Cumulative dose of bevacizumab associates with albuminuria rather than podocyturia in cancer patients.

Angiogenesis inhibition with bevacizumab, a monoclonal antibody against vascular endothelial growth factor A (VEGF-A), is an anticancer treatment associated with hypertension and renal glomerular toxicity referred to as a preeclampsia-like syndrome. In preeclampsia, podocyturia predates proteinuria and clinical features of preeclampsia, and is regarded as a biomarker of ongoing glomerular injury. Using a quantitative polymerase chain reaction of the podocyte-specific molecules nephrin, podocin, and VEGF-A in the urine, we examined whether podocyturia is present in bevacizumab-treated cancer patients, and whether it relates to proteinuria and the cumulative dose of bevacizumab. Urine samples were cross-sectionally collected from 43 bevacizumab-treated patients, 21 chemotherapy-treated patients, and 7 healthy controls. Urinary protein-to-creatinine ratio (mean and range) was 32.0 mg/mmol (5.2-284.4) in the bevacizumab group, compared with 11.4 mg/mmol (1.1-21.0) in the chemotherapy group and 7.4 mg/mmol (3.9-16.5) (P < .05) in healthy controls, whereas urinary albumin-to-creatinine ratio values in the three groups were, respectively, 18.9 mg/mmol (0.1-227.7), 1.5 mg/mmol (0.2-3.5), and 0.2 mg/mmol (0.1-0.4) (P < .05). The cumulative dose of bevacizumab ranged from 550 to 93,628 mg. Urinary podocin mRNA expression was undetectable in 59% of participants, urinary nephrin mRNA expression per mmol creatinine ranged from 0.0 to 5.3 and urinary VEGF-A mRNA expression from 0.0 to 2.7. Urinary nephrin mRNA expression did not correlate to the albumin-to-creatinine ratio or the cumulative dose of bevacizumab, whereas the latter correlated with the albumin-to-creatinine ratio (r = 0.77; P < .001). Our results demonstrate that the cumulative dose of bevacizumab is closely correlated with albuminuria but not with podocyturia as measured with the quantitative polymerase chain reaction technique, challenging the feasibility of this measurement to monitor ongoing glomerular injury in patients chronically treated with bevacizumab.

Greater Sensitivity of Blood Pressure Than Renal Toxicity to Tyrosine Kinase Receptor Inhibition With Sunitinib.

Hypertension and renal injury are off-target effects of sunitinib, a tyrosine kinase receptor inhibitor used for the treatment of various tumor types. Importantly, these untoward effects are accompanied by activation of the endothelin system. Here, we set up a study to explore the dose dependency of these side effects. Normotensive Wistar Kyoto rats were exposed to 3 different doses of sunitinib or vehicle. After 8 days, rats were euthanized. Telemetrically measured blood pressure rose dose dependently, from 13 to 30 mm Hg. Proteinuria was present at all doses, but a rise in cystatin C occurred only at the intermediate and high doses. Compared with vehicle circulating endothelin-1 increased dose dependently, whereas 24-hour urinary endothelin excretion decreased. Light and electron microscopy revealed glomerular endotheliosis and ischemia with the intermediate and high doses of sunitinib but completely absent histological abnormalities with the low dose. Podocyte number per glomerular circumference did not change. Glomerular nephrin, Neph1, podocin, and endothelin-converting enzyme gene expression were downregulated in a dose-dependent manner. We conclude that the sunitinib-induced rise in blood pressure requires lower doses than its induction of renal function impairment and that functional changes in glomerular filtration barrier contribute to the occurrence of proteinuria, given the lack of histopathologic changes with the low dose of sunitinib.

[Treatment options for resistant hypertension: from pseudoresistant to refractory hypertension]. / Behandelopties voor therapieresistente hypertensive: van pseudoresistentie tot refractaire hypertensive.

Resistant hypertension is defined as blood pressure above the target level despite treatment with 3 classes of antihypertensive drugs, including a diuretic. A large number of patients meeting the definition of TRH actually have 'pseudoresistant hypertension': there is either a secondary cause of the hypertension, non-adherence, high dietary salt intake, or use of interfering co-medication or recreational drugs. Treating pseudoresistant hypertension is just as challenging as 'true' resistant hypertension since causes of resistance cannot always be eliminated and elimination of causes will not necessarily lead to blood pressure normalization. It is estimated that only 10% of patients with TRH have 'true' resistant hypertension. A very small proportion of these patients is defined as having 'refractory hypertension' because their blood pressure still remains uncontrolled despite extending their medication to five or more agents, including an aldosterone receptor blocker. At present, non-pharmacological, invasive interventions should be considered only in patients with refractory hypertension.

