Coronaviruses are positive-strand RNA viruses with features attractive for oncolytic therapy. To investigate this potential, we redirected the coronavirus murine hepatitis virus (MHV), which is normally unable to infect human cells, to human tumor cells by using a soluble receptor (soR)-based expression construct fused to an epidermal growth factor (EGF) receptor targeting moiety. Addition of this adapter protein to MHV allowed infection of otherwise nonsusceptible, EGF receptor (EGFR)-expressing cell cultures. We introduced the sequence encoding the adaptor protein soR-EGF into the MHV genome to generate a self-targeted virus capable of multiround infection. The resulting recombinant MHV was viable and had indeed acquired the ability to infect all glioblastoma cell lines tested in vitro. Infection of malignant human glioblastoma U87ΔEGFR cells gave rise to release of progeny virus and efficient cell killing in vitro. To investigate the oncolytic capacity of the virus in vivo, we used an orthotopic U87ΔEGFR xenograft mouse model. Treatment of mice bearing a lethal intracranial U87ΔEGFR tumor by injection with MHVsoR-EGF significantly prolonged survival compared to phosphate-buffered saline-treated (P = 0.001) and control virus-treated (P = 0.004) animals, and no recurrent tumor load was observed. However, some adverse effects were seen in normal mouse brain tissues that were likely caused by the natural murine tropism of MHV. This is the first demonstration of oncolytic activity of a coronavirus in vivo. It suggests that nonhuman coronaviruses may be attractive new therapeutic agents against human tumors.
Oncolytic viruses aim to specifically kill tumor cells. A major challenge is the effective targeting of disseminated tumors in vivo. We retargeted herpes simplex virus (HSV) tropism to HER-2 oncoprotein p185, overexpressed in ovary and breast cancers. The HER-2-retargeted R-LM249 exclusively infects and kills tumor cells expressing high levels of human HER-2. Here, we assessed the efficacy of systemically i.p. delivered R-LM249 against disseminated tumors in mouse models that recapitulate tumor spread to the peritoneum in women. The human ovarian carcinoma SK-OV-3 cells implanted intraperitoneally (i.p.) in immunodeficient Rag2−/−;Il2rg−/− mice gave rise to a progressive peritoneal carcinomatosis which mimics the fatal condition in advanced human patients. I.p. administration of R-LM249 strongly inhibited carcinomatosis, resulting in 60% of mice free from peritoneal diffusion, and 95% reduction in the total weight of neoplastic nodules. Intraperitoneal metastases are a common outcome in breast cancer: i.p. administration of R-LM249 strongly inhibited the growth of ovarian metastases of HER-2+ MDA-MB-453 breast cells. Brain metastases were also reduced. Cumulatively, upon i.p. administration the HER-2-redirected oncolytic HSV effectively reduced the growth of ovarian and breast carcinoma disseminated to the peritoneal cavity.
In the genome of human herpes simplex virus (HSV) we have replaced part of the sequences encoding the receptor-binding glycoprotein with antibody fragments directed against the oncoprotein HER-2, overexpressed in human breast and ovarian cancers. The retargeted HSV only infects and kills human cancer cells expressing high levels of HER-2. Earlier experiments showed that the retargeted HSV injected inside localized tumors inhibits human tumor growth in immunodeficient mice. As tumor dissemination is the major cause of cancer mortality, we have now used the HER-2-retargeted HSV to treat disseminated HER-2+ ovarian and breast cancer in immunodeficient mice. Intraperitoneal treatments significantly inhibited the peritoneal spread (carcinomatosis and ascites formation) of ovarian cancer cells and the metastatic growth of breast cancer cells in the ovaries. Brain metastases were also inhibited. Our results showed that a HER-2-redirected oncolytic HSV is an effective therapeutic agent against metastatic HER-2+ cancers and, more generally, provide the first demonstration of the efficacy of a systemically-administered, retargeted oncolytic HSV.
