In many ways the most significant bioscience advances rely not only on the design and discovery of new materials chemistries but on how these are arranged in 3D space to interact with a 3D cellular system. Less obvious than the science but equally critical to creating new medical solutions is the ability to work with and listen to others who understand the biological and medical challenges at hand. These others have the ability to identify the most useful bits of this tsunami of knowledge that can be diverted into selected medical challenges.
As we work together to find the technical solution, it must be something that is scalable and deployable, something that meets all of the commercial and economic requirements to be sustainable. This translation to scaling, manufacture and deployment will depend on the availability of records maintained under an appropriate quality management system. These are processes alien to the ingrained thinking of most traditional researchers. This progress requires a change in environment and culture, and is evoking a change in mindset.
We can explore this translational journey through the lens of some of our recent projects.
The Axcelda system was developed to regenerate cartilage in defects in the knee. This collaboration was initiated through interactions with Professor Peter Choong, an orthopaedic surgeon at St Vincent’s Hospital in Melbourne. The team has now grown to include Associate Professor Claudia Di Bella (surgeon) and an amazing research team drawn from researchers at the universities of Wollongong and Melbourne, Swinburne University of Technology and RMIT University.
Hyaline cartilage is characterised by low cell numbers and consists of an avascular, non-neural aliphatic environment. The distinct collagen fibre structure confers appropriate mechanical properties. The regeneration of cartilage remains a challenge. The use of an approved clinical product failed in more than 60% of patients. This inability to effectively repair cartilage leads to the development of osteoarthritis, a condition that contributes greatly to healthcare expenditure.
We have developed optimal ink formulations to protect adipose stem cells during 3D printing and to ensure differentiation into cartilage. As this project progressed, the delivery (printing system) evolved to meet the needs of the bioink properties and the structural requirements of the print. Innovative approaches to stem cell isolation from the fat pad below the kneecap have been developed, enabling a rapid turnaround time relevant to clinical deployment. Being a trailblazer, to deploy a 3D bioprinting strategy in clinic for cartilage regeneration, this project extends back over an extended time period during which we have learnt a lot.
In retrospect, a blueprint for the clinical deployment plan, developed in the early stages of our collaborative work would have accelerated translation. It would have helped forge the most appropriate research/experimental pathway.
Adhering to an appropriate quality management system to document progress and changes in that pathway (as we now have) would also have facilitated translation and strengthened our IP position and strategy at an earlier stage.
In terms of bioink formulations, simplest is best to enable effective scale-up. In choosing the biomaterial components for bioinks, consideration of issues such as storage conditions required, sterilisation and packaging for deployment need to be considered. Of course, the use of biomaterials already approved for implantation will help in navigating the regulatory pathway.
We also did some things right! Building an interdisciplinary research team that brought together the right mix of skills and personalities has been critical to our success to date. In interdisciplinary research with a view to translation, the composition of the team needs to be dynamic and has a critical temporal dimension. In this case, perhaps injection of regulatory and commercial expertise earlier in the journey would have positioned the project a bit better for translation – you live, you experiment, you adjust, you learn.
3D printed ears
We are endeavouring to 3D print prosthetic and living ears to treat patients with microtia. This collaborative project has clinical input from Professor Payal Mukherjee, an ENT surgeon at the Royal Prince Alfred Hospital in Sydney and a prosthetist, Sophie Fleming. The research team comprises researchers with expertise in imaging, mechatronic engineers to design and build printers, materials scientists and biologists.
Bioprinting is an emerging technology that integrates cellular delivery while ensuring structural support is pre-ordained through 3D-printed scaffolds that are personalised to the patient’s needs. The final outcome is to have a specialised 3D-printed bioactive implant customised to a patient’s biology and anatomy, so that the scaffold material in the implant will eventually be absorbed within the body, leaving behind the patient’s own tissue-engineered cartilage in the shape of their own anatomy.
The need for prosthetic ears
Worldwide, about one in 5000 babies are born with microtia/anotia, a congenital birth defect in which the external ear framework is poorly developed or completely missing. This condition is mainly treated by reconstructive surgery or external prostheses, with the latter being favoured significantly. Prosthetic reconstruction is less intensive and expensive, and provides a more aesthetic shape of the natural ear than surgery. Currently, most prosthetics are manufactured by a method that requires a highly trained prosthetist and large time investments in addition to high processing cost. 3D printing has been extensively used in various fields, most notably the biomedical domain to overcome the limitations of conventional manufacturing techniques. With this technique, complex structures can be produced more quickly and the production technology is deployable, enabling service in remote areas.
