Viral Oncolysate Research Articles

A Novel Approach to Cervical Cancer Therapy: A Short Review

Cervical cancer is one of the most common cancers affecting women worldwide, and it occurs following persistent infection. Cervical cancer incidence is tightly linked to HPV infection (human papillomavirus), and particularly, type 16 and type 18 viruses cause the majority of cases. The common therapies for cervical cancer include surgery, chemotherapy, and radiation therapy, which are often invasive or unbearable treatment methods with many side effects. They just probably slow down the disease progression or alleviate any comorbid conditions, including vaginal bleeding and pain, which is called palliative care, while novel treatment approaches, especially virus-like particles and viral oncolysate, could eliminate these complications. In this review study, we have proposed a novel approach to cervical cancer therapy focused on utilizing Newcastle disease virus as viral oncolysate with a high potential of immunity induction and low side effects. Furthermore, we have attempted to shed some light on the perspectives of novel virus-based cervical cancer immunotherapy. Finally, we review the recent findings from basic and clinical studies and also discuss the usefulness and limitations of this approach, as well as the reasons why it is believed that viral oncolysate immunotherapy may be of relevance in the treatment of human cervical cancer.

Antitumor immunity enhancement through Newcastle viral oncolysate in mice model: A promising method to treat tumors.

A Newcastle disease virus (NDV) oncolysate has been established as a unique and effective immune-stimulatory root for tumor treatment. Thus, the aim of the current study was to investigate the effects of intratumoral administration of NDV oncolysate on immune response and tumor regression of C57BL/6 mouse model of human papillomavirus (HPV) related transplanted with TC-1 syngeneic cancer cells. To further investigate the mechanism underlying the antitumor response, cytolytic and lymphocyte proliferation responses in splenocytes were measured using lactate dehydrogenase (LDH) release and MTT assays, respectively. In this regard, levels of IL-10, IFN-γ, and IL-4 were measured using ELISA after re-stimulation. The immune responses efficacy was evaluated by in vivo tumor regression assay. The results showed that immunization with the different titers of NDV lysate significantly reduced tumor volume in comparison with a combination of virus lysate and tumor cell lysate. Also, virus lysate could significantly enhance cytotoxic T lymphocyte production and lymphocyte proliferation rates versus tumor cell lysate. Also, our major findings are that the peritumorally injection of NDV oncolysate effectively induces antitumor immune responses through increased levels of IL-4, IFN-γ, and reduction of IL-10. These results indicate that this treatment is a specific, active immune mechanism stimulator, and may prove to be a useful therapeutic for a treatment against cervical cancers and merits further investigation.

The First Human Immunotherapy Trial with a Viral Oncolysate Vaccine Induced a Large Number of Granular Lymphocytes Later Recognized as 'Natural Killer Cells'

The First Human Immunotherapy Trial with a Viral Oncolysate Vaccine Induced a Large Number of Granular Lymphocytes Later Recognized as 'Natural Killer Cells'

Oncolytic Newcastle Disease Virus as Cutting Edge between Tumor and Host.

Oncolytic viruses (OVs) replicate selectively in tumor cells and exert anti-tumor cytotoxic activity. Among them, Newcastle Disease Virus (NDV), a bird RNA virus of the paramyxovirus family, appears outstanding. Its anti-tumor effect is based on: (i) oncolytic activity and (ii) immunostimulation. Together these activities facilitate the induction of post-oncolytic adaptive immunity. We will present milestones during the last 60 years of clinical evaluation of this virus. Two main strategies of clinical application were followed using the virus (i) as a virotherapeutic agent, which is applied systemically or (ii) as an immunostimulatory agent combined with tumor cells for vaccination of cancer patients. More recently, a third strategy evolved. It combines the strategies (i) and (ii) and includes also dendritic cells (DCs). The first step involves systemic application of NDV to condition the patient. The second step involves intradermal application of a special DC vaccine pulsed with viral oncolysate. This strategy, called NDV/DC, combines anti-cancer activity (oncolytic virotherapy) and immune-stimulatory properties (oncolytic immunotherapy) with the high potential of DCs (DC therapy) to prime naive T cells. The aim of such treatment is to first prepare the cancer-bearing host for immunocompetence and then to instruct the patient’s immune system with information about tumor-associated antigens (TAAs) of its own tumor together with danger signals derived from virus infection. This multimodal concept should optimize the generation of strong polyclonal T cell reactivity targeted against the patient’s TAAs and lead to the establishment of a long-lasting memory T cell repertoire.

