Melanoma is one of the fastest-growing cancers and has the highest mortality rate of skin cancers 1–3. More than half of melanoma patients experience progression of the disease within 3 years of diagnosis 4,5. Current clinical practice guidelines for stage I–II melanoma provide few, if any, recommendations for treatment 6–9. The oncolytic property of viruses has been observed for over a century and is presently being studied intensively 10–16. An oncolytic, nonpathogenic ECHO-7 virus, adapted and selected for melanoma that has not been genetically modified (Rigvir), was approved and registered in 2004 in Latvia for melanoma therapy 17–27. The effect of viruses on cancers, including melanoma, has been tested in clinical trials; however, the effectiveness of an approved and marketed virus has not yet been shown in a clinical setting 12,14,16.
In oncolytic virotherapy, Rigvir is a first-in-class. At a later time, a genetically modified adenovirus was approved for head and neck cancer 28,29. Melanoma is staged with substages from 0 to IV by measuring the thickness of the tumour according to Breslow, by assessment of ulceration, mitotic rate and metastases and by collecting pathologic information on regional lymph nodes 30,31. On the basis of the stage of the disease, treatment is currently performed according to published guidelines 4,6–9.
The aim of the present study is to test the effectiveness of Rigvir in a retrospective study in substage IB, IIA, IIB and IIC melanoma patients on time to progression and overall survival.
Retrospective clinical study patients
White patients (N=79) who had undergone surgical excision of melanoma and diagnosis verified histologically during the 4 years between January 2008 and December 2011 were included in this study. All patients were free of disease after surgery and were classified into substages IB, IIA, IIB and IIC according to the American Joint Committee on Cancer 30,31. For disease progression, all were followed for a minimum of 3 months until January 2014. The overall survival was checked on 5 June 2014 and considered to reflect the status by 27 May 2014. The detailed study population characteristics of this retrospective study are shown in Table 1.
Current guidelines for melanoma advise no treatment postsurgery for patients who are classified into substages IB and IIA. Patients in substages IIB and IIC are provided three options: participation in a clinical trial, observation and interferon 7,8. In the absence of strict guidelines, treatment with Rigvir was offered. Thus, 52 study participants received Rigvir and 27 were observed according to the guidelines. The patients who had been treated with interferon were excluded from the present analysis as, in the registry, they were too few to allow for any comparison.
As a part of the safety assessment, serum clinical chemistry parameters were recorded.
The patients in this study were treated in the Latvian Oncology Center of Riga Eastern Clinical University Hospital, the Latvian Virotherapy Center in Riga and the Oncology Clinic of Piejūras Hospital in Liepāja, Latvia.
The study was approved by the respective ethics committee.
Rigvir is a 2 ml frozen solution of an adapted and selected ECHO-7 virus strain, Picornaviridae family, Enterovirus genus, Enteric Cytopathic Human Orphan (ECHO) type 7, group IV, positive-sense single-stranded RNA virus produced under GMP. The titre is not less than 106 TCID50/ml in sodium chloride for injection.
Method of Rigvir administration
Treatment was started after surgical excision of the primary melanoma tumour when the wound had healed. First, Rigvir (2 ml) was administered intramuscularly regionally for 3 consecutive days. After about 4 weeks, administration was repeated for three consecutive days and repeated about 4 weeks later. Subsequently, a single administration of Rigvir (2 ml, intramuscularly) was performed at monthly intervals during the first year, at 6-week intervals during the first half of the second year, at 2-month intervals during the second half of the second year and at 3-month intervals in the third year. Rigvir is not to be used during an acute infection.
Statistical analysis of the data was carried out using the SPSS statistical software, V.20 (SPSS Inc., Chicago, Illinois, USA). Mann–Whitney U-test and Wilcoxon tests (for continuous variables), Fisher’s exact test and the χ2-test (for categorical variables) were used to test differences between and within groups. Cox proportional hazard survival regression analysis was carried out, which is the most commonly used multivariate model in survival analysis. Thus, any difference between the groups, for example, in age, has been taken into account in the Cox analysis. (This is in contrast to Kaplan–Meier analysis, which is a bivariate analysis that only takes into account one predictor at a time). Hazard ratios (HRs) and 95% confidence intervals were calculated using bivariate and multivariate Cox regression analysis on survival. Endpoints were occurrence of metastases or disease recurrence for time to progression, and death from any cause for analysis of overall survival. Predictors (covariates) used in regression analysis were tumour stages, treatment (Rigvir, observation), sex and age. A P value less than 0.05 from a two-sided test was established to indicate statistical significance.
