EGF Receptor Inhibition by Erlotinib Increases Aquaporin 2–Mediated Renal Water Reabsorption

Abstract

Nephrogenic diabetes insipidus (NDI) is caused by impairment of vasopressin (VP) receptor type 2 signaling. Because potential therapies for NDI that target the canonical VP/cAMP/protein kinase A pathway have so far proven ineffective, alternative strategies for modulating aquaporin 2 (AQP2) trafficking have been sought. Successful identification of compounds by our high-throughput chemical screening assay prompted us to determine whether EGF receptor (EGFR) inhibitors stimulate AQP2 trafficking and reduce urine output. Erlotinib, a selective EGFR inhibitor, enhanced AQP2 apical membrane expression in collecting duct principal cells and reduced urine volume by 45% after 5 days of treatment in mice with lithiuminduced NDI. Similar to VP, erlotinib increased exocytosis and decreased endocytosis in LLC-PK1 cells, resulting in a significant increase in AQP2 membrane accumulation. Erlotinib increased phosphorylation of AQP2 at Ser-256 and Ser-269 and decreased phosphorylation at Ser-261 in a dose-dependent manner. However, unlike VP, the effect of erlotinib was independent of cAMP, cGMP, and protein kinase A. Conversely, EGF reduced VP-induced AQP2 Ser-256 phosphorylation, suggesting crosstalk between VP and EGF in AQP2 trafficking and a role of EGF in water homeostasis. These results reveal a novel pathway that contributes to the regulation of AQP2-mediated water reabsorption and suggest new potential therapeutic strategies for NDI treatment.

Inhibition of non‐receptor tyrosine kinase Src induces phosphoserine 256‐independent aquaporin‐2 membrane accumulation

“…AQP2 membrane trafficking can also be facilitated by other receptor ligands, including angiotensin, prostaglandin, and calcitonin (Jensen et al 2009(Jensen et al , 2010Li et al 2009;Bouley et al 2011), which affect the cAMP/PKA signalling pathway. In addition, we found that epidermal growth factor receptor (EGFR) inhibition can induce AQP2 phosphorylation in a similar pattern to VP, but independently of cAMP and PKA, suggesting a crosstalk between EGFR and VP signalling in AQP2 regulation (Cheung et al 2016). One possible common denominator in this crosstalk is Src (pp60 c-SRC ).…”

“…, ; Cheung et al . ). After 5 washes in PBS, tissues were cryoprotected in PBS, 30% sucrose overnight before being embedded in Tissue‐Tek O.C.T.…”

“…; Cheung et al . ). In brief, exocytosis was quantified in LLC‐PK1 cells stably expressing both c‐myc‐tagged AQP2 and the soluble yellow fluorescent protein (ssYFP) (LLC‐AQP2‐ssYFP cells) or ssYFP alone (LLC‐ssYFP cells).…”

“…In addition, we found that epidermal growth factor receptor (EGFR) inhibition can induce AQP2 phosphorylation in a similar pattern to VP, but independently of cAMP and PKA, suggesting a crosstalk between EGFR and VP signalling in AQP2 regulation (Cheung et al . ). One possible common denominator in this crosstalk is Src (pp60 c‐SRC ).…”

Metformin improves urine concentration in rodents with nephrogenic diabetes insipidus

“…Although the disease is better understood and complications are less often seen currently due to our increased understanding of their causes, the necessity for frequent and high-volume urination is still a problem for survival and quality of life. There are several chemicals, including erlotinib, sildenafil, and simvastatin, that have been reported to activate urine-concentrating ability through nonvasopressin pathways in vivo and in vitro (7–10), but none of these studies showed complete restoration of urine-concentrating ability.…”