Periodic Reporting for period 1 - BEACONSANDEGG (Mechanobiology of cancer progression)
Período documentado: 2022-09-01 hasta 2025-02-28
Female breast cancer is currently the most diagnosed cancer, with an estimated 2.3 million new cases per year. Breast cancer is known to be initiated by the mutation of specific oncogenes. It has been recently proven, however, that progression of the tumour is dictated by changes in the mechanical microenvironment of the mutated cells. Breast cancer aggressiveness correlates specifically with fibrosis involving the deposition of a collagen matrix by stromal fibroblasts.
When primary breast tumours form stiffer matrices, they also metastasize more. Tumour stage progresses with fibrosis that modulates tumour entry by immune cells, nutrients, gases and anticancer drugs carried by the tumour microvascular network. Pharmaceutical companies are investing billions on the development of personalised treatments for patients who develop recurrent disease after radiation, surgery and/or chemotherapy. To overcome the progressive tumour resistance to anticancer treatments, it would be transformative for the field to understand and control how the tumour fibrotic environment evolves.
The development of tissue is significantly faster in embryonic organisms. The embryonated chicken embryo model has also been used to monitor in vivo the invasive features of human ovarian, thyroid, and skin cancer cells. No one has ever replicated the human cancer fibrotic niche and relevant druggability using this model.
My general goal is to bioengineer an array of tumour fibrotic microenvironments in vivo with varying levels of matrix stiffness and vascularity, and predict mass transport within such environments. My specific goals are: (a) To model 3D tumour micro environments with variable levels of fibrotic progression. (b) To develop an imaging window that incorporates the micro scaffolds, also transparent, oxygen-permeable, biocompatible and implantable in vivo (c) To create an experimental model of cancer fibrotic stiffening in vivo, in the chorioallantoic membrane of an embryonated avian egg. (d) To monitor the fibrotic progression of the tumour micro environments in vivo, in real time. (e) To predict mass transport in the tumour micro environments. (f) To validate the platform, in its ability to reproduce on breast tumours with different fibrotic progression the effect of anticancer drugs, both approved and investigational.
When primary breast tumours form stiffer matrices, they also metastasize more. Tumour stage progresses with fibrosis that modulates tumour entry by immune cells, nutrients, gases and anticancer drugs carried by the tumour microvascular network. Pharmaceutical companies are investing billions on the development of personalised treatments for patients who develop recurrent disease after radiation, surgery and/or chemotherapy. To overcome the progressive tumour resistance to anticancer treatments, it would be transformative for the field to understand and control how the tumour fibrotic environment evolves.
The development of tissue is significantly faster in embryonic organisms. The embryonated chicken embryo model has also been used to monitor in vivo the invasive features of human ovarian, thyroid, and skin cancer cells. No one has ever replicated the human cancer fibrotic niche and relevant druggability using this model.
My general goal is to bioengineer an array of tumour fibrotic microenvironments in vivo with varying levels of matrix stiffness and vascularity, and predict mass transport within such environments. My specific goals are: (a) To model 3D tumour micro environments with variable levels of fibrotic progression. (b) To develop an imaging window that incorporates the micro scaffolds, also transparent, oxygen-permeable, biocompatible and implantable in vivo (c) To create an experimental model of cancer fibrotic stiffening in vivo, in the chorioallantoic membrane of an embryonated avian egg. (d) To monitor the fibrotic progression of the tumour micro environments in vivo, in real time. (e) To predict mass transport in the tumour micro environments. (f) To validate the platform, in its ability to reproduce on breast tumours with different fibrotic progression the effect of anticancer drugs, both approved and investigational.
At the end of the second project year, we achieved the following results.
We want to recapitulate progressive stages of fibrosis by varying the geometrical features of a 3D micro scaffold. The geometry we chose is a 3D micro lattice with variable spatial arrangement into several repeated levels, with pores on each level of variable dimension. Upon implantation in a living chicken embryo, the scaffold geometry was able to modulate the infiltration in the 3D scaffold spaces by the host’s vascular, stromal and immune cells involved in the foreign-body reaction to the synthetic structure.
We fabricated the scaffolds in a biocompatible polymer by a micro-stereolithography technique called two-photon laser polymerisation (2PP). We fabricated ultra-precise micro scaffolds of maximum 100 microns in thickness, at a spatial resolution down to 300 nm. We integrated the micro scaffolds in an imaging window, able to directly interface the micro scaffolds with the avian chorioallantoic membrane in vivo. We structured the micro scaffolds on glass coverslips.
We selected human breast cancer cell lines carrying specific mutations and with genomic similarities to metastatic breast cancer patient samples. Our primary study model is infiltrating ductal carcinoma. We plan to use the same cell lines for all the project tasks in vitro, in vivo, and for the validation tasks foreseen for the platform. We selected cell lines according to clinical markers of prognosis that are estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2).
In the first half of the project, we obtained a working prototype of the platform that we will use for the in vivo quantifications in the second half of the project.
