Periodic Reporting for period 2 - InCanTeSiMo (Intelligent cancer therapy with synthetic biology methods)
Período documentado: 2022-12-01 hasta 2024-05-31
Cancer is still one of the major causes of death worldwide. For women, breast cancer is the second most deadly type of cancer after lung cancer. Surgery, radio and chemotherapy remain the most common practices to date to fight this disease. While surgery and radiotherapy are local treatments –that is, they are directed only to the affected tissue–, they are rarely applied alone. Typically, chemotherapy is additionally prescribed to patients either because the cancer is too advanced or as prevention against metastases. As a systemic treatment, chemotherapy leads to severe side effects by damaging not only cancerous cells but also healthy ones. Moreover, even radiotherapy is often applied to areas of the body suspected to be susceptible to metastases, even before they become visible in a CT scan or MRI, damaging potentially healthy sites.
The ideal cancer therapy should be targeted only to the cancer cells, should be able to act to the same extent on the primary tumour and the metastases, should be effective and affordable.
In this project InCanTeSiMo, we apply synthetic biology approaches to develop a cancer therapy meeting several of the requirements listed above: targeted, effective, affordable.
To make the treatment targeted to cancer cells, we exploit the enhanced permeability and retention effect typical of the cancer microenvironment that results in a natural accumulation of nanoparticles of sizes ranging from 10-500 nm at the site of cancer. These nanoparticles carry on their surface specific ligands binding to receptors overexpressed on the surface of cancer cells. Once internalized by the cancer cells, the nanoparticles deliver their cargo, which is an agent able to trigger the destruction of the cancer cells. To improve on specificity, our aim is to use cargo able to distinguish cancer cells from healthy ones and act only there, leaving healthy cells in which the nanoparticles might accidentally enter unaffected. To make the treatment effective, we aim to evoke the patient’s own immune system, by employing nanoparticles that can activate the immune system. To make the treatment affordable, we aim to develop novel, more cost-effective strategies to obtain the nanoparticles and to substitute the antibodies generally used to target cancer cells with small peptides or protein binders such as nanobodies, monobodies or repebodies.
The ideal cancer therapy should be targeted only to the cancer cells, should be able to act to the same extent on the primary tumour and the metastases, should be effective and affordable.
In this project InCanTeSiMo, we apply synthetic biology approaches to develop a cancer therapy meeting several of the requirements listed above: targeted, effective, affordable.
To make the treatment targeted to cancer cells, we exploit the enhanced permeability and retention effect typical of the cancer microenvironment that results in a natural accumulation of nanoparticles of sizes ranging from 10-500 nm at the site of cancer. These nanoparticles carry on their surface specific ligands binding to receptors overexpressed on the surface of cancer cells. Once internalized by the cancer cells, the nanoparticles deliver their cargo, which is an agent able to trigger the destruction of the cancer cells. To improve on specificity, our aim is to use cargo able to distinguish cancer cells from healthy ones and act only there, leaving healthy cells in which the nanoparticles might accidentally enter unaffected. To make the treatment effective, we aim to evoke the patient’s own immune system, by employing nanoparticles that can activate the immune system. To make the treatment affordable, we aim to develop novel, more cost-effective strategies to obtain the nanoparticles and to substitute the antibodies generally used to target cancer cells with small peptides or protein binders such as nanobodies, monobodies or repebodies.
We made good progress by establishing a robust method to produce the nanoparticles (NPs), albeit we still have to optimize it to reduce costs and usage of chemical substances, which we would like to avoid. We also established a microfluidic setup which seems promising in the separation of NPs of the desired size range; however, it must still be optimized and the throughput increased.
We have established a robust method to decorate the NPs with any ligand of choice. Actually, we achieved more than we originally set ourselves to do, because the method we eventually adopted is very modular and allows us to change the ligand in an easy way. We started analysing the tumour tissue of the mouse model we plan to use in the last phase of the project to check if the EGF receptor (EGFR) is indeed overexpressed compared to healthy tissue. This analysis was performed on a microarray dataset previously obtained from our collaborator. To our surprise, EGFR is not differently expressed in cancerous and healthy tissue. We found some other promising candidates, which need validation via immunohistochemistry on tissue slices to see if the receptors are indeed present on the surface of the mouse cancer cells.
To obtain the cargo able to distinguish cancer from healthy cells we set up a collaboration with the metabolomics group to measure metabolites in tumour and healthy tissues from the specific mouse model we plan to use in the future to find potential metabolites that differentiate cancerous from healthy tissue in this mouse model.
We have established a robust method to decorate the NPs with any ligand of choice. Actually, we achieved more than we originally set ourselves to do, because the method we eventually adopted is very modular and allows us to change the ligand in an easy way. We started analysing the tumour tissue of the mouse model we plan to use in the last phase of the project to check if the EGF receptor (EGFR) is indeed overexpressed compared to healthy tissue. This analysis was performed on a microarray dataset previously obtained from our collaborator. To our surprise, EGFR is not differently expressed in cancerous and healthy tissue. We found some other promising candidates, which need validation via immunohistochemistry on tissue slices to see if the receptors are indeed present on the surface of the mouse cancer cells.
To obtain the cargo able to distinguish cancer from healthy cells we set up a collaboration with the metabolomics group to measure metabolites in tumour and healthy tissues from the specific mouse model we plan to use in the future to find potential metabolites that differentiate cancerous from healthy tissue in this mouse model.
So far, the NPs we decided to use have been decorated with antibodies, which are expensive and laborious to obtain. We have moved the technique farther by establishing a convenient way to decorate the NPs with any ligand of choice, mostly peptides and small proteins binders. We are currently testing the ability of NPs decorated with these various ligands to get internalized in a set of cancer cell lines. We also started developing a microfluidics device that can separate the NPs of the desired size from all others. The results are promising and we expect to have a final prototype by the end of the project, which will work at the desired high throughput. One of our aims was to improve on the production of the NPs in terms of costs and we feel rather confident that we will achieve this goal by the end of the project. More generally, the targeted cancer treatment we are working on is not represented in any clinical trials in Europe so far. There are clinical trials testing it in the USA –with substantial differences, such as the targeting strategy and the cargos delivered. Our aim is to bring this therapy to the clinical trial stage in Europe. This will likely take much more time than the 5 years of this project. The goal is to obtain enough evidence of the specificity and efficacy of the treatment in the mouse model to spark future work in other animals and, eventually, patients.