Clinical systemic lupeol administration for canine oral malignant melanoma

  • Authors:
    • Inoru Yokoe
    • Kazuo Azuma
    • Keishi Hata
    • Toshiyuki Mukaiyama
    • Takahiro Goto
    • Takeshi Tsuka
    • Tomohiro Imagawa
    • Norihiko Itoh
    • Yusuke Murahata
    • Tomohiro Osaki
    • Saburo Minami
    • Yoshiharu Okamoto
  • View Affiliations

  • Published online on: October 30, 2014     https://doi.org/10.3892/mco.2014.450
  • Pages:89-92
0

Abstract

Canine oral malignant melanoma (COMM) is the most aggressive malignant tumor in dogs. Lupeol is a triterpene extracted from various fruits and vegetables that reportedly inhibits melanoma cell proliferation in vitro and in vivo. In this study, the efficacy of subcutaneous lupeol for spontaneous COMM was evaluated. A total of 11 dogs (3, 5 and 3 dogs diagnosed with clinical stage I, II and III melanoma, respectively) were evaluated. Subcutaneous lupeol (10 mg/kg) was administered postoperatively at various time points to treat these 11 COMM cases. of the 11 subjects, 7 exhibited no local recurrence 180 days postoperatively and no severe adverse effects were observed in any of the cases. Furthermore, no distant metastasis was observed during the experimental period. Therefore, systemic lupeol may prevent local tumor progression and distant metastasis and may be a novel adjuvant treatment for the treatment of COMM.

Introduction

Canine oral malignant melanoma (COMM) is the most common malignancy in dogs, accounting for 30–40% of all oral tumors (1, 2). COMM is characterized by extensive local invasion, as well as distant metastasis (2). Melanoma treatment includes radiation therapy and chemotherapy (CT), either alone, as an adjuvant therapy following surgery, or in combination (2). Although radiation therapy provides effective local tumor control, it is only performed in selected facilities (35). In addition, radiation therapy may be costly and, hence, beyond the financial means of some owners. Several reports have investigated the efficacy of CT in COMM, but there is no definitive evidence regarding its effectiveness (5,6,79). Therefore, there is a need for an alternative treatment for COMM.

Lupeol is a triterpene found in fruits such as olives, mangoes, strawberries, grapes and figs, numerous vegetables and several medicinal plants (10). Previous studies reported that lupeol has antitumor properties (11, 12) and several reports indicated that lupeol inhibits melanoma cell proliferation in vitro and in vivo (13, 14). Recently, Nitta et al (14) reported that systemic lupeol administration inhibited tumor growth in a melanoma-bearing mouse model; however, there is no report evaluating its clinical efficacy in COMM. In this study, we aimed to evaluate the efficacy of subcutaneous lupeol as an adjuvant therapy for spontaneous COMM.

Subjects and methods

Subjects

A total of 11 dogs were included in this study. The characteristics of the sublects are summarized in Table I. The represented breeds included 2 miniature Dachshunds, 2 Beagles, 2 miniature Schnauzers, 1 Golden Retriever, 1 Labrador Retriever, 1 American Cocker Spaniel, 1 Cavalier King Charles Spaniel and 1 mixed-breed dog. The dogs ranged in age between 8 and 17 years. The tumors in all 11 dogs were classified according to the TNM classification (1, 2) as follows: 3 dogs had stage I, 5 dogs had stage II and 3 dogs had stage III disease (Table I). Two cases of cancer recurrence were included (C04 and C07). This study was conducted between April, 2010 and March, 2013 in animal hospitals, including the Veterinary Teaching Hospital of Tottori University, Japan. Treatment was administered to dogs whose owners agreed to this study. The owners were informed of the risk of recurrence and other available treatment options, including surgery, radiation therapy and CT, and they all declined radiation therapy. The owners were offered the alternative treatment comprising surgery, subcutaneous lupeol and CT and they were all informed that lupeol was an experimental therapy. All the owners consented to the enrolment of their dogs in this clinical trial.

Table I

Clinical case characteristics.

