Will Stub1 Be the Next Target for Pulmonary Hypertension Therapy Development?

Pulmonary hypertension (PH) is a vascular disorder characterized by elevated mean pulmonary artery pressure (mPAP) exceeding 20 mm Hg, accompanied by pulmonary vascular remodeling, particularly the muscularization of small pulmonary arteries (1, 2). PH is classified into five groups (3), with group 3 PH (PH caused by lung diseases and/or hypoxia) receiving growing attention, largely because of the rising prevalence of chronic lung diseases and the limited availability of effective therapies (4). A key challenge in developing treatments targeting pulmonary vascular remodeling in established PH is an incomplete understanding of the mechanisms driving the proliferation of pulmonary arterial vascular cells, including pulmonary arterial smooth muscle cells (PASMCs) and pulmonary arterial endothelial cells (PAECs).

Previous research highlights the critical roles of hypoxia-inducible factor-1α (HIF-1α) and HIF-2α in the progression of PH (5, 6). Although HIF-1α may be the primary driver of hypoxia-induced PASMC hyperproliferation and HIF-2α is responsible for PAEC proliferation and survival, growing evidence suggests that HIF-2α may also regulate PASMCs. However, the mechanisms governing HIF-2α remain poorly understood. STIP1 homology and U-box containing protein 1 (Stub1/CHIP) is a chaperone-dependent E3 ubiquitin ligase with activity that is largely modulated by post-translational modifications, including phosphorylation, ubiquitylation, ISGylation (a process that attaches IFN-stimulated gene 15 to proteins), and acetylation (7). Activated Stub1 can regulate cellular processes such as protein degradation, autophagy, and mitochondrial biogenesis. Research on the regulation of HIF by Stub1 has focused on cancer and infectious diseases (8, 9). Given the shared signaling pathways that promote PH-associated pulmonary arterial cell and cancer cell proliferation, Stub1 regulation of HIF proteins warrants further investigation.

In this issue of the Journal, Afolayan and colleagues (pp. 369–382) reveal a noncanonical role of K287-acetylated Stub1 in PAECs and PASMCs during hypoxia (10). This modification, regulated by CREB-binding protein (CBP)/p300, selectively suppresses HIF-2α activity, mitigates right ventricular remodeling, reduces pulmonary vascular resistance, and attenuates experimentally induced PH in mice. The authors demonstrate that hypoxia-induced K287 acetylation is directly mediated by CBP/p300. Interestingly, although K287 acetylation inactivates Stub1, it does not impair its ability to bind substrates in PAECs and PASMCs. Instead, it triggers Stub1 nuclear localization, upregulating glycolysis- and cell cycle–related genes while suppressing tumorigenesis-related pathways. Furthermore, using a novel mouse model overexpressing human Stub1 in PAECs, they confirmed that K287 acetylation significantly reduces HIF-2α mRNA, protein concentrations, and activity in PAECs. This results in attenuated PH compared with wild-type mice after 4 weeks of hypoxia exposure. The study also revealed that mitochondrial redox signaling, including superoxide dismutase-2, regulates Stub1 acetylation by CBP/p300, whereas histone deacetylase 9 deacetylates Stub1, sustaining HIF-2α activity during chronic hypoxia. Their exciting findings expand mechanistic understanding of PAEC and PASMC proliferation in PH.

The only U.S. Food and Drug Administration–approved medication targeting pulmonary vascular remodeling in PH is sotatercept-csrk (Winrevair; Merck), a homodimeric recombinant fusion protein that binds to activin A and inhibits activin-mediated pulmonary vascular cell proliferation. Although researchers suggest that sotatercept-csrk could be used across PH categories, it is only approved for group 1 PH. This highlights the lack of therapies that specifically target pulmonary vascular remodeling in PH. In established PH, PAECs and PASMCs exhibit numerous dysregulated features. PAECs are often associated with impaired regulation of vascular tone, increased proliferation, and enhanced endothelial-to-mesenchymal transition (11). Conversely, PASMCs display hyperproliferation, hypertrophy, and altered contractility (2, 12). A major challenge in developing new therapies lies in identifying safe treatments capable of reversing or attenuating pulmonary vascular remodeling driven by the proliferative profiles of these pulmonary resident cells.

