In Vivo Efficacy of Nano Hyaluronan-Conjugated Cisplatin for Treatment of Murine Melanoma
March 2014 | Volume 13 | Issue 3 | Original Article | 283 | Copyright © March 2014
Qiuhong Yang MS,a* Daniel J. Aires MD JD,b* Shuang Cai PhD,a Garth R. Fraga MD,b
Da Zhang MD,b Cicy Z. Li MS,b and M. Laird Forrest PhDa
aUniversity of Kansas, Lawrence, KS
bUniversity of Kansas Medical Center, Kansas City, KS
*Contributed equally to this article
BACKGROUND: Melanoma is a deadly skin cancer with rapidly rising incidence. While localized melanoma can be treated with excision, there are at present no similarly effective treatments for regional and distant disease, so survival rates are low. One problem is that melanoma is chemo-resistant, and most chemotherapy doses are limited by systemic toxicity. A method for delivering high-dose chemotherapy directly to tumors and draining lymph nodes could have the advantage of allowing much higher effective doses with reduced systemic exposure.
METHODS: Human melanoma cell line A-2058 tumor cells were injected into athymic mice. After tumors grew to 50~100 mm3 mice were divided into five groups: (1) nontreated (2) intravenous (i.v.) cisplatin, (3) i.v. nano hyaluronan-conjugated cisplatin (HA-Pt), (4) subcutaneous (s.c.) peri-tumoral cisplatin, and (5) s.c. peri-tumoral HA-Pt. All treatment groups received 3 weekly doses of 10 mg/kg.
RESULTS: Tumors grew progressively in all control, i.v. cisplatin, and s.c. cisplatin groups. Tumors showed a trend toward slower growth in the i.v. HA-Pt group, but all animals died or were euthanized per protocol within 3 weeks of treatment. Tumors showed shrinkage only in the subcutaneous peri-tumoral HA-cisplatin group; one of these mice appeared to be cured.
CONCLUSIONS: Peri-tumoral HA-cisplatin may be shown potential as a therapeutic option in treatment of certain types of melanoma. J Drugs Dermatol.
Melanoma is a deadly skin cancer, killing more than 9,000 Americans in 2012. Incidence is rising rapidly, to the point where 1 in 50 Americans will develop melanoma.1 Stage at diagnosis is the main determinant of survival. While 5-year survival for localized melanoma is 98%, involvement of regional lymph nodes drops survival to 62%, and in distant metastatic disease survival is only 15%.1
The main reason is that very early disease can generally be successfully treated with simple excision. However, once it has spread to the lymph nodes and beyond, melanoma becomes very hard to treat and survival decreases accordingly. This applies to “locally advanced” melanoma with lymph node involvement as well as widespread metastatic disease. One important reason for this is that melanoma is notoriously resistant to chemotherapy. Conventional chemotherapy does not result in high levels of penetration into tumors or lymph nodes. Therefore efficacy is limited by systemic toxicity. It has long been a goal to increase relative penetration of chemotherapy into tumors and lymph nodes. Here we report the first use of a novel peri--tumor injectable chemotherapy compound in an in-vivo murine model for locally advanced melanoma. We seek to answer the question of whether increased peri-tumoral dose translates into a measurable in vivo response.
MATERIALS AND METHODS
All chemicals were obtained from commercial suppliers and used without further purification unless otherwise noted. Hyaluronan (HA; 35 kDa) was purchased from Lifecore Biomedical (Chaska, MN) as sodium hyaluronate, which was cultured and produced by a microbial fermentation process. Cisplatin (CDDP) was obtained from AK Scientific (Union, CA). All other chemicals and cell culture
supplies were purchased from Sigma-Aldrich Co (St. Louis, MO) or Fisher Scientific (Pittsburgh, PA). Distilled water was used in syntheses, cell culture (sterilized by autoclaving) and animal experiments (sterilized by autoclaving). Human melanoma cell line A-2058 was obtained from American Type Culture Collection (ATCC, MA) and cultured according to ATCC protocol.
Synthesis of Hyaluronan-Cisplatin Conjugates
HA-Cisplatin (HA-Pt) conjugate was prepared as previously described.2 Briefly, HA (50 mg) and CDDP (40 mg) were dissolved in a total of 80 mL double distilled water (ddH2O) and stirred in the dark for 96 hr at ambient temperature (~25°C). By the end of the reaction, the mixture was filtered through a 0.22-μm nylon membrane filter (Fisher Scientific; Pittsburgh, PA), followed by dialysis (MWCO 10,000 Da; Pierce, IL) against ddH2O for 24 hr in dark with 4 water changes. The crude HA-Pt conjugate was concentrated by evaporation under reduced pressure and then stored at room temperature in dark.