Cancer gene therapy consists of numerous approaches where the common denominator is utilization of vectors for achieving therapeutic effect. A particularly potent embodiment of the approach is virotherapy, in which the replication potential of an oncolytic virus is directed towards tumor cells to cause lysis, while normal cells are spared. Importantly, the therapeutic effect of the initial viral load is amplified through viral replication cycles and production of progeny virions. All cancer gene therapy approaches rely on a sufficient level of delivery of the anticancer agent into target cells. Thus, enhancement of delivery to target cells, and reduction of delivery to non-target cells, in an approach called transductional targeting, is attractive. Both genetic and non-genetic retargeting strategies have been utilized. However, in the context of oncolytic viruses, it is beneficial to have the specific modification included in progeny virions and hence genetic modification may be preferable. Serotype chimerism utilizes serotype specific differences in receptor usage, liver tropism and seroprevalence in order to gain enhanced infection of target tissue. This review will focus on serotype chimeric adenoviruses for cancer gene therapy applications.
adenoviruses; serotype chimerism; neutralizing antibodies; tumor targeting; liver detargeting
Gene targeting to tumors using adenoviral vectors holds great potential for cancer imaging and therapy, but the limited efficacy of current methods used to improve delivery to target tissues and reduce unwanted interactions remain substantial barriers to further development. Progress in characterizing the set of molecular interactions used by adenoviral vectors to infect particular tissues has aided the development of novel strategies for retargeting vectors to tumor cells. One method is chemical retargeting of adenovirus using bispecific antibodies against both viral capsid proteins and tumor-specific cell surface molecules. This approach can be combined either with competitive inhibitors designed to reduce viral tropism in undesired tissues, or with traditional therapeutics to increase the expression of surface molecules for improved tumor targeting. Ablating liver cell-specific interactions through mutation of capsid proteins or chemical means are promising strategies for reducing adenovirus-induced liver toxicity. The nature of tumor neovasculature also influences adenoviral delivery, and the use of vascular disrupting agents such as combretastatin can help elucidate these contributions. In this investigation, we evaluate a variety of these methods for retargeting adenoviral vectors to tumor cells in vitro and in vivo, and assess the contributions of specific molecular and tissue interactions that affect adenoviral transgene delivery.
adenovirus; Bavituximab; bispecific antibody; fiber; targeting
Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality worldwide, and novel treatment modalities to improve the prognosis of patients with advanced disease are highly desirable. Oncolytic virotherapy is a promising approach for the treatment of advanced NSCLC. MicroRNAs (miRNAs) may be a factor in the regulation of tumor-specific viral replication. The purpose of this study was to investigate whether miRNA-145 regulated oncolytic herpes simplex virus-1 (HSV-1) can selectively kill NSCLC cells with reduced collateral damage to normal cells.
We incorporated 4 copies of miRNA-145 target sequences into the 3′-untranslated region of an HSV-1 essential viral gene, ICP27, to create AP27i145 amplicon viruses and tested their target specificity and toxicity on normal cells and lung cancer cells in vitro.
miRNA-145 expression in normal cells was higher than that in NSCLC cells. AP27i145 replication was inversely correlated with the expression of miRNA-145 in infected cells. This oncolytic HSV-1 selectively reduced cell proliferation and prevented the colony formation of NSCLC cells. The combination of radiotherapy and AP27i145 infection was significantly more potent in killing cancer cells than each therapy alone.
miRNA-145-regulated oncolytic HSV-1 is a promising agent for the treatment of NSCLC.
Oncolytic virus; MicroRNA; Lung cancer; Radiotherapy
Envelope virus replication begins with receptor binding, followed by fusion of the viral envelope with the cell membrane. The binding and fusion steps are usually mediated by envelope proteins. The ability of envelope proteins of a particular virus to bind and fuse with target cells defines the host range of the virus, known as “viral tropism”. The mechanism(s) of fusion by the viral envelope is largely categorized as either pH-dependent or –independent. By redirecting the binding specificities of envelope proteins to desired target molecules while maintaining fusion activity, it is possible to redirect the tropisms of virus and viral vectors, enabling specific killing and/or transduction of desired cells in vivo. Recently, a lipid, phosphatidylserine, was also shown to mediate binding of virus, which affects the tropisms of viruses and viral vectors.
