Therapeutic Tissue Regeneration by a Macrophage Colony-Stimulating Factor Fc Conjugate
Authors: Jasper G van den Boorn, Gunther Hartmann
Journal: Molecular Therapy (2014)

Macrophage colony-stimulating factor 1 (CSF1) is a hematopoietic growth factor involved in the proliferation, differentiation, and survival of monocytes, macrophages, and bone marrow progenitor cells. Current clinical interest in CSF1 lies in relatively short-term treatments using recombinant protein for applications such as tissue regeneration. However, because the small size of the CSF1 protein ensures rapid renal clearance, a continuous intravenous application is needed to maintain an effective dose. In this issue of Molecular Therapy, Gow et al.1Gow DJ Sauter KA Pridans C Moffat L Sehgal A Stutchfield BM et al.Characterisation of a novel Fc conjugate of macrophage colony-stimulating factor.Mol Ther. 2014; 22: 1580-1592Abstract Full Text Full Text PDF PubMed Scopus (70) Google Scholar describe how they addressed this issue by developing an Fc conjugate of CSF1. They then go on to dissect its biological effects and capacity to induce tissue regeneration, showing that the conjugate augments the circulating half-life of the protein, which in turn stimulates the capacity of the mononuclear phagocyte system (MPS) to induce tissue regeneration in the liver. Although this provides us with an exceptional new therapeutic tool in situations in which liver homeostasis is compromised, caution is required before moving into clinical application, because of the possible effects on other organs and tissues. The MPS is a part of the immune system that is composed primarily of monocytes and macrophages that accumulate in lymph nodes and the spleen, and it includes the Kupffer cells of the liver. In many tissues, macrophages are derived from progenitor cells of fetal origin2Ginhoux F Greter M Leboeuf M Nandi S See P Gokhan S et al.Fate mapping analysis reveals that adult microglia derive from primitive macrophages.Science. 2010; 330: 841-845Crossref PubMed Scopus (3174) Google Scholar or, possibly, circulating monocytes,3van Furth R Cohn ZA The origin and kinetics of mononuclear phagocytes.J Exp Med. 1968; 128: 415-435Crossref PubMed Scopus (970) Google Scholar and these tissue-resident macrophages may self-renew by proliferation.4Davies LC Jenkins SJ Allen JE Taylor PR Tissue-resident macrophages.Nat Immunol. 2013; 14: 986-995Crossref PubMed Scopus (1265) Google Scholar During infection or inflammation there is generally an influx of monocytes that will also differentiate to become macrophages.3van Furth R Cohn ZA The origin and kinetics of mononuclear phagocytes.J Exp Med. 1968; 128: 415-435Crossref PubMed Scopus (970) Google Scholar CSF1 is essential to macrophage survival and function,5Hamilton JA Colony-stimulating factors in inflammation and autoimmunity.Nat Rev Immunol. 2008; 8: 533-544Crossref PubMed Scopus (979) Google Scholar and there are profound macrophage deficiencies in op/op mice carrying an inactive Csf1 gene.6Yoshida H Hayashi S-I Kunisada T Ogawa M Nishikawa S Okamura H et al.The murine mutation osteopetrosis is in the coding region of the macrophage colony stimulating factor gene.Nature. 1990; 345: 442-444Crossref PubMed Scopus (1518) Google Scholar,7Wiktor-Jedrzejczak WW Ahmed A Szczylik C Skelly RR Hematological characterization of congenital osteopetrosis in op/op mouse. Possible mechanism for abnormal macrophage differentiation.J Exp Med. 1982; 156: 1516-1527Crossref PubMed Scopus (270) Google Scholar Besides playing a vital role in innate immunity, the MPS is involved in other essential processes, such as the regulation of inflammation, scavenging of tissue debris, angiogenesis, and facilitating tissue repair.8Newson J Stables M Karra E Arce-Vargas F Quezada S Motwani M et al.Resolution of acute inflammation bridges the gap between innate and adaptive immunity.Blood. 