Abstract 1569: Multivalent melanoma vaccine using primary human melanoma cells and a vaccinia virus based antigen retrieval system

Abstract Melanoma is the fifth most diagnosed cancer in men and seventh most diagnosed cancer in women and it is estimated that there will be over 70,000 new cases and 8,000 deaths in 2011. Prevention of both primary and secondary melanomas requires the development of a melanoma specific multi-valent vaccine. We have been involved in developing a vaccinia virus based vaccine consisting of melanoma cell membrane antigens and integrated vaccinia viral proteins, which serve as a potent adjuvant. This melanoma vaccine, referred to as viral membrane oncolysate (VMO), is composed of five primary cell lines derived from patient tumors each independently infected with vaccinia virus and has been shown to induce anti-cancer immune response in clinical trials. Notable results were obtained in node positive subsets of patients that showed a positive response in survival distribution function suggesting that this preparation is an active therapeutic immunogen but the mechanism of immune activation needs to be elucidated. In order for us to define the immunogens that impart the clinical efficacy of the this melanoma VMO vaccine, it is essential to characterize the primary cell lines with respect to their phenotypic characteristics as well as to define the melanoma antigen repertoire so that effective vaccine strategies can be designed that do not rely exclusively on tumor derived primary cells. Our study reveals that all the five primary cell lines expressed antigens, such as gp100, CD146, and tyrosinase, similar to several commercially available melanoma cell lines although at varying concentrations. In addition, the five primary cell lines demonstrated varying migratory/invasive properties using a Boyden Chamber assay and their tumorigenic potential is being tested in nude mice. The cells were also characterized with respect to BRAF mutations. We observed several similarities between the antigenic profile of the primary and established cell lines as well as some unique characteristics. We conclude that using a combination of defined antigens from primary tumor cells is useful in designing a pan-antigen immunotherapeutic vaccine. This melanoma pan antigen vaccine can be an effective tool in primary prevention of melanoma presumably having the highest clinical efficacy when combined with unique antigens from primary cells. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1569. doi:1538-7445.AM2012-1569

Natural and genetically engineered viral agents for oncolysis and gene therapy of human cancers

Based on personal acquaintances and experience dating back to the early 1950s, the senior author reviews the history of viral therapy of cancer. He points out the difficulties encountered in the treatment of human cancers, as opposed by the highly successful viral therapy of experimentally maintained tumors in laboratory animals, especially that of ascites carcinomas in mice. A detailed account of viral therapy of human tumors with naturally oncolytic viruses follows, emphasizing the first clinical trials with viral oncolysates. The discrepancy between the high success rates, culminating in cures, in the treatment of tumors of laboratory animals, and the moderate results, such as stabilizations of disease, partial responses, very rare complete remissions, and frequent relapses with virally treated human tumors is recognized. The preclinical laboratory testing against established human tumor cell lines that were maintained in tissue cultures for decades, and against human tumors extricated from their natural habitat and grown in xenografts, may not yield valid results predictive of the viral therapy applied against human tumors growing in their natural environment, the human host. Since the recent discovery of the oncosuppressive efficacy of bacteriophages, the colon could be regarded as the battlefield, where incipient tumor cells and bacteriophages vie for dominance. The inner environment of the colon will be the teaching ground providing new knowledge on the value of the anti-tumor efficacy of phage-induced innate anti-tumor immune reactions. Genetically engineered oncolytic viruses are reviewed next. The molecular biology of viral oncolysis is explained in details. Elaborate efforts are presented to elucidate how gene product proteins of oncolytic viruses switch off the oncogenic cascades of cancer cells. The facts strongly support the conclusion that viral therapy of human cancers will remain in the front lines of modern cancer therapeutics. It may be a combination of naturally oncolytic viruses and wild-type viruses rendered oncolytic and harmless by genetic engineering, that will induce complete remissions of human tumors. It may be necessary to co-administer certain chemotherapeutic agents, advanced cancer vaccines, or even immune lymphocytes, and targeted therapeuticals, to ascertain, that remissions induced by the viral agents will remain complete and durable; will co-operate with anti-tumor host immune reactions, and eventually will result in cures of advanced metastatic human cancers.