Table I

Clinical case characteristics.

Case no.BreedAge, yearsGenderWeight, kgStage
C01Golden Retriever11M39I
C02Cavalier King Charles Spaniel17M7I
C03Labrador Retriever13M28I
C04Miniature Dachshund13M5II
C05Mixed-breed15F10II
C06Miniature SchnauzerNDM6II
C07Miniature Dachshund12M5II
C09Beagle8M14II
C10American Cocker Spaniel14F12III
C11Beagle12M8III
C12Miniature Schnauzer14M10III

[i] M, male; F, female; ND, no data.

Surgical excision

Surgery was performed after the tumors were histopathologically confirmed as malignant melanomas. Complete excision was performed in 1 case (C05), in which the tumor could be excised with sufficient surgical margins. Partial excision without complete margins was performed in the remaining 10 cases; these tumors were difficult to excise completely due to their size or location (Table II).

Table II

Clinical outcomes following melanoma therapy.

Table II

Clinical outcomes following melanoma therapy.

Case no.StageSurgical excisionCombined therapyLocal recurrenceDistant metastasisDFT, daysST, daysLong-term outcome
C01IPartial---210210Alive
C02IPartial---210210Alive
C03IPartial-Yes-110180Alive
C04IIPartialMelphalan, piroxicamYes-170186Alive
C05IIComplete---187187Alive
C06IIPartial---180180Alive
C07IIPartialPHCT--120420Alive
C08IIPartial---201201Alive
C09IIIPartial---248248Alive
C10IIIPartialPHT--189189Alive
C11IIIPartial---3939Alive

[i] DFT, disease-free time; ST, survival time; PHCT, photodynamic hyperthermal chemotherapy; PHT, photodynamic hyperthermal therapy.

Lupeol therapy

Lupeol solution was prepared as previously described (15). Briefly, lupeol was extracted from Indian lettuce (Lactua indica) and dissolved in olive oil (Wako Pure Chemical Industries Ltd., Osaka, Japan) using heat (37°C) and sonification (3 h). The concentrated lupeol solution was diluted to 5 mg/ml.

Lupeol was administered subcutaneously at 10 mg/kg as an adjuvant therapy at least 1 week postoperatively to prevent local recurrence or metastasis. Lupeol was initially administered twice a week for 2 weeks (4 administrations in total). Treatment was then decreased to once a week for a total of 4 weeks; alternate weeks for 8 weeks (4 administrations in total); once a month for several months (≥2 administrations); and, finally, discontinued. However, this standard protocol was increased or decreased in frequency, depending on the owner's request, travel restrictions, or the physical condition of the subject. Follow-up examinations to detect recurrence and metastasis were performed once monthly during the trial period.

Combination therapy

Photodynamic hyperthermal therapy (PHT) (16, 17), photodynamic hyperthermal CT (PHCT) (18, 19) and CT were performed, with lupeol administered as an adjuvant therapy. PHT is a local treatment combining photodynamic therapy and hyperthermia that uses indocyanine green as a photosensitizer (16, 17). PHCT is an advanced form of PHT, in combination with carboplatin CT (18, 19).

Results

Clinical outcome

The clinical outcomes are summarized in Table II. There were no severe adverse effects, such as local pain, diarrhea, or vomiting, in any of the subjects during lupeol treatment. Local tumor recurrence occurred in 1 case with stage I disease (C03) at 110 days and in 1 case with stage II disease (C04), which underwent partial surgical excision initially, at 170 days. Case C03 was immediately restarted on lupeol therapy alone every 2 weeks. After the additional treatment, tumor progression was arrested in case C03; and the disease condition has remained stable in both cases. In case C03, the tumor decreased in size and the owner reported that the dog's quality of life had improved.

Survival

At the end of the experimental period, all the dogs remained alive. Of the 12 dogs, 10 had survived for >180 days after surgery. Moreover, no distant metastasis was observed during the experimental period.