Treatment of hypertension and renal injury induced by the angiogenesis inhibitor sunitinib: preclinical study.

Common adverse effects of angiogenesis inhibition are hypertension and renal injury. To determine the most optimal way to prevent these adverse effects and to explore their interdependency, the following drugs were investigated in unrestrained Wistar Kyoto rats exposed to the angiogenesis inhibitor sunitinib: the dual endothelin receptor antagonist macitentan; the calcium channel blocker amlodipine; the angiotensin-converting enzyme inhibitor captopril; and the phosphodiesterase type 5 inhibitor sildenafil. Mean arterial pressure was monitored telemetrically. After 8 days, rats were euthanized and blood samples and kidneys were collected. In addition, 24-hour urine samples were collected. After sunitinib start, mean arterial pressure increased rapidly by ≈30 mm Hg. Coadministration of macitentan or amlodipine largely prevented this rise, whereas captopril or sildenafil did not. Macitentan, captopril, and sildenafil diminished the sunitinib-induced proteinuria and endothelinuria and glomerular intraepithelial protein deposition, whereas amlodipine did not. Changes in proteinuria and endothelinuria were unrelated. We conclude that in our experimental model, dual endothelin receptor antagonism and calcium channel blockade are suitable to prevent angiogenesis inhibition-induced hypertension, whereas dual endothelin receptor antagonism, angiotensin-converting enzyme inhibitor, and phosphodiesterase type 5 inhibition can prevent angiogenesis inhibition-induced proteinuria. Moreover, the variable response of hypertension and renal injury to different antihypertensive agents suggests that these side effects are, at least in part, unrelated.

Hypertension during vascular endothelial growth factor inhibition: focus on nitric oxide, endothelin-1, and oxidative stress.

SIGNIFICANCE: Angiogenesis inhibition with humanized antibodies targeting vascular endothelial growth factor (VEGF) or orally active small tyrosine kinase inhibitors targeting VEGF receptors has become an established treatment modality for various forms of cancer. A common side effect of angiogenesis inhibition is the development of sometimes severe hypertension, which simultaneously appears to be predictive for a favorable antitumor response. RECENT ADVANCES: Since VEGF increases the expression and activity of endothelial nitric oxide synthase, it has been assumed that the mean blood pressure (MAP) rise during angiogenesis inhibition is caused by a decrease in nitric oxide bioavailability. Yet, the results from experimental and clinical studies exploring this possibility are conflicting. Recent studies provided evidence that the MAP rise during angiogenesis inhibition rather is mediated by activation of the endothelin-1 (ET-1) axis, which, among others, induces oxidative stress. Nevertheless, conclusive evidence for the involvement of reactive oxygen species in the MAP rise could not be obtained so far. CRITICAL ISSUES: The mechanism underlying activation of the ET-1 axis during angiogenesis inhibition is unclear, and this activation was not anticipated in view of studies showing that VEGF stimulates both the expression and production of ET-1 by endothelial cells. FUTURE DIRECTIONS: In fact, this activation of the ET-1 axis may support the use of ET receptor antagonists for the treatment of angiogenesis inhibition-induced hypertension, especially because ET receptor stimulation in vascular smooth muscle cells results in VEGF production and mitogenesis in a mitogen-activated protein kinase pathway-dependent manner.

Mechanism of hypertension and proteinuria during angiogenesis inhibition: evolving role of endothelin-1.

Angiogenesis inhibition by blocking vascular endothelial growth factor (VEGF)-mediated signalling with monoclonal antibodies or tyrosine kinase inhibitors has become an established treatment of various forms of cancer. This treatment is frequently associated with the development of hypertension and proteinuria. As VEGF increases the expression and the activity of nitric oxide synthase in endothelial cells, a decrease in the bioavailability of nitric oxide has been proposed as a key mechanism leading to hypertension during angiogenesis inhibition. However, results of clinical and experimental studies exploring this possibility are conflicting. Rarefaction, that is a structural decrease of microcirculatory vessels, has been reported during antiangiogenic treatment, but evidence that it plays a role in development of hypertension is lacking. Elevated circulating and urinary levels of endothelin-1 have been observed in clinical and experimental studies with angiogenesis inhibitors. Furthermore, the observation that endothelin receptor blockers can prevent or revert the rise in blood pressure during angiogenesis inhibition and attenuate proteinuria provides strong evidence that an activated endothelin-signalling pathway is a final common mediator of angiogenesis inhibition-induced rise in blood pressure and renal toxicity.

