Fabrication of antibody functionalized silk fibroin micro-well arrays using reactive inkjet printing for circulating tumour cell capture

癌症是一组由细胞异常生长引起的疾病，并可能扩散到身体的其他部位。它是全世界人类健康的主要威胁，死亡人数达820万（2012年）。在英国，每年有30万新的癌症患者患有200多种癌症，其中乳腺癌，肺癌，结肠癌和前列腺癌是最常见的类型。一些癌症，如前列腺癌，没有早期症状，使预后非常困难。因此，非常需要用于早期检测癌症的有效工具，并且可以大大增加成功治疗的机会或延长寿命。循环肿瘤细胞（CTC）是从原发性肿瘤脱落并在血流中循环的细胞种子，随后可以生长成其他肿瘤。CTC的分离和表征已被证明在癌症预后中是有效的。然而，由于血液中CTC的显著低丰度（在含有数百万个白色和红细胞的毫升血液中有几个到几十个），该任务极具挑战性。目前，CellSearch是唯一一种FDA批准的CTC捕获设备，需要特定的设备来进行分析，并且不能区分不同类型的癌症。此外，需要将滤膜上捕获的CTC转移到细胞培养板中进行计数/表征。因此，迫切需要开发高效的高通量筛查设备/方法，可以在一次测试中以优异的灵敏度筛查多种癌症。拟议的研究设想了这一具有挑战性的任务和迫切需求，试图利用新兴的增材制造技术（反应喷墨打印，RIJ）和FDA批准的生物材料（再生丝纤蛋白，RSF）用于制造用于捕获和分选CTC的抗体功能化微孔阵列。RIJ的优点不仅是计算机辅助设计（CAD），其在预定位置提供皮升（1 e-12升）油墨的精确递送，允许制造复杂的微阵列，而且还提供不同抗体油墨的交替递送（通过不同的打印头），其允许用不同的癌症生物标志物抗体功能化不同的微孔。制造的设备不仅使我们能够有效地捕获罕见的CTC，而且还使我们能够区分不同类型的癌细胞，为有效的癌症诊断提供了一种有前途的工具。该项目还扩展了采用新兴增材制造技术的知识和技能，以实现高价值产品的成本效益制造。因此，该项目与EPSRC的医疗保健技术（新型诊断工具）和未来制造的优先主题保持一致，并将对癌症诊断和医疗器械制造业产生重大影响。

Cancer is a group of diseases caused by the abnormal growth of cells and can potentially spread to other parts of the body. It has been the major threat to human health worldwide, counting for 8.2 million deaths (2012). In UK, there are 0.3 million new cancer patients every year with more than 200 types of cancers of which breast, lung, colon and prostate cancers are the most common types. Some of the cancers such as prostate cancer have no early symptoms making the prognosis extremely difficult. Therefore, effective tools for early detection of cancer are highly desired and can greatly increase the chances for successful treatment or extend the lifetime. Circulating tumour cells (CTCs) are cell seeds that have shed from a primary tumour and circulate in the blood stream and can subsequent growth into additional tumours. Isolation and characterization of CTCs have been proved to be effective in cancer prognosis. However, this task is extremely challenging due to the significant low abundance of CTCs in blood (a few to tens in a millilitre of blood which contains millions of white and red blood cells). Currently, CellSearch is the only FDA approved CTC capture device that require specific equipment to perform the analysis and are not able to discriminate from different types of cancers. Furthermore, the captured CTCs on the filter membrane need to be transferred to a cell culture plate for enumeration/characterization. Hence, there is an urgent need to develop efficient high throughput screen devices/methods that can screen multiple cancers in one test with excellent sensitivity.The proposed research, envisaging this challenging task and urgent needs, seeks to harness the emerging additive manufacturing technology (reactive inkjet printing, RIJ) and the FDA approved biomaterial (regenerated silk fibroin, RSF) for the fabrication of antibody functionalized micro-well arrays for the capture and sorting of CTCs. The advantages of RIJ are not only the computer assisted design (CAD) that offers the precise delivery of pico-litres (1e-12 litre) of ink at predetermined locations allowing the fabrication of complex micro arrays but also the alternate delivery of different antibody inks (through different print heads) that allows the functionalization of different micro-wells with different cancer biomarker antibodies. The fabricated devices will not only allow us to efficiently capture the rare CTCs but also allow us to discriminate different types of cancer cells, providing a promising tool for efficient cancer diagnosis. The project also extends the knowledge and skills of employing emerging additive manufacturing technology for cost-effective manufacturing of high value products. Therefore, this project firmly aligns with EPSRC priority themes of Healthcare technologies (novel diagnostic tools) and Manufacturing the future and will have significant impact to cancer diagnosis and medical device manufacturing industry.