C-C motif chemokine receptor 2 and phosphodiesterase type 5 inhibition in immune subphenotypes: a step toward personalized treatment for pulmonary arterial hypertension?

This editorial refers to ‘C-C motif chemokine receptor-2 blockade ameliorates pulmonary hypertension in rats and synergizes with a pulmonary vasodilator’, by Tsuboya et al., https://doi.org/10.1093/cvr/cvae244. It is broadly accepted that pulmonary arterial hypertension (PAH) is a rare yet devastating disease that is characterized by lung vascular remodelling, increased pulmonary vascular resistance (PVR) and right heart failure with no known cure to date. Indeed, PAH is characterized in humans by vascular remodelling, and formation of complex vascular lesions characterized by a pro-inflammatory milieu, accelerated proliferation and migration of pulmonary endothelial (ECs) and smooth muscle cells (SMCs) and, in turn, increased PVR.1 Although multiple United States Food and Drug Administration (FDA) approved vasodilator treatments that target PVR, i.e. prostaglandin I2 (PGI2) agonist, endothelin-1 (ET-1) antagonist, phosphodiesterase type 5 (PDE5) inhibitor, soluble guanylate cyclase (sGC) stimulator, or a combination thereof, have improved healthspan and extended lifespan, they are not curative, and their durable effects on pulmonary vascular remodelling remain uncertain.2 Thus, much effort has gone into identifying novel therapeutic targets to reverse or halt vascular remodelling or extend healthspan in PAH patients. Despite the large body of research on PAH, modelling of the disease in mice/rats remains a challenge and, predictably, no one model fully recapitulates the human disease. Indeed, the disease is multi-factorial and instigators of PAH vary among subpopulations of patients.1–3 A unique strength of the study in this issue: ‘C-C motif chemokine receptor-2 blockade ameliorates pulmonary hypertension in rats and synergizes with a pulmonary vasodilator’ by Tsuboya et al.4 is that, unlike many other pre-clinical studies, it employs several pre-clinical models of PAH to test the hypothesis that disruption or blockade of the C-C motif chemokine receptor 2 (CCR2) could potentially ameliorate PAH in a subset of the human population. Specifically, the models approximating PAH used in this study4 include chronic hypoxia (CH), Sugen hypoxia (Su/Hx), and monocrotaline (MCT), all in rats. In spite of and because previous studies targeting CCR2 in CH mice (least definitive for PAH per se) yielded disappointing results, the authors employed no fewer than the three models to test their hypothesis. Importantly, CH in rats causes a reversible increase in pulmonary pressure due to minimal vascular remodelling and RV hypertrophy, while Su/Hx, a combination of hypoxia and Su5416 (VEGF receptor tyrosine kinase inhibitor) results in more severe, persistent PAH with significant vascular remodelling including severe RV dysfunction/hypertrophy and development of occlusive plexiform-like vascular lesions (Figure 1).3 Lastly, rat MCT-induced PAH despite not producing plexiform lesions, introduces an additional feature that neither CH nor Su/Hx possesses that is salient in a subset of human PAH cohorts; that is, increased expression of pro-inflammatory cytokines, i.e. transforming growth factor beta 1 (TGF-β1) and interleukin-6 (IL-6) in lung tissue, which are reportedly both elevated in PAH.3,5 As such, the spotlighted study4 examines CCR2 knockout (KO) rats (CCR2−/−) in each of these models to provide as comprehensive an evaluation, to our knowledge, of the effect of CCR2 KO in pre-clinical PAH to date, and, importantly, investigate its potential therapeutic benefit of targeting CCR2 in human PAH. Graphical hypothesis. Hypothesis of CCR2 and PDE5 inhibition effectiveness in immune subphenotype of monocrotaline-treated rats. SMCs, smooth muscle cells; ECs, endothelial cells; L, lumen; WT, wild type; CH, chronic hypoxia; MCT, monocrotaline; Su/Hx, sugen hypoxia; RV, right ventricle; LV, left ventricle; CCR2, C-C motif chemokine receptor 2; KO, knockout; PAH, pulmonary arterial hypertension; PDE5, phosphodiesterase type 5. Indeed, CCR2, the main receptor of the C-C motif chemokine ligand 2 (CCL2) as well as several other chemokine ligands including CCL7, CCL8, CCL12, CCL13, and CCL16, is expressed on the surface of human pulmonary artery SMCs6 (PASMCs) and pulmonary artery ECs7 (PAECs). Moreover, recent studies have shown that high levels of inflammatory cytokines induce proliferation of PASMCs8 and/or proliferation and dysfunction of PAECs.9 Consequently, several pre-clinical studies10,11 of PAH manipulating effects of cytokines including tumour necrosis factor-α (TNF-α) or cytokine receptors including interleukin-6 receptor (IL-6R) and interleukin-1 receptor (IL-1R) in PAH showed promise for these cellular targets. Despite these attempts, in the pre-clinical CH PAH mouse model CCR2 receptor deletion (CCR2−/−) showed inconsistent and counterintuitive results (little, no, or detrimental effect) in previously published reports.12–14 In spite of the above-mentioned inconclusive pre-clinical outcomes,12–14 Tsuboya et al.4 continued with the study and chose to more broadly investigate CCR2 receptor deletion in PAH. In so doing, their three-pronged investigation using CH, Su/Hx, and MCT rats revealed novel provocative findings. The authors found that CCR2 receptor deletion (CCR2−/−) attenuated hallmark parameters elevated in PAH, i.e. mean pulmonary artery pressure (mPAP), right ventricle systolic pressure (RVSP), Fulton index (RV/LV + Septum), among other parameters in rat MCT PAH, whereas it elicited no effect in CH or Su/Hx (Figure 1). Further, they showed in rat MCT PAH, CCR2 receptor blockade led to a reversal in several pro-inflammatory cascades including CCL2, IL-6, TNF-α, interleukin-1β (IL-1β), and TGF-β. Along with these outcomes, CCR2 receptor blockade reduced macrophage infiltration, as well as reversed bone morphogenetic protein receptor type 2 signalling (BMPR2, genetic factor in PAH1) and improved nitric oxide (NO) signalling. Moreover, and most germane to this discussion CCR2 receptor blockade increased PDE5 expression in lungs from MCT-treated rats. Adding to these findings, disruption of CCR2 signalling in PASMCs from MCT rats triggered PDE5 upregulation, consistent with results in vivo. On the contrary, these anti-inflammatory effects and PDE5 upregulation did not occur in CH or Su/Hx rat models. While it may seem peculiar to the uninitiated that CCR2 receptor blockade would attenuate PAH in MCT rats, but not CH rats or Su/Hx rats, it is indeed the centrepiece of the findings by these authors.4 In fact, these studies rather uniquely imply that perhaps a more nuanced approach should be taken in the treatment of PAH. With the knowledge that a multitude of signalling networks are at play in PAH1 particularly in distinct cohorts of PAH patients, it makes sense that a precision medicine approach could be beneficial. Importantly, in this regard, several CCR2-regulated genes, including CCL2, IL-6, TNF-α, and IL-1β are upregulated and specifically clustered to certain subpopulations of PAH patients.15 As such, Tsuboya et al.4 tested CCR2 receptor deletion in conjunction with tadalafil (PDE5 inhibitor, FDA-approved for PAH in 20091) in MCT rats with a view toward a potential future application of targeting CCR2 in a cluster/subpopulation of PAH patients who would display an increase in PDE5.15 Intriguingly, tadalafil administration ameliorated PAH and improved survival in MCT CCR2-blocked rats markedly better than in wildtype MCT rats. The authors thus theorize that if used for human PAH treatment, CCR2 receptor blockade could potentially prove clinically relevant in the subpopulation of patients with elevated cytokines in combination with tadalafil, or perhaps even more in the recently approved combination therapy of macitentan and tadalafil.1 By investigating CCR2 blockade in an unbiased manner, with several different models of PAH, Tsuboya et al.4 identified a subpopulation of PAH patients15 that could benefit from a breakthrough adjunctive treatment of PAH. Nevertheless, future pre-clinical studies investigating the effects of cell-specific targeting of CCR2 deletion in vascular endothelial or smooth muscle cells could be more informative as to the role of CCR2 in PAH pathophysiology. Importantly, this study was unable to determine whether tadalafil alters downstream signalling and proliferation in PASMCs with CCR2 blockade, a crucial step in understanding the mechanism behind CCR2 receptor blockade in PAH. Furthermore, future studies testing the effect of a small molecule CCR2 antagonist in MCT-treated rats could lay the groundwork for a druggable CCR2-targeted therapy in PAH. One potential improvement of the current study for comprehensiveness and balance could have been employment of the MCT + hypoxia rat model of PAH. In that model, rats develop occlusive plexiform lesions (PLs) seen in human PAH, with the added inflammatory effects of MCT.3 In that vein, the ability of this model to perhaps more accurately recapitulate the subpopulation of PAH patients with increased inflammation could prove more beneficial to our appreciation of the advantage of CCR2 receptor blockade in PAH. This work was supported by R01 HL142248 (P.J.P.), AHA 18TPA34170069 (P.J.P.), P01 HL103455 (P.J.P.), R35 HL161177 (C.J.G.), and F31 1HL167563 (C.J.G.). No new data were generated or analysed in support of this research.