Effectiveness in patients: time to progression
Melanoma patients of substages IB, IIA, IIB and IIC were studied according to the postsurgery management that they had received. One group was treated with Rigvir and the other was managed according to current guidelines by observation (the control group is called ‘observation’) 6–9. The follow-up period was not statistically different between both treatment groups (Table 1).
Patients who were free of melanoma after surgical excision and were treated with Rigvir appeared to remain disease free (free of metastases and/or recurrence) for a longer period of time compared with a similar group of patients who did not receive Rigvir. The difference between the treatment groups did not, however, reach statistical significance (Table 2).
Effectiveness in patients: overall survival
The survival of patients who were treated with Rigvir was significantly (P<0.05) longer compared with a similar group of patients who did not receive Rigvir (Fig. 1 and Table 3). The difference between both treatment groups was statistically significant on analysing all four substages together (IB, IIA, IIB, IIC) (Table 3) and on analysing stage II together (substages IIA, IIB, IIC). Adjusting for patient age, sex and substage of disease, the HR was calculated in multivariate Cox regression analysis. The HR for patients treated according to current guidelines by observation versus treated with Rigvir was 6.27 (P<0.005) for all patients, 4.39 (P<0.032) for substage IIA–IIC patients and 6.57 (P<0.014) for substage IIB–IIC patients (Fig. 1). This indicates that the patients who were treated with Rigvir had a 4.39–6.57-fold lower mortality than those treated using current guidelines by observation.
In the previous clinical studies, a few side effects were reported, for example subfebrile temperature (37.5°C for a couple of days), pain in the tumour area, sleepiness and diarrhoea. In this retrospective study, however, there was no record of any untoward side effect from Rigvir treatment or its discontinuation.
Serum clinical chemistry parameters were recorded and graded according to NCI CTCAE 32 (Table 4). In the observation group, grade 1–3 values were obtained. All grade 3 samples were from two patients obtained within the last few months of life. In one of these patients, progression of the disease was reported simultaneously. In contrast, in the Rigvir-treated patients, values above grade 2 were not observed.
Oncolytic virotherapy is one of three forms of virotherapy (the other two being viral vectors for gene therapy and viral immunotherapy, respectively). Early observations of tumour regressions after virus infections have been published starting from the late 19th century (cf. 10–16). Recently, several oncolytic viruses have been tested clinically 33–35 and Science named cancer immunotherapy the breakthrough of the year of 2013 36. The melanoma adapted and selected ECHO-7 virus Rigvir is first-in-class in oncolytic virotherapy; it is approved as therapy for melanoma.
The present results show that in substage IB, IIA, IIB and IIC melanoma patients, Rigvir administration after surgery significantly (P<0.05) prolongs survival compared with patients who were managed according to current published guidelines 6–9. For the Rigvir-treated patients, the HR (risk of death) is 4.39–6.57-fold lower than for the control group treated according to current guidelines by observation. The HR was calculated in multivariate Cox regression analysis adjusting for patient age, sex and substage of disease.
In this study, there was no record of any untoward side effect from Rigvir treatment, which is in agreement with clinical studies using other oncolytic viruses 14,16,33,34,37. Moreover, no value higher than grade 2 was recorded in Rigvir-treated patients. This is in contrast to most other cancer therapies, where grades 3 and 4 are frequently observed (cf. 38).
Administration of virus induces the formation of neutralising antibodies that might potentially influence the efficiency of Rigvir. In previous studies, the titre of neutralising antibodies against ECHO-7 was determined in both healthy individuals and patients before administration of Rigvir. In 94 healthy adult participants tested, the titres were found to be low (1 : 20 to 1 : 62) 39,40. When tested in 155 adult cancer patients who had not been treated with Rigvir, neutralising antibodies against ECHO-7 were detected in ∼50% of the patients 41. In a local study of 472 individuals, the presence of ECHO-7 antibodies was shown to increase with age in children and level off to a plateau of around 75% in adults 42. To our knowledge, the prevalence of neutralising antibodies against the ECHO-7 virus in the general adult population has not been reported.
Rigvir is an immunomodulator that affects both the humoral, antibody-mediated, and the cellular immune systems 20–22. When virus adsorption and penetration to tumour tissue were measured, it was shown that they are not influenced by the presence of neutralising antibodies (titre 1 : 10) 43,44. Furthermore, in a preliminary study, the levels of neutralising antibodies to Rigvir during the first 18 months of treatment of melanoma patients did not appear to correlate with time to progression after 3 years of follow-up 40. In that study, the neutralising antibody titre was 1 : 10 before the start of treatment (N=34). After the first dose, the titre was 1 : 25 to 1 : 91 (determined 24–48 h after administration). A month later, before the second dose, the titre was 1 : 250 to 1 : 320 (N=30); after the second dose, it was 1 : 510 to 1 : 850. Two months later, before the third administration, the titre was 1 : 160 to 1 : 895 (N=26) and after the eighth dose, 18 months after the first dose, it was 1 : 280 to 1 : 1350 40.
