In vitro and in vivo delivery of the secretagogue diadenosine tetraphosphate from conventional and silicone hydrogel soft contact lenses

Purpose To evaluate the possible use of soft contact lenses (CL) to improve the secretagogue role of diadenosine tetraphosphate (Ap4A) promoting tear secretion. Methods Two conventional hydrogel CL (Omafilcon A and Ocufilcon D) and two silicone hydrogel (SiH) CL (Comfilcon A and Balafilcon A) were used. Ap4A was loaded into the lenses by soaking in a 1 mM Ap4A solution during 12 h. In vitro experiments were performed by placing the lenses in multi-wells during 2 h containing 1 ml of ultrapure water. 100 μl aliquots were taken at time zero and every minute for the first 10 min, and then every 15 min. In vivo experiments were performed in New Zealand rabbits and both the dinucleotide release from SiH and tear secretion were measured by means of Schirmer strips and high-pressure liquid chromatography (HPLC) analysis. Results Ap4A in vitro release experiments in hydrogel CL presented a release time 50 (RT50) of 3.9 ± 0.2 min and 3.1 ± 0.1 min for the non-ionic and the ionic CL, respectively. SiH CL released also Ap4A with RT50 values of 5.1 ± 0.1 min for the non-ionic and 2.7 ± 0.1 min for the ionic CL. In vivo experiments with SiH CL showed RT50 values of 9.3 ± 0.2 min and 8.5 ± 0.2 min for the non-ionic and the ionic respectively. The non-ionic lens Ap4A release was able to induce tear secretion above baseline tear levels for almost 360 min. Conclusion The delivery of Ap4A is slower and the effect lasts longer with non-ionic lenses than ionic lenses.

Efficient in vivo antitumor effect of an immunotoxin based on ribotoxin α-sarcin in nude mice bearing human colorectal cancer xenografts

Tagging of RNases, such as the ribotoxin α-sarcin, with the variable domains of antibodies directed to surface antigens that are selectively expressed on tumor cells endows cellular specificity to their cytotoxic action. A recombinant single-chain immunotoxin based on the ribotoxin α-sarcin (IMTXA33αS), produced in the generally regarded as safe (GRAS) yeast Pichia pastoris, has been recently described as a promising candidate for the treatment of colorectal cancer cells expressing the glycoprotein A33 (GPA33) antigen, due to its high specific and effective cytotoxic effect on in vitro assays against targeted cells. Here we report the in vivo antitumor effectiveness of this immunotoxin on nude mice bearing GPA33-positive human colon cancer xenografts. Two sets of independent assays were performed, including three experimental groups: control (PBS) and treatment with two different doses of immunotoxin (50 or 100 μg/ injection) (n = 8). Intraperitoneal administration of IMTXA33αS resulted in significant dose-dependent tumor growth inhibition. In addition, the remaining tumors excised from immunotoxin-treated mice showed absence of the GPA33 antigen and a clear inhibition of angiogenesis and proliferative capacity. No signs of immunotoxin-induced pathological changes were observed from specimens tissues.Overall these results show efficient and selective cytotoxic action on tumor xenografts, combined with the lack of severe side effects, suggesting that IMTXA33αS is a potential therapeutic agent against colorectal cancer.