Discussion

COMM is an extremely malignant tumor, with a high propensity for local invasion and metastasis (1, 2). The treatment options for COMM include surgery, radiation therapy, CT and immunotherapy comprising xenogeneic DNA vaccination. MacEwen et al (20) reported that the median survival time (ST) in dogs with oral malignant melanoma that underwent complete surgical excision was ~17–18 months, 5–6 months and 3 months for stage I, II and III disease, respectively. There may be a potential to prolong ST beyond these reported durations, as all the subjects remained alive during the most recent examination. In stage III cases, the median ST increased in subjects administered adjuvant lupeol treatment (>6 vs. 3 months). The reported median ST for dogs receiving radiotherapy ranges between 5.3 and 11.9 months (35, 7, 9); however, as the radiotherapy equipment, treatment protocols and supplementary therapies differ between studies, it is difficult to directly compare previously reported results. Regardless, the results of previous reports suggest that systemic lupeol administration may help prolong the median ST in COMM cases and may prove efficacious as an adjuvant postoperative treatment.

Distant metastasis was not observed in any of the cases, including the 3 stage III cases, which were at highest risk for metastasis. Furthermore, there was no local recurrence for 180 days postoperatively in 7 of the 11 dogs. The median disease-free time (DFT) of dogs undergoing radiation therapy reportedly ranges between 5 and 7.9 months (5, 20). In a study evaluating radiation therapy in 38 dogs with COMM, Theon et al (21) reported a median DFT of 11.3 months in stage T1 cases, 6.0 months in stage T2 cases and 6.7 months in stage T3 cases. The results of the present study suggest that surgery combined with lupeol administration achieves local tumor control equivalent to that of radiation therapy. Saleem et al (13) reported that lupeol inhibits the growth of highly aggressive human metastatic melanoma cells in vitro and in vivo by inducing apoptosis. The results of the present study suggest that, in addition to preventing local progression, systemic postoperative adjuvant lupeol administration may also prevent the development of distant tumor metastasis.

For the local treatment of COMM, aggressive complete surgical excision, such as partial mandibulectomy and maxillectomy is the most effective option (5). Radiation therapy also plays an important role and is an effective treatment for achieving local tumor control; however, radiation therapy is associated with several adverse effects, including dermal desquamation, alopecia and bone necrosis, which may negatively affect the quality of life (7). The subjects must be anesthetized during radiation therapy, which itself is accompanied by serious risks. In addition, therapy is only available at selected facilities and may not be affordable for all owners; therefore, radiation therapy cannot be easily performed in all cases. CT combined with radiation therapy and surgery has been used as an alternative systemic therapy to prevent local recurrence and distant metastasis. Although several reports have evaluated the efficacy of CT in COMM as a sole or adjuvant therapy, there is no clear evidence that CT decreases the risk of local recurrence or metastasis, or that it prolongs median ST (59). These limitations highlight the need to develop an effective systemic therapy for COMM.

PHT, PHCT, melphalan and piroxicam were administered in combination, but there is no evidence that these treatments are effective in controlling COMM progression. PHT and PHCT were performed with owner consent after explaining the experimental nature of these treatments. Proulx et al (5) reported that melphalan administered for COMM exerted no beneficial effects. There is limited evidence that non-steroidal anti-inflammatory drugs (NSAIDs) may exert a protective effect against malignant melanoma development in humans (22); however, in a study evaluating NSAID therapy in dogs, there was no significant difference in survival attributed to NSAID administration and NSAID therapy did not provide any survival benefit or achieved an improved therapeutic response in the treated subjects (7). Therefore, these adjuvant therapies were unlikely to confer any beneficial treatment effect in this study.

No severe adverse effects, such as local pain, were observed in any of the dogs during lupeol treatment. There are no known reports of adverse effects associated with lupeol administration, which indicates that lupeol is safe when administered at the dosage recommended in the present study. Notably, this study was only a limited clinical trial and evaluated lupeol as a postoperative adjuvant therapy. Prospective controlled studies are required to assess the clinical efficacy of subcutaneous lupeol administration for the treatment of COMM, as well as in combination with other modalities, such as radiotherapy and CT.

