The Vasopressin Receptor Antagonist Tolvaptan Counteracts Tumor Growth in a Murine Xenograft Model of Small Cell Lung Cancer.

We have previously demonstrated that the vasopressin type 2 receptor (AVPR2) antagonist tolvaptan reduces cell proliferation and invasion and triggers apoptosis in different human cancer cell lines. To study this effect in vivo, a xenograft model of small cell lung cancer was developed in Fox1nu/nu nude mice through the subcutaneous inoculation of H69 cells, which express AVPR2. One group of mice (n = 5) was treated with tolvaptan for 60 days, whereas one group (n = 5) served as the control. A reduced growth was observed in the tolvaptan group in which the mean tumor volume was significantly smaller on day 60 compared to the control group. In the latter group, a significantly lower survival was observed. The analysis of excised tumors revealed that tolvaptan effectively inhibited the cAMP/PKA and PI3K/AKT signaling pathways. The expression of the proliferative marker proliferating cell nuclear antigen (PCNA) was significantly lower in tumors excised from tolvaptan-treated mice, whereas the expression levels of the apoptotic marker caspase-3 were higher than those in control animals. Furthermore, tumor vascularization was significantly lower in the tolvaptan group. Overall, these findings suggest that tolvaptan counteracts tumor progression in vivo and, if confirmed, might indicate a possible role of this molecule as an adjuvant in anticancer strategies.

Juxtaglomerular apparatus-mediated homeostatic mechanisms: therapeutic implication for chronic kidney disease.

INTRODUCTION: Juxtaglomerular apparatus (JGA)-mediated homeostatic mechanism links to how sodium-glucose cotransporter 2 inhibitors (SGLT2is) slow progression of chronic kidney disease (CKD) and may link to how tolvaptan slows renal function decline in autosomal dominant polycystic kidney disease (ADPKD). AREA COVERED: JGA-mediated homeostatic mechanism has been hypothesized based on investigations of tubuloglomerular feedback and renin-angiotensin system. We reviewed clinical trials of SGLT2is and tolvaptan to assess the relationship between this mechanism and these drugs. EXPERT OPINION: When sodium load to macula densa (MD) increases, MD increases adenosine production, constricting afferent arteriole (Af-art) and protecting glomeruli. Concurrently, MD signaling suppresses renin secretion, increases urinary sodium excretion, and counterbalances reduced sodium filtration. However, when there is marked increase in sodium load per-nephron, as in advanced CKD, MD adenosine production increases, relaxing Af-art and maintaining sodium homeostasis at the expense of glomeruli. The beneficial effects of tolvaptan on renal function in ADPKD may also depend on the JGA-mediated homeostatic mechanisms since tolvaptan inhibits sodium reabsorption in the thick ascending limb.The JGA-mediated homeostatic mechanism regulates Af-arts, constricting to relaxing according to homeostatic needs. Understanding this mechanism may contribute to the development of pharmacotherapeutic compounds and better care for patients with CKD.

Visceral Adiposity and Progression of ADPKD: A Cohort Study of Patients From the TEMPO 3:4 Trial.