Oncolytic viruses are being developed as anticancer drugs. They propagate selectively in tumor tissue and destroy it without causing excessive damage to normal non-cancerous tissues. When used as drugs, they must meet stringent criteria for safety and efficacy and be amenable to pharmacological study in human subjects. Specificity for neoplastic tissue is the key to safety, and this goal can be achieved through a variety of ingenious virus-engineering strategies. Antiviral immunity remains a significant barrier to the clinical efficacy of oncolytic viruses but this is being addressed by using novel immune-evasive delivery strategies and immunosuppressive drugs. Noninvasive pharmacokinetic monitoring is facilitated by engineering marker genes into the viral genome. Clinical data on the pharmacokinetics of oncolytic viruses will be the key to accelerating their development and approval as effective anticancer drugs. This review introduces concepts relevant to the use of viruses as anticancer drugs, emphasizing targeting mechanisms as well as safety and efficacy issues that are currently limiting their clinical success.
A novel frontier in the treatment of tumors that are difficult to treat is oncolytic virotherapy, in which a replication-competent virus selectively infects and destroys tumor cells. Herpes simplex virus (HSV) represents a particularly attractive system. Effective retargeting to tumor-specific receptors has been achieved by insertion in gD of heterologous ligands. Previously, our laboratory generated an HSV retargeted to human epidermal growth factor receptor 2 (HER2), a receptor overexpressed in about one-third of mammary tumors and in some ovarian tumors. HER2 overexpression correlates with increased metastaticity and poor prognosis. Because HER2 has no natural ligand, the inserted ligand was a single-chain antibody to HER2. The objective of this work was to genetically engineer an HSV that selectively targets the HER2-expressing tumor cells and that has lost the ability to enter cells through the natural gD receptors, HVEM and nectin1. Detargeting from nectin1 was attempted by two different strategies, point mutations and insertion of the single-chain antibody at a site in gD different from previously described sites of insertion. We report that point mutations at gD amino acids 34, 215, 222, and 223 failed to generate a nectin1-detargeted HSV. An HSV simultaneously detargeted from nectin1 and HVEM and retargeted to HER2 was successfully engineered by moving the site of single-chain antibody insertion at residue 39, i.e., in front of the nectin1-interacting surface and not lateral to it, and by deleting amino acid residues 6 to 38. The resulting recombinant, R-LM113, entered cells and spread from cell to cell solely via HER2.
Oncolytic virotherapy is emerging as a promising approach for the treatment of several neoplasms. The term “oncolytic viruses” is generally employed to indicate naturally occurring or genetically engineered attenuated viral particles that cause the demise of malignant cells while sparing their non-transformed counterparts. From a conceptual standpoint, oncolytic viruses differ from so-called “oncotropic viruses” in that only the former are able to kill cancer cells, even though both display a preferential tropism for malignant tissues. Of note, such a specificity can originate at several different steps of the viral cycle, including the entry of virions (transductional specificity) as well as their intracellular survival and replication (post-transcriptional and transcriptional specificity). During the past two decades, a large array of replication-competent and replication-incompetent oncolytic viruses has been developed and engineered to express gene products that would specifically promote the death of infected (cancer) cells. However, contrarily to long-standing beliefs, the antineoplastic activity of oncolytic viruses is not a mere consequence of the cytopathic effect, i.e., the lethal outcome of an intense, productive viral infection, but rather involves the elicitation of an antitumor immune response. In line with this notion, oncolytic viruses genetically modified to drive the local production of immunostimulatory cytokines exert more robust therapeutic effects than their non-engineered counterparts. Moreover, the efficacy of oncolytic virotherapy is significantly improved by some extent of initial immunosuppression (facilitating viral replication and spread) followed by the administration of immunostimulatory molecules (boosting antitumor immune responses). In this Trial Watch, we will discuss the results of recent clinical trials that have evaluated/are evaluating the safety and antineoplastic potential of oncolytic virotherapy.
GM-CSF; HSV; immunotherapy; JX594; reolysin; talimogene laherparepvec
Murine hepatitis coronavirus (MHV)-A59 infection depends on the interaction of its spike (S) protein with the cellular receptor mCEACAM1a present on murine cells. Human cells lack this receptor and are therefore not susceptible to MHV. Specific alleviation of the tropism barrier by redirecting MHV to a tumor-specific receptor could lead to a virus with appealing properties for tumor therapy. To demonstrate that MHV can be retargeted to a nonnative receptor on human cells, we produced bispecific adapter proteins composed of the N-terminal D1 domain of mCEACAM1a linked to a short targeting peptide, the six-amino-acid His tag. Preincubation of MHV with the adapter proteins and subsequent inoculation of human cells expressing an artificial His receptor resulted in infection of these otherwise nonsusceptible cells and led to subsequent production of progeny virus. To generate a self-targeted virus able to establish multiround infection of the target cells, we subsequently incorporated the gene encoding the bispecific adapter protein as an additional expression cassette into the MHV genome through targeted RNA recombination. When inoculated onto murine LR7 cells, the resulting recombinant virus indeed expressed the adapter protein. Furthermore, inoculation of human target cells with the virus resulted in a His receptor-specific infection that was multiround. Extensive cell-cell fusion and rapid cell killing of infected target cells was observed. Our results show that MHV can be genetically redirected via adapters composed of the S protein binding part of mCEACAM1a and a targeting peptide recognizing a nonnative receptor expressed on human cells, consequently leading to rapid cell death. The results provide interesting leads for further investigations of the use of coronaviruses as antitumor agents.
Current treatments for high-grade lymphoma often have curative potential, but unfortunately many cases relapse and develop therapeutic resistance. Thus, there remains a need for novel therapeutics that can target the residual cancer cells whose phenotypes are distinct from the bulk tumor and that are capable of reforming tumors from very few cells. Oncolytic viruses offer an approach to destroy tumors by multiple mechanisms, but they can not effectively reach residual disease or micrometastases, especially within the lymphatic system. To address these limitations, we have generated immune cells infected with oncolytic viruses as a therapeutic strategy that can combine effective cellular delivery with synergistic tumor killing. In this study, we tested this approach against minimal disease states of lymphomas characterized by the persistence of cancer cells which display stem cell-like properties and resistance to conventional therapies. We found that the immune cells were capable of trafficking to and targeting residual cancer cells. The combination biotherapy used prevented relapse by creating a long-term, disease-free state, with acquired immunity to the tumor functioning as an essential mediator of this effect. Immune components necessary for this acquired immunity were identified. We further demonstrated that the dual-biotherapy could be applied before or after conventional therapy. Our approach offers a potentially powerful new way to clear residual cancer cells, showing how restoring immune surveillance is critical for maintenance of a disease-free state.
oncolytic vaccinia; lymphoma; CIK cells; Immune cell therapy
Members of the rodent subgroup of the genus Parvovirus exhibit lytic replication and spread in many human tumor cells and are therefore attractive candidates for oncolytic virotherapy. However, the significant variation in tumor tropism observed for these viruses remains largely unexplained. We report here that LuIII kills BJ-ELR ‘stepwise-transformed’ human fibroblasts efficiently, while MVM does not. Using viral chimeras, we mapped this property to the LuIII capsid gene, VP2, which is necessary and sufficient to confer the killer phenotype on MVM. LuIII VP2 facilitates a post-entry, pre-DNA-amplification step early in the life cycle, suggesting the existence of an intracellular moiety whose efficient interaction with the incoming capsid shell is critical to infection. Thus targeting of human cancers of different tissue-type origins will require use of parvoviruses with capsids that effectively make this critical interaction.
LuIII; Minute Virus of Mice (MVM); parvovirus; oncolytic; oncolysis; virotherapy; tropism; VP2; qPCR; stepwise transformation
The prognosis of malignant brain tumors remains extremely bad in spite of moderate improvements of conventional treatments. A promising alternative approach is the use of oncolytic viruses. Strategies to improve viral toxicity include the combination of oncolytic viruses with standard therapies. Parvovirus H-1 (H-1PV) is an oncolytic virus with proven toxicity in glioma cells. Recently it has been demonstrated that the combination of ionizing radiation (IR) with H-1PV showed promising results. Previously irradiated glioma cells remained fully permissive for H-1PV induced cytotoxicity supporting the use of H-1PV for recurrent gliomas, which typically arise from irradiated cell clones. When glioma cells were infected with H-1PV shortly (24 h) after IR, cell killing improved and only the combination of both treatments lead to complete long-term tumor cell killing. The latter finding raises the question whether IR in combination with H-1PV exerts an additional therapeutic effect on highly resistant glioma stem cells. A likely translation into current clinical treatment protocols is to use stereotactic radiation of non-resectable recurrent gliomas followed by intratumoral injection of H-1PV to harvest the synergistic effects of combination treatment.
oncolytic virus; radiation therapy; malignant glioma; stem cells; parvovirus H-1; combination therapy
Oncolytic viruses consist of a diverse range of DNA and RNA viruses traditionally thought to mediate their effects by exploiting aberrations in tumor pathways, allowing preferential viral replication in, and killing of, tumor cells. Clinical development has progressed to late phase trials, potentially heralding their introduction into clinical practice. However, despite this promise, the activity of oncolytic viruses has yet to achieve the potential suggested in preclinical models. To address this disparity, we need to recognise the complex interaction between oncolytic viruses, tumor, chemotherapy, host immune system, and appreciate that direct oncolysis may not be the only factor to play an important role in oncolytic virus-mediated anti-tumor efficacy.
Although key in inactivating viruses, the host immune system can also act as an ally against tumors, interacting with oncolytic viruses under the right conditions to generate useful and long-lasting anti-tumor immunity.
Preclinical data also suggest that oncolytic viruses demonstrate synergy with standard therapies, which may offer improved clinical response rates. Here we explore clinical and preclinical data on clinically relevant oncolytic viruses, highlighting areas of progress, uncertainty and translational opportunity, with respect to immune recruitment and therapeutic synergy.
Oncolytic virus; oncolysis; anti-tumor immune response; chemotherapy; synergy
Cancer stem cells (CSC), also termed “cancer initiating cells” or “cancer progenitor cells”, which have the ability to self-renew, proliferate, and maintain the neoplastic clone, have recently been discovered in a wide variety of pediatric tumors. These CSC are thought to be responsible for tumorigenesis, tumor maintenance, aggressiveness and recurrence due to inherent resistance to current treatment modalities such as chemotherapy and radiation. Oncolytic virotherapy offers a novel, targeted approach for eradicating pediatric CSC by utilizing mechanisms of cell killing that differ from conventional therapies. Moreover, oncolytic viruses have the ability to target specific features of CSC such as cell surface proteins, transcription factors, and the CSC microenvironment. Through genetic engineering, a wide variety of foreign genes may be expressed by oncolytic viruses to augment the oncolytic effect. We review the current data regarding the ability of several types of oncolytic viruses (herpes simplex virus-1 (HSV-1), adenovirus, reovirus, Seneca Valley virus, vaccinia virus, Newcastle disease virus, myxoma virus, vesicular stomatitis virus) to target and kill both CSC and tumor cells in pediatric tumors. We highlight advantages and limitations of each virus and potential ways next-generation engineered viruses may target resilient CSC.
Oncolytic measles virus (MV) induces cell fusion and cytotoxicity in a CD46 dependent manner. Development of fully retargeted oncolytic MVs would improve tumor selectivity. The urokinase receptor (uPAR) is a tumor and stromal target overexpressed in multiple malignancies. MV-H glycoproteins fully retargeted to either human or murine uPAR were engineered and their fusogenic activity was determined. Recombinant human (MV-h-uPA) and murine (MV-m-uPA) uPAR retargeted MVs expressing eGFP were rescued and characterized. Viral expression of chimeric MV-H was demonstrated by RT-PCR and Western Blot. In vitro viral replication was comparable to MV-GFP control. Receptor and species specificity of MV-uPAs were demonstrated in human and murine cells with different levels of uPAR expression. Removal of the ATF ligand from MV-uPA -by Factor Xa treatment- ablated MV-uPA’s functional activity. Cytotoxicity was demonstrated in uPAR expressing human and murine cells. MV-h-uPA efficiently infected human endothelial cells and capillary tubes in vitro. Intravenous administration of MV-h-uPA delayed tumor growth and prolonged survival in the MDA-MB-231 breast cancer xenograft model. Viral tumor targeting was confirmed by immunohistochemistry. MV-m-uPA transduced murine mammary tumors (4T1) in vivo, after intratumor administration. MV-m-uPA targeted murine tumor vasculature after systemic administration, as demonstrated by dual (CD31 and MV-N) staining of tumor capillaries in the MDA-MB-231 model. In conclusion, MV-uPA is a novel oncolytic MV associated with potent and specific antitumor effects and tumor vascular targeting. This is the first retargeted oncolytic MV able to replicate in murine cells and target tumor vasculature in an uPAR dependent manner.
The treatment of patients with malignant brain tumors remains a major oncological problem. The median survival of patients with glioblastoma multiforme (GBM), the most malignant type, is only 15 months after initial diagnosis and even less after tumor recurrence. Improvements of standard treatment including surgery and radio-chemotherapy have not lead to major improvements. Therefore, alternative therapeutics such as oncolytic viruses that specifically target and destroy cancer cells are under investigation. Preclinical data of oncolytic parvovirus H-1 (H-1PV) infection of glioma cells demonstrated strong cytotoxic and oncosuppressing effects, leading to a phase I/IIa trial of H-1PV in patients with recurrent GBM (ParvOryx01). ParvOryx01 is the first trial with a replication competent oncolytic virus in Germany.
ParvOryx01 is an open, non-controlled, two groups, intra-group dose escalation, single center, phase I/IIa trial. 18 patients with recurrent GBM will be treated in 2 groups of 9 patients each. Treatment group 1 will first receive H-1PV by intratumoral injection and second by administration into the walls of the tumor cavity during tumor resection. In treatment group 2 the virus will initially be injected intravenously and afterwards, identical to group 1, into the surrounding brain tissue during tumor removal. Main eligibility criteria are: age of 18 years, unifocal recurrent GBM, amenable to complete or subtotal resection. Dose escalation will be based on the Continual Reassessment Method. The primary objective of the trial is local and systemic safety and tolerability and to determine the maximum tolerated dose (MTD). Secondary objectives are proof of concept (PoC) and Progression-free Survival (PFS) up to 6 months.
This is the first trial with H-1PV in patients with recurrent GBM. The risks for the participants appear well predictable and justified. Furthermore, ParvOryx01 will be the first assessment of combined intratumoral and intravenous application of an oncolytic virus. Due to its study design the trial will not only generate data on the local effect of H-1PV but it will also investigate the penetration of H-1PV into the tumor after systemic delivery and obtain safety data from systemic delivery possibly supporting clinical trials with H-1PV in other, non-CNS malignancies.
ClinicalTrials.gov Identifier: NCT01301430
Cancer is one of the major causes of death worldwide. In spite of achieving significant successes in medical sciences in the past few decades, the number of deaths due to cancer remains unchecked. The conventional chemotherapy and radiotherapy have limited therapeutic index and a plethora of treatment related side effects. This situation has provided an impetus for search of novel therapeutic strategies that can selectively destroy the tumour cells, leaving the normal cells unharmed. Viral oncotherapy is such a promising treatment modality that offers unique opportunity for tumour targeting. Numerous viruses with inherent anti-cancer activity have been identified and are in different phases of clinical trials. In the era of modern biotechnology and with better understanding of cancer biology and virology, it has become feasible to engineer the oncolytic viruses (OVs) to increase their tumour selectivity and enhance their oncolytic activity. In this review, the mechanisms by which oncolytic viruses kill the tumour cells have been discussed as also the development made in virotherapy for cancer treatment with emphasis on their tumour specific targeting.
Apoptosis; oncolytic viruses; tumour targeting; viral oncotherapy
Oncolytic viruses are genetically modified viruses that preferentially replicate in host cancer cells, leading to the production of new viruses and, ultimately, cell death. Currently, no oncolytic viruses that are able to kill only tumor cells while leaving normal cells intact are available. Using T-REx (Invitrogen, Carlsbad, CA) gene switch technology and a self-cleaving ribozyme, we have constructed a novel oncolytic HSV-1 recombinant, KTR27, whose replication can be tightly controlled and regulated by tetracycline in a dose-dependent manner. Infection of normal replicating cells as well as multiple human cancer cell types with KTR27 in the presence of tetracycline led to 1,000- to 250,000-fold-higher progeny virus production than in the absence of tetracycline, while little viral replication and virus-associated cytotoxicity was observed in infected growth-arrested normal human cells. We show that intratumoral inoculation with KTR27 markedly inhibits tumor growth in a xenograft model of human non-small-cell lung cancer in nude mice. It is shown further that replication of KTR27 in the inoculated tumors can be efficiently controlled by local codelivery of tetracycline to the target tumors at the time of KTR27 inoculation. Collectively, KTR27 possesses a unique pharmacological feature that can limit its replication to the targeted tumor microenvironment with localized tetracycline delivery, thus minimizing unwanted viral replication in distant tissues following local virotherapy. This regulatory mechanism would also allow the replication of the virus to be quickly shut down should adverse effects be detected.
Re-engineering the tropism of viruses is an attractive translational strategy for targeting cancer cells. The Ras signal transduction pathway is a central hub for a variety of pro-oncogenic events with a fundamental role in normal and neoplastic physiology. In this work we were interested in linking Ras activation to HSV-1 replication in a direct manner in order to generate a novel oncolytic herpes virus which can target cancer cells. To establish such link, we developed a mutant HSV-1 in which the expression of ICP4 (infected cell protein-4, a viral protein necessary for replication) is controlled by activation of ELK, a transcription factor down-stream of the Ras pathway and mainly activated by ERK (extracellular signal-regulated kinase, an important Ras effector pathway). This mutant HSV-1 was named as Signal-Smart 1 (SS1). A series of prostate cells were infected with the SS1 virus. Cells with elevated levels of ELK activation were preferentially infected by the SS1 virus, as demonstrated by increased levels of viral progeny, herpetic glycoprotein C and overall SS1 viral protein production. Upon exposure to SS1, the proliferation, invasiveness and colony formation capabilities of prostate cancer cells with increased ELK activation were significantly decreased (p<0.05), while the rate of apoptosis/necrosis in these cells was increased. Additionally, high Ras signaling cells infected with SS1 showed a prominent arrest in the G1 phase of the cell cycle as compared to cells exposed to parental HSV-1. The results of this study reveal the potential for re-modeling the host-herpes interaction to specifically interfere with the life of cancer cells with increased Ras signaling. SS1 also serves as a “prototype” for development of a family of signal-smart viruses which can target cancer cells on the basis of their signaling portfolio.
Genetic instability of cancer cells generates resistance after initial responses to chemotherapeutic agents. Several oncolytic viruses have been designed to exploit specific signatures of cancer cells, such as important surface markers or pivotal signaling pathways for selective replication. It is less likely for cancer cells to develop resistance given that mutations in these cancer signatures would negatively impact tumor growth and survival. However, as oncolytic viral vectors are large particles, they suffer from inefficient extravasation from tumor blood vessels. For larger particles, such as viral vectors, their ability to reach cancer cells is an important consideration in achieving specific oncolytic targeting and potential vector replication. Our previous studies indicated that the Sindbis viral vectors target tumor cells via the laminin receptor (LAMR). Here, we present evidence that modulating tumor vascular leakiness, using VEGF and/or metronomic chemotherapy regimens significantly enhances tumor vascular permeability and directly enhances oncolytic Sindbis vector targeting in tumor models. Since host-derived vascular endothelium cells are genetically stable and less likely to develop resistance to chemotherapeutics, a combined metronomic chemotherapeutics and oncolytic viruses regimen should provide a new approach for cancer therapy. This mechanism could explain the synergistic treatment outcomes observed in clinical trials of combined therapies.
Sindbis virus; viral vector; cancer; vascular leakiness; molecular imaging; chemotherapy
Oncolytic viruses refer to those that are able to eliminate malignancies by direct targeting and lysis of cancer cells, leaving non-cancerous tissues unharmed. Several oncolytic viruses including adenovirus strains, canine distemper virus and vaccinia virus strains have been used for canine cancer therapy in preclinical studies. However, in contrast to human studies, clinical trials with oncolytic viruses for canine cancer patients have not been reported. An 'ideal' virus has yet to be identified. This review is focused on the prospective use of oncolytic viruses in the treatment of canine tumors - a knowledge that will undoubtedly contribute to the development of oncolytic viral agents for canine cancer therapy in the future.
cancer; canine cancer therapy; oncolytic virus; oncolysis; target molecule, combination therapy
Tremendous advances have been made in developing oncolytic viruses (OVs) in the last few years. By taking advantage of current knowledge in cancer biology and virology, specific OVs have been genetically engineered to target specific molecules or signal transduction pathways in cancer cells in order to achieve efficient and selective replication. The viral infection and amplification eventually induces cancer cells into cell death pathways and elicits host anti-tumor immune responses to further help eliminate cancer cells. Specifically targeted molecules or signaling pathways (such as RB/E2F/p16, p53, IFN, PKR, EGFR, Ras, Wnt, anti-apoptosis or hypoxia) in cancer cells or tumor microenvironment have been studied and dissected with a variety of OVs such as adenovirus, herpes simplex virus, poxvirus, vesicular stomatitis virus, measles virus, Newcastle disease virus, influenza virus and reovirus, setting the molecular basis for further improvements in the near future. Another exciting new area of research has been the harnessing of naturally tumor-homing cells as carrier cells (or cellular vehicles) to deliver OVs to tumors. The trafficking of these tumor-homing cells (stem cells, immune cells and cancer cells), which support proliferation of the viruses, is mediated by specific chemokines and cell adhesion molecules and we are just beginning to understand the roles of these molecules. Finally, we will highlight some avenues deserving further study in order to achieve the ultimate goals of utilizing various OVs for effective cancer treatment.
cancer; oncolytic virus; oncolysis; selective replication; target molecule; signaling pathway; chemokine; drug
Unresectable hepatic colorectal cancer (CRC) metastases are a leading cause of cancer mortality. These tumors, and other epithelial tumors, often express both cyclooxygenase 2 (COX-2) and carcinoembryonic antigen (CEA). Because adenovirus vectors infect liver and lack tumor tropism, they cannot be utilized for systemic therapy of hepatic metastases. We used COX-2 transcriptional restriction, in combination with transductional adenovirus hepatic untargeting and tumor retargeting by a bispecific adapter, sCARhMFE, composed of sCAR (the Coxsackie and Adenovirus Receptor ectodomain) and MFE-23 (a single-chain anti-CEA antibody), to untarget liver following intravenous administration of adenovirus vectors expressing firefly luciferase and to retarget virus to hepatic colorectal tumor xenografts and non-small cell lung tumor xenografts. To improve both liver untargeting and tumor retargeting, we developed sCARfMFE, a trimerized sCARhMFE adapter. Trimerization greatly improves both untargeting of CAR-dependent adenovirus infection and CEA-dependent virus retargeting, in culture and in vivo.
Combining sCARfMFE bispecific adapter transductional liver untargeting and transductional tumor retargeting with COX-2 transcriptional tumor-restricted transgene expression increases systemically-administered adenovirus therapeutic efficacy for hepatic CRC tumors, using Herpes Virus Type 1 thymidine kinase as a therapeutic gene in conjunction with the prodrug ganciclovir. Both transductional untargeting and COX-2 transcriptional restriction also reduce HSV1-tk/GCV hepatic toxicity. In addition, transductional sCARfMFE untargeting reduces the innate immune response to systemic adenovirus administration. Combined transductional liver Ad untargeting, transductional tumor retargeting and transcriptional transgene restriction suggests a means to engineer practical, effective therapeutic agents for hepatic CRC metastases in particular, as well as hepatic metastases of other epithelial cancers.
adenovirus; colorectal cancer; COX-2 transcriptional restriction; transductional retargeting
Prostate cancer is the most frequently diagnosed cancer and the second leading cause of cancer deaths in men today. Although virus-based gene therapy is a promising strategy to combat advanced prostate cancer, its current effectiveness is limited partially due to inefficient cellular transduction in vivo. To overcome this obstacle, conditional oncolytic viruses (such as conditional replication adenovirus (CRAD)) are developed to specifically target prostate without (or with minimal) systemic toxicity due to viral self-replication. In this study, we have analyzed and compared three prostate-specific promoters (PSA, probasin, and MMTV LTR) for their specificity and activity both in vitro and in vivo. Both mice model with xenograft prostate tumor model and canine model were used. The best PSP was selected to construct a prostate-specific oncolytic adenovirus (CRAD) by controlling the adenoviral E1 region. The efficacy and specificity of CRAD on prostate cancer cells were examined in cell culture and animal models.