Also, after intravenous administration, the correlation between antibody titres varies from one virus to another, and neutralising antibodies do not affect efficacy when local or regional administration is used 14,45,46.
An estimated 14.1 million new cancer cases were diagnosed worldwide in 2012, the latest available. The number is expected to increase to 24 million by 2035. About 232 000 patients are estimated to be diagnosed with melanoma in 2014 3. In the 20-year survival data analysis of the American Joint Committee on Cancer (cf. Figure 31.1 of 31), the majority of all melanoma patients belonged to stage I and stage II, 47 and 24%, respectively 31. However, at present, clinical practice guidelines suggest postsurgery therapy only for late-stage melanoma (radiation therapy and interferon α) 6–9.
Rigvir has also been used in other types of cancer. In vitro, it reduces the viability of melanoma, as well as pulmonary, gastric, pancreatic, bone, and breast cancer cell cultures 47,48. It is oncolytic in melanoma and rectum cancer patients 49,50 (26, p. 115) and has been shown to improve the 5-year survival in rectum cancer patients 24.
Taken together, the results suggest that a significant number of melanoma patients would benefit from prolonging the survival with Rigvir treatment. The results also show that this can be achieved without side effects. Results suggest that Rigvir could also be tested in the treatment of other types of cancer.
Rigvir is an oncolytic, nonpathogenic ECHO-7 virus that significantly prolongs survival in early-stage melanoma patients without any side effect.
The authors are indebted to Anna Krilova, Oncology Clinic of Piejūras Hospital, Liepāja, Latvia, for sharing patient information, and Oksana Holodņuka, Riga Eastern Clinical University Hospital, and Linda Brokāne, Latvian Virotherapy Center, for technical assistance, and Vaira Saulīte, Institute of Microbiology and Virology, Riga Stradiņš University, for expert advice.
Conflicts of interest
Aina Muceniece, Dite Venskus, Jurgis Auziņš and Pēteris Alberts are past and present employees of the Latvian Virotherapy Center. For the remaining authors there are no conflicts of interest.
1. Siegel R, Ma J, Zou Z, Jemal A. Cancer statistics, 2014. CA Cancer J Clin 2014; 64:9–29.
2. World Health Organization (WHO). Skin cancers. Available at: http://www.who.int/uv/faq/skincancer/en/index1.html
. [Accessed 8 January 2015].
3. World Health Organization. GLOBOCAN 2012: estimated cancer incidence, mortality and prevalence worldwide in 2012. Available at: http://globocan.iarc.fr/Pages/fact_sheets_population.aspx
. [Accessed 10 January 2015].
4. Marsden JR, Newton-Bishop JA, Burrows L, Cook M, Corrie PG, Cox NH, et al.. Revised U.K. guidelines for the management of cutaneous melanoma
2010. Br J Dermatol 2010; 163:238–256.
5. Tas F. Metastatic behavior in melanoma
: timing, pattern, survival, and influencing factors. J Oncol 2012; 2012:647684.
6. Garbe C, Peris K, Hauschild A, Saiag P, Middleton M, Spatz A, et al.. Diagnosis and treatment of melanoma
. European consensus-based interdisciplinary guideline – update 2012. Eur J Cancer 2012; 48:2375–2390.
7. Dummer R, Hauschild A, Guggenheim M, Keilholz U, Pentheroudakis G. Cutaneous melanoma
: European Society for Medical Oncology (ESMO) Clinical Practice Guidelines for diagnosis, treatment and follow-up. Ann Oncol 2012; 23 (Suppl 7):86–91.
8. National Comprehensive Cancer Network (NCCN). Clinical practice guidelines in oncology. Melanoma
9. National Cancer Institute (NCI). Melanoma
. Treatment option overview. Available at: http://www.cancer.gov/cancertopics/pdq/treatment/melanoma/HealthProfessional/page1/AllPages#4
. [Accessed 8 January 2015].
10. Muceniece A. Oncotropism of viruses and the problem of virotherapy
of malignant tumours (in Russian). Riga: Zinatne; 1972. 1–443.