Dendritic cells pulsed with viral oncolysates potently stimulate autologous T cells from cancer patients.

We demonstrated before that primary operated breast cancer patients contain in their bone marrow (BM) cancer reactive memory T cells (MTC) which have to be re-activated to become tumor infiltrating effector cells. The aim of this study was to optimize an ex vivo stimulation protocol for MTC based on autologous dendritic cells (DC). As source of tumor antigens we used lysates from unmodified tumor cells or from tumor cells infected with Newcastle Disease Virus (NDV) which contain IFN-alpha inducing viral dsRNA as one danger signal. DC from breast cancer patients were pulsed with lysates from the MCF-7 breast cancer line (Tu-L) or from NDV infected MCF-7 cells (TuN-L, viral oncolysates) and compared for stimulatory capacity in an ELISPOT response of autologous BM derived MTC. To analyze potential further danger signals derived from NDV infection, we employed MALDI mass spectrometry, Western blots, FACS cytometry and ELISA tests. DC pulsed with viral oncolysates showed increased expression of co-stimulatory molecules in comparison to Tu-L pulsed DC and induced significantly higher ELISPOT MTC responses. Supernatants from co-cultures of MTC and TuN-L pulsed DC contained increased titers of IFN-alpha and IL-15. NDV infection of tumor cells resulted in a number of differences in protein expression including a heat-shock protein (HSP27) which became phosphorylated. The results suggest that a DC preparation pulsed with viral oncolysate includes danger signals (e.g. dsRNA, cytokines, HSP molecules) and is superior for MTC stimulation to a DC preparation pulsed with lysate from non-infected tumor cells.

Tumor vaccines: a role in preventing recurrence in melanoma?

Patients presenting with thick primary melanomas or those with regional nodal metastases have a high risk of recurrence after surgery alone. Chemotherapy has limited efficacy in the adjuvant setting, and while the use of high-dose interferon in the adjuvant setting has been reported to improve survival, treatment with interferon is not without significant cost and toxicity. Mounting evidence suggests a prominent role for the immune system in the natural history of melanoma, and the clinical success of interferon highlights the potential for immunotherapy to prevent recurrence. Many researchers hope to use melanoma vaccines to reduce recurrence without significant toxicity, and many different vaccine strategies are under investigation. Peptide vaccines attempt to induce immunity to melanoma MHC-restricted peptide antigens by delivering the peptide to the patient along with an immune adjuvant meant to induce inflammation and stimulate immunity. While peptide vaccines have advantages with regard to cost and feasibility, it is still unclear whether highly purified peptides will stimulate an adequate immune response. An alternative approach is the use of cellular vaccines. Autologous cellular vaccines present all biologically relevant antigens to the immune system, but this is limited to individuals with sufficient tumor to prepare a vaccine. Allogeneic cellular vaccines are based on the fact that melanoma-associated antigens are shared among a large number of patients, so a vaccine prepared from a cultured cell line could stimulate an anti-tumor immune response in many patients. Allogeneic vaccines are available for all patients, and can be standardized, preserved and distributed in a manner akin to any other therapeutic agent. Because of this, they are more readily available for evaluation in large trials, and there are two major allogeneic vaccines presently being evaluated as an adjuvant therapy for melanoma. Several additional approaches to vaccine therapies are being investigated including among others ganglioside vaccines, viral oncolysates, cytokine gene-modified tumor cell vaccines, dendritic cell vaccines, anti-idiotype antibodies and DNA vaccines. While there appears to be tremendous potential for vaccines, it must be remembered that there has been significant interest in immunotherapy for melanoma for over 50 years and, to date, no large prospective, randomized trial has shown a survival benefit.