In conclusion, to the best of our knowledge, this study is the first clinical trial to evaluate systemic lupeol therapy for COMM. Postoperative lupeol is easily administered, is not associated with severe adverse effects and was shown to prevent local tumor progression and distant metastasis. Therefore, the results of the present study suggest that lupeol is a potential novel adjuvant treatment option and may be used as systemic therapy for COMM.

References

1 

Smith SH, Goldschmidt MH and McManus PM: A comparative review of melanocytic neoplasms. Vet Pathol. 39:651–678. 2002. View Article : Google Scholar : PubMed/NCBI

2 

Bergman PJ: Canine oral melanoma. Clin Tech Small Anim Pract. 22:55–60. 2007. View Article : Google Scholar

3 

Bateman KE, Catton PA, Pennock PW and Kruth SA: 0-7-21 radiation therapy for the treatment of canine oral melanoma. J Vet Intern Med. 8:267–272. 1994. View Article : Google Scholar : PubMed/NCBI

4 

Blackwood L and Dobson JM: Radiotherapy of oral malignant melanomas in dogs. J Am Vet Med Assoc. 209:98–102. 1996.PubMed/NCBI

5 

Proulx DR, Ruslander DM, Dodge RK, Hauck ML, Williams LE, Horn B, Price GS and Thrall DE: A retrospective analysis of 140 dogs with oral melanoma treated with external beam radiation. Vet Radiol Ultrasound. 44:352–359. 2003. View Article : Google Scholar : PubMed/NCBI

6 

Rassnick KM, Ruslander DM, Cotter SM, Al-Sarraf R, Bruyette DS, Gamblin RM, Meleo KA and Moore AS: Use of carboplatin for treatment of dogs with malignant melanoma: 27 cases (1989–2000). J Am Vet Med Assoc. 218:1444–1448. 1996.PubMed/NCBI

7 

Murphy S, Hayes AM, Blackwood L, Maglennon G, Pattinson H and Sparkes AH: Oral malignant melanoma - the effect of coarse fractionation radiotherapy alone or with adjuvant carboplatin therapy. Vet Comp Oncol. 3:222–229. 2005. View Article : Google Scholar

8 

Dank G, Rassnick KM, Sokolovsky Y, Garrett LD, Post GS, Kitchell BE, Sellon RK, Kleiter M, Northrup N and Segev G: Use of adjuvant carboplatin for treatment of dogs with oral malignant melanoma following surgical excision. Vet Comp Oncol. 12:78–84. 2014. View Article : Google Scholar : PubMed/NCBI

9 

Freeman KP, Hahn KA, Harris FD and King GK: Treatment of dogs with oral melanoma by hypofractionated radiation therapy and platinum-based chemotherapy (1987–1997). J Vet Intern Med. 17:96–101. 2003.PubMed/NCBI

10 

Saleem M, Kaur S, Kweon MH, Adhami VM, Afaq F and Mukhtar H: Lupeol, a fruit and vegetable based triterpene, induces apoptotic death of human pancreatic adenocarcinoma cells via inhibition of Ras signaling pathway. Carcinogenesis. 26:1956–1964. 2005. View Article : Google Scholar

11 

Chaturvedi PK, Bhui K and Shukla Y: Lupeol: connotations for chemoprevention. Cancer Lett. 263:1–13. 2008. View Article : Google Scholar : PubMed/NCBI

12 

Saleem M: Lupeol, a novel anti-inflammatory and anti-cancer dietary triterpene. Cancer Lett. 285:109–115. 2009. View Article : Google Scholar : PubMed/NCBI

13 

Saleem M, Maddodi N, Abu Zaid M, Khan N, bin Hafeez B, Asim M, Suh Y, Yun JM, Setaluri V and Mukhtar H: Lupeol inhibits growth of highly aggressive human metastatic melanoma cells in vitro and in vivo by inducing apoptosis. Clin Cancer Res. 14:2119–2127. 2008. View Article : Google Scholar : PubMed/NCBI

14 

Nitta M, Azuma K, Hata K, Takahashi S, Ogiwara K, Tsuka T, Imagawa T, Yokoe I, Osaki T, Minami S and Okamoto Y: Systemic and local injections of lupeol inhibit tumor growth in a melanoma-bearing mouse model. Biomed Rep. 1:641–645. 2013.PubMed/NCBI

15 

Hata K, Hori K, Mukaiyama T, Sakamoto K and Takahashi S: Activators of melanin biosynthesis from Lactuca indica. Nat Med. 57:238–241. 2003.

