

Discriminating experiment

Validation §1.32 tests an empirically confirmed transporter-orphan, membrane-impermeant tracer in matched pore-on/off conditions. KPV is a comparator in a pore-on/off × PepT1-on/off design, not an efficacy endpoint. Any conclusion is bounded to the exact tracer, concentration, cell model, and time window tested.

Evidence and reproduction

• In Vitro: Dalmasso et al. reported PepT1-mediated KPV uptake and nanomolar extracellular activity in PepT1-positive cell assays.
• In Vitro / structural: Sborgi et al. and Xia et al. provide the GSDMD pore-geometry anchors.
• Mechanistic Extrapolation (computational): permeability, equilibration, route proxy ratios, and the A2 response surface.
• Named gaps: synovial-macrophage PepT1 function, route-specific synovial KPV exposure, per-cell pore count, intracellular KPV degradation, and matched total-cell accumulation.

The exact code, inputs, generated outputs, and independent lifecycle receipts are in etc/experiments/comp-042-kpv-gsdmd-pore-influx/.