Chronic hypoxia, a hallmark of established PH, exacerbates pulmonary vascular cell proliferation. HIFs regulate cellular responses to hypoxia, with HIF-1α and HIF-2α exhibiting distinct functions and tissue distributions (Figure 1). In PASMCs, PH-associated HIF-1α activation promotes glycolysis and hyperproliferation. Increased HIF-1α stability in PH PAECs is linked to hypoxia-associated endothelial nitric oxide synthase downregulation and decreased nitric oxide (NO) release. On the other hand, in PAECs, HIF-2α governs cell proliferation and angiogenesis in PH (13). There is significant progress in unraveling the interplay between HIF-1α and HIF-2α, and Afolayan and colleagues advance understanding by demonstrating that Stub1, particularly its K287-acetylated form, maintains HIF-1α activity while suppressing HIF-2α in PAECs and PASMCs. Importantly, their in vivo studies show that Stub1 acetylation-mediated suppression of HIF-2α in PAECs attenuates PASMC proliferation, offering new insights into potential therapeutic strategies for PH.

Figure 1.

Hypoxia induces hypoxia-inducible factor (HIF) pathways in pulmonary arterial endothelial cell (PAECs) and pulmonary arterial smooth muscle cell (PASMCs) in pulmonary hypertension (PH). PH progression is closely linked to hypoxia-induced dysfunction in PAECs and PASMCs. HIF proteins, including HIF-1α and HIF-2α, are critical regulators in the cellular response to hypoxia and exhibit distinct regulatory roles. Dysregulated signaling pathways in PAECs, such as reduced NO production and elevated secretion of ET-1, growth factors, and cytokines, contribute to PASMC proliferation. These interactions drive pulmonary vascular remodeling, a hallmark of PH. Ac = acetyl-; ET-1 = endothelin-1; NO = nitric oxide; Stub1 = STIP1 homology and U-box containing protein 1. Figure created in BioRender.

Cell–cell communication, particularly between PAECs and PASMCs, plays a critical role in the progression of PH and presents a valuable target for therapeutic development. In PH, dysregulated PAECs exhibit impaired NO production and increased secretion of endothelin-1, growth factors (e.g., vascular endothelial growth factor), cytokines (e.g., interleukin-6), and microRNA-containing extracellular vesicles. Metabolic signaling shifts in PAECs also exacerbate PASMC hyperproliferation (5). HIF-2α signaling is dysregulated in PAEC communication with PASMCs (Figure 1). For instance, HIF-2α activation in PAECs from PH animal models is associated with decreased NO production, partly because of the upregulation of arginase-1 (14). Afolayan and colleagues provide compelling evidence that Stub1 K287 acetylation and the resulting suppression of HIF-2α in PAECs significantly attenuate PASMC proliferation in vivo. The authors also show that Stub1 acetylation downregulates PASMC HIF-2α.

Despite these insights, this research does not fully elucidate whether and how Stub1/HIF-2α signaling mediates communications between PAECs and PASMCs. In addition, it remains unclear whether Stub1 acetylation and HIF-2α suppression effectively reduce PASMC proliferation in PH animal models. Thus, a more comprehensive understanding of PAEC–PASMC communication is essential before developing HIF-2α–targeted therapies for clinical application. It is also important to note that although several experimentally induced PH animal models exist, no single model fully replicates all the phenotypes of PH in patients (15). Afolayan and colleagues used a transgenic mouse model with chronic hypoxia-induced PH; however, mice exposed to chronic hypoxia are known to develop mild PH, which is reversible upon reexposure to normoxic conditions. Therefore, the development of future therapeutic agents targeting the Stub1/HIF-2α pathway should be validated in multiple animal models, particularly those that mimic severe and irreversible PH, to better evaluate the potential clinical applicability of these treatments.

In conclusion, Afolayan and colleagues provide novel insight into PH pathological cell signaling and lay the groundwork for further research. Specifically, their work deepens our understanding of the distinct roles of HIF-1α and HIF-2α in PH progression and highlights a promising avenue for therapeutic development. Given the critical involvement of HIF proteins in PH, these findings have the potential to extend beyond adult PH to pediatric cases, such as congenital diaphragmatic hernia-associated PH, and even to other PH classifications beyond group 3.