The Herpes virus strikes again, though not in the way you might think. In one of my earlier blog (Who Says Herpes Is All Bad), the Herpes Simplex Virus (HSV) was exposed for being more than a menace but actually an innovative tool for marking cancerous cells with Gaussia Luciferase. This marking enables cancer cells that would normally go undetected to glow and be seen. I came across the article, Targeting HSV-1 virions for specific binding to epidermal growth factor receptor-vIII bearing tumor cells, where Paola Grandi and her fellow researchers have found a way to effectively target and destroy deadly cancerous cells through the use of this multi-faced Herpes Simplex Virus (HSV). Grandi found a way to modify the HSV viral envelope glycoprotein’s, by swapping out the normal heparan sulfate binding domain with a tumor-specific immunotoxin. This immunotoxin, not only has the ability to target this modified virus to the naturally found tumor cell biomarkers, but in other research has also shown the capacity to mediate cell death on its own. This type of immunotherapy is a promising alterative to current methods of treatment, especially with their focus on glioblastoma multiform, one of the “most common primary brain tumors [which] are almost universally fatal despite aggressive therapies, including surgery, radiotherapy, and chemotherapy”(1).
What Did Grandi Set Out To Target, With What, and Why:
Grandi and her fellow researchers focused on the biomarker EGFR vIII, a mutant form of the tyrosine kinase receptor EGFR. But why use EGFR vIII as a target for this viral vector therapeutic treatment for glioblastoma multiform (GMB)? Well, the wild type EGFR is a type of proto-oncogen, meaning that if mutated in a certain way it could lead to the cell acquiring cancer like properties. The wild type EGFR elicits a signal transduction cascade that lead to cell proliferation, DNA synthesis, and acquisition of new phenotypes like cell migration. All these properties, if left unregulated can lead to cancer. As we have learned in class one of the reasons for signal transduction pathways being permanently activated is through the truncation of the extracellular portion of the receptor. This phenomna is seen in EGFR vIII. Because of an in-frame deletion in the mRNA, the extracellular receptor portion is truncated causing the kinase to be continually activated due to the ligand independent binding. This has been shown to enhance the tumorigenicity and apoptosis resistance of cells containing the EGFR vIII mutant. (2) So, why is this important? EGFR vIII is associated with increased invasiveness and growth rate of tumors. Mutations in the EGFR, especially the EGFR vIII mutant, are some of the most common mutations found in GMB cells, leading to higher concentrations of the EGFR vIII mutant on the cell surface compared to normal cells, which have virtually none. Also, it has been shown that “several antibodies have been described that are specific to EGFR vIII and do not cross-react with wild-type EGFR”(1).
To target EGFRvIII, Grandi and her fellow researchers, used a modified version of the oncolytic virus HSV. An oncolytic virus is a type of virus that causes the infected cell to lyses. As the virus replicates it not only destroys the host cell but also keeps this destructive cycle going since it infects adjacent cells causing subsequent destruction. Though there are multiple types of viral vector therapies in clinical trials, oncolytic viruses like HSV show the most promise. The key to Grandi research was finding a way to engineer the HSV virus to effectively target only the cell specific surface receptors EGFR VIII, unique to cancerous GMB cells, leaving the normal cells unharmed. To generate this tumor selectivity Grandi implemented the single strand monoclonal antibody MR1-1. In previous studies MR1-1 has been shown to have high binding affinity accompanied with long lasting infectivity with EGFR vIII causing it to be extremely potent to cells with this mutation. Instead of having a heparan sulfate binding domain, which allows for the viruses initial binding with the cell surface, the virus was modified so it would only bind to the EGFR vIII receptor. Because Grandi created a specific immunotherapeutic target, EGFR vIII, it not only concentrated the viral vectors specifically to the EGFR vIII containing cells but it also increases the safety of the treatment and lessens the needed dose.
Testing Method and Results:
What Grandi proposes would increase the potency of these vectors, by enhancing targeting and increasing vector infectivity to tumor cells. To test that attaching MR1-1 to HSV actually does this, Grandi first did a comparative study to see different monoclonal antibodies: MR1-1 (high affinity expected), MRB (low affinity expected), and the normal gC binding domain without the HS-binding domain relative binding affinity. Each of these “were inserted in the pCONG amplicon which also contains an expression cassette for green fluorescent protein (GFP) to monitor amplicon vector infection”(1). Grandi found that the MR1-1 attachment increased the infectivity rate of the human glioblastomas cells with EGFR vIII 5x. I should note that this was preformed at low M.O.I (multiplicity of infection) to imitate the expected in vivo ratio of vectors to tumor cells. To test infectivity of the of MR1-1 modified HSV and make made sure the MR1-1 modified HSV would only bind to glioma cells with the EGFR vIII receptors Grandi implanted two sets of U87 cells, a human primary glioblastoma cell line, with and without the EGFR vIII in nude mice and let them grow. She then infected the modified virus in each set of mice and monitored the progression over a week as seen in the picture. The infectivity was monitored by using fluorescent or lacZ reporter genes in vivo that were incorporated into a virus. A set of infected mice were injected with the normal gC binding domain lacking the Herparan sufalte binding domain as a control. Grandi observed that not only did the tumor shrink but also the modified virus had longer infectivity. This shows incredible promise for future use in cancer therapies.
