Category: Editorials

Introduction

The ‘scientific method’ begins with a hypothesis, which is the critical keystone in forming a well-designed study. As important as it is to ask the correct questions to form the hypothesis, it is equally important to be aware of the available tools to derive the answers.

Experimental models provide a crucial platform on which to interrogate cells, tissues, and even whole animals. They broadly serve two important purposes: investigation of biological mechanisms to understand diseases and the opportunity to perform preclinical trials of new therapies.

Here, an overview of experimental models based on animals commonly used in research is provided. Limitations which may impact clinical translation of findings from animal experiments are discussed, along with strategies to overcome this. Additionally, stem cells present a novel human-derived model, with great potential from both scientific and clinical viewpoints. These perspectives should draw attention to the incredible value of model systems in biomedical research, and provide an exciting view of future directions.

Animal models – a palette of choices

Animal models provide a ‘whole organism’ context in studying biological mechanisms, and are crucial in testing and optimising delivery of new therapies before the commencement of human studies. They may be commonly referred to under the classification of invertebrates (flies, worms) and vertebrates (fish, rodents, swine, primates); or small animal (fish, rodents) and large animal (swine, primates, sheep).

Whilst organisms have their own niche area of research, the most frequently used is the humble mouse. Its prominence is attested by the fact that it was only the second mammalian species after humans to have its genome sequenced, demonstrating that both species share 99% of their genes. [1] Reasons for the popularity of mice as a choice include that mice share many anatomical and physiological similarities with humans. Other advantages include that they are small, hardy, cheap to maintain and easy to breed with a short lifespan (approximately three years), [2] allowing experiments to gather results more quickly. Common human diseases such as diabetes, heart disease, and cancer affect mice, [3] hence complex pathophysiological mechanisms such as angiogenesis and metastasis can be readily demonstrated. [2] Above all, the extraordinary ease with which mice are manipulated has resulted in the widespread availability of inbred, mutant, knockout, transgenic or chimeric mice for almost every purpose conceivable. [3] By blocking or stimulating the overexpression of specific genes, their role in developmental biology and disease can be identified and even demonstrated in specific organs. [4]

Humanised mice are another step closer in representation of what happens in the human body, thereby increasing the clinical value of knowledge gained from experiments. Humanised mice contain either human genes or tissue allowing the investigation of human mechanisms whilst maintaining an in vivo context within the animal. Such approaches are also available in other organisms such as rats, but are often adapted from initial advances in mice, and hardly mirror the ease and diversity with which humanised mice are produced.

Aside from the mouse, invertebrates such as the Drosophila vinegar fly [5] and Caenorhabditis elegans worm [6] are also widely used in research of genetics or developmental biology studies. They are particularly easy to maintain and breed and therefore large stocks can be kept. Furthermore, there are fewer ethical dilemmas and invertebrates have a genome simple enough to be investigated in its entirety without being cost-prohibitive or requiring an exhaustive set of experiments. Their anatomies are also distinct and simple, allowing developmental changes to be readily visualised.

Another alternative is the Zebrafish, which shares many of the advantages offered by Drosophila and C. elegans. Additionally, it offers greater scope for investigating more complex diseases like spinal cord injury and cancer, and possesses advanced anatomical structures as a vertebrate. [7] Given the inherent capacity of the Zebrafish for cardiac regeneration, it is also of interest in regenerative medicine as we seek to harness this mechanism for human therapy. [8]

Large animals tend to be prohibitively expensive, time-consuming to manage and difficult to manipulate for use in basic science research. Instead, they have earned their place in preclinical trials. Their relatively large size and physiological similarity to humans provides the opportunity to perform surgical procedures and other interventions on a scale similar to that used clinically. Disease models created in sheep or swine are representative of the complex biological interactions that are present in highly evolved mammals; hence may be suitable for vaccine discovery. [9] Furthermore, transgenic manipulation is now possible in non-human primates, presenting an opportunity to develop humanised models. [10] Despite this, there are obvious limitations confining their use to specialised settings. Large animals need more space, are difficult to transport, require expert veterinary care, and their advanced psychosocial awareness raises ethical concerns. [9]

The clinical context of animal experimentation

A major issue directly relevant to clinicians is the predictive value of animal models. Put simply, how much of research using animals is actually clinically relevant? Although most medical therapies in use today were initially developed using animal models, it is also recognised that many animal experiments fail to reproduce their findings when translated into clinical trials. [11] The reasons for this are numerous, and require careful analysis.

