Growth factors: the journey continues

Growth Factors is approaching its 30th year of publication (in 2018) and during that period has been expertly guided by the commitment of its inaugural Editor-in-Chief Tony Burgess. When Tony penned his introduction to Growth Factors in 1988 (A. W. Burgess, Growth Factors: The Beginnings, Vol. 1;1-6, 1988) (Burgess, 1988) this area was at a formative, but exciting stage. In the 28 years since there has been unprecedented advances in the understanding of growth factors, their receptors and signaling machinery, and their application to human development and disease. It has also been a period of significant change in how scientific discoveries and data are published, including the number, quality, and role of these journals. Re-reading Tony’s article (and I would highly recommend to all that you do) highlights the key role of earlier biologists and biochemists in establishing the field. Their efforts to isolate and characterize these often elusive “activities” using cell-based assays spanned decades of painstaking work. This was then followed by the role of the biochemist/protein chemist to purify and concentrate the activity with the hope of gaining initial primary amino acid sequence data for further identification. At the onset of Growth Factors, a number of revolutions were about to impact biological sciences and transform the humble beginnings of this field.

Around 1988, biology/biochemistry was preparing to fully embrace a number of important technologies. One being monoclonal antibodies (mAbs), a discovery that gained Kohler and Milstein the Nobel prize in 1982, was revolutionizing the characterization and understanding of growth factors and receptors in general (Kohler & Milstein, 1975). This technological advance allowed an almost unambiguous identification of particular protein chains through immunoprecipitation and the new approach of Western blotting, and provided a renewable handle for biochemists to purify proteins through affinity chromatography (Williams & Barclay, 1988). While antisera would take a backseat to mAbs for the next 30 years as investigative reagents, the role of mAbs in human therapy would take another decade to overcome the hurdles of clinical application to become powerful therapeutic agents in their own right (Scott et al., 2012).

Around the same time, recombinant DNA technology was about to transform biology. The groundwork for this was developed in the 1970s. Fred Sanger and Alan Coulson described a rapid method for sequencing DNA using DNA polymerase (Sanger & Coulson, 1975; Sanger et al., 1977) and this was followed by a chemical sequencing method described by Allan Maxam and Walter Gilbert (Maxam & Gilbert, 1977). At a similar time-point, scientists led by Paul Berg, Stanley Cohen, and Herbert Boyer would devise ways to splice together different pieces of DNA using restriction enzymes, introduce genetic material from one organism into the genome of another, and then allow replication and expression of the protein. Despite significant resistance to the development of this technology due to concerns about cloning and mixing of DNA from different species (Grobstein, 1977), the benefits of gene technology were soon realized through the commercial production of recombinant protein growth factors such as human somatostatin and insulin in the late 1970s. Human insulin (Humulin) was the first biotechnology product to appear on the market, made by the then new biotechnology company Genentech, and was approved for use by the FDA in 1982. Gene technology was also taking a firm hold in research laboratories, and the so-called “Molecular Biologists” or “Gene Jockeys” were fast becoming the pivotal research personal, as individual genes were being sequenced, and new genes being discovered at an express rate due to homology with preexisting genes. The rapid growth in availability of primary amino acid sequence data from proteins purified by affinity chromatography with the aid of mAbs, or through new expression cloning approaches (Aruffo & Seed, 1987), was a key driver to this process.

A good example of the rapid expansion of knowledge during this time comes directly from the area of growth factors and their receptors. Andrew Wilks used polymerase chain reaction (PCR), another exciting new technology at the time, to devise a method to amplify novel members of the receptor tyrosine kinase family through a system of degenerate oligonucleotide primers which would amplify sequences of related genes from mRNA extracted from specific tissue sources (e.g. colorectal cancer tissue). This approach identified a range of novel cell surface receptors and kinases located in the cytoplasm including the JAK family (Wilks, 1989). Approaches like this, combined with a growth in computing and automation of DNA sequencing and bioinformatics, quickly revealed unknown members in families of proteins long thought to have been completely described (Robinson et al., 2000). Interestingly, in 1988 the US Government funded both the NIH and the Department of Energy to “coordinate research and technical activities related to the human genome”, with the initial planning phase finalized soon after in 1990. This “human genome” project, completed around the turn of the new millennia, finally brought us close to the complete list of expressed proteins encoded by human genes (Venter et al., 2001).

