Menu

Application scenario

Our application scenario is summarized in this video. If you would like to know more about the application scenario, read the text below!

Introduction

Cancer is one of the leading causes of death worldwide. In 2012, 14.1 million new cancer cases occurred and the estimation is, that this number will rise to 22 million within the next two decades.^[1] Malignant tumors associated with cancer are known to have the ability to invade tissues and generate metastases.^[2-3] Matrix Metallo Proteinases (MMPs) are one of the components responsible for this behaviour of tumors. MMPs are able to degrade the extracellular matrix (ECM) surrounding the tumor, and thus permitting the invasion of nearby tissues.^[2-4] Next to that, MMPs are also involved in the regulation of bioactive molecules, such as cytokines and growth factors. These bioactive molecules play a role in cellular processes which are involved in promoting aspects of tumor growth, such as cell proliferation, angiogenesis and apoptosis.^[2] Therefore, MMPs are a useful target for blocking cancer progression.

Besides the fact that MMPs play a role in cancer progression, another important aspect of the usefulness of blocking MMPs is the stage of cancer progression in which the MMPs play a significant role. Research showed that for the administration of synthetic metalloproteinase inhibitors (MPIs) in advanced tumor stages, no efficacy was observed, indicating that MMPs are important in the early stages of tumor progression.^[5-6] Once metastases have been generated, the MMPs seem to be less important. Therefore, next to being a useful target for blocking cancer progression, MMPs are also a useful tool for detecting early stage cancer.

Current MMP targeting therapies

Due to the pivotal role that MMPs play in early stages of cancer, it should be no surprise that the field of cancer therapy already shows interest in the multiple approaches that target these MMPs.^[2]

One such approach is the inhibition of MMP gene transcription, which can be achieved with different methods, the first of which is the targeting of extracellular factors. Although difficult, studies have shown that certain interferons should be capable of blocking transcription of multiple MMPs. Apart from this, there is also potential in targeting growth factors and cytokines. Another method focuses on inhibiting the signaling pathway that induces the transcription. Halofuginone, for example, targets such a pathway and thereby inhibits the expression of MMP2, which cleaves collagen and is linked to different types of cancer.^[7] In this method it is important that other signaling pathways are not affected. However, this is difficult to achieve. A third way of inhibiting MMP gene transcription is targeting nuclear factors that are involved in the genetic activation. Once again it must be stressed that these factors may be involved in multiple processes and therefore yield difficulties when it comes to creating applicable treatments.

A second approach is based on blocking the activation of proMMPs. Normally these inactive proMMPs are cleaved by other proteases such as MT-MMPs, turning them into active MMPs. Antibodies or other molecules may be used to prevent this from happening, thus preventing MMPs from breaking down the matrix.

A third alternative would be blocking the active site of an MMP, using aforementioned metalloproteinase inhibitors (MPIs). So far MPIs have shown low efficacies and pharmacokinetic values, thus also yielding poor results.

Overall, the results of these therapies have shown only varying degrees of success and exposed numerous difficulties, which is why we introduce a novel approach to impairing the detrimental effect of MMPs in cancer.

Our solution

Our system, GUPPI, works in a different way than current strategies as it uses the presence of the MMPs. GUPPI consists of a 14-3-3 tetramer, which functions as a scaffold protein, and its binding partner CT33. Both protein constructs have a different valency which makes network formation possible. This network provides a gel-like structure which can be used to encapsulate targeted tissue.

In case of cancer, GUPPI can use the presence of MMPs in an activating way. The system can be inhibited using a bivalent ExoS peptide.^[8] The linker that connects the 14-3-3 construct with the ExoS contains a protease recognition site, which can be cleaved in presence of that specific protease. If the linker is cleaved by a protease, the binding between the ExoS and 14-3-3 will be broken and the binding potency of 14-3-3 and CT33 increases. In our system this protease recognition site is an MMP recognition site, providing that the gel can be formed only in the presence of an MMP.

Concluding, GUPPI can be used in a way that makes use of the properties of tumor cells. The excreted proteases can cleave a linker after which inhibition of the system is eliminated, leading to gel formation around the tumor cells.

Figure 1: Encapsulation of tumor cells by GUPPI, triggered by MMPs

Integrated Human Practices

To see whether this application scenario is viable in cancer treatment, we talked to experts: Ignace de Hingh from the Catharina Hospital in Eindhoven and Arnoud Sonnenberg from the Netherlands Cancer Institute. More information about these talks can be found here.

