Periodic Reporting for period 3 - 3DBrainStrom (Brain metastases: Deciphering tumor-stroma interactions in three dimensions for the rational design of nanomedicines)
Reporting period: 2022-04-01 to 2023-09-30
Brain metastases are one of the leading causes of mortality among patients with metastatic cancer, in particular, melanoma, breast and lung cancer. Despite significant breakthroughs in targeted therapies in the post-genomic era, survival rates of patients diagnosed with brain metastases remain poor1. Nowadays, discovery, development and evaluation of new therapies is performed on human cancer cells grown in “2D-culture research techniques” on rigid plastic plates followed by in vivo testing in immunodeficient mice. These experimental settings are lacking and constitute a fundamental hurdle for the translation of preclinical discoveries into clinical practice. The knowledge we acquired in my previous ERC Consolidator grant (POLYDORM) regarding the events involved in tumor initiation as well as our synthetic expertise in polymeric nanotechnology laid the foundation for this 3DBrainStrom proposal. We propose to establish state-of-the-art 3D-printed models of brain metastases (Aim 1), which include brain extracellular matrix, stroma and serum containing immune cells flowing in functional tumor vessels. Our unique models better capture the clinical physio-mechanical tissue properties, signaling pathways, hemodynamics, cellular heterogeneity, diverse cellular populations and drug responsiveness. Using our 3D-printed models, we aim to develop two new fronts for identifying novel clinically-relevant molecular markers (Aim 2) followed by the development of precision nanomedicines (Aim 3). We will exploit our vast experience in anticancer nanomedicines to design three therapeutic approaches that target various cellular compartments involved in brain metastases: 1) Prevention of brain metastatic dissemination and brain colonization using targeted nano-vaccines, which elicit antitumor immune response; 2) Intervention of tumor-brain stroma cells crosstalk when brain micrometastases establish; 3) Regression of macrometastatic disease by selectively targeting tumor cells. These approaches will materialize using our libraries of polymeric nanocarriers that selectively accumulate in tumors.
This research project will result in a paradigm shift by generating new preclinical cancer models that bridge the translational gap in cancer therapeutics. I believe that our unique multidisciplinary background2 position us in a perfect state to provide the much needed breakthrough in clinically-relevant anticancer therapies at the nano-scale. The insights and tumor-stroma-targeted nanomedicines developed here will pave the way for prediction of patient outcome, revolutionizing our perception of tumor modelling and consequently, the way we prevent and treat cancer.
This research project will result in a paradigm shift by generating new preclinical cancer models that bridge the translational gap in cancer therapeutics. I believe that our unique multidisciplinary background2 position us in a perfect state to provide the much needed breakthrough in clinically-relevant anticancer therapies at the nano-scale. The insights and tumor-stroma-targeted nanomedicines developed here will pave the way for prediction of patient outcome, revolutionizing our perception of tumor modelling and consequently, the way we prevent and treat cancer.
In the first reporting period we continued working on the establishment and characterization of 3D preclinical models of melanoma, breast and lung cancer brain metastases (Aim 1).
(a) Tumor spheroids- We created 3D spheroids from 131/4-5B1 melanoma cells with human cerebral microvascular endothelial cells (hCMEC/D3) and human astrocytes. The spheroids were used for invasion assays in matrigel and for internalization kinetics studies of our nanomedicines (Fig 1A).
(b) 3D-printed tumor models- One of our proposed bio-ink formulations is comprised of fibrinogen and thrombin, which form fibrin hydrogel. The stiffness of the hydrogel can be manipulated by changing the initial concentration of gelatin (7.5 15 or 30% w/v) to resemble the physiological stiffness of different tissues such as lung, brain or breast (Fig 1B). Bio-ink formulated with 15% gelatin, which shares the physiological stiffness of the brain, was used to print a 3D brain model (Fig 1C). Exploiting our fibrin hydrogel as bio-ink and the sacrificable pluronic hydrogel, we printed a tumor model composed of tumor cells, brain stroma, and a functional vascular network connected to a microfluidic pump (Fig 1D). The inner surface of the resulting vascular network was coated with endothelial cells, which successfully formed a vascular lumen (Fig 1E-F).
(a) Tumor spheroids- We created 3D spheroids from 131/4-5B1 melanoma cells with human cerebral microvascular endothelial cells (hCMEC/D3) and human astrocytes. The spheroids were used for invasion assays in matrigel and for internalization kinetics studies of our nanomedicines (Fig 1A).
(b) 3D-printed tumor models- One of our proposed bio-ink formulations is comprised of fibrinogen and thrombin, which form fibrin hydrogel. The stiffness of the hydrogel can be manipulated by changing the initial concentration of gelatin (7.5 15 or 30% w/v) to resemble the physiological stiffness of different tissues such as lung, brain or breast (Fig 1B). Bio-ink formulated with 15% gelatin, which shares the physiological stiffness of the brain, was used to print a 3D brain model (Fig 1C). Exploiting our fibrin hydrogel as bio-ink and the sacrificable pluronic hydrogel, we printed a tumor model composed of tumor cells, brain stroma, and a functional vascular network connected to a microfluidic pump (Fig 1D). The inner surface of the resulting vascular network was coated with endothelial cells, which successfully formed a vascular lumen (Fig 1E-F).
The 3DBrainStrom research program is directed to provide the scientific and clinical communities with new experimental tools and accurate 3D cancer models. We are fully aware of the complexity and highly multidisciplinary nature of this proposal. By complementing basic, translational and clinical approaches, we believe the proposed research will have an enormous impact on the whole field of cancer therapy. This proposal will potentially affect the lives of patients with brain metastasis through the development of much needed new efficacious and non-toxic nanomedicines delivered selectively to brain tumor and stroma cells. These smart nanomaterials are especially appealing due to their controlled polymer synthesis, advanced recognition moieties, inherent biocompatibility, biodegradability and structural diversity allowing tailor-made environmental-responsiveness. Besides the novel targeted nanotherapies designed and synthesized by us, the proposed research will hopefully reveal new and profound insights on key players in the brain metastatic milieu that will be further developed into new potential therapeutic applications. Our vision is that our 3D models combined with our nanomedicines will motivate the design of novel approaches to target anticancer agents to tumor cells and their stroma, treating cancer in new multimodal and synergistic ways tackling tumor-host interactions. By implementing our patient-specific 3D models into clinical practice, we can help physicians choose the right therapy for the right patient rapidly, efficiently and safely.