RATIONALE & OBJECTIVE: Body mass index (BMI) is an independent predictor of kidney disease progression in individuals with autosomal dominant polycystic kidney disease (ADPKD). Adipocytes do not simply act as a fat reservoir but are active endocrine organs. We hypothesized that greater visceral abdominal adiposity would associate with more rapid kidney growth in ADPKD and influence the efficacy of tolvaptan. STUDY DESIGN: A retrospective cohort study. SETTING & PARTICIPANTS: 1,053 patients enrolled in the TEMPO 3:4 tolvaptan trial with ADPKD and at high risk of rapid disease progression. PREDICTOR: Estimates of visceral adiposity extracted from coronal plane magnetic resonance imaging (MRI) scans using deep learning. OUTCOME: Annual change in total kidney volume (TKV) and effect of tolvaptan on kidney growth. ANALYTICAL APPROACH: Multinomial logistic regression and linear mixed models. RESULTS: In fully adjusted models, the highest tertile of visceral adiposity was associated with greater odds of annual change in TKV of≥7% versus<5% (odds ratio [OR], 4.78 [95% CI, 3.03-7.47]). The association was stronger in women than men (interaction P<0.01). In linear mixed models with an outcome of percent change in TKV per year, tolvaptan efficacy (% change in TKV) was reduced with higher visceral adiposity (3-way interaction of treatment ∗ time ∗ visceral adiposity, P=0.002). Visceral adiposity significantly improved classification performance of predicting rapid annual percent change in TKV for individuals with a normal BMI (DeLong's test z score: -2.03; P=0.04). Greater visceral adiposity was not associated with estimated glomerular filtration rate (eGFR) slope in the overall cohort; however, visceral adiposity was associated with more rapid decline in eGFR slope (below the median) in women (fully adjusted OR, 1.06 [95% CI, 1.01-1.11] per 10 unit increase in visceral adiposity) but not men (OR, 0.98 [95% CI, 0.95-1.02]). LIMITATIONS: Retrospective; rapid progressors; computational demand of deep learning. CONCLUSIONS: Visceral adiposity that can be quantified by MRI in the coronal plane using a deep learning segmentation model independently associates with more rapid kidney growth and improves classification of rapid progression in individuals with a normal BMI. Tolvaptan efficacy decreases with increasing visceral adiposity. PLAIN-LANGUAGE SUMMARY: We analyzed images from a previous study with the drug tolvaptan conducted in patients with autosomal dominant polycystic kidney disease (ADPKD) to measure the amount of fat tissue surrounding the kidneys (visceral fat). We had previously shown body mass index can predict kidney growth in this population; now we determined whether visceral fat was an important factor associated with kidney growth. Using a machine learning tool to automate measurement of fat in images, we observed that visceral fat was independently associated with kidney growth, that it was a better predictor of faster kidney growth in lean patients than body mass index, and that having more visceral fat made treatment of ADPKD with tolvaptan less effective.

Long-Term Effects of Tolvaptan in Autosomal Dominant Polycystic Kidney Disease: Predictors of Treatment Response and Safety over 6 Years of Continuous Therapy.

Tolvaptan, an oral vasopressin V2 receptor antagonist, reduces renal volume expansion and loss of renal function in patients with autosomal dominant polycystic kidney disease (ADPKD). Data for predictive factors indicating patients more likely to benefit from long-term tolvaptan are lacking. Data were retrospectively collected from 55 patients on tolvaptan for 6 years. Changes in renal function, progression of renal dysfunction (estimated glomerular filtration rate [eGFR], 1-year change in eGFR [ΔeGFR/year]), and renal volume (total kidney volume [TKV], percentage 1-year change in TKV [ΔTKV%/year]) were evaluated at 3-years pre-tolvaptan, at baseline, and at 6 years. In 76.4% of patients, ΔeGFR/year improved at 6 years. The average 6-year ΔeGFR/year (range) minus baseline ΔeGFR/year: 3.024 (-8.77-20.58 mL/min/1.73 m2). The increase in TKV was reduced for the first 3 years. A higher BMI was associated with less of an improvement in ΔeGFR (p = 0.027), and family history was associated with more of an improvement in ΔeGFR (p = 0.044). Hypernatremia was generally mild; 3 patients had moderate-to-severe hyponatremia due to prolonged, excessive water intake in response to water diuresis-a side effect of tolvaptan. Family history of ADPKD and baseline BMI were contributing factors for ΔeGFR/year improvement on tolvaptan. Hyponatremia should be monitored with long-term tolvaptan administration.

Vasopressin V2 receptor, tolvaptan, and ERK1/2 phosphorylation in the renal collecting duct.

Tolvaptan, a vasopressin antagonist selective for the V2-subtype vasopressin receptor (V2R), is widely used in the treatment of hyponatremia and autosomal-dominant polycystic kidney disease (ADPKD). Its effects on signaling in collecting duct cells have not been fully characterized. Here, we perform RNA-seq in a collecting duct cell line (mpkCCD). The data show that tolvaptan inhibits the expression of mRNAs that were previously shown to be increased in response to vasopressin including aquaporin-2, but also reveals mRNA changes that were not readily predictable and suggest off-target actions of tolvaptan. One such action is activation of the MAPK kinase (ERK1/ERK2) pathway. Prior studies have shown that ERK1/ERK2 activation is essential in the regulation of a variety of cellular and physiological processes and can be associated with cell proliferation. In immunoblotting experiments, we demonstrated that ERK1/ERK2 phosphorylation in mpkCCD cells was significantly reduced by vasopressin, in contrast to the increases seen in non-collecting-duct cells overexpressing V2R in prior studies. We also found that tolvaptan has a strong effect to increase ERK1/ERK2 phosphorylation in the presence of vasopressin and that tolvaptan's effect to increase ERK1/ERK2 phosphorylation is absent in mpkCCD cells in which both protein kinase A (PKA)-catalytic subunits have been deleted. Thus, it appears that the tolvaptan effect to increase ERK activation is PKA-dependent and is not due to an off-target effect of tolvaptan. We conclude that in cells expressing V2R at endogenous levels: 1) vasopressin decreases ERK1/ERK2 activation; 2) in the presence of vasopressin, tolvaptan increases ERK1/ERK2 activation; and 3) these effects are PKA-dependent.NEW & NOTEWORTHY Vasopressin is a key hormone that regulates the function of the collecting duct of the kidney. ERK1 and ERK2 are enzymes that play key roles in physiological regulation in all cells. The authors used collecting duct cell cultures to investigate the effects of vasopressin and the vasopressin receptor antagonist tolvaptan on ERK1 and ERK2 phosphorylation and activation.