Newcastle disease virus infection induces B7-1/B7-2-independent T-cell costimulatory activity in human melanoma cells.

Viral oncolysates of Newcastle disease virus (NDV) have been widely used for the treatment of malignant melanoma. Apparently, this nononcogenic and apathogenic paramyxovirus can alter the immunogenicity of tumor cells. To determine the influence of NDV infection on a tumor-specific T-cell response on a functional level, we used autologous primary melanoma cells infected with the NDV-strain Ulster. Therefore, melanoma cells and tumor-infiltrating lymphocytes were prepared from a freshly resected tumor, and tumor-infiltrating lymphocytes were subjected to limited dilution cloning. Proliferation assays of the T-helper cell clone sTS3 (CD4+) showed that the T-cell clone was rendered nonreactive against its autologous major histocompatibility complex II+, B7-1/B7-2- melanoma SMS, even remaining unresponsive to subsequent stimulation by interleukin-2. NDV infection of the SMS melanoma cell line not only completely restored the proliferative response of sTS3 to SMS, comparable with stimulation by cross-linking of anti-CD3/anti-CD28 monoclonal antibodies, but also inhibited the induction of anergy. Electrophoretic mobility shift assays of sTS3 cell lysates revealed the induction of the CD28-responsive complex by coincubation with NDV-infected melanoma cells. Because the induction of this complex of nuclear proteins shows specificity for the activation of the CD28 pathway but functional B7-1/B7-2 expression was not detectable on SMS melanoma cells at any timepoint, we propose the induction of a costimulatory factor different from B7 by NDV viral proteins.

Vaccination against human cancers (review).

Classical and molecular immunological means of active tumor-specific immunization against human cancers yielded whole cell or tumor cell lysate vaccines of preventive value (reduced relapse rates) and dendritic cell-peptide or genetically engineered vaccines that may induce remissions even in metastatic disease. Active tumor-specific immunization was often successful in the past 50 years against experimental tumors maintained in the laboratory. During the epochs of classical and molecular immunology several vaccines were generated and used for the reduction of relapse rates of human cancer after surgical removal of the primary or metastatic tumors. Whole cell vaccines consist of X-irradiated autologous or allogeneic tumor cells coadministered with immunostimulants (BCG, Detox). Tumor cells haptenized biologically (as in viral oncolysates) or chemically were also used. Dendritic cell vaccines are prepared by transfection or transduction with tumor antigen-encoding DNA or by pulsing the cells with antigenic peptides in vitro; or collecting dendritic cells that engulfed apoptotic tumor cell DNA and/or peptide antigens in vivo for reinjection into the patient. Genetically engineered tumor cells are prepared in vitro to express MHC and peptides, costimulatory molecules (B7.1) and cyto- or lymphokines (interferons, interleukins, hematopoietic growth factors) for vaccination of patients. Antibody- and immune T cell-mediated immune reactions to autologous tumor cells are newly generated and/or quantitatively increased in immunized patients but do not always correlate with clinical response. Most vaccines are claimed to have reduced relapse rates presumably by inducing effective host immunity against micrometastases. Dendritic cell-peptide vaccines could induce partial or occasionally complete remissions in metastatic disease. The wrong antigenic presentation may result in tolerance induction toward the tumor; occasionally tumor enhancement may occur. Human tumor antigens when presented appropriately (with costimulatory molecules and with IL-2, IL-12) break the host's natural tolerance toward its tumor and induce rejection strength immune reactions even in patients with metastatic disease. Immune T cells thus generated could be collected for adoptive immunotherapy. For successful active specific immunization against human cancers the understanding of the immunoevasive maneuvers of the tumor cell (through FasL --> Fas; TRAIL; CD40L --> CD40; TGFbeta etc. systems) is essential.

Cancer vaccines with emphasis on a viral oncolysate melanoma vaccine.