We have demonstrated for the first time the development of a portable and low-cost 3D printing system for the manufacturing of complex structures from different grades of silicone materials.
This approach allows an ear to be scanned by a handheld scanner, mirrored in virtual space and reproduced symmetrically in polydimethylsiloxane (PDMS). The rheological properties of PDMS were modified without damaging the inherent properties to make a suitable ink for 3D printing complex structures. This system can be adapted to 3D print many types of body shapes besides facial prosthetics through minor hardware modifications.
Our system was demonstrated to be suitable for fabricating facial prostheses. However, improvements in both hardware and software in order to 3D print prosthetics in larger dimensions and higher resolution are required. Broader conversing with the prosthetic community earlier in the project would have resulted in very useful inputs towards the functional requirements and printer design.
Scanning ears to obtain 3D images using mobile phones or handheld scanners is a procedure that has a high error rate with limited resolution. Developing an efficient mobile application and computer software, as well as creating a standard protocol for ear scanning that clinicians, and preferably the patients, can perform with high accuracy and repeatability is a must. Using machine-learning techniques in this field can be very helpful. However, many 3D scans are required to be taken from different participants for this purpose. Building links with machine-learning experts earlier would have meant many images in the correct format could have been obtained.
The need for bioactive implants
In the head and neck region, elastic cartilage is key to the skeletal support of the ear, nose and throat, which are also complex 3D shapes. Elastic cartilage has limited self-regenerative capacity. Repair and reconstruction is therefore challenged by limited donor sites in the body and donor site morbidity. Alloplastic implants (which use synthetic material) can avoid donor site morbidity, shorten surgery time and improve size and contour matching, but reported rates of fracture and exposure are high. Autologous tissue reconstruction is therefore considered superior. Tissue engineering on its own has had little clinical impact in the head and neck, as solutions do not address the anatomical 3D complexity of the face.
Using 3D printing techniques, we have solved the engineering complexities needed to address the 3D shape demands and mechanical properties of the cartilage implants, using polycaprolactone-hydrogel scaffolds. We have overcome several translational milestones. We have:
- created a customised bioprinter (3D Alek)
- validated a novel technique in bioprinting that allows the scaffolds and desired cells to be co-printed without compromising cell viability
- confirmed through in vitro studies that cell survival and differentiation are maintained when both cells and scaffold are printed in this manner
- delivered 3D Alek into a clinical environment at the Royal Prince Alfred Hospital
- performed the first sheep trial for scaffold assessment in microtia, using polycaprolactone-hydrogel scaffolds printed through 3D Alek
- studied the impact of mesenchymal stem cells (the precursors of structural cell types) by co-culturing them with donor cells able to make cartilage.
Our collaborative efforts to date have enabled progress from discovery to animal trials in four years.
Effective communication between engineers designing the prototype printer and biologists printing the scaffold cannot be underestimated. Throughout the project, several modifications of the bioprinting process were required to ensure structural integrity of the framework and viability of the cells. However, this was not discussed immediately with the engineers, resulting in multiple iterations that severely delayed crucial experiments. It is also important to ensure successful transition between team members leaving or joining the team, to avoid miscommunication or a knowledge gap.
Similar to the Axcelda project, an appropriate quality management system should have been implemented very early on in the project. This would have facilitated translation and an understanding of the challenges in the regulatory pathway.
Animal trials are one of the most crucial stages in gaining regulatory approval or commercialisation with the appropriate tissue model. This is expensive, time consuming, but has the lowest level of success in grant proposals. Including any animal trials in the initial developmental grants or prototypes will significantly avoid the ‘valley of death’ in funding in later stages.
An interdisciplinary research team with the right skills is everything to a project’s success. A project of this calibre requires a clinical mentor, a material scientist, a mechatronic/
software engineer and a biologist. The team needs to be agile in making changes at all stages of the project and have a full understanding of the final goal in translation.
3D printed skin
Using in operando 3D printing to facilitate skin regeneration, in particular, in people with burns, is a collaborative project with Professor Fiona Wood at the Royal Perth Hospital.
The need for prosthetic skin
Skin loss through burns and chronic conditions such as skin cancer, diabetic ulcers and genetic blistering diseases cause significant patient morbidity and mortality worldwide. In Australia, 200 000 people suffer burns annually, resulting in a total cost of more than $150 million. Half of hospital admissions are children. Chronic wounds are also a major contributor to mortality, morbidity and permanent disability worldwide. There are more than 400 000 chronic wound cases in Australia at any time, with an estimated direct healthcare cost of $2.85 billion per annum.