Prioritized polycystic kidney disease drug targets and repurposing candidates from pre-cystic and cystic mouse Pkd2 model gene expression reversion.

BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) is one of the most prevalent monogenic human diseases. It is mostly caused by pathogenic variants in PKD1 or PKD2 genes that encode interacting transmembrane proteins polycystin-1 (PC1) and polycystin-2 (PC2). Among many pathogenic processes described in ADPKD, those associated with cAMP signaling, inflammation, and metabolic reprogramming appear to regulate the disease manifestations. Tolvaptan, a vasopressin receptor-2 antagonist that regulates cAMP pathway, is the only FDA-approved ADPKD therapeutic. Tolvaptan reduces renal cyst growth and kidney function loss, but it is not tolerated by many patients and is associated with idiosyncratic liver toxicity. Therefore, additional therapeutic options for ADPKD treatment are needed. METHODS: As drug repurposing of FDA-approved drug candidates can significantly decrease the time and cost associated with traditional drug discovery, we used the computational approach signature reversion to detect inversely related drug response gene expression signatures from the Library of Integrated Network-Based Cellular Signatures (LINCS) database and identified compounds predicted to reverse disease-associated transcriptomic signatures in three publicly available Pkd2 kidney transcriptomic data sets of mouse ADPKD models. We focused on a pre-cystic model for signature reversion, as it was less impacted by confounding secondary disease mechanisms in ADPKD, and then compared the resulting candidates' target differential expression in the two cystic mouse models. We further prioritized these drug candidates based on their known mechanism of action, FDA status, targets, and by functional enrichment analysis. RESULTS: With this in-silico approach, we prioritized 29 unique drug targets differentially expressed in Pkd2 ADPKD cystic models and 16 prioritized drug repurposing candidates that target them, including bromocriptine and mirtazapine, which can be further tested in-vitro and in-vivo. CONCLUSION: Collectively, these results indicate drug targets and repurposing candidates that may effectively treat pre-cystic as well as cystic ADPKD.

Nutritional Status is Associated With Preserved Kidney Function in Patients With Autosomal Dominant Polycystic Kidney Disease.

OBJECTIVE: Malnutrition is a common complication in autosomal dominant polycystic kidney disease (ADPKD). We examined whether nutritional status is associated with the preservation of kidney function, using a cohort of typical ADPKD. METHODS: We enrolled ambulatory ADPKD patients in 9 tertiary medical centers in Korea from May 2019 to December 2021. We excluded patients who were aged less than 18 years, who had known end-stage kidney disease at the time of enrollment, who had a diagnosis of atypical ADPKD, and who were Tolvaptan users. The primary outcome was an estimated glomerular filtration rate (eGFR) decline >3 mL/min/1.73 m2, based on nutritional status assessed by subjective global assessment (SGA). We also evaluated an eGFR decline >1 mL/min/1.73 m2, an increase in urine protein-creatinine ratio (UPCR) > 0, and an increase in UPCR >0.3 as secondary outcomes, based on SGA after the 1-year follow-up. A logistic regression (LR) model was used to calculate the odds ratio (OR) for the primary outcome. Because there were differences in several baseline variables, such as Mayo classification, serum hemoglobin, serum creatinine, and UPCR between SGA groups, we matched propensity scores. RESULTS: In total, 805 patients were prospectively enrolled. Among them, 236 patients who had 1-year follow-up data and typical imaging findings were analyzed to evaluate the effect of nutritional status on kidney function. SGA was used to assess the nutritional status. The mean age was 45.0 ± 13.3 years, and 49.6% of the patients were female. The mean eGFR was 81.9 mL/min/1.73 m2. Among the 236 patients, 91 (38.6%) experienced a 1-year eGFR decline >3 mL/min/1.73 m2. When a multivariable LR was applied, SGA 3-6 was identified as a significant factor related to a 1-year eGFR decline >3 mL/min/1.73 m2 (adjusted OR = 1.22 [1.04-1.43]; P = .017). Despite matching propensity scores, the 1-year eGFR decline >3 mL/min/1.73 m2 was still higher in the SGA 3-6 group regardless of proteinuria. CONCLUSION: Good nutritional status is associated with better-preserved kidney function in non-obese typical ADPKD patients who do not take Tolvaptan.