The most obvious is that despite some similarities, animals are still animals and humans are humans. Genetic similarities between species as seemingly disparate as humans and mice may lead to assumptions of conserved function between humans and other animal species that are not necessarily correct. Whilst comparing genomes can indicate similarities between two species such studies are unable to capture differences in expression or function of a gene across species that may occur at a molecular level. [12]

The effectiveness and clinical relevance of experimental animal trials is further complicated by epigenetics. Epigenetics is the modification of genetic expression due to environmental or other cues without actual change in DNA sequence. [13] These changes are now considered just as central to the pathogenesis of cancer and other conditions as genetic mutations.

It is also important to consider the multi-factorial nature of human diseases. Temporal patterns such as asymptomatic or latent phases of disease can further complicate matters. Patients have co-morbidities, risk factors, and family history, all of which contribute to disease in a way that we may still not completely understand. With such complexity, animal models do not encapsulate the overall pathophysiology of human disease. Animals may be too young, too healthy, or too streamlined in sex or genetics. [14] To obtain animals with specific traits, they are often inbred such that two animals in the same experiment will have identical genetic make-up – like twins, hardly representative of the diversity present in nature. Understandably, it can be an extraordinary challenge to incorporate all these dimensions into one study. This is especially so when the very principles of scientific method dictate that variables except for the one under experimentation should be minimised as much as possible.

A second area of concern is the sub-optimal rigour and research design of animal experiments. Scientists who conduct animal experiments and clinicians who conduct clinical trials often have different goals and perspectives. Due to ethical and cost concerns, the sample size of animal experiments is often kept to a minimum, and studies are prolonged no more than necessary, often with arbitrarily determined end-points. [14] Randomisation, concealed allocation, and blinded outcome of assessment are poorly enforced, leading to concerns of experimental bias. [11] Additionally, scientific experiments are rarely repeated due to an emphasis on originality, whereas clinical trials are often repeated (sometimes as multi-centre trials) in order to assess reproducibility of results. Furthermore, clinical trials are more likely to be published regardless of the nature of results; in contrast, scientific experiments with negative findings or low statistical significance often fail to be reported. These gaps highlight the fact that preclinical trials should be expected to adhere to the same standards and principles of clinical trials in order to improve the translatability of results between the two settings.

Although deficiencies in research conduct is a concern, the fundamental issue that remains is that even the best-designed preclinical study cannot overcome the inherent differences that exist between animal models and ‘real’ human patients. However, it is reassuring to know that we are becoming better at manipulating animal models and enhancing their compatibility with their human counter-parts. As such, this drive towards increasingly sophisticated animal models will provide more detailed and clinically relevant answers. Additionally, with the recognition that a single animal model is inadequate on its own, experiments may be repeated in multiple models. Each model will provide a different perspective and lead to the formation of a more comprehensive and balanced conclusion. A suggested structure is to start initial proof-of-principle experiments in small, relatively inexpensive and easily manipulated animals, and then scale up to larger animal models.

