Belle Creith, Dr Claire Clarkin, Professor Sumeet Mahajan and Professor Richard Oreffo 

University of Southampton 

Part funded PhD project (2019 - 2023)

BACKGROUND TO THE RESEARCH PROJECT

This research project aims to introduce a new technology for the diagnosis and examination of osteosarcoma tumours. Current diagnosis of osteosarcoma is a very time-consuming process involving a number of imaging techniques and a confirmative biopsy. 

Cutting-edge laser microscopes that allow the imaging of biological samples in a non-invasive manner are being widely explored in the field of cancer research as they do not require the use of any special dyes of labels. 

This research project utilises one such laser microscope – known as second harmonic generation microscopy – being used as a new method for the diagnosis of osteosarcoma by examining abnormal collagen – a protein in our bones  which is thought to be changed drastically within bone tumours. 

KEY RESULTS

During this research project, we developed an optimised imaging and analysis methodology that allowed accurate examination of collagen in human bone and osteosarcoma biopsies. Our findings provide proof-of-concept for osteosarcoma diagnosis by identifying abnormally short collagen fibres within osteosarcoma tumours compared to healthy bone. We also identified that these changes to collagen length become more pronounced with progression of osteosarcoma. Furthermore, we discovered that the collagen of osteosarcoma tumours also shows some differences from the collagen in other bone cancers. Together, our results demonstrate the diagnostic ability of SHG microscopes by identifying diseased collagen. This could help with both diagnosis of the disease as well as prediction of osteosarcoma progression and response to treatment. Moreover, our findings also highlights diseased collagen as a biological molecule that may be targeted in future osteosarcoma treatment. 

Figure: The collagen matrix of clinical healthy bone and primary bone cancer biopsies visualised by label-free SHG/TPEF imaging. SHG (green) and TPEF (blue) images were acquired of male: healthy bone (A), stage IA osteosarcoma (B), stage IA malignant giant-cell sarcoma (C) and stage IA chondrosarcoma (D) biopsies and overlaid for visualisation of the collagen matrix interface. Scale bars correspond to 200 µm (A-D), 40 µm (Ai-Di).

OUTPUTS & KNOWLEDGE AND FUTURE STEPS

Publications:

Hornsey T, Sharma A, Michels L, Kerns J , Clarkin CE (2024). Machine Learning predicts unique extracellular matrix characteristics between osteosarcoma cell phenotypes. Submitted for oral and poster presentation at the European Calcified Tissue Congress, Marseille, France, May 2024.

Sharma A, Oreffo ROC, Mahajan S, Beers S, Kanczler J and Clarkin CE (2024). Raman spectroscopy reveals the association between nanomolecular matrix signatures with pro-angiogenic potential in osteosarcoma. Accepted for poster presentation at the Bone Cancer Research Trust Meeting: Advancing diagnosis of bone and soft tissue sarcomas, Leeds, UK, January 2024.

Sharma A, Oreffo ROC, Mahajan S, Beers S, Kanczler J and Clarkin CE (2023). Characterisation of osteosarcoma matrix signatures reveals nanoscale molecular composition is linked to pro-angiogenic potential. Accepted for oral and poster presentation at the Bone Research Society Annual Meeting and European Calcified Tissue Society Congress 2023, Liverpool, UK, April 2023.

Sharma A, Oreffo ROC, Mahajan S, Beers S, Kanczler J & Clarkin CE (2022). Nanoscale characterisation of osteosarcoma cell matrix signatures link to pro-angiogenic potential. Accepted for poster presentation at the Gordon Research Seminar and Conference on Musculoskeletal Biology and Bioengineering: Multi-scale Approaches to Understanding Musculoskeletal Tissues, New Hampshire, USA, August 2022.

Presentations:

Creith B, Johnson PB, Harrison J, Oreffo ROC, Mahajan S and Clarkin CE (2021). Prospects of multi-modal non-linear microscopy for research and diagnosis of osteosarcoma. Accepted for oral presentation. Bone Research Society Annual Meeting (virtual program), 28-30 June.

Creith B, Johnson PB, Harrison J, Oreffo ROC, Mahajan S and Clarkin CE (2022). Characterisation of Diagnostic Collagen Phenotypes in Human Osteosarcoma Biopsies Using Second Harmonic Generation Imaging. Accepted for poster presentation. Bone Research Society Annual Meeting, Manchester, 6-8 July.

Creith B, Johnson PB, Harrison J, Oreffo ROC, Mahajan S and Clarkin CE (2022). Disclosing Diagnostic Collagen Phenotypes in Osteosarcoma Biopsies with Second Harmonic Generation Imaging. Accepted for oral presentation. Gordon Research Seminar on Musculoskeletal Biological and Bioengineering, New Hampshire, USA, 6-7 August.

Creith B, Johnson PB, Harrison J, Oreffo ROC, Mahajan S and Clarkin CE (2022). Disclosing Diagnostic Collagen Phenotypes in Osteosarcoma Biopsies with Second Harmonic Generation Imaging. Accepted for poster presentation. Gordon Research Conference on Musculoskeletal Biological and Bioengineering, New Hampshire, USA, 7-12 August.

Creith B, Johnson PB, Harrison J, Oreffo ROC, Mahajan S and Clarkin CE (2022). Prospects of Label-Free Microscopy for Research and Diagnosis of Osteosarcoma. Invited for Talk. North Bone  Cancer Research Trust Conference, Leeds, UK, 14-15 October.

Our future funding strategy will utilise the methodologies developed herein to form the basis of a larger grant application (Medical Research Council, Cancer Research UK, Bone Cancer Research Trust) with focus on improving osteosarcoma diagnosis and treatment using multidisciplinary approaches.