Structure-Affinity and Structure-Kinetic Relationship Studies of Benzodiazepine Derivatives for the Development of Efficacious Vasopressin V2 Receptor Antagonists.

Vasopressin V2 receptors (V2R) are a promising drug target for autosomal dominant polycystic kidney disease (ADPKD). As previous research demonstrated that the residence time of V2R antagonists is critical to their efficacy in both ex vivo and in vivo models of ADPKD, we performed extensive structure-kinetic relationship (SKR) analyses on a series of benzodiazepine derivatives. We found that subtle structural modifications of the benzodiazepine derivatives dramatically changed their binding kinetics but not their affinity. Compound 18 exhibited a residence time of 77 min, which was 7.7-fold longer than that of the reference compound tolvaptan (TVP). Accordingly, compound 18 exhibited higher efficacy compared to TVP in an in vivo model of ADPKD. Overall, our study exemplifies a kinetics-directed medicinal chemistry effort for the development of efficacious V2R antagonists. We envision that this strategy may also have general applicability in other therapeutic areas.

MD simulation-based screening approach identified tolvaptan as a potential inhibitor of Eg5.

We discovered tolvaptan as a new Eg5 inhibitor using molecular dynamics simulation-based virtual screening. The Eg5-monastrol, Eg5-ispinesib, and Eg5-STLC complexes with "closed" L5 conformation obtained in MD simulation were used to generate a combined pharmacophore model, and this model was used during the process of virtual screening. Further, the MD simulation for 1 µs showed that the binding of tolvaptan to Eg5 was stable due to the closure of the α2/L5/α3 pocket. Tolvaptan belongs to the class of drugs called vaptans which are non-peptide vasopressin receptor antagonists. Since our virtual search for mitotic inhibitors identified tolvaptan as a potential candidate, we were interested in unraveling its antimitotic mechanism. Tolvaptan bound to purified Eg5-437H with a dissociation constant of 27 ± 3.8 µM. Tolvaptan inhibited the growth of HeLa cells through the mitotic block, and around 70% of these mitotic cells exhibited a characteristic monopolar spindle. Tolvaptan bound to goat brain tubulin with a dissociation constant of 103 ± 13 µM. The binding location of tolvaptan on tubulin overlapped with that of colchicine, according to molecular docking analysis. The combination of tolvaptan with STLC augmented mitotic bock with monopolar cells, whereas its combination with vinblastine increased mitotic block with bipolar cells. Since tolvaptan is found to have a significant cytotoxic effect on HeLa cells, it can be developed as a prospective anticancer agent either alone or in combination with other antimitotic drugs. Tolvaptan was identified as an inhibitor of Eg5 in a MD simulation-based virtual screening using a combined pharmacophore model.

The inhibition of MDM2 slows cell proliferation and activates apoptosis in ADPKD cell lines.