‘Human’ experimental models – the revolution of stem cells

Given the intrinsic differences between animals and humans, it is crucial to develop experimental systems that simulate human biology as much as possible. Stem cells are ‘master cells’ with the potential to differentiate into more mature cells, and are involved in the development and maintenance of organs through all stages of life from an embryo (embryonic stem cells) to adult (tissue-specific stem cells). [15] With the discovery of human embryonic stem cells [16] and other tissue-specific stem cells [17] it is now possible to appreciate the developmental biology of human tissues and organs in the laboratory. Stem cells may be studied under various controlled conditions in a culture dish, or even implanted into an animal to recapitulate in vivo conditions. Furthermore, stem cell transplantation has been used in animal models of disease to replace lost or damaged tissue. These methods are now commencing high-profile clinical trials with both embryonic stem cells [18] and tissue-specific stem cells. [19] Although stem cells hold great potential, translating this into the clinical environment has been hindered by several obstacles. Chiefly, tissue- specific stem cells are rare and difficult to isolate, while embryonic stem cells can only be created by destroying an embryo. In order to generate personalised embryonic stem cells for cell therapy or disease modelling, they need to be created via ‘therapeutic cloning.’ The considerable ethical quandary associated with this resulted in a field mired in controversy and political debate. This led to research coming almost to a standstill. Fortunately, stem cell research was rejuvenated in 2007 with the revolutionary discovery of induced pluripotent stem (iPS) cells – a discovery notable enough to be awarded the 2012 Nobel Prize in Physiology/Medicine.

Induced pluripotent stem (iPS) cells are created by reprograming mature cells (such as skin fibroblasts) back into a pluripotent ‘stem cell’ state, which can then re-differentiate into cells of any of the three germ layers irrespective of what its original lineage was. [20] Cells from patients with various diseases can be re-programmed into iPS cells, examined and compared to cells from healthy individuals to understand disease mechanisms and identify therapeutic opportunities. Rather than using models created in animals, this approach represents a ‘complete’ model where all genes contributing to a specific disease are present. Crucially, this enables the previously inconceivable notion of deriving patient-specific ‘disease in a dish’ models, which could be used to test therapeutic response. [21] It also provides unprecedented insight into conditions such as those affecting the heart [22] or brain, [23] which have been difficult to study due to limitations accessing tissue specimens and conducting experiments in live patients.

However, if a model system rests purely on stem cells alone this would relegate the approach to in vitro analysis without the whole organism outlook that animal experiments afford us. Accordingly, by combining this with rapidly evolving cell transplantation techniques it is possible to derive stem-cell based animal models. Although this field is flourishing at an exponential rate it is still in its infancy. It remains to be seen how the actual translation of iPS technology will fit into the pharmacological industry, and whether personalised drug screening assays will become adopted clinically.

Conclusion

Experimental models provide us with insight into human biology in ways that are more detailed and innovative than ever before, with a dazzling array of choices now available. Although the limitations of animal models can be sobering, they remain highly relevant in biomedical research. Their contribution to clinical knowledge can be strengthened by refining models to mimic human biology as closely as possible, and by modifying research methods to include protocols similar to that used in clinical trials. Additionally, the emergence of stem cells has shifted current paradigms by introducing patient-specific models of human development and disease. However, it should not be seen as rendering animal models obsolete, but rather a complementary methodology that should improve the predictive power of preclinical experiments as a whole.

Understanding and awareness of these advances is imperative in becoming an effective researcher. By applying these models and maximising their potential, medical students, clinicians and scientists alike will enter a new frontier of scientific discovery.

Conflict of interest

None declared.

Correspondence

k.yap@amsj.org

Introduction

Growing in the world of academic medicine is a new generation of doctors known as “clinician-scientists”. Trained in both science and medicine, with post-graduate research qualifications in addition to their medical degree, they serve as an essential bridge between the laboratory and clinic.

The development of sophisticated experimental approaches has created opportunities to investigate clinical questions from a basic science perspective, often at a cellular and molecular level previously impossible. With new and detailed understanding of disease mechanisms, we are rapidly accelerating the discovery of new preventative measures, diagnostic tools, and importantly, novel therapeutic approaches. In these emerging avenues there is not just a need for collaboration between scientists and clinicians, but a need for individuals who are fluent in both science and medicine – hence, the advent of clinician-scientists. The terms “translational research” or “translational medicine” are often associated with clinician-scientists, alluding to the notion that these people facilitate the two-way process of translating scientific findings into clinical applications (bench-to-bedside), and provide clinical data and specimens back to the laboratory to investigate underlying disease processes (bedside-to-bench).