A significant transformation of our understanding of cells and their specialized and coordinated function in tissues is the concept of signal transduction. This was an area that was in its infancy during the 1980s but that matured greatly due to the ability to identify and study novel proteins, driven by the technology advances. Signal transduction is fundamental to understanding how the binding of an extracellular growth factor to a cell surface receptor can ultimately trigger a cascade that involves a myriad of intermediates, chemical modifications and physical adjustments which then interplay with other signaling systems and genetic material to drive cellular outcomes (Ullrich & Schlessinger, 1990). The understanding of signal transduction is epitomized by the work of scientists like Tony Hunter who discovered that the addition of phosphate groups to tyrosine residues in protein chains was a key mechanism to drive protein activation and connection to downstream signaling cascades (Hanks et al., 1988; Hunter, 1987; Sefton et al., 1980). This work would provide insight into key human diseases such as cancer where gene mutations that caused constitutive phosphorylation would leave a protein in a state of constant activation driving the transformed phenotype of the cancer cells. Others such as Tony Pawson defined the prototypic modules that proteins use for interaction with each other, e.g. the so-called phosphotyrosine-binding Src homology domain (SH2 domain) that is employed by multiple different protein families for protein:protein interactions (Sadowski et al., 1986). Since this time, a plethora of different protein domains have been defined, allowing the connections between proteins and signal transduction pathways to be established and the question of signaling networks to be fully realized (Pawson, 1995; Pawson & Nash, 2003).

The need to understand and dissect the physiological role of these growth factors and receptors in the whole animal saw the use of genetic modification techniques to generate transgenic mice or gene-targeting techniques to produce gene knockout mice by homologous recombination in embryonic stems cells (Thomas & Capecchi, 1987). These models gave unique insight into the individual contribution of genes and protein products, at times showing redundancy of activity within families as well as recognizing critical temporal and spatial requirements within the developing animal. These early studies have given rise to more sophisticated mouse models where genes are deleted in a tissue- or cell-specific manner to highlight precise roles for growth factors in cellular systems (Gu et al., 1993). More recently the Clustered regularly interspaced short palindromic repeats (CRISPR/Cas9) interference technique for editing the germline of cells or animals through guided RNA has revolutionized the methods for rapid modification of the genome (Sander & Joung, 2014).

While the generic term “Growth Factor” implies effects on the proliferation of a cell, this perhaps more reflects the readouts of initial assays of the time which were designed to monitor important parameters such as increased cell number, mitotic index, or survival (Burgess, 1988; Morstyn & Burgess, 1988). Importantly many growth factors have been shown over the past three decades to have quite diverse roles, often including driving migration, chemotaxis, localization, and differentiation (Tammela et al., 2005). Therefore, molecules often discovered and named according to their initial activity have evolved by name and function. An example is the aptly named vascular permeability factor (VPF) observed by Harold Dvorak in the ascites of cancer patients (Senger et al., 1983). Its potent vascular permeability activity was a dramatic feature of its first described properties. When cloned and further biochemically characterized in the late 1980s, its role as an endothelial mitogen was revealed and its potential to drive angiogenesis (the formation of blood vessels) was demonstrated. This finding was highly relevant to Judah Folkman’s theory of anti-angiogenesis treatment for cancer (Folkman, 1972). This molecule was subsequently renamed as vascular endothelial growth factor (VEGF) and was seen as the centerpiece to signaling required for the growth and development of vascular structures. Work led by Kari Alitalo (Joukov et al., 1996), Ulf Eriksson (Olofsson et al., 1996), Salvatore Oliviero (Orlandini et al., 1996), and Marc Achen (Achen et al., 1998) showed the existence of a broader VEGF family of related factors encoded by separate genes, with a common cysteine-knot structural motif, also shared by the TGFβ family (McDonald & Hendrickson, 1993). This diversity of the VEGF family of growth factors and the receptors that interact with them is typical of how families of growth factors grew in number and complexity through advances in biology, biochemistry, genetics, and bioinformatics.