INTRODUCTION: Autosomal dominant polycystic kidney disease (ADPKD) is characterised by progressive cysts formation and renal enlargement that in most of cases leads to end stage of renal disease (ESRD). This pathology is caused by mutations of either PKD1 or PKD2 genes that encode for polycystin-1 (PC1) and polycystin-2 (PC2), respectively. These proteins function as receptor-channel complex able to regulate calcium homeostasis. PKD1/2 loss of function impairs different signalling pathways including cAMP and mTOR that are considered therapeutic targets for this disease. In fact, Tolvaptan, a vasopressin-2 antagonist that reduces cAMP levels, is the only drug approved for ADPKD treatment. Nevertheless, some ADPKD patients developed side effects in response to Tolvaptan including liver damage. Conversely, mTOR inhibitors that induced disease regression in ADPKD animal models failed the clinical trials. RESULTS: Here, we show that the inhibition of mTOR causes the activation of autophagy in ADPKD cells that could reduce therapy effectiveness by drug degradation through the autophagic vesicles. Consistently, the combined treatment with rapamycin and chloroquine, an autophagy inhibitor, potentiates the decrease of cell proliferation induced by rapamycin. To overcome the dangerous activation of autophagy by mTOR inhibition, we targeted MDM2 (a downstream effector of mTOR signalling) that is involved in TP53 degradation by using RG7112, a small-molecule MDM2 inhibitor used for the treatment of haematologic malignancies. The inhibition of MDM2 by RG7112 prevents TP53 degradation and increases p21 expression leading to the decrease of cell proliferation and the activation of apoptosis. CONCLUSION: The targeting of MDM2 by RG7112 might represent a new therapeutic option for the treatment of ADPKD.

Combination of tolvaptan and valsartan improves cardiac and renal functions in doxorubicin-induced heart failure in mice.

Heart failure (HF) is often complicated by renal dysfunction. Tolvaptan and valsartan are two well-known agents for the treatment of HF. However, the role of tolvaptan/valsartan combination on HF with renal dysfunction remains unclear. To establish a mice model with HF with renal dysfunction, mice were intraperitoneally injected with doxorubicin (Dox). Echocardiogram was applied to assess the left ventricular function. Additionally, serum aldosterone (ALD) and angiotensin II (Ang II) level in mice were determined by ELISA. Meanwhile, western blot assay was used to evaluate the expressions of B cell lymphoma-2 (Bcl-2), Bcl-2 associated X (Bax) and cleaved caspase 3 in the heart and kidney tissues of mice. In this study, we found that compared to tolvaptan or valsartan alone treatment group, tolvaptan/valsartan combination obviously improved the left ventricular ejection fraction (LVEF) and the left ventricular fractional shortening (LVFS), and reduced serum ALD and Ang II level in Dox-treated mice. Additionally, tolvaptan/valsartan combination significantly prevented the inflammation and fibrosis of heart and kidney tissues in Dox-treated mice. Meanwhile, tolvaptan/valsartan combination notably inhibited the myocardial and renal cell apoptosis in Dox-treated mice via upregulation of Bcl-2 and downregulation of Bax and cleaved caspase 3, compared to the single drug treatment. Collectively, tolvaptan/valsartan combination could improve cardiac and renal functions, as well as prevent the fibrosis, inflammation and apoptosis of heart and kidney tissues in Dox-treated mice. Taken together, combining tolvaptan with valsartan might be a promising approach to achieve enhanced therapeutic effect for treatment of HF with renal dysfunction.

[Tolvaptan, a vasopressin V2 receptor antagonist, is the world's first approved drug for treatment of autosomal dominant polycystic kidney disease (ADPKD)].

Autosomal dominant polycystic kidney disease (ADPKD) is the most common genetic kidney disease. Fluid-filled cysts develop and enlarge in both kidneys, eventually leading to kidney failure. Tolvaptan is a selective vasopressin V2 receptor antagonist and the first and only drug approved for treatment of ADPKD. It blocks binding of arginine vasopressin (AVP) to V2 receptors in the collecting duct of kidney, thereby inducing water diuresis (aquaresis) without losing electrolytes. Therefore, tolvaptan was originally developed and approved as the first oral aquaretic agent for treatment of hyponatremia and fluid volume overload in heart failure and cirrhosis. During the development of tolvaptan as aquaretics, efficacy of V2 antagonist in polycystic kidney animal model was reported and then the development of tolvaptan for ADPKD was also initiated. Cyclic adenosine monophosphate (cAMP) plays an important role in cyst growth by promoting cell proliferation and fluid secretion. Tolvaptan showed suppression of cyst growth through inhibiting AVP-induced cAMP production and delayed the onset of end-stage renal disease in an animal model. In the phase 3 clinical trial in ADPKD patients (TEMPO 3:4 trial), 3-year treatment with tolvaptan slowed the disease progression including increase of kidney volume and decline in renal function. Efficacy of tolvaptan in patients with late-stage ADPKD was confirmed in another 1-year phase 3 REPRISE trial. Tolvaptan is approved for treatment of ADPKD in more than 40 countries and we expect it can contribute to more ADPKD patients worldwide. We also expect that drugs with new mechanisms will be available in the near future.