From a student’s perspective however, these concepts can be confusing and finding their way through the breadth and categories of research conducted in academic institutions and hospitals may prove daunting. A discussion of the clinician-scientist niche and some of the challenges and opportunities faced may prove helpful.

Defining the clinician-scientist

Most clinicians at an academic hospital are engaged in research to some extent, but this tends to be mainly clinically-oriented, with patient care, treatment outcomes, and population health being broad areas commonly involved. Their day-to-day job is mostly defined by their clinical duties, often with some teaching responsibilities involved. Clinician-scientists, by contrast, dedicate a significant proportion of their time to research, typically spending ≥50% protected time in order to be remain academically competitive. [1] Whilst still loosely defined, in a purist sense this is a clinician who is involved in research at an organ, tissue, cellular, or molecular level, as opposed to focussing solely on whole patients as a clinical subject. Such research may not always have clinical findings that are directly relevant to everyday medical practice but the difference from a pure basic scientist is that the science has been approached with clinical relevance in mind. Interestingly, on the other hand, science itself has become inter-disciplinary and is recognising the importance of clinical relevance and translation with new ventures such as the Stanford University PhD in Stem Cell Biology where graduate science students interested in involvement with translational research in regenerative medicine undertake rotations shadowing clinicians in order to develop a clinical perspective to their research. [2] These developments indicate that not only are the frontiers between science and medicine becoming blurred, but that translational research is the exciting intersection where clinician-scientists, as well as scientists well-attuned to clinical practice, are uniquely poised to thrive.

The clinician-scientist niche

Clinician-scientists possess a distinctive set of skills, being trained as a clinician to apply scientific knowledge to patient care, and trained as a scientist with an enquiring mind designed to test hypotheses. Understanding the clinical relevance of observations in science and the ability to translate this back into clinical practice is truly the domain of the clinician-scientist, and uniquely so.

The pursuit of additional post-graduate research qualification such as a Masters or PhD has traditionally been the main pathway to becoming a clinician-scientist in Australia, unlike in the United States where combined MD-PhD programs have been well established in the past. However, the recent development of similar combined MBBS-PhD and MD-PhD programs in Australia is likely be instrumental in building a body of clinician-scientists that have been moulded specifically for this task. [3] Skills developed in scientific training essential for success in research include literature appraisal, manuscript and grant writing, and mastery of laboratory techniques, all of which are life-long skills honed over time, and which are rarely acquired in medical school.

It goes without saying that clinician-scientists are expected to be experts in both medicine and science. Anything subpar of clinical competence would pose a threat to patient safety and cannot be compromised. On the other hand without a solid commitment in research with the appropriate output in terms of publications, conference attendance, and grant proposals, a career in research will not take off since a track record is something that needs to be built on constantly. Given that clinical training itself takes a good number of years before being able to practice as an independent clinician it is little wonder that many are unwilling to tackle both clinical and scientific careers at once. Again, this lends further credence to the MD-PhD path where scientific training would have already been completed by the end of the program, although this itself has its drawbacks, since the science gained can become neglected in the last clinical years and will need to be polished again upon completion. [4]

But where lie challenges also lie opportunities: for the determined few, funding statistics indicate that the rigorous training is entirely worthwhile. Clinician-scientists have been found to consistently perform better in national funding programs such as the National Institutes of Health Research Project Grants (United States) than their pure clinician (MD only) and basic science (PhD only) counterparts. [5] Although the pool of clinician-scientists in Australia is significantly smaller than that of the United States and data on funding trends are less widely discussed in literature, it is generally acknowledged that clinician-scientists also do well in obtaining NHMRC funding. This may be due partly to the fact that clinician-scientists are afforded more flexibility in labelling their projects as “basic science” or “clinical”, and therefore have access to funds for both basic science and clinical projects, whereas pure clinicians and scientists are generally limited to their own funding areas.

