Human Lipocalin-Type Prostaglandin D Synthase-Based Drug Delivery System for Poorly Water-Soluble Anti-Cancer Drug SN-38

<div><p>Lipocalin-type prostaglandin D synthase (L-PGDS) is a member of the lipocalin superfamily, which is composed of secretory transporter proteins, and binds a wide variety of small hydrophobic molecules. Using this function, we have reported the feasibility of using L-PGDS as a novel drug delivery vehicle for poorly water-soluble drugs. In this study, we show the development of a drug delivery system using L-PGDS, one that enables the direct clinical use of 7-ethyl-10-hydroxy-camptothecin (SN-38), a poorly water-soluble anti-cancer drug. In the presence of 2 mM L-PGDS, the concentration of SN-38 in PBS increased 1,130-fold as compared with that in PBS. Calorimetric experiments revealed that L-PGDS bound SN-38 at a molecular ratio of 1:3 with a dissociation constant value of 60 μM. The results of an <i>in vitro</i> growth inhibition assay revealed that the SN-38/L-PGDS complexes showed high anti-tumor activity against 3 human cancer cell lines, i.e., Colo201, MDA-MB-231, and PC-3 with a potency similar to that of SN-38 used alone. The intravenous administration of SN-38/L-PGDS complexes to mice bearing Colo201 tumors showed a pronounced anti-tumor effect. Intestinal mucositis, which is one of the side effects of this drug, was not observed in mice administered SN-38/L-PGDS complexes. Taken together, L-PGDS enables the direct usage of SN-38 with reduced side effects.</p></div

Systemic anaphylactic reaction in mice.

<p>Change in body temperature of mice. Mice were sensitized with OVA or L-PGDS, and then challenged with OVA (○) or L-PGDS (●). The body temperature was measured for 60 min. The data are expressed as the mean ± SE of 6 independent experiments.</p

Effects of SN-38/L-PGDS complexes on the small intestines.

<p>(A) Histology of the ileal mucosa. Inset shows the villi architecture. Scale bar represents 100 μm. The left panel shows the ileal mucosa of the mice administered SN-38/L-PGDS complexes; and the right one, that of the mice administered PBS. The ileal mucosa in both groups was almost the same. (B) Expression levels of inflammatory cytokines in the ileal mucosa. Each bar represents the mean ± SD (<i>n</i> = 3).</p

Structures of L-PGDS and compounds.

<p>(A) Crystal structure of human L-PGDS (molecular mass: 18777.7, PDB ID: 3O2Y). (B, C) Chemical structures of SN-38 (relative molecular mass: 392.4) and CPT-11 (relative molecular mass: 677.2).</p

<i>In vitro</i> growth inhibition effect of SN-38/L-PGDS complex (○), SN-38 (●) and CPT-11 (■) on human colorectal cancer cell line Colo201 (A), breast cancer cell line MDA-MB-231 (B), and prostate cancer cell line PC-3 (C).

<p>The data are expressed as the mean ± SE. (<i>n</i> = 6).</p

Characterization of the SN-38/L-PGDS complex.

<p>(A) Calorimetric titration of SN-38 with L-PGDS. L-PGDS in the injection syringe was reversely titrated to SN-38 in the cell. The upper panel shows the change in heat over time as L-PGDS was titrated into SN-38. The lower panel shows the normalized change in heat after subtracting reference data of L-PGDS injections into PBS. The one-set of independent binding sites model was used to fit the binding isotherms. (B) SAXS profiles of L-PGDS (red line) and SN-38/L-PGDS complex (blue line). These profiles were obtained by extrapolating all data at different concentrations (12, 9.0, 6.0, and 3.0 mg/ml) to the zero concentration. The logarithm of scattering intensity is shown as a function of the reciprocal vector (<i>S</i>). The inset shows the logarithm of scattering intensity in the small <i>S</i> region.</p