Biotherapy of malignant diseases has become the fourth treatment modality besides surgery, chemo- and radiotherapy. Whole cell melanoma vaccines with or without BCG and other adjuvants, purified ganglioside and shed antigens, recombinant viruses carrying tumor antigens, dendritic cells pulsed with antigenic peptides etc. are in clinical trials. Efficacious viral oncolysate vaccines induce the host to mount tumor-specific cytotoxic T-cell response and prevention of relapses is supported by clinical trials. The use of "polyvalent" whole cell vaccines vs. purified or genetically engineered single antigen vaccines is justified as i. only very few single tumor antigens are present in all tumors of a given histological type; and ii. antigen modulation occurs in tumors rendering them resistant to immune attack generated by vaccine against a single antigen. Thus polyvalent vaccines immunize against several antigens vs. against a selected antigen.

Indium-111 labeled leukocytes in evaluation of active specific immunotherapy responses.

Active immunization of patients utilizing viral oncolysates (VO) has been studied in clinical trials. VO are extracts of cultured tumor cells that have been infected with certain types of viruses, particularly surface budding varieties. The objectives of the current studies were to examine the trafficking patterns of Indium (In)-111-labeled leukocytes or lymphocytes in two groups of patients with gynecologic malignancies to determine whether these cells migrate to sites of active immunization after VO. Eight patients with ovarian cancer received VO intraperitoneally followed by In-111-labeled leukocytes or lymphocytes (500 &mgr;Ci) intravenously. In a separate trial, three patients with cervical cancer received In-111-labeled lymphocytes after they had been treated with VO administered by the intralymphatic route. Gamma camera imaging was performed to evaluate the distribution patterns of the labeled cells at several time intervals after injection. Results indicate that metastatic tumor sites exposed to VO therapy show significant uptake of In-111 cells. These sites of malignancies were confirmed by computerized tomography and ultrasound scans. In patients with ovarian cancer no uptake of the radiolabeled cells was observed in metastatic tumors of the liver and lymph nodes. In patients with cervical cancer, lymph node metastases exposed to intralymphatic VO therapy were visualized very well. Other known tumor sites not exposed to VO therapy showed no uptake of radioactivity. These findings confirm that VO induces immune responses. This diagnostic nuclear medicine technique may prove to be a useful method for following-up responses to immunotherapy.

Can virus therapy of human cancer be improved by apoptosis induction?

Direct virus inoculations and viral oncolysates may induce temporary remissions and prolong life with reduced tumor burden, or decrease relapse rates but fall short of curing human cancers. We propose: (i) investigations of Cassel's 73-T, an Ehrlich's mouse ascites carcinoma-adapted Newcastle disease virus (NDV) strain that so effectively reduced relapse rates in malignant melanoma if it is an admixture or a recombinant with a murine parvovirus; (ii) transfection of prostatic carcinoma cells with the TRMP gene; (iii) transfection of sarcoma cells with the fas gene followed by treatment with anti-fas monoclonal antibodies, and (iv) treatment of metastatic tumors with a parvovirus incorporating the apoptosis-inducer Ad5 E1A gene. Thus, replicating virions and haphazard generation of cytokines in the inoculated host could be replaced with transfection of single genes of well-defined, limited but selected efficacy.

Active intralymphatic immunotherapy of uterine cervical carcinoma with viral oncolysate: a pilot study.

A pilot clinical trial was conducted in patients with squamous carcinoma of the uterine cervix to evaluate the clinical and biologic effects of active intralymphatic immunotherapy (AILI) with allogenic viral oncolysate (VO) prior to radiation therapy. Sixteen patients with advanced primary squamous carcinoma of the uterine cervix and lymph node metastases underwent bipedal intralymphatic injections of VO. VO was derived from lysates of cervical carcinoma cells that had been infected with influenza A virus. AILI was repeated after 2 weeks and followed one week later by standard or extended-field radiation therapy (RT). The first seven patients were treated at one of the three dose levels: 6 mg (three patients), 12 mg (three patients) and 18 mg (one patient). Remaining patients were treated at the 12 mg dose level. Sixteen patients received 63 injections (one patient received three of four doses) of AILI-VO without significant toxicity. Eleven patients have died of persistent or recurrent carcinoma with a total median survival of 19.4 months. Examination of humoral and cellular immunity during AILI-VO showed an increase in the serum liters of antibodies to a surface antigen on cervical carcinoma cells and to the influenza virus. Increased non-MHC restricted lymphocyte cytotoxicity was exhibited by three of four patients treated above the first dose level. Two of the three patients are survivors. By contrast, lymphocytes of patients treated with AILI-VO exhibited either an increase or a decrease in proliferation responses to cervical carcinoma cells. Similarly, post-treatment lymphocytes exhibited either helper or suppressor inducer effects on pre-treatment lymphocytes.