16 

Radzi R, Osaki T, Tsuka T, Imagawa T, Minami S, Nakayama Y and Okamoto Y: Photodynamic hyperthermal therapy with indocyanine green (ICG) induces apoptosis and cell cycle arrest in B16F10 murine melanoma cells. J Vet Med Sci. 74:545–551. 2012. View Article : Google Scholar : PubMed/NCBI

17 

Radzi R, Osaki T, Tsuka T, Imagawa T, Minami S and Okamoto Y: Morphological study in B16F10 murine melanoma cells after photodynamic hyperthermal therapy with indocyanine green (ICG). J Vet Med Sci. 74:465–472. 2012. View Article : Google Scholar : PubMed/NCBI

18 

Onoyama M, Tsuka T, Imagawa T, Osaki T, Minami S, Azuma K, Kawashima K, Ishi H, Takayama T, Ogawa N and Okamoto Y: Photodynamic hyperthermal chemotherapy with indocyanine green: a novel cancer therapy for 16 cases of malignant soft tissue sarcoma. J Vet Sci. 15:117–123. 2014. View Article : Google Scholar : PubMed/NCBI

19 

Onoyama M, Azuma K, Tsuka T, Imagawa T, Osaki T, Minami S, Ogawa N and Okamoto Y: Effects of photodynamic hyperthermal therapy with indocyanine green on tumor growth in a colon 26 tumor-bearing mouse model. Oncol Lett. 7:1147–1150. 2014.PubMed/NCBI

20 

MacEwen EG, Patnaik AK, Harvey HJ, et al: Canine oral melanoma: comparison of surgery versus surgery plus Corynebacterium parvum. Cancer Invest. 4:397–402. 1986. View Article : Google Scholar : PubMed/NCBI

21 

Theon AP, Rodriguez C and Madewell BR: Analysis of prognostic factors and patterns of failure in dogs with malignant oral tumors treated with megavoltage irradiation. J Am Vet Med Assoc. 210:778–784. 1997.PubMed/NCBI

22 

Boria PA, Murry DJ, Bennett PF, Glickman NW, Snyder PW, Merke BL, Schlittler DL, Mutsaers AJ, Thomas RM and Knapp DW: Evaluation of cisplatin combined with piroxicam for the treatment of oral malignant melanoma and oral squamous cell carcinoma in dogs. J Am Vet Med Assoc. 224:388–394. 2004. View Article : Google Scholar : PubMed/NCBI

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APA
Yokoe, I., Azuma, K., Hata, K., Mukaiyama, T., Goto, T., Tsuka, T. ... Okamoto, Y. (2015). Clinical systemic lupeol administration for canine oral malignant melanoma. Molecular and Clinical Oncology, 3, 89-92. https://doi.org/10.3892/mco.2014.450
MLA
Yokoe, I., Azuma, K., Hata, K., Mukaiyama, T., Goto, T., Tsuka, T., Imagawa, T., Itoh, N., Murahata, Y., Osaki, T., Minami, S., Okamoto, Y."Clinical systemic lupeol administration for canine oral malignant melanoma". Molecular and Clinical Oncology 3.1 (2015): 89-92.
Chicago
Yokoe, I., Azuma, K., Hata, K., Mukaiyama, T., Goto, T., Tsuka, T., Imagawa, T., Itoh, N., Murahata, Y., Osaki, T., Minami, S., Okamoto, Y."Clinical systemic lupeol administration for canine oral malignant melanoma". Molecular and Clinical Oncology 3, no. 1 (2015): 89-92. https://doi.org/10.3892/mco.2014.450