NCRC supported researchers at the Royal College of Surgeons in Ireland and colleagues at Johns Hopkins University, Texas Tech University, and Monash University have developed a 3D experimental model of neuroblastoma invasion and shown that the composition of the extracellular matrix influences the growth, viability, and local invasion of patient-derived neuroblastoma cells.
Neuroblastoma is childhood cancer caused by the abnormal growth and development of immature nerve cells (neuroblasts) of the sympathetic nervous system. This part of our nervous system is most commonly associated with our flight-fight response. Around 90% of neuroblastomas are seen in children under the age of 5. At the time of diagnosis, metastatic spread is seen in half of the patients with primary neuroblastomas, which are in a high-risk category. Despite recent advances in the treatment available to combat neuroblastoma, only 1 in 5 children with drug-resistant and/or recurrent neuroblastoma will survive.
One of the difficulties in studying neuroblastoma, and paediatric cancers in general, is the lack of experimental models that can recreate the local environment in which neuroblastoma cells will develop and grow. Typically, in the lab, neuroblastoma cells are grown as monolayers in 2D culture systems that don’t accurately recreate the 3D architecture of a solid tumour, where individual neuroblastoma cells will be exposed to different biochemical (nutrients, oxygen) and biophysical (stiffness and topography of the local environment) stimuli depending on their location within the tumour mass. 2D culture systems also result in cellular homogeneity (a population of cells with identical properties) while a tumour will be composed of a heterogenous mass of cancer cells with different properties (morphology, motility, gene expression, drug resistance and metastatic potential). To date, the vast majority of preclinical studies on neuroblastoma have relied on 2D experimental models and for the above reasons, have often failed to predict the clinical efficacy of targeted anti-cancer therapies used in clinical trials
In their newly published study, Cian Gavin, Dr Olga Piskareva, and colleagues have aimed to develop a 3D experimental model for studying the local tumour invasion in neuroblastoma. To do this. neuroblastoma organoids (tumour fragments of ~30-60 cells) were obtained from patient-derived xenografts (models of cancer where patient-derived neuroblastoma cells are implanted into and a mouse and allowed to develop into a tumour) and embedded in 3D Matrigel (a gelatinous protein mixture containing laminin, collagen, nidogen, heparan sulphate proteoglycan, and entactin) or collagen. Matrigel and collagen mimic the extracellular matrix (ECM), the non-cellular component of tissue that provides a scaffold for cells to grow on and regulates a number of important physiological process such as cell adhesion, growth, and migration. The team then used time lapse video-microscopy to study the behaviour of neuroblastoma cells in real time.
In 3D assays, neuroblastoma organoids displayed heterogenous morphologies with both non-invasive (cyst, spheroid) and invasive (collective mesenchymal, elongated, neuronal, protrusive) phenotypes identified. The composition of the ECM influenced the growth, migration, viability, and local invasion of the neuroblastoma organoids. Cell invasion was influenced by the addition of growth factors or the repression of prognostic biomarkers (MYCN gene) showing that the model is suitable for gene knockdowns/induction studies or to test the efficacy of anti-cancer agents. The combination of neuroblastoma organoids, real-time imaging, and the novel 3D culture assays developed in the study will enable rapid progress in elucidating the molecular mechanisms that control neuroblastoma invasion.
Figure 1: Time lapse microscopy video showing neuroblastoma organoids growing in 3D extracellular matrix cultures.
On publication of the study results, Dr. Olga Piskareva said “I am puzzled with the fact that half of the children with neuroblastoma have the disease spread at the time of diagnosis. Thanks to continuous support by NCRC and a Fulbright-HRB Health Impact Scholar Award, I travelled to Johns Hopkins University to adapt their 3D models to learn how neuroblastoma spreads. The data generated there was exciting and promising, so Cian (Gavin) spent almost a year systematising, characterising it, and placing it into a context. My team is now opening a new research question to understand cellular players behind neuroblastoma invasion and how we can target them to stop the spread. It won’t be a short and sweet journey, but we are ready for it!”
The paper was published in the journal “Cancers”. The complete publication can be found through the following link: https://www.mdpi.com/2072-6694/13/4/736
Cian Gavin, Nele Geerts, Brenton Cavanagh, Meagan Haynes, C. Patrick Reynolds, Daniela Loessner, Andrew J. Ewald and Olga Piskareva. Neuroblastoma invasion strategies are regulated by the extracellular matrix. Cancers 2021, 13(4), 736; DOI: 10.3390/cancers13040736