Sunitinib-induced systemic vasoconstriction in swine is endothelin mediated and does not involve nitric oxide or oxidative stress.

Angiogenesis inhibition with agents targeting tyrosine kinases of vascular endothelial growth factor receptors is an established anticancer treatment, but is, unfortunately, frequently accompanied by systemic hypertension and cardiac toxicity. Whether vascular endothelial growth factor receptor antagonism also has adverse effects on the pulmonary and coronary circulations is presently unknown. In chronically instrumented awake swine, the effects of the vascular endothelial growth factor receptor antagonist sunitinib on the systemic, pulmonary, and coronary circulation were studied. One week after sunitinib (50 mg PO daily), mean aortic blood pressure (MABP) had increased from 83±5 mm Hg at baseline to 97±6 mm Hg (P<0.05) because of a 57±20% increase in systemic vascular resistance as cardiac output decreased. In contrast, sunitinib had no discernible effects on pulmonary and coronary hemodynamics or cardiac function. We subsequently investigated the mechanisms underlying the sunitinib-induced systemic hypertension. Intravenous administration of NO synthase inhibitor N(G)-nitro-l-arginine increased MABP by 24±1 mm Hg under baseline conditions, whereas it increased MABP even further after sunitinib administration (32±3 mm Hg; P<0.05). Reactive oxygen species scavenging with a mixture of antioxidants lowered MABP by 13±2 mm Hg before but only by 5±2 mm Hg (P<0.05) after sunitinib administration. However, intravenous administration of the dual endothelin A/endothelin B receptor blocker tezosentan, which did not lower MABP at baseline, completely reversed MABP to presunitinib values. These findings indicate that sunitinib produces vasoconstriction selectively in the systemic vascular bed, without affecting pulmonary or coronary circulations. The sunitinib-mediated systemic hypertension is principally attributed to an increased vasoconstrictor influence of endothelin, with no apparent contributions of a loss of NO bioavailability or increased oxidative stress.

Sunitinib-induced hypothyroidism is due to induction of type 3 deiodinase activity and thyroidal capillary regression.

CONTEXT: Anticancer treatment with the tyrosine kinase inhibitor sunitinib causes thyroid dysfunction. OBJECTIVE: Our objective was to investigate the time course and underlying mechanisms of sunitinib-induced thyroid dysfunction. DESIGN: Thyroid function tests of 83 patients on sunitinib were collected retrospectively for their total treatment duration between January 2006 and November 2009 and prospectively in 15 patients on sunitinib for 10 wk. Additionally, thyroid function and histology were assessed in rats on sunitinib (8 d; n = 10) and after sunitinib withdrawal (11 d; n = 7) and compared with controls (n = 7). SETTING: Patients were seen at a university outpatient oncology clinic. Patients and Animals: Patients with metastatic renal cell carcinoma or gastrointestinal stromal tumors participated in the clinical study and Wistar Kyoto rats were used in the rat study. INTERVENTION: Sunitinib was taken according to a 4 wk "on," 2 wk "off" treatment regimen. Blood samples for measurement of thyroid function were collected at baseline and at wk 4 and 10. In rats, blood, liver, and thyroid were collected to assess thyroid hormones, deiodinase activity, and thyroid histology. MAIN OUTCOME MEASURES: TSH and free T(4) levels, deiodinase activity, and thyroid histology were assessed. RESULTS: Forty-two percent of patients in the retrospective study developed elevated TSH levels. Prospective analysis showed increased TSH levels within 10 wk of treatment, accompanied by a decreased T(3)/rT(3) ratio. In rats, serum T(4) and T(3) decreased, hepatic type 3 deiodinase activity increased, and thyroid histology showed marked capillary regression, which all but thyroid hormones reversed after sunitinib withdrawal. CONCLUSION: Sunitinib induces hypothyroidism due to alterations in T(4)/T(3) metabolism as well as thyroid capillary regression.

The vascular endothelial growth factor receptor inhibitor sunitinib causes a preeclampsia-like syndrome with activation of the endothelin system.

Angiogenesis inhibition is an established treatment for several tumor types. Unfortunately, this therapy is associated with adverse effects, including hypertension and renal toxicity, referred to as "preeclampsia." Recently, we demonstrated in patients and in rats that the multitarget tyrosine kinase inhibitor sunitinib induces a rