Oncolytic Viruses: the homing missiles of cancer therapy

...or evil.

I think viruses are cool. I mean, really cool. I listen to a weekly podcast on viruses when I’m in the shower, lab, car, or on a run and I keep up with a couple of virus blogs that talk about viral global health concerns and related research. I don’t really have a good reason for my viral fascination other than that they’re a cultural taboo. They’re portrayed in the general public as vectors of disease and death, but in reality viruses are so diverse, so versatile, and so omnipresent that their pathogenicity is incredibly variable. Viruses like smallpox and HIV can ravage a society, but some, like the set I will be taking about, can be used to fight a variety of cancers...

So without further ado, I am excited to present my first post on the fascinating world of Oncolytic (onco=“cancer” and lytic=“bursting”) Viruses.



Oncolytic viruses were first discovered in the early 19th century when cancer regression was clinically reported in patients that were naturally infected with certain viruses (this was even before the discovery of the first virus, the tobacco mosaic virus, by Martinus Beijerinck in 1898). The basic theory was that infected patients had something in them that preferred or "targeted" cancer cells (Figure 1).  The implementation of oncolytic viruses into the clinical setting, however, has been hindered due to a significant lack of research and knowledge in the fields of virology, oncology, and molecular biology. When cancer patients were first therapeutically infected with these viruses, either the host immune response would prevail and destroy the virus or the virus would be too "general" in its targeting and end up attacking the patient systemically. Targeting has been one of the biggest setbacks in the use of oncolytic virotherapy, for decades scientists did not have enough information on the determinants of viral tropism (tropism: viral specificity for certain tissues). These viruses are naturally oncotropic, but their efficacy was not sufficient enough. However, due to the advancements in the three fields mentioned earlier, improving the targeting efficiency of oncolytic viruses allows for highly cancer-specific pathogens, possibly resulting in a safe and efficacious therapeutic. 

Figure 1. Basic principle of oncolytic virotherapy.

There are three modifications being studied to make naturally oncolytic viruses more efficacious: 

• targeting (increasing cancer specificity)
• arming (increasing oncolytic potency), and 
• shielding (protecting the virus from immunorecognition/suppression)

I've rewritten this blog post twice now, so for the sake of brevity, I will focus on one of four current research strategies being utilized for enhanced viral targeting of cancer cells.

Before going on about how oncolytic viruses target cancer cells, however, I should take a second to discuss tropism. Viruses are highly specific regarding which cells they can reproduce in. For example, rabies targets neurons, HIV targets helper T lymphocytes, hepatitis B targets hepatocytes, and so on. But, viruses don't "target" cancer as one might think. The use of the anthropomorphic description of these non-living agents describes how each viral family has a specific tropic environment that is prime for infection and reproduction. In the case of oncolytic viruses, that just happens to be cancer cells. Cancer targeting is possible because in the creation of cancer cells, specific genes that promote the hallmarks of cancer (i.e. metastasis, angiogenesis, evading immune suppression, etc.) compromise the cell's ability to fight off certain viral infection. An understanding of viral tropism for each of the oncolytic viral families (adenovirus (AD), herpes simplex virus (HSV1), vaccinia virus, myxoma virus, measles virus, Newcastle disease virus (NWD), reovirus, poliovirus, vesicular stomatitis virus (VSV)) has been critical for knowing how to modify these viruses to increase their cancer specificity.


In researching oncolytic viruses I have come across a plethora of journal articles and over a dozen review papers published in "big" journals. My guiding source though has been an extensive review paper published in Nature Reviews: Microbiology (Cattaneo et al., 2008). This review identifies four "layers" of modifications in viruses that can enhance viral oncotropism: host-cell entry through targeting cancer-specific viral glycoprotein proteases, entry through cancer-cell-specific receptors, enhanced viral replication through the utilization of tissue- or cancer-specific promoters, and finally I will be discussing viral exploitation of specific cancer cell defects.

Exploiting cancer cell defects
The goal in designing any cancer treatment is to create a therapeutic that is maximally potent and maximally safe. All too often though, these two goals are mutually exclusive. In the case of oncolytic virotherapeutics, the most potent cancer treatment is a toxic dose of wild-type virus. Seeing as this treatment is unreasonable, a lot of research is being to done to reduce toxicity of these wild-type by attenuating (weakening) the wild-type strains. As the review paper states, however, "attenuation is always relative to the target cell." These attenuated viruses being constructed are unable to reproduce in and kill normal cells, however, in cancer cells they proliferate. Viruses selectively replicate in cancer cells because these cells sacrifice a great deal of their immune function in order to avoid cell death. Thereby, these weakened viruses are still relatively effective in infecting and replicating in cancer cells. 

Figure 2 illustrates IFN's role in fighting a viral infection in normal cells, however, one of the key immune defects in cancer cells is in their expression of interferon (IFN). Cancer cells can neither produce nor respond to the IFN cytokine, making them highly susceptible to viral infection and ultimately cell death. At a quick glance, this would appear to be too easy a solution to an incredibly complicated issue, almost "too good to be true." It is. Kind of.

Figure 2. Outline of IFN synthesis and mechanism of action.  When a virus infects a normal cell (1), the virus stimulates expression of IFN. In a noble effort, this cell then dies resulting in the local release of IFN to warn nearby cells of a viral infection. When IFN binds to IFN receptors on nearby normal cells, the cell will activate the expression of antiviral proteins (2) that will inhibit viral protein synthesis, specifically break down viral mRNA, and halt overall viral reproduction (3).

Viruses have evolved a variety of mechanisms to evade or halt the IFN antiviral pathway. Meaning that IFN producing and sensing cells can be just as susceptible to infection as those that can do neither. IFN-blocking viruses drive the toxicity of oncolytic viruses much too high. In order to make this treatment viable, the virus must be relatively attenuated, it has to be made IFN susceptible again. An article (Devaux et al., 2006) published in Virology details how a measles virus (MV) was attenuated. 

MV is a fairly small virus with five genes in total. One of these genes, the P gene, codes for three proteins (P, C, and V proteins) that are responsible for bypassing the IFN mechanism of action by disrupting the pathway downstream of the IFN receptor, inhibiting the phosphorylation, activation, and nuclear translocation of a protein called STAT (signal transduction and activator of transcription), which is a key transcription factor for the antiviral proteins described in Figure 2. Devaux's strategy in attenuating the virus is to knockout each gene and see its effects on STAT inhibition. Each gene was independently knocked out and ultimately resulted in no effect. Figure 3 shows that when the C and V proteins were knocked and the P protein was mutated at residue 110 (P protein functionally important, not possible to replicate if fully knocked out), STAT shows significantly higher nuclear translocation in Figure 3A than in IFN resistant wild-type MV.

Figure 3. STAT translocation for P gene mutants. Human HeLa cells were inoculated with (A) recombinant MV with double-knockout C and V proteins and a mutation in the P protein (Y110H), MVvacP(Y110H)CV(ko) and (B) a wild-type MV, MV-NSe, then stimulated with IFNα. STAT stained red, nuclei stained with DAPI, overlapping appears as pink.

A really interesting side note on IFN modification is that many human ovarian carcinoma cancers actually promote IFN, limiting the application of the aforementioned recombinant strains. However, to increase oncolytic activity in these unique cancer cells, scientists have purposely engineered IFN susceptible MV strains, which strangely sacrifices the goal of specified targeting for a more blunt aim of attacking the cancer.

The field oncolytic viruses is prime for further investigation. These viruses are proving incredible promise and have even spawned the Journal of Oncolytic Virotherapy, a small publication started in late 2012. There is still however a great deal of research left to be conducted, not only in targeting and safety of the virus, but in designing viruses that maintain their oncolytic properties when supplemented with other cancer therapeutics, such as an MV recombinant (MV-NIS) that is fully functional when combined with cyclophosphamide. Currently, clinical trials for oncolytic viruses have been much faster in China than in the United States. China has several viruses in all stages of clinical trials, along with the only approved therapeutic of the group, an adenovirus that goes by the name Oncorine (H101) that can be functionally used in treatment with cisplatin. As viruses advance through clinical trials into approval in China, their efficacy is driving the United States to greater research in this fascinating field that is sure to become a tool in the treatment of cancer for years to come.

References

1. Cattaneo, R., Miest, T., Shashkova, E. V., & Barry, M. A. (2008). Reprogrammed viruses as cancer therapeutics: targeted, armed and shielded. Nature Reviews. Microbiology, 6(7), 529–540.
2. Devaux, P., Messling, von, V., Songsungthong, W., Springfeld, C., & Cattaneo, R. (2007). Tyrosine 110 in the measles virus phosphoprotein is required to block STAT1 phosphorylation. Virology, 360(1), 72–83.
3. Hammill, A. M., Conner, J., & Cripe, T. P. (2010). Oncolytic virotherapy reaches adolescence. Pediatric Blood & Cancer, 55(7), 1253–1263. 
4. Miest, T. S., & Cattaneo, R. (2014). New viruses for cancer therapy: meeting clinical needs. Nature Reviews. Microbiology, 12(1), 23–34. 
5. Workenhe, S. T., & Mossman, K. L. (2014). Oncolytic virotherapy and immunogenic cancer cell death: sharpening the sword for improved cancer treatment strategies. Molecular Therapy, 22(2), 251–256.