The most important detail to make our application scenario more viable is the kind of MMP that is to be targeted. The presence or increased expression of MMP-1, 2, 3, 7, 9, 13 and -14 is associated with tumor progression, but only a few MMPs have been identified for a specific type of cancer.^[9] In most of the cases, elevated plasma levels of gelatinases correlate with higher occurrence of metastases, indicating that MMP-2 and MMP-9 play an important role, since these are gelatinases. Moreover, MMP-9 is shown to also play a role in upregulating the release of growth factors (VEGF), to alter local microenvironments to attract circulating tumor cells and a major role as a pro-angiogenic molecule.^[9] MMP-9 also plays a critical role in tumor-induced angiogenesis.^[9] So, by using MMP-9 as a signal for gel formation, metastasis could be prevented and the upregulation of growth factors can be altered.

MMP-2 and MMP-9 are the most studied MMPs, but MMP-10 could also be a good target for our GUPPI therapy. A paper of Zhang et al.^[10] shows that MMP-10 also promotes tumor progression for tumor cells of epithelial origin, via the regulation of angiogenic and apoptotic pathways. So, MMP-10 might also be a good target for our GUPPI system.

Apart from the aforementioned proteases, MMP-1 also shows a promising target for preventing metastasis.^[11] This specific protease is shown to be important in vascular intravasation, which indicates that it might be a promising target, since this would lead to targeting the tumor early in the process of metastasis. So, summarizing, MMP-1, 2, 9 and 10 could be potential MMPs to target.

Arnoud Sonnenberg expressed his concern whether tumors can be encapsulated entirely by our gel. Looking further into this, tumor cells indeed do have a invasive front, rather than releasing proteases all around the tumor area.^[9] This might lead to incomplete encapsulation of the tumor cell, but it is unclear whether this would influence the ability of our gel to prevent metastasis. However, to prevent incomplete encapsulation, another application of our GUPPI system might be the encapsulation of metastatic tumor cells already in the vascular system. This way, the tumor cells are still small enough to fully encapsulate, but instead of encapsulating the cells to stop them from spreading, it can be used as a diagnostic tool to detect metastases early on in the process.

^[1]“World cancer factsheet,” vol. 2012, no. 2012, pp. 2012–2015, 2014.
^[2]C. M. Overall and C. López-otín, “Strategies for MMP inhibition in cancer: innovations for the post-trial era”, Nat. Rev. Cancer, vol. 2, no. September, 2002.
^[3]D. Bourboulia and W. G. Stetler-stevenson, “Seminars in Cancer Biology Matrix metalloproteinases ( MMPs ) and tissue inhibitors of metalloproteinases ( TIMPs ): Positive and negative regulators in tumor cell adhesion,” Semin. Cancer Biol., vol. 20, no. 3, pp. 161–168, 2010.
^[4]L. M. Coussens, B. Fingleton, and L. M. Matrisian, “Matrix Metalloproteinase Inhibitors and Cancer : Trials and Tribulations,” Science (80-. )., vol. 295, no. March, pp. 2387–2393, 2002.
^[5]B. M. Fingleton, K. J. H. Goss, H. C. Crawford, and L. M. Matrisian, “Matrilysin in early stage intestinal tumorigenesis,” APMIS, no. 3, pp. 102–110, 1999.
^[6]G. Bergers, K. Javaherian, and K. Lo, “Effects of Angiogenesis Inhibitors on Multistage Carcinogenesis in Mice,” Science (80-. )., vol. 284, no. April, pp. 808–813, 1999.
^[7]M. Bjo and E. Koivunen, “Gelatinase-mediated migration and invasion of cancer cells,” Biochim. Biophys. Acta, vol. 1755, pp. 37–69, 2005.
^[8]S.J.A. Aper, “Engineering protein switches for sensing and actuation“, Ph.D. dissertation, Dept. Biomed. Eng., TU/e, 2016.
^[9]E. I. Deryugina and J. P. Quigley, “Matrix metalloproteinases and tumor metastasis,” Cancer Metastasis Rev., vol. 25, no. 1, pp. 9–34, 2006.
^[10]G. Zhang, M. Miyake, A. Lawton, S. Goodison, and C. J. Rosser, “Matrix metalloproteinase-10 promotes tumor progression through regulation of angiogenic and apoptotic pathways in cervical tumors,” BMC Cancer, vol. 14, pp. 1–14, 2014.
^[11]A. Juncker-jensen, E. I. Deryugina, I. Rimann, E. Zajac, T. A. Kupriyanova, L. H. Engelholm, and J. P. Quigley, “Tumor MMP-1 Activates Endothelial PAR1 to Facilitate Vascular Intravasation and Metastatic Dissemination,” Cancer Res., vol. 73, no. 14, 2013.

Footer