When describing the clinician-scientist niche, an aspect of research “translation” that is often neglected is the importance of the delivery of research-based medicine into actual practice. The classic bench-to-bedside process refers to the invention of a new drug, device, or diagnostic tool where the hope is that it will undergo clinical evaluation in a controlled setting with a specific patient cohort. But bringing a discovery into the market is simply the beginning, and to bring this to the general public a much more concerted effort is required involving collaboration between public health experts, policy makers, and clinicians amongst others. So drawn-out and complex is the process that it is well acknowledged that this area of “translational” research often fails, with many potentially important discoveries unable to make changes to everyday medical practice.

[6] However, clinician-scientists are well suited to play an active role in negotiating the many hurdles in this endeavour by facilitating communication between the various experts involved, whilst providing a first-hand inventor as well as treating clinician’s perspective that is not only unique but critical in ensuring that an invention is appropriately implemented and evaluated. In the Australian context, the National Health and Medical Research Council (NHMRC) has recognised this gap in research translation and the Centres for Research Excellence and Translating Research Into Practice (TRIP) Fellowships are specific measures aimed to address this issue. [7]

Wearing two hats: double the challenges?

A commonly quoted recommended research:non-research ratio for workload is 75:25, with the majority of time devoted to research in order to succeed as a clinician-scientist. [8] In reality this is more likely to be exactly opposite the case, where a 75:25 ratio in favour of clinical work becomes the norm instead. [9] This may be particularly so in the early years after graduation when specialist training is being undertaken, despite the fact that this is also the time when a solid research foundation needs to be built in order to establish a clinician-scientist’s academic presence. As pressing as clinical demands may be, it is widely recognised that a research career cannot flourish without negotiating some protected time from clinical duties with the hospital department.

The biggest challenge for clinician-scientists is therefore time management. In addition to patient care, clinical training, and teaching responsibilities, clinician-scientists are expected to undertake labwork, keep abreast of advances in both scientific and medical literature, and engage in professional development and conferences on both fronts. They must maintain manuscript preparation and grant proposals, complete administrative duties, and often lead research teams. To realistically keep up with these demands of juggling a dual career, the ability to delegate and seek cooperation from scientist and clinician colleagues is critical. The lack of a supportive environment and a suitable mentor who can share their experiences and show the way can present an impossible struggle to the time-constrained clinician-scientist.

On the clinical front, to manage their workload clinician-scientists may tightly focus their interests to subspecialised areas to maintain an adequate caseload and expertise without stretching oneself too thin. This depends however on working in an environment where the volume and diversity of patients permits such subspecialisation, with appropriate facilitation by supervisors such as Department Heads. Unfortunately these conditions tend to be found only in major tertiary hospitals, relegating clinician-scientists to these settings.

Additionally, a research career is often less financially rewarding than clinical work particularly when private practice may need to be sacrificed in order to undertake lab work. This can pose a significant barrier particularly because the number of years required to gain appropriate training results in clinician-scientists being likely to be older than their scientist and clinician counterparts and may therefore have family commitments, and have often also accumulated student debts that need to be repaid. [10] Some solutions to this may be the Practitioner and Career Development Fellowships offered by the NHMRC aimed at clinicians involved in research, [11] as well as hospital and philanthropic organisation funding specifically for buying time out from clinical practice for research.

Opportunities for the clinician-scientist

For any researcher, securing funding is a lifeline in continuing their work and burnishing a track record, and it is here where clinician-scientists can be creative in sourcing their benefactors. Philanthropic organisations often affiliated with a disease or clinical cause, specialist training colleges like the Royal Australasian College of Surgeons, hospital based foundations, pharmaceutical companies, and fundraising from patient advocates are all important and significant funding avenues that clinician-scientsts may find more accessible than pure scientists. [12] These grants often allow pilot projects to be undertaken in order to generate sufficient amount of preliminary data to become competitive for major research funding such as from the NHMRC. Additionally, a number of these organisations offer clinician-scientist fellowships similar to the NHMRC.