The V2 receptor antagonist tolvaptan counteracts proliferation and invasivity in human cancer cells.

PURPOSE: Hyponatremia, the most frequent electrolyte alteration in clinical practice, has been associated with a worse prognosis in cancer patients. On the other hand, a better outcome has been related to serum sodium normalization. In vitro studies have shown that low extracellular sodium promotes cancer cells proliferation and invasiveness. Tolvaptan, a selective vasopressin receptor type 2 (V2) antagonist, has been effectively used in the last decade for the treatment of hyponatremia secondary to the Syndrome of Inappropriate Antidiuresis. A few in vitro data suggested a direct role of tolvaptan in counteracting cancer progression, so far. The aim of this study was to evaluate the effect and the mechanism of action of tolvaptan in cell lines from different tumours [i.e. colon cancer (HCT-8), hepatocarcinoma (HepG2), neuroblastoma (SK-N-AS)]. METHODS AND RESULTS: First, we showed that these cell lines express the V2 receptor. Tolvaptan significantly reduced cell proliferation with an IC50 in the micromolar range. Accordingly, reduced levels of cAMP, of the catalytic α subunit of PKA, and a reduced pAKT/AKT ratio were observed. Tolvaptan effectively inhibited cell cycle progression, whereas it induced apoptotis. Furthermore, it reduced cell invasiveness. In particular, anchorage-independent growth and the activity of collagenases type IV were blunted in the three cell lines. Accordingly, tolvaptan counteracted the RhoA/ROCK1-2 pathway, which has a pivotal role in regulating cell movement. CONCLUSIONS: Overall, these findings indicate that tolvaptan effectively inhibits tumour progression in vitro. Further studies should clarify whether the V2 receptor might be considered a possible target in anti-cancer strategies in the future.

Initial decline in eGFR to predict tolvaptan response in autosomal-dominant polycystic kidney disease.

BACKGROUND: Tolvaptan, a vasopressin V2 receptor antagonist, is used to treat autosomal-dominant polycystic kidney disease (ADPKD). Although tolvaptan curbs disease progression, a few reports have examined factors related to treatment response. The estimated glomerular filtration rate (eGFR) decreases soon after tolvaptan is initiated. We investigated whether initial eGFR decline affects renal prognosis of patients. METHODS: This was a single-center, retrospective observational cohort study. Eighty-three patients with ADPKD who initiated tolvaptan were selected. We analyzed the relationship of the initial eGFR change with clinical parameters and analyzed the annual eGFR change in terms of renal prognostic value using univariable and multivariable linear regression analyses. RESULTS: The initial eGFR change was - 4.6 ± 8.0%/month. The initial eGFR change correlated significantly with the annual eGFR change in multivariable analysis, suggesting that the larger decline in the initial eGFR change, the better the renal prognosis. Furthermore, the change in fractional excretion (FE) of free water (FEH2O) correlated positively with initial eGFR change. FEH2O and urea nitrogen FE (FEUN) increased significantly; however, sodium FE (FENa) level remained unchanged. In approximately half of the patients, FENa unexpectedly decreased. CONCLUSIONS: The initial eGFR decline might be caused by suppressing glomerular hyperfiltration, due to the pharmacological effect of tolvaptan, and/or by reducing renal plasma flow, due to potential volume depletion. The initial eGFR change reflects the tolvaptan effect, can be easily evaluated in clinical practice, and may be useful as one of the clinical indicator for predicting renal prognosis in patients under tolvaptan.

Low sodium and tolvaptan have opposite effects in human small cell lung cancer cells.