Despite progress on developing treatment methods to facilitate skin regeneration, the current gold standard treatment for full thickness wounds remains with skin autografting. This involves harvesting healthy skin from other body sites to graft onto wounds, with severe limitations such as insufficient donor sites in extensive burns cases and the creation of additional injuries with further risk of infection and scarring.
We have focused on developing bioink formulations, printing hardware and organotypic printing protocol for printing skin cells directly into wounds to produce full-thickness skin structures. For instance, we have developed biologically active ink formulations that support 3D printing dermal-like structures and subsequently promote organotypic interactions with skin epidermal keratinocytes to form full thickness skin-like structures. These include bioink formulations using a sulfate- and rhamnose-rich polysaccharide that resembles mammalian glycosaminoglycans being involved in wound healing and tissue matrix structure and function. These molecules are extracted from seaweed in a collaborative project with Dr Pia Winberg and Venus Shell Systems.
To improve the biological activities of ink formulation, we have developed bioinks based on human platelet lysate, a rich source of growth factors that play an important role in regulating the healing cascade. More recently, we have progressed to investigations into the role of different skin cell types in facilitating repair of full thickness skin defects in a preclinical pig model, and used the knowledge accrued to guide development of a clinical prototype 3D bioprinting platform for use in surgical theatre.
There are urgent needs for the development of technologies to improve the functionality of regenerated skin, such as vascularisation and innervation, as well as to reduce processing time consumption and production cost. Bioprinting brings unique opportunities to tackle these critical challenges; however, to realise its full potential, a number of technical hurdles must be overcome. These include the development of safe, effective and affordable stem cells to reduce reliance on human donors and reduce the risk of infections and thus chronic wounds. It is critical to develop a knowledge base on how bioink formulations interact with key skin cell types and sources, and how 3D-printed structures regulate self-assembly of these cell types to enable optimal tissue morphogenesis and further development of skin organ. The clinical and regulatory needs for bioink constituents should be considered at the early stage of bioink development. Last, but not least, there is an increasing appreciation of the need to develop intraoperative skin bioprinters that can print multiple biomaterial–cell combinations into large and complex wound defects at high speed and with high resolution.
The Translational Research Initiative for Cell Engineering and Printing (TRICEP) at the University of Wollongong is focused on developing materials synthesis protocols and bioink formulations as well as hardware manufacturing governed by an auditable quality management system that pertains to ISO accreditation. Attracting support for development and resourcing of such entities is challenging. The transition of mainstream researchers into such an environment is not without challenge and brings a new dimension to the level of collaboration required to ensure success. Facilities such as TRICEP are critical to this.
Positioning to translate
As we clamour to engage and develop more effective deployment of fundamental research findings, a new ecosystem is emerging in (pockets of) our research organisations. There is a growing recognition of the need to partner early to bring in the
non-technical skills required to translate research. There is also an acknowledgement that such skills will most likely not be found or retained within research organisations. There is a growing recognition of the need to train our researchers to be ‘technology jockeys’ – to take ideas to industries and drive the process of translation – dropping technology off at the door doesn’t work.
Encouraging industry into research laboratories will also help this process – not just for fleeting visits but to work alongside researchers – experience their journey and the complexities associated with it.
For effective translation in the areas discussed here, an insight into health economics (value-based care), regulatory issues and commercialisation pathways is essential. As an important element of this, positioning the project to create protectable intellectual property that will warrant investment is essential. IP portfolio needs to be developed in partnership with commercial partners. Universities do not have the expertise nor the resources to build portfolios across the whole gamut of university activities. All of these inputs are required early. They are no longer an afterthought in today’s research ecosystem.
We are well placed in Australia to tap into an amazing ecosystem, including Australian Research Council Centres of Excellence, Corporate Research Centres and Growth Centres. Of particular relevance here is MTP (medical technologies and pharmaceuticals) Connect. All provide access to individuals and teams committed to translation. We are also fortunate to have an amazing array of national facilities supported by the national collaborative Research Infrastructure Strategy (NCRIS). Of particular relevance here is the ANFF Materials Node, providing facilities and expertise for materials synthesis scale-up and access to advanced fabrication capabilities. We also have an exciting new opportunity to facilitate interactions of younger clinicians and researchers through the Beyond Science Program.
The research community in Australia can stand tall – we have listened, we have learnt, we have changed and we are ready to play our part in an emerging ecosystem that supports us in taking ideas to industries.
Thank you to all of our collaborators who continue to inspire us, provide incredible intellectual capital and energise through their enthusiasm.