Prospects for gene therapy and lymphokine therapy for metastatic melanoma.

Conventional treatment of cancer, especially for patients with metastatic melanoma tumor, is often ineffective. Immunotherapy and recently introduced gene therapy have revolutionized the treatments of patients with metastatic melanoma tumor. Use of biological response modifiers, such as interleukins and interferons, have been found to enhance therapeutic benefits to patients with malignant melanoma. Initial studies with a high-dose interleukin-2 (IL-2) therapy have proved effective in patients with melanoma tumor, although a variety of systemic toxicities were observed. A low-dose IL-2 continuous infusion has shown a similar response in patients with melanoma tumor, but produced lesser toxicity. The low-dose IL-2 therapy has been studied with an adoptive transfer combined with either autologous lymphokine activated killer cells or autologous tumor infiltrating lymphocytes (TIL). IL-2 in combination with chemotherapeutic agents such as flavone acetic acid, dacarbazine, and cyclophosphamide have also been studied in patients with metastatic melanoma. Results have shown a moderate response in patients with metastatic melanoma. TIL therapy, however, has been shown to result in higher objective regression due to potent tumor-specific killing and tumor-specific targeting characters of the TIL. The tumor targeting nature of the TIL creates the possibility of using TIL as a vehicle to deliver gene product specifically to tumor tissue. Safety and toxicity of gene-transduced TIL were addressed by the use of neomycin-resistant, gene-transduced TIL in patients with metastatic melanoma. We also investigated the use of vaccinia oncolysate therapy by using the viral oncolysate prepared with IL-2 gene encoded vaccinia virus. Preliminary studies with murine hepatic metastases colon model have shown encouraging results.

Active specific adjuvant immunotherapy with vaccinia melanoma oncolysate.

Vaccinia melanoma oncolysate is an investigative agent for active specific adjuvant immunotherapy. Viral oncolysate work in animals has led to application of this treatment in humans. Success of initial trials in treatment of melanoma has led to a phase III randomized, prospective, double-blind, multi-institutional trial, which is currently under way.

Experimental trials of immunotherapy for malignant melanoma

Conventional treatment for metastatic melanoma consists of surgical resection, chemotherapy, and radiation therapy. New approaches toward treatment of this disease include the development of passive and active immunotherapeutic regimens. Malignant melanoma is particularly amendable to immunotherapy since the tumor is relatively immunogenic, expressing unique cell surface protein and lipid antigens. Clinical trials investigating the benefit of active specific immunotherapy documented increased survival of invasive Stage 1 and metastatic Stage 2 melanoma patients following immunization with tumor cell vaccines and BCG. Additional trials showed that the development of specific antibodies after immunization of Stage 2 patients with a viral oncolysate was correlated with an increased survival compared to matched controls given only BCG. Passive immunotherapy approaches using either lymphokine-activated killer (LAK) cells or tumor-infiltrating lymphocytes (TIL) administered with interleukin 2 have also resulted in regression of disease with complete or partial remissions occurring in 25% of the patients. Additional studies have focused on the generation of specific cytotoxic T lymphocytes by stimulation with autologous tumor in vivo. Future trials will evaluate the therapeutic efficacy of these specific cytologic T lymphocytes relative to LAK and TIL.