Apart from funding success, it has also been found that many clinician-scientists opt to apply for and are successful in obtaining university academic positions. [12,13] Such engagement in academia provides synergy for research efforts by opening up institutional resources often more diverse than hospital settings, prospects for networking with likeminded professionals and mentors.

Additionally, the scope translational research itself is widening. An increasing number of academic hospitals are dedicating departments to translational research, with clinician-scientists often taking the lead. The need to prioritise translational research has been further underlined by the Chief-Scientist of Australia’s recent speech calling for increase in research funding for this area. [14] Whilst these are positive developments, further input from clinician-scientists themselves is required to shape policy changes and design steps to increase their numbers.

Moving forward

An apt saying may be, “Clinicians know all of the problems, but none of the solutions; scientists know all of the solutions, but none of the problems”. [15] This is where clinicianscientists represent a unique breed suited to fulfil this vacant niche, and are absolutely necessary in forging the next success stories of medicine. Despite the complexities of a dual career, the rewards and satisfaction in pursuing this path are evident and meaningful, and can lead to tangible health outcomes in patients. Although it is important to maintain a realistic notion that being a clinician-scientist is by no means an easy feat, it is equally important to take hope that the best of both worlds can be experienced. These perspectives are increasingly acknowledged in the form of progresses being made in the right direction to encourage clinician-scientists. In light of this, perhaps it is well worth noting that there may never be a better time than now to venture into, and indeed take charge in riding this next wave of medical evolution.

References

[1] Archer SL. The making of a physician-scientist–the process has a pattern: lessons from the lives of Nobel laureates in medicine and physiology. Eur Heart J. 2007 Feb;28(4):510-2

[2] Stanford University School of Medicine, Institute for Stem Cell Biology and Regenerative Medicine. PhD Program: Curriculum Overview. [updated February 17th 2009; cited March 8th 2012]; Available from: http://stemcell.stanford.edu/education/phd/curriculum.html

[3] Power BD, White AJ, Sefton AJ. Research within a medical degree: the combined MB BS-PhD program at the University of Sydney. Med J Aust. 2003 Dec 1-15;179(11-12):614-6

[4] Marban E, Braunwald E. Training the clinician investigator. Circ Res. 2008 Oct 10;103(8):771-2

[5] Dickler HB, Fang D, Heinig SJ, Johnson E, Korn D. New physician-investigators receiving National Institutes of Health research project grants: a historical perspective on the “endangered species”. JAMA. 2007 Jun 13;297(22):2496-501

[6] Woolf SH. The meaning of translational research and why it matters. JAMA. 2008 Jan 9;299(2):211-3

[7] McCallum J, Forster R. From the NHMRC: Research translation network targets the evidence-practice lag. Med J Aust. 2011;195(5):252

[8] Tai IT. Developing a clinician-scientist career. Clin Invest Med. 2008;31(5):E300-1

[9] Bosse D, Milger K, Morty RE. Clinician-scientist trainee: a German perspective. Clin Invest Med. 2011;34(6):E324

[10] Lander B, Hanley GE, Atkinson-Grosjean J. Clinician-scientists in Canada: barriers to career entry and progress. PLoS One. 2010;5(10)

[11] National Health and Medical Research Council. Fellowship Awards. [Internet] [updated December 21st 2011; cited March 1st 2012]; Available from: http://www.nhmrc.gov.au/grants/apply-funding/fellowship-awards

[12] Hayward CP, Danoff D, Kennedy M, Lee AC, Brzezina S, Bond U. Clinician investigator training in Canada: a review. Clin Invest Med. 2011;34(4):E192

[13] Toouli J. Training surgeon scientists. ANZ J Surg. 2003 Aug;73(8):630-2

[14] Australian Government – Chief Scientist of Australia 2012. Can Australia afford to fund translational research? [updated April 3rd 2012]; Available from: http://www.chiefscientist.gov.au/2012/04/can-australia-afford-to-fund-translational-research/

[15] Hait WN. Translating research into clinical practice: deliberations from the American Association for Cancer Research. Clin Cancer Res. 2005 Jun 15;11(12):4275-7