One of the major advances seen over the past three decades is the successful translation of growth factors and their inhibitors for clinical practice which was built on a foundation of knowledge generated by biologists, biochemists, and geneticists. Development of molecular modeling techniques, small molecule inhibitors and a greater understanding of how to manufacture and administer therapeutic mAbs have added substantially to the application of growth factors and their inhibitors to human disease. Some excellent examples of this are: the epidermal growth factor receptor, for which small molecule inhibitors to the kinase domain (gefitinib, erlotinib, lapatinib, and brigatinib) and mAbs (cetuximab and panitumumab) inhibiting the receptor–ligand interaction have been developed and are in widespread clinical use as anti-cancer agents (Burgess, 2008; Dancey & Sausville, 2003; Hynes & Lane, 2005; Imai & Takaoka, 2006; Yarden & Schlessinger, 1987); recombinant human granulocyte colony-stimulating factor (Filgrastim), a growth factor used clinically in cancer patients to accelerate their recovery from chemotherapy-induced neutropenia (Lieschke & Burgess, 1992a, 1992b), and in more recent times as we have learnt more about its mechanisms of action, for mobilization of blood stem cells for transplantation (Petit et al., 2002).

There are still many questions that need to be resolved about growth factors, both at a basic and a translational level. Some families of growth factors, receptors or signaling intermediates have been left in the “too hard basket” due to difficulties in working with the proteins, or poor immunogenicity leading to inadequate research tools. Some receptors lack classical means of activation and therefore will require greater examination to reveal their modus operandi. Additionally, there are still complex problems for the most well-known growth factors: diversity of expression, action and function, which are still being revealed, their role in an overall cellular network and the synergistic or antagonistic effects of other cellular outputs and signaling thresholds, etc. Systems biology, the study of systems of biological components such as molecules, cells, and organisms has now taken center stage as a way to quantify the interaction of the biological components that have been discovered through individual experimentation and large “omic” platforms (Kitano, 2002). How factors can be applied to tissues through gene delivery techniques or pathways corrected through genome editing are all longer-term questions that deserve our attention. The recent discovery of miRNA and the role of non-coding parts of the genome add to the complexity, as do understanding fully the implications of mutated or altered signaling pathways in diseases such as cancer. As a famous statesman once said “Now this is not the end. It is not even the beginning of the end. But it is perhaps the end of the beginning.” So we are now ready to launch into the next phase of more complex questions, which will give us deeper insight into the biology of the animal kingdom.

Over the past three decades, the model for scientific publishing has also been transformed. Print is giving way to electronic versions of journals, libraries are giving way to mobile devices, and journals are being diversified to cope with the increase of data in areas like bioinformatics, genomics and translational medicine. Open-access journals such as PLoS One have provided a different mechanism for publishing from diverse areas, and their role in science still needs time to be fully evaluated. While the number of scientific articles is still doubling approximately every nine years, it is difficult to determine whether this represents a real scientific advance or merely the splitting of larger studies into smaller segments as scientists respond to the “publish or perish” pressures for career preservation, as well as advancement, and for obtaining critical institutional, government, or grant funding. Methods to assess those parts of published science that sustain scientific knowledge and development are critical. While citations are a key measure of the interest in an individual paper, we are yet to generate universally accepted metrics for evaluating the quality of a journal, which would seem an important component of evaluating the worth of a paper in the early part of its lifecycle.

In the first edition of Growth Factors, Tony Burgess remarked that projects such as a new journal can only be judged over many years. That Growth Factors has stood the test of time and is still an important part of this scientific community now almost 30 years on, is a testament to the importance of the growth factor research field. I hope, with the support of Taylor & Francis and the Editorial team, to continue Tony’s virtuous work and maintain Growth Factors as an enduring and relevant part of the scientific literature.

Acknowledgements

Thank you to Margaret Hibbs and Marc Achen for their helpful input on the content and emphasis of the article. S.A.S. is supported by funding from the National Health and Medical Research Council of Australia.

Declaration of interest

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of this article.