PURPOSE: Hyponatraemia is frequently observed in cancer patients and can be due to the syndrome of inappropriate anti-diuresis (SIAD), related to ectopic vasopressin secretion, particularly in small cell lung cancer (SCLC). Hyponatraemia is associated with a worse outcome in cancer patients. The vasopressin receptor antagonist tolvaptan effectively corrects hyponatraemia secondary to SIAD and there is in vitro evidence that it has also an antiproliferative effect in cancer cells. The purpose of this study was i) to analyse the effect of low serum sodium concentrations ([Na+]) in SCLC cells and ii) to determine whether tolvaptan counteracts tumor progression. METHODS: We evaluated cell proliferation, cell cycle, apoptosis, oxidative stress, invasivity in low [Na+] as well as after exposure to tolvaptan. We also analysed the intracellular signalling pathways involved. RESULTS: In reduced [Na+] cell proliferation was significantly increased compared to normal [Na+] and cells were mostly distributed in the G2/M phase. Apoptosis appeared reduced. In addition, the ability to cross matrigel-coated membranes markedly increased. As observed in other cancer cell models, the expression of the heme-oxigenase-1 gene was increased. Finally, we found that in cells cultured in low [Na+] the RhoA/ROCK1/2 pathway, which is involved in the regulation of actin cytoskeleton, was activated. On the other hand, we found that tolvaptan effectively inhibited cell proliferation, anchorage-independent growth, invasivity and promoted apoptosis. Accordingly, the RhoA/ROCK-1/2 pathway was inhibited. CONCLUSIONS: These findings demonstrate for the first time that low [Na+] favours tumor progression in SCLC cells, whereas tolvaptan effectively inhibits cell proliferation, survival and invasivity.

Regulatory guidelines do not accurately predict tolvaptan and metabolite interactions at BCRP, OATP1B1, and OAT3 transporters.

Tolvaptan (TLV) was US Food and Drug Administration (FDA)-approved for the indication to slow kidney function decline in adults at risk of rapidly progressing autosomal dominant polycystic kidney disease in 2018. In vitro, TLV was a breast cancer resistance protein (BCRP) inhibitor, whereas the oxobutyric acid metabolite of TLV (DM-4013) was an inhibitor of organic anion transport polypeptide (OATP)1B1 and organic anion transporter (OAT)3. Based on the 2017 FDA guidance, potential for clinically relevant inhibition at these transporters was indicated for the highest TLV regimen. Consequently, two postmarketing clinical trials in healthy subjects were required. In trial 1, 5 mg rosuvastatin calcium (BCRP and OATP1B1 substrate) was administered alone, with 90 mg TLV or 48 h following 7 days of once daily 300 mg TLV (i.e., in the presence of DM-4103). In trial 2, 40 mg furosemide (OAT3 substrate) was administered alone and in presence of DM-4103. For BCRP, rosuvastatin geometric mean ratios (90% confidence intervals [CIs]) for maximum plasma concentration (Cmax ) were 1.54 (90% CI 1.26-1.88) and for area under the concentration-time curve from time 0 to the time of the last measurable concentration (AUCt ) were 1.69 (90% CI 1.34-2.14), indicating no clinically significant interaction. DM-4103 produced no clinically meaningful changes in rosuvastatin or furosemide concentrations, indicating no inhibition at OATP1B1 or OAT3. The BCRP prediction assumed the drug dose is completely soluble in 250 ml; TLV has solubility of ~0.01 g/250 ml. For OATP1B1/OAT3, if fraction unbound for plasma protein binding (PPB) is less than 1%, then 1% is assumed. DM-4103 has PPB greater than 99.8%. Use of actual drug substance solubility and unbound fraction in plasma would have produced predictions consistent with the clinical results.

Preservation of kidney function irrelevant of total kidney volume growth rate with tolvaptan treatment in patients with autosomal dominant polycystic kidney disease.

BACKGROUND: Tolvaptan slowed the rates of total kidney volume (TKV) growth and renal function decline over a 3-year period in patients with autosomal dominant polycystic kidney disease (ADPKD) enrolled in the Tolvaptan Efficacy and Safety in Management of Autosomal Dominant Polycystic Kidney Disease and Its Outcomes (TEMPO) 3:4 trial (NCT00428948). In this post hoc analysis of Japanese patients from TEMPO 3:4, we evaluated whether the effects of tolvaptan on TKV and on renal function are interrelated. METHODS: One hundred and forty-seven Japanese patients from TEMPO 3:4 were included in this analysis (placebo, n = 55; tolvaptan, n = 92). Tolvaptan-treated patients were stratified into the responder group (n = 37), defined as tolvaptan-treated patients with a net decrease in TKV from baseline to year 3, and the non-responder group (n = 55), defined as tolvaptan-treated patients with a net increase in TKV. RESULTS: Mean changes during follow-up in the placebo, responder, and non-responder groups were 16.99%, - 8.33%, and 13.95%, respectively, for TKV and - 12.61, - 8.47, and - 8.58 mL/min/1.73 m2, respectively, for estimated glomerular filtration rate (eGFR). Compared with the placebo group, eGFR decline was significantly slowed in both the responder and non-responder groups (P < 0.05). CONCLUSION: Tolvaptan was effective in slowing eGFR decline, regardless of TKV response, over 3 years in patients with ADPKD in Japan. Treatment with tolvaptan may have beneficial effects on slowing of renal function decline even in patients who have not experienced a reduction in the rate of TKV growth by treatment with tolvaptan.