T Cell Responses to Ovarian Tumor Vaccines: Identification and Significance for Future Immunotherapy

The cellular immune responses to ovarian cancer patients treated with viral oncolysates (VO) ovarian tumor vaccines to vaccines are described. CD3+ cells proliferated after stimulation with the tumor vaccines in a dose-dependent manner. The proliferation of CD3+ cells stimulated with the tumor vaccine was blocked by anti-HLA-DR monoclonal antibody and anti-CD4 mAb indicating that CD3+ CD4+ cells from the blood of the patients treated with VO recognize tumor derived determinants in conjunction with MHC class II antigens. The regulatory activity of the T cells collected after VO treatment was assayed by co-cultivation with PBMC collected before VO treatment. These cells demonstrated increased helper activity for immunoglobin production by cells collected before vaccination and secreted IL-2 in response to stimulation by vaccine. Finally, when biochemical fractionation of the components of VO was attempted, PBMC from VO treated patients responded by proliferation to several fractions suggesting that they recognize multiple epitopes in the ovarian tumor vaccine. Therefore, these data provide novel evidence for the involvement of the T cells in response to ovarian tumor vaccines.

Viral oncolysates as human tumor vaccines.

Postoncolytic immunity entails immune reactions acquired through an oncolytic virus infection or through repeated immunizations with viral oncolysates (or virally modified tumor cell membranes) that are valid and operational also against virally not modified tumor cells of the same type. NK cells react to budding virions, induce target cell lysis primarily but not exclusively by the production of granzymes and pore-forming proteins and operate without direction from memory cells. In contrast, immune T cells (including some TIL) are MHC-restricted, act under the direction of memory cells and lyse target cells primarily but not exclusively by the release of lymphotoxin (TNF beta) causing programmed cell death (apoptosis) through endonuclease activation and target cell DNA fragmentation. This author proposes that it is not NK, but the immune T cells that mediate postoncolytic immunity. Oncogene amplification may protect immortalized tumor cells even when expressing peptide antigens through MHC molecules against lymphotoxin-mediated apoptosis; but virally-infected tumor cells releasing budding virions remain susceptible to NK cells. Highly immunogenic viral oncolysates should present both budding virions for NK cells and processed viral and tumoral peptide antigens co-jointly for immune T cells.

Active specific immunotherapy with vaccinia colon oncolysate enhances the immunomodulatory and antitumor effects of interleukin-2 and interferon alpha in a murine hepatic metastasis model.

The role of cytokines as primary or adjuvant antineoplastic agents has been well established. Interleukin-2 (IL-2) and the interferons have, particularly, proven to be effective antitumor agents when given alone, and seem to act synergistically on the eradication of metastases from immunogenic tumors. Active specific immunotherapy, in the form of viral oncolysates, has also shown effectiveness in cancer therapy. Bearing this in mind, we decided to combine these agents in an adjuvant triple regimen and compare their effectiveness to other treatments in terms of tumor burden and survival in a murine colon cancer hepatic metastases model. BALB/c mice were injected with CC-36, a weakly immunogenic murine colon adenocarcinoma, intrasplenically, to produce artificial liver metastases. The animals were divided into one control group and seven treatment groups receiving either vaccinia colon oncolysate (VCO), IL-2, interferon-alpha (IFN alpha) alone, or combinations of these agents. Half the animals were followed for survival and the other half were sacrificed at the end of the experiment for quantification of tumor burden. The blood of the sacrificed animals was utilized in a series of immunological tests in order to demonstrate the cytolytic potential of the peripheral blood lymphocytes (PBL) in each treatment group, as well as to characterize phenotypically the cells acting as effectors. The triple-adjuvant regimen group was by far the most effective treatment group, demonstrating 100% survival and a significant reduction in tumor burden when compared to other groups. Furthermore, the PBL from the animals in this group showed 69.4% lysis of the CC-36 target cells in vitro. These effector lymphocytes were characterized as ASMG1-/Lyt2.2+ cytolytic lymphocytes. We conclude that these lymphocytes were stimulated by the administration of VCO and further augmented by the immunomodulation of the cytokines given in the triple regimen, and that such a regimen might prove beneficial in the treatment of established hepatic metastases from weakly immunogenic tumors.