Critique and Concerns: This research certainly provides a creative approach to target cancer cells for virus-mediated destruction. GMB is such a fast paced disease, having a median survival rate of 12-18 months after diagnosis, making research into creating effective treatments like this one necessary. Though this article did provide me with a way that viruses could actually destroy cancerous cells it still left me with many concerns and causes for future research opportunities. First of all this study did not specify the necessary dosage of the virus and at what concentration of EGFR vIII mutant on the cell surface is necessary for the modified virus to be able to target and actually have the desired effect on the cancer cell. Another major aspect that I feel was left out of this study is the implication of the blood brain barrier. Grandi did not present any data on how the blood brain barrier would affect the MR1-1 EGFR vIII accessibility to the GMB tumor. Even though the wild type HSV virus is known to be able to penetrate the blood brain barrier it was not mentioned if the attachment of the monoclonal antibody would have any effect on the modified virus passing this barrier. Because this study has only been done in nude mice, and even in these studies the tumor was not grafted in the cranial portion of the mouse, the effect of this modification has not been shown in respect of it passing the blood brain barrier.
Since this process is dependent on the presence of the specific mutant EGFR vIII being present on the cell surface, I am skeptical that cancer cells with prolonged exposure to this treatment will not mutate and become resistant to this treatment. As discussed in class there have been multiple other drugs that target oncoproteins that after an extended length of time are found to be useless because resistance has been confirmed in the cancerous cells. Two of the most relatable drugs to this are Herceptin and Gleevec. As we have learned Herceptin uses a Monoclonal antibody to target and block function of the HER2, a type of Epidermal growth factor receptor, by preventing dimerization. But upon tumor relapse, Herceptin elicits no response. This is because the tumor cells that some how survived the first round of Herceptin treatment managed to mutate and developed alternate means to propel growth independent of the HER2 oncogene expression. Or, in the case of Gleevec, the oncoprotein it was designed to bind to mutated causing a change of various amino acids in the binding domain, resulting in the inability for the treatment to bind and target the cell. So what is to say the same thing does not happen to the MR1-1 HSV treatment? If either of these processes occur, finding another path to cause cell growth while avoiding the expression of EGFR vIII or modifying the binding domain of EGFR vIII, this therapy in its current state would be rendered useless. Therefore there needs to further research into other monoclonal antibodies attachment to the virus allowing for the continuation of binding to EGFR vIII and or new cell surface kinase targets for the HSV virus. Also, additional research should explore how the addition of multiple monoclonal antibodies to the HSV virus would accommodate for these changes in the cancer cell’s morphology.
Since this treatment uses a virus, I am left to question how the body’s immune response will affect the efficacy of this therapy. As with increased exposure to any virus the body’s natural immune system starts to acquire, for the lack of a better word, memory of the virus, making the virus a key target for immune systems attack. If this treatment calls for the virus therapy to be administered frequently in a short length of time, I am concerned that the bodys’ own immune system will start attacking the virus and gain immunity to it, greatly diminishing the effects of the treatment. If this happened, it would not matter that the virus can target the cancer; the virus would be prevented from propagating leaving the cancer cell unharmed and the patient in no better condition then before treatment
In this study Grandi experimented on U87 cells, which were taken from a stage 4 GMB tumor cell lineage, but the article does not specify at what stage the EGFR vIII mutation begins to appears on the cancer cell’s surface. This would definitely need to be explored because if it is only expressed in certain stages this would dramatically effect the window of opportunity for the drug. And since the ERGF vIII is seen on multiple cancer types and is not limited to GMB I feel it would be beneficial to explore the use of this treatment for metastasized cancer. And lastly, I feel there will be public and ethical barriers to overcome. Intentionally injecting a virus into someone does not come without hostile response and apprehension.
Grandi, Paola, et al. “Targeting HSV-1 virions for specific binding to epidermal growth
factor receptor-vIII bearing tumor cells.” Cancer Gene Ther. 2010 September ; 17(9): 655–663. Web. 20 May, 2012 <https://www.dropbox.com/s/hcydsgsh7bz1gqe/nihms-194496.pdf>
Kuan, C-T. , et al. “EGF mutant receptor vIII as a molecular target in cancer therapy.” Endocrine-Related Cancer. 2001 June; 8: 83–96. Web. 20 May, 2012
Introduction picture from:
Simula, Pekka. “Maturing data in 2011 will get us focused on the oncolytic virus industry.” Developing oncolytic virus therapies. Web. 22 May, 2012. <http://oncolyticvirus.files.wordpress.com/2010/12/oncolytic-virus-in-action.jpg>