A metabolically stable apelin-17 analog decreases AVP-induced antidiuresis and improves hyponatremia.

Apelin and arginine-vasopressin (AVP) are conversely regulated by osmotic stimuli. We therefore hypothesized that activating the apelin receptor (apelin-R) with LIT01-196, a metabolically stable apelin-17 analog, may be beneficial for treating the Syndrome of Inappropriate Antidiuresis, in which AVP hypersecretion leads to hyponatremia. We show that LIT01-196, which behaves as a potent full agonist for the apelin-R, has an in vivo half-life of 156 minutes in the bloodstream after subcutaneous administration in control rats. In collecting ducts, LIT01-196 decreases dDAVP-induced cAMP production and apical cell surface expression of phosphorylated aquaporin 2 via AVP type 2 receptors, leading to an increase in aqueous diuresis. In a rat experimental model of AVP-induced hyponatremia, LIT01-196 subcutaneously administered blocks the antidiuretic effect of AVP and the AVP-induced increase in urinary osmolality and induces a progressive improvement of hyponatremia. Our data suggest that apelin-R activation constitutes an original approach for hyponatremia treatment.

Expression of aquaporin-2 in the collecting duct and responses to tolvaptan.

Tolvaptan, a vasopressin type-2 receptor antagonist, is indicated for fluid retention. It is considered that the response to tolvaptan reduces as renal function deteriorates, whereas we sometimes experience "non-responders" to tolvaptan despite well-preserved renal function. While the expression of aquaporin-2 might be a key to response to tolvaptan, detailed mechanism of refractoriness to tolvaptan remains unknown. We experienced two patients with congestive heart failure and diabetic nephropathy, in whom the responses to tolvaptan were uniquely opposite. In one case, immunohistochemical staining showed expression of aquaporin-2 in the collecting duct despite severely reduced renal function, followed by the good response to tolvaptan with increased urine output. In another case, immunohistochemical staining showed absence of aquaporin-2 with infiltration of inflammatory cells in the kidney medulla despite relatively preserved renal function, followed by refractoriness to tolvaptan without any increase in urine output. Inactivated aquaporin-2 expression in the collecting duct, which was for example caused by pre-clinical urinary infection as our latter case, might have an association with refractoriness to tolvaptan.

Autosomal-dominant polycystic kidney disease: tolvaptan use in adolescents and young adults with rapid progression.

BACKGROUND: The phase 3 Tolvaptan Efficacy and Safety in Management of Autosomal Dominant Polycystic Kidney Disease and Its Outcomes (TEMPO 3:4) clinical trial demonstrated the beneficial effect of tolvaptan on kidney growth and function in subjects aged 18-50 years over a 3-year period. However, it did not specifically assess the use of tolvaptan in adolescents and young adults (AYAs) with ADPKD. METHODS: A post hoc analysis of the TEMPO 3:4 trials was performed for patients aged 18-24 years. The primary outcome was the annual rate of change in total kidney volume (TKV). The secondary outcome was to evaluate long-term safety of tolvaptan using Hy's law of hepatotoxicity. RESULTS: A total of 51 patients in the 18-24 age group were analyzed (tolvaptan: 29, placebo: 22). The tolvaptan group had a lower mean percentage of TKV growth per year compared to the placebo group (3.9% vs. 6.5%, P = 0.0491). For secondary outcomes, 63 patients in the AYA subgroup were evaluated. In both the AYA and adult groups, none of the patients met the criteria for Hy's law of hepatotoxicity. CONCLUSIONS: This post hoc analysis suggests that tolvaptan, with appropriate patient selection and management, can provide effective and acceptably safe treatment in AYAs with ADPKD. IMPACT: Tolvaptan slows the increase in total kidney volume in patients aged 18-24 years with ADPKD. Tolvaptan posed no risk of potential liver injury measured via Hy's law of hepatotoxicity in the AYA stratum. This study suggests that tolvaptan has beneficial outcomes in AYAs. This post hoc analysis suggests the need for additional studies with a larger pediatric patient population. The impact is significant as tolvaptan had not been specifically examined in the AYA patient population previously.