11. Ferdats A. Cancer virus hunting (in Latvian). Riga: Zinatne; 1977. 1–108.
12. Mullen JT, Tanabe KK. Viral oncolysis. Oncologist 2002; 7:106–119.
13. Kelly E, Russell SJ. History of oncolytic
viruses: genesis to genetic engineering. Mol Ther 2007; 15:651–659.
14. Liu TC, Galanis E, Kirn D. Clinical trial results with oncolytic virotherapy
: a century of promise, a decade of progress. Nat Clin Pract Oncol 2007; 4:101–117.
15. Bartlett DL, Liu Z, Sathaiah M, Ravindranathan R, Guo Z, He Y, Guo ZS. Oncolytic
viruses as therapeutic cancer vaccines. Mol Cancer 2013; 12:103.
16. Lichty BD, Breitbach CJ, Stojdl DF, Bell JC. Going viral with cancer immunotherapy
. Nat Rev Cancer 2014; 14:559–567.
17. Muceniece A, Ferdats A. Cancer virotherapy
. Virology (in Latvian). Riga: Zvaigzne; 1985. 187.
18. Grigalinovich G, Rudzitis M, Skudra M, Popena B, Desjatnikova I, Garklava R. Effect of a viral immunomodulator
(Rigvir®) on the morphology and survival of cutaneous melanoma
patients (in Russian). Proc Latv Acad Sci 1988; 497:72–75.
19. Heisele O, Glinkina L, Muceniece A, Garklava R. The effect of a viral immunomodulator
, Rigvir, on the parameters of humoral immunity in malignant skin melanoma
patients. Proc Latv Acad Sci 1991; 533:64–67.
20. Glinkina LS, Heisele OG, Garklava RR, Muceniece AJ. The humoral immunity indices of patients with malignant skin melanoma
using the viral immunomodulator
Rigvir. Vopr Onkol 1992; 38:534–540.
21. Glinkina LS, Bruvere RZ, Venskus DR, Garklava RR, Muceniece AJ. The cellular immunity indices of patients with malignant melanoma
using the viral immunomodulator
Rigvir. Vopr Onkol 1992; 38:540–547.
22. Glinkina LS, Bruvere RZ. The reaction of the T-immunity system in patients with malignant skin melanoma
and stomach cancer to active nonspecific immunotherapy
. Vopr Onkol 1992; 38:659–666.
23. Muceniece A. Rigvir – development of a viral immunomodulator
and cancer virotherapy
clinical trials (in Latvian). Riga: The 4th Latvian Congress of Physicians; 2001. 126–127.
24. Bruvere R, Heisele O, Ferdats A, Rupais A, Muceniece A. Echovirus-mediated biotherapy for malignant tumours: 40 years of investigation. Acta Med Litu 2002; 9 (Suppl 9):97–100.
25. Bruvere R, Feldmane G, Ferdats A, Heisele O, Muceniece A. Adjuvant immunotheraphy with virus-mediated biomodulators developed in Latvia: experimental and clinical data. Abstracts of the Perspectives in Melanoma
X and The Third Annual International Melanoma
Research Congress 14–16 September, Noordwijk, The Netherlands. Melanoma
Res 2006; 16 (Suppl 1):S33–S34.
26. Muceniece A, Venskus D. How to assess immunity – the melanoma
model (in Latvian). Riga: Ainas Mucenieces society for cancer immunotherapy
; 2007. 1–199.
27. Chumakov PM, Morozova VV, Babkin IV, Baikov IK, Netesov SV, Tikunova NV. Oncolytic
enteroviruses. Mol Biol (Mosk) 2012; 46:639–650.
28. Garber K. China approves world’s first oncolytic
virus therapy for cancer treatment. J Natl Cancer Inst 2006; 98:298–300.
29. Frew SE, Sammut SM, Shore AF, Ramjist JK, Al-Bader S, Rezaie R, et al.. Chinese health biotech and the billion-patient market. Nat Biotechnol 2008; 26:37–53.
30. Balch CM, Gershenwald JE, Soong SJ, Thompson JF, Atkins MB, Byrd DR, et al.. Final version of 2009 AJCC melanoma
staging and classification. J Clin Oncol 2009; 27:6199–6206.
31. Edge SB, Byrd DR, Compton CC, Fritz AG, Greene FL, Trotti A. AJCC cancer staging manual, 7th ed. New York, NY: Springer; 2010. 1–648.
32. U.S. Department of Health and Human Services, National Institutes of Health, National Cancer Institute. Common terminology criteria for adverse events (CTCAE) V4. Bethesda, MD: National Institutes of Health; 2010. 1–79.
