Kamali Dashtarzheneh, Ashkan; (2018) Development of 3D cancer models and targeted fluorescent gold nanoclusters. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

The aim of this thesis was to study and develop suitable natural binary hydrogels to generate and isolate 3D spheroids from cancer cell lines (HT29 & MCF7) initially grown on 2D surfaces. The isolated spheroids were then used for fluorescent gold nanocluster (AuNCs) targeting studies. The major novel findings of this work are: 1. The low melting temperature agar in combination with either collagen or egg albumen were slow in generating spheroids and at times inhibited their formation. Isolating any formed spheroids from the preparation was difficult with disruption and loss of the cluster formation. This combination format was found to be unsuitable in the study. 2. The natural macromolecule based carboxymethyl cellulose in combination with gelatine as a gel, gave the ideal outcome in generating spheroids in a short time. Its porous structure can be fine-tuned by varying the composition ratio of CMC and gelatine and autoclaving the mixture at 121oC. The swelling capacity and gelling properties of the CMC+gelatine mixture was ideal for generating cancer cell spheroids and harvesting by low centrifugation without destroying the clusters. 3. Spheroids greater than 200µm in diameter displayed hypoxic environment at the core detected using hypoxic fluorescent probes. 4. A novel specific cancer marker calreticulin was detected at increased levels on the surface membranes of spheroids using fluorescent gold nanoclusters conjugated to anti-calreticulin antibodies. 5. Specificity of anti-calreticulin antibodies via translocation for surface calreticulin was established by testing in a 3D co-culture of preformed endothelial cells (HUVECs) on membrane inserts with harvested fixed spheroids at the basolateral surface. 6. The obtained results suggest that the porous natural polymer carboxymethyl cellulose in combination with gelatine shows great promise in generating harvestable cancer cell spheroids reminiscent of tumours in vivo at low cost, high efficiency and robustness.

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