Effects of Topically Applied Betulinic Acid and NVX-207 on Melanocytic Tumors in 18 Horses

Abstract

The naturally occurring betulinic acid (BA) and its derivative NVX-207 induce apoptosis in equine melanoma cells in vitro. After topical application, high concentrations of the substances can be reached in healthy equine skin. With the aim to investigate the effect and safety of topically applied BA and NVX-207 in horses with melanocytic tumors, the longitudinal, prospective, randomized, double-blind, placebo-controlled study protocol included eighteen Lipizzaner mares with early-stage cutaneous melanoma assigned to three groups. Melanocytic lesions were topically treated either with a placebo, 1% BA or 1% NVX-207 twice a day for 91 days. Caliper measurements, clinical examinations and blood tests were performed to assess the effects and safety of the medication. The topical treatment was convenient and safe. The volumes of tumors treated with BA were significantly reduced over time as compared to tumors treated with the placebo from day 80 of the study. Although treatment with NVX-207 seemed to decrease tumor volume, these results did not reach statistical significance. The findings must be regarded as preliminary due to the limited group size and need to be replicated in a larger cohort with modified pharmaceutical test formulations. Accordingly, the treatment protocol cannot yet be recommended in its current form.

In Vitro Insights Regarding the Role of Melanin in Melanoma Cells’ Response to Betulinic Acid Treatment

“…In a recent study by Coricovac et al, BA is shown to induce a dose-dependent cytotoxicity in A375 MM cells after 24 h, accompanied by apoptotic nuclear features (shrinkage, chromatin condensation), up-regulation of pro-apoptotic markers (Bax, Bad, and Bak), inhibition of mitochondrial respiration and glycolysis, and decrease in the mitochondrial membrane potential [21]. A very recent study showed that BA proved to be very active against equine melanoma cells [35].…”

Potent drug delivery enhancement of betulinic acid and NVX-207 into equine skin in vitro – a comparison between a novel oxygen flow-assisted transdermal application device and microemulsion gels

“…Time- and dose-dependent [ 23 , 33 ] cytotoxicity of BA and NVX-207 against melanoma and various cancer cell lines from different species (e.g. human, horse, dog, mouse) has been repeatedly demonstrated in vitro and in vivo [ 23 , 31 , 33 – 49 ]. The cytotoxic mechanism of BA and its derivates is based on a CD95- and p53-independent induction of apoptosis [ 50 , 51 ].…”

“…In Franz-type diffusion cell (FDC) trials, Weber et al examined equine skin permeation of formulations containing BA and NVX-207, which exceeded the half maximal inhibitory concentrations (IC 50 ) of equine melanoma cells in required skin layers [ 33 , 38 ]. The authors also performed the first in vivo experiments in healthy and melanoma-bearing horses, suggesting the efficient distribution of the active ingredients BA and NVX-207 even in deeper skin layers than studied in vitro [ 49 , 54 ]. As previously demonstrated in other studies [ 23 , 32 , 54 ], good tolerability and safety were confirmed [ 49 , 54 ].…”

“…The authors also performed the first in vivo experiments in healthy and melanoma-bearing horses, suggesting the efficient distribution of the active ingredients BA and NVX-207 even in deeper skin layers than studied in vitro [ 49 , 54 ]. As previously demonstrated in other studies [ 23 , 32 , 54 ], good tolerability and safety were confirmed [ 49 , 54 ]. In the study of melanoma-bearing horses, in all three groups (formulations containing 1% BA, 1% NVX-207 or a placebo) reduction in tumor size was observed, whereby the effect was only significant in the BA group [ 49 ].…”

“…In the study of melanoma-bearing horses, in all three groups (formulations containing 1% BA, 1% NVX-207 or a placebo) reduction in tumor size was observed, whereby the effect was only significant in the BA group [ 49 ]. Considering this, and with regard to the long and intensive treatment protocol (application of the formulation twice a day for three consecutive months), the authors recommended further studies with adapted pharmaceutical formulations [ 49 ].…”