33. Bourke MG, Salwa S, Harrington KJ, Kucharczyk MJ, Forde PF, de Kruijf M, et al.. The emerging role of viruses in the treatment of solid tumours. Cancer Treat Rev 2011; 37:618–632.
34. Russell SJ, Peng KW, Bell JC. Oncolytic virotherapy
. Nat Biotechnol 2012; 30:658–670.
35. Vacchelli E, Eggermont A, Sautes-Fridman C, Galon J, Zitvogel L, Kroemer G, Galluzzi L. Trial watch: oncolytic
viruses for cancer therapy. Oncoimmunology 2013; 2:e24612.
36. Couzin-Frankel J. Breakthrough of the year 2013. Cancer immunotherapy
. Science 2013; 342:1432–1433.
37. Kaufman HL, Andtbacka RHI, Collichio FA, Amatruda T, Senzer NN, Chesney J, et al.. Primary overall survival (OS) from OPTiM, a randomized phase III trial of talimogene laherparepvec (T-VEC) versus subcutaneous (SC) granulocyte-macrophage colony-stimulating factor (GM-CSF) for the treatment (tx) of unresected stage IIIB/C and IV melanoma
. J Clin Oncol 2014; 32:5s.
38. Hauschild A, Gogas H, Tarhini A, Middleton MR, Testori A, Dreno B, Kirkwood JM. Practical guidelines for the management of interferon-alpha-2b side effects in patients receiving adjuvant treatment for melanoma
: expert opinion. Cancer 2008; 112:982–994.
39. Heisele O. The effect of a viral immunomodulator
(Rigvir®) on the immune reactivity of patients with skin malignant melanoma
[Thesis] (Riga). 1987; 1–166 (in Russian).
40. Glinkina L. Effect of Rigvir® on systemic and local manifestations of immunity in patients with melanoma
and gastric cancer [Thesis] (Riga). 1993; 1–171 (in Russian).
41. Volrate A, Stefanovich HL. Sturis T, Muceniece A, Aleksandrova M, Černobajeva I, Volrate A. Study of humoral immunity to enteroviruses in cancer patients (in Russian). Viral oncotropism (in Russian). Riga: Zinatne; 1969. 215–219.
42. Henigst WW, Gelfand HM, Leblanc DR, Fox JP. ECHO virus type 7 infections in a continuously observed population group in Southern Louisiana. Am J Trop Med Hyg 1961; 10:759–766.
43. Garklava R. Determination of oncotropism of enteroviruses in human tumours by adsorption [Thesis] (Riga). 1968; 1-246 (in Russian).
44. Garklava R. Sturis T, Muceniece A, Aleksandrova M, Černobajeva I, Volrate A. The adsorption of some enteroviruses in the tissues of gastric cancer and the human breast (in Russian). Viral oncotropism (in Russian). Riga: Zinatne; 1969. 41–52.
45. Melcher A, Parato K, Rooney CM, Bell JC. Thunder and lightning: immunotherapy
viruses collide. Mol Ther 2011; 19:1008–1016.
46. Hwang TH, Moon A, Burke J, Ribas A, Stephenson J, Breitbach CJ, et al.. A mechanistic proof-of-concept clinical trial with JX-594, a targeted multi-mechanistic oncolytic
poxvirus, in patients with metastatic melanoma
. Mol Ther 2011; 19:1913–1922.
47. Grigalinovich G, Petrovska R. Morphological changes of human osteosarcoma cells in cell culture caused by a viral immunomodulator
(Rigvir®) (in Russian). Proc Latv Acad Sci 1988; 497:69–71.
48. Golubs G, Veinalde R, Petrovska R, Bruvere R, Pjanova D. Oncolytic
activity of Rigvir® in various cell lines (Abstract). Riga, Latvia: 12th Joint symposium Riga-Rostock; 2014. 13–14.
49. Garklava R, Bruvere R, Vitolina L, Priedite I, Muceniece A. Muceniece A, Augstkalne M, Volrate A, Bruvere R, Ferdats A, Heisele O. Morphological and clinical parallels of changing rectal cancer during combined treatment. Immunological aspects of viral oncotropism (in Russian). Riga: Zinatne; 1979. 114–120.
50. Bruvere R, Vitolina L, Garklava R, Priedite I, Muceniece A. Influence of a viral immunomodulator
(Rigvir®) on cellular composition and topographic characteristics of the infiltration of the stroma of primary tumours of colorectal cancer (in Russian). Proc Latv Acad Sci 1980; 396:137–142.