We want to set-up computational models of individual tumour micro environments, in a network with all the tumour micro environments of the 3D array. In the second project year, we started with modelling the individual tumour microenvironments. In each tumour micro environment, we assumed boundary conditions read from the microscopy images.
We set the main simulation parameters based on actual measurements of vascularity, collagen density and orientation, and blood velocity taken on the microscope images. Our preliminary results show a slower diffusion of Doxorubicin, one of the chemotherapy agents that we want to test in the platform, in function of increasing levels of collagen content and vascularisation of the microenvironments.
We want to recapitulate progressive stages of fibrosis by varying the geometrical features of a 3D micro scaffold. The geometry we chose is a 3D micro lattice with variable spatial arrangement into several repeated levels, with pores on each level of variable dimension. Upon implantation in a living chicken embryo, the scaffold geometry was able to modulate the infiltration in the 3D scaffold spaces by the host’s vascular, stromal and immune cells involved in the foreign-body reaction to the synthetic structure.
We fabricated the scaffolds in a biocompatible polymer by a micro-stereolithography technique called two-photon laser polymerisation (2PP). We fabricated ultra-precise micro scaffolds of maximum 100 microns in thickness, at a spatial resolution down to 300 nm. We integrated the micro scaffolds in an imaging window, able to directly interface the micro scaffolds with the avian chorioallantoic membrane in vivo. We structured the micro scaffolds on glass coverslips.
We selected human breast cancer cell lines carrying specific mutations and with genomic similarities to metastatic breast cancer patient samples. Our primary study model is infiltrating ductal carcinoma. We plan to use the same cell lines for all the project tasks in vitro, in vivo, and for the validation tasks foreseen for the platform. We selected cell lines according to clinical markers of prognosis that are estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2).
In the first half of the project, we obtained a working prototype of the platform that we will use for the in vivo quantifications in the second half of the project.
We want to set-up computational models of individual tumour micro environments, in a network with all the tumour micro environments of the 3D array. In the second project year, we started with modelling the individual tumour microenvironments. In each tumour micro environment, we assumed boundary conditions read from the microscopy images.
We set the main simulation parameters based on actual measurements of vascularity, collagen density and orientation, and blood velocity taken on the microscope images. Our preliminary results show a slower diffusion of Doxorubicin, one of the chemotherapy agents that we want to test in the platform, in function of increasing levels of collagen content and vascularisation of the microenvironments.
During the first half of the project, we experienced significant developments.
One was the discovery of an anti-cancer mechanism for statins, that emerged from a database analysis that we conducted in the aim to find drugs approved for other applications, which could be repurposed to treat breast cancer.
Another one was the invention of an artificial egg shell that constitutes part of the platform planned in this project. I managed to find a start-up company interested in testing their product on our platform using the protocols developed in this project. Thanks to this connection, I submitted an ERC PoC proposal to investigate the use of our platform on their product, as a first step for a valorisation strategy for our invention.
Our finding that the micro scaffolds condition stromal stem cells towards maintenance of all the main aspect of their function, due to massive deregulation of around eight hundred genes related to their mechanics, advances the field of cell culture significantly beyond the state of the art. This finding drove me by curiosity to test the effect of our micro scaffolds also on stem cells that are not considered adherent. I started a collaboration with a group that tested the micro scaffolds to expand human hematopoietic stem and progenitor cells (HSPC), also in a gene-edited version designed to be used as advanced therapy medicinal product on patients, with impressive results.
Another significant breakthrough concerns the synthetic eggshell platform developed and patented within this project. Stimulated by a group of colleagues, we discovered that such synthetic eggshell can be used to monitor chick embryonic development, and to test the safety and efficacy of drugs, not only in fluorescence microscopy, as studied in this project, but also using other non-invasive diagnostic methods.
One was the discovery of an anti-cancer mechanism for statins, that emerged from a database analysis that we conducted in the aim to find drugs approved for other applications, which could be repurposed to treat breast cancer.
Another one was the invention of an artificial egg shell that constitutes part of the platform planned in this project. I managed to find a start-up company interested in testing their product on our platform using the protocols developed in this project. Thanks to this connection, I submitted an ERC PoC proposal to investigate the use of our platform on their product, as a first step for a valorisation strategy for our invention.
Our finding that the micro scaffolds condition stromal stem cells towards maintenance of all the main aspect of their function, due to massive deregulation of around eight hundred genes related to their mechanics, advances the field of cell culture significantly beyond the state of the art. This finding drove me by curiosity to test the effect of our micro scaffolds also on stem cells that are not considered adherent. I started a collaboration with a group that tested the micro scaffolds to expand human hematopoietic stem and progenitor cells (HSPC), also in a gene-edited version designed to be used as advanced therapy medicinal product on patients, with impressive results.
Another significant breakthrough concerns the synthetic eggshell platform developed and patented within this project. Stimulated by a group of colleagues, we discovered that such synthetic eggshell can be used to monitor chick embryonic development, and to test the safety and efficacy of drugs, not only in fluorescence microscopy, as studied in this project, but also using other non-invasive diagnostic methods.