2014; (e-pub ahead of print 8 July 2014)PubMed Google Scholar Initial human studies have indicated that recombinant CSF1 exhibits a broad action profile by activating the MPS and influencing a variety of other processes, such as enhancing antibody-dependent cytotoxicity, augmenting macrophage-mediated microbial killing, and lowering of blood cholesterol and circulating platelets.9VandePol CJ Gemick MB Clinical applications of recombinant macrophage-colony stimulating factor (rhM-CSF).Biotechnol Ther. 1991; 2: 231-239PubMed Google Scholar As illustrated by Hume and MacDonald,10Hume DA MacDonald KPA Therapeutic applications of macrophage colony-stimulating factor-1 (CSF-1) and antagonists of CSF-1 receptor (CSF-1R) signaling.Blood. 2012; 119: 1810-1820Crossref PubMed Scopus (449) Google Scholar a number of studies have since established the pleiotropic effects of CSF1. These findings have sparked interest in CSF1 as a therapeutic agent. During wound healing, tissue-resident macrophages expand in response to CSF1. Together with recently recruited macrophages, they engulf apoptotic cells and clear cell debris, forming a first line of immune defense and fostering tissue repair. The significance of their role in tissue repair has been established by depletion studies showing failed regeneration in the absence of macrophages.11Lucas T Waisman A Ranjan R Roes J Krieg T Müller W et al.Differential roles of macrophages in diverse phases of skin repair.J Immunol. 2010; 184: 3964-3977Crossref PubMed Scopus (754) Google Scholar Gow et al. observed similar regenerative effects in the livers of CSF1-Fc–treated animals. The engineered Fc-bound form gave rise to a 10- to 100-fold greater concentration of CSF1 in vivo that was maintained for up to 72 hours. As depicted in Figure 1a, the Fc-fusion protein was able to achieve this by avoiding renal clearance and via protection from endosomal degradation in the MPS through binding of the Brambell receptor (neonatal Fc receptor) upon internalization, resulting in recycling into the extracellular milieu. As illustrated in Figure 1b, liver-resident Kupffer cells were activated by CSF1-Fc to support matrix remodeling and hepatocyte proliferation, probably by expression of the urokinase plasminogen activator (Plau) and production of tumor necrosis factor-α (TNF-α and interleukin 6 (IL-6). This was accompanied by increased spleen size, which was due mainly to extramedullary hematopoiesis and not directly related to the tissue-regeneration response. It is both surprising and highly clinically relevant that CSF1-Fc is able to increase the size of a healthy liver by induction of a local tissue-repair response. CSF1-Fc may be applicable in several clinical situations. Certain chemotherapeutics, for example, rely on active liver metabolism for clearance or activation and can induce liver damage as a side effect. Peritherapeutic administration of CSF1-Fc could improve liver function, thereby allowing higher, prolonged, or more frequent dosing. Additionally, CSF1-Fc could help restore liver function following partial hepatectomy or in inherited or acquired liver pathologies, and it might be useful in situations of drug overdosing or toxin-induced liver injury. The study by Gow et al. suggests that CSF1 might also be beneficial in the resolution phase of heart diseases, spinal cord injury, fertility issues, or bone repair. A major issue to be addressed is the targeting of the agent. As noted above, CSF1 has a broad activity spectrum, and targeting specific tissues might help to focus these activities to address clinical needs. In addition to extending the compound's half-life, conjugation to Fc appears to provide partial targeting to the liver, particularly to Kupffer cells, the primary blood-exposed phagocytes of the MPS. Interestingly, a previous study using recombinant CSF1 (ref. 12) indicated that the kidney, not the liver, was the primary target organ of CSF1 in mice, and it showed efficacy of recombinant CSF1 on kidney tissue repair after ischemic reperfusion injury. At first, this seems contradictory, but it actually shows that tissue targeting of CSF1 is possible and effective; in the study by Alikhan et al.12Alikhan MA Jones CV Williams TM Beckhouse AG Fletcher AL Kett MM et al.Colony-stimulating factor-1 promotes kidney growth and repair via alteration of macrophage responses.Am J Pathol. 