Friday, May 4, 2012

Markers For Life


  To de diagnosed with cancer is bad enough, but to find out it has metastasized is often deemed to be a death sentence. With the survival rate of those with metastatic cancer slim and the fact that, “American men and women have an 38-44% chance of developing invasive cancers in their life time”(1) there has to be something done. The article published in the American Chemical Society, Reprogramming Urokinase into an Antibody-Recruiting Anticancer Agent by Spiegel and colleges, provides insight into research creating a synthetic compound to target the natural biomarker of many metastatic cancers. With this technology of biomarker targeting synthetic compounds Spiegel and his fellow colleagues are able to take advantage of the bodies native immunological response. This could hold the potential of returning hope to those who thought there was none. Spiegel’s research not only targets metastasized cancer cells, but is more likely to possess the potential for long lasting immunity in addition to the added benefit of eliciting less immunological side effects for the reason that this process takes advantage of natural processes and substances.



What is Spiegel Targeting and Why:
Spiegel and his fellow researchers focuse on the biomarkers the urokinase-type plasminogen activator (uPA) and its receptor urokinase-type plasminogen activator receptor (uPAR). But why use uPA and uPAR as a target for new types of therapeutic treatments for metastatic cancer. Tumor metastasis begins with cancer cells invading surrounding tissues which is frequently accelerated by cell-surface proteases. uPA, is a type of cell surface protease, meaning it is involved in a variety of basic cell activities and possesses influences on growth factors that could have very drastic and rapid effects on the cell. In this case uPA has the potential to activate migration-inducing signal transduction cascades. This explains why uPA and uPAR is found at substantially higher levels on metastatic cancer cells. Due to this, “high levels of both uPA and uPAR are used as a diagnostic markers for metastatic potential and poor clinical outcome in numerous malignancies”(1). Also previous studies that target the uPA- uPAR system cause a reduction in both the size and invasive potential of cancer cells without extensive damage to healthy tissue. This is beneficial since the majority of classic treatments, such as chemotherapy and radiation leave normal tissue harmed by the indiscriminate cancer treatment.   
 New Technology For Targeting uPA and uPar: 
 To target uPA and uPAR, Spiegel and his fellow researchers used concepts from the field of synthetic immunology. Research in this field attempts to modify biological processes that would otherwise not normally function as in nature through the use of synthetic materials. Spiegel came up with two biofuntional molecules that allows for the immune system to recognize cancer cells that would have otherwise avoided detection. These two molecules “can convert uPA into catalytically inactive, bifunctional constructs (ARM-Us) that are capable of both recruiting antibodies and directing antibody-dependent immune responses against uPAR- expressing cancer cells”(1).  These two molecules posses the power to covalently bond to its active site by either a fluorescein label or to 2,4-dinitrophenyl (DNP) moiety. The covalent linkage of DNP offers a way for the body to recruit antibodies to attack the cancer cells because in humans there are natural found anti-DNP antibodies. This means that when the ARM-U binds with high affinity to the cells with high uPAR levels, malignant cancer cells, the simultaneously covalent attachment of DNP triggers the natural immune response of phagocytosis and cytotoxicity because of the signaling of anti-DNP antibodies.
Testing and results:
To perform this experiment the researchers used chloromethyl ketones 1,2. Ketones 1 can simulate ARM-U attached to a fluorescein label while ketone 2 can simulates ARM-U bonded to DNP.  Methyl Ketone 3 acted as a negative control because of it lack of ability to form a covalent bond. While ketone 3 showed no bonding to uPA as predicted, the results did show that there was a 97% reduction in uPA activity with the use of Ketone 1 and the fluorescein label when compared to uPA and buffer activity alone. Further testing showed that ARM-Ufluor targets cells via the desired uPAR interactions, and that the attachment of the fluorescence does not affect the initial binding process. After determining that ARM-U actually binds to uPA and can decrease its activity it was time to test the capability of the ARM-U mediated killing via DNP. The ARM-UDNP complex was shown to have high levels of cellular cytotxicity and to be very efficient in the mediated phagocytosis. The subsequent increase in the presence of anti-DNP antibodies in turn decreased the cell viability revealing the importance and actual viability of the attachment of the DNP antibodies. As hoped the negative controls did not elicit an immune response. Lastly, when both the fluorescein label and DNP were combined the immune response increased even more.



Critiques and Concerns: 
           This research is certainly a novel set up capable of targeting cancer cells for immune mediated cancer destruction found in numerous malignant cancer types. As shown in Spiegel’s research this method of therapeutic treatment is unlikely to affect other cells, which is definitely a step in the right direction compared to other current methods of cancer treatment.  An added bonus is that the FDA has already approved processes using uPA. This means that if further clinical trials continue to show this progress there will be less time to wait for this technology to come to market. By having other applications targeting uPA, potential side effects can be explored further to minimize and prevent them.
           Because cancer is such a prevalent and menacing disease, research into creating effective treatments like this one is necessary, however, I did come across some limitations. Since this process is dependent on abnormally high concentration of uPA and uPAR, I am left with the question that if cancer cells find a way to either mutate or bypass these cell surface proteases,  thus leading to lower levels of uPA, would this therefore make this model ineffective? We have learned in class that as cancer cells try to avoid detection from the immune system it modifies or even down regulates signaling that would otherwise act as an alarm. For example if RasGTD is constantly active due to some mutation, normally the cell would sense this and start to senescence or even apoptosis. To prevent this from occurring cancer cells will try to reduce the concentration of RasGTD. If this same process occurs with uPA and cell surface uPAR, and the cancer cells still manage to spread, then this innovative therapeutic will be rendered ineffective.  This is why I feel it is necessary to have further research into at what concentration levels of uPA and uPAR will ARM-U be able to effectively target and activate and immune response. Another follow up research idea is to look into applying this technology to target other cell surface proteases involved in the spread of cancer, subsequently if cancer cell do find a way of flourishing without uPA or uPAR there will be a back up target. Because the biomarker they use is one found in malignant cells due to its connection with migration-inducing signal transduction cascades, there needs to be future research into how this technology could be applied to pre malignant forms of cancer.

            Additionally, I found to have questions with the claim made that this process can provide a longer lasting immune response. Even though this course of action uses natural antiagents, the antiagents used are not normally associated with that cancer on its own. So, if the cancer were to come out of remission how would the body be able to take use of that immunological memory on its own with out another treatment?  Due to this there would also need to be research into how long the synthetic molecules stay in the body, this way we could not only see how long the treatment is initially needed for, but also to potentially see how long its presence could keep cancer in remission, and if it does stay in the body for long periods of time will the cancer find ways around it. 



Work Cited:

Spiegel ,David A, et al. “Reprogramming Urokinase into an Antibody-Recruiting Anticancer Agent.” ACS Chemical Biology (18 November, 2011). Web 30 April, 2012. <http://pubs.acs.org/doi/abs/10.1021/cb200374e>

Introduction picture from: 
n.p. “Breast Cancer Metastasis Could be Predicted” TopCancer News. Web. 3 May, 2012.     <http://www.topcancernews.com/news/2073/breast-cancer-metastasis-could-be-predicted.html>