2011; 179: 1243-1256Abstract Full Text Full Text PDF PubMed Scopus (117) Google Scholar the recombinant CSF1 was secreted primarily by the kidneys, which therefore were exposed to the highest concentrations of CSF1. By contrast, in the study by Gow et al.,1Gow DJ Sauter KA Pridans C Moffat L Sehgal A Stutchfield BM et al.Characterisation of a novel Fc conjugate of macrophage colony-stimulating factor.Mol Ther. 2014; 22: 1580-1592Abstract Full Text Full Text PDF PubMed Scopus (70) Google Scholar the Fc conjugate rescued CSF1 from endosomal degradation in Kupffer cells, ensuring accumulation of high local CSF1 concentrations in liver sinusoids. Renal excretion was excluded by the size of the conjugate, thereby rendering the liver the primary target organ. These results invite further investigation into antibody- and Fc-receptor-mediated targeting of CSF1 to different tissues, with the aim of resolving specific pathological situations. A major limitation of prolonged CSF1 use is its effects on the immune system: CSF1 reduces the expression of Toll-like receptors 1, 2, 6, and 9 on induced monocytes or macrophages,13Chitu V Stanley ER Colony-stimulating factor-1 in immunity and inflammation.Curr Opin Immunol. 2006; 18: 39-48Crossref PubMed Scopus (465) Google Scholar thereby diminishing effective immune sensing of bacteria, fungi, and viruses. Additionally, the expression of the purinergic P2x7 receptor is augmented, which can exaggerate NLRP3 inflammasome-dependent inflammatory responses driven by extracellular adenosine triphosphate, such as those that occur during bacterial infections or contact hypersensitivity responses. Moreover, the differentiation and expansion of dendritic cells (DCs) is under CSF1 control.14MacDonald KP Rowe V Bofinger HM Thomas R Sasmono T Hume DA et al.The colony-stimulating factor 1 receptor is expressed on dendritic cells during differentiation and regulates their expansion.J Immunol. 2005; 175: 1399-1405Crossref PubMed Scopus (153) Google Scholar CSF1-Fc administration can induce CSF1 levels 1,000-fold the physiological concentration. This carries the risk of inflating DC activity and thereby unwillingly inducing tolerogenic DC populations or exacerbating T-cell responses. Importantly, persistently elevated CSF1 levels are found in many diseases associated with immune system dysfunction, such as cancer, arthritis, and certain autoimmune disorders.13Chitu V Stanley ER Colony-stimulating factor-1 in immunity and inflammation.Curr Opin Immunol. 2006; 18: 39-48Crossref PubMed Scopus (465) Google Scholar It should be noted, however, that no adverse effects were observed in any of the animals treated in the study by Gow et al. The study by Gow et al. also raises questions about the exact mechanism of action of CSF1 as compared with CFS1-Fc. Liver-tissue regeneration was mediated by hepatocyte- and Kupffer cell activation, associated with local TNF-α and IL-6 production, but not with an acute-phase reaction. Importantly, there was no evidence for immunotoxicity in vivo, which would occur as a consequence of substantial TNF-α and IL-6 levels. The cellular source of TNF-α and IL-6 has not been determined. Furthermore, CSF1-Fc induced interferon signaling–associated genes (such as Ifit1, Irf7, and Cxcl10, Il6), which would be expected in M1-like proinflammatory macrophages (exhibiting a cytokine expression profile supporting cytotoxic immune responses), but not in M2-like tissue-regenerating macrophages. Thus, further investigation is warranted into whether a change in the balance of M1 vs. M2 occurs in response to CSF-Fc treatment. Another concern is the other ligand of the CSF1 receptor (CSF1R): IL-34. Persistent occupation of the CSF1R by exogenous CSF1 or CSF1-Fc might outcompete IL-34 for CSF1R interaction. This could lead to overactivation of as-yet-unknown alternative IL-34 targets or attenuation of IL-34–driven immune activity through CSF1R. Finally, the use of a fusion protein, even when fully humanized, raises the possibility of generating a (neutralizing) immune response, which is important for applications involving prolonged or repetitive use. These issues notwithstanding, the study by Gow et al. establishes CSF1-Fc as a novel therapeutic agent for possible use in liver regeneration that also holds promise for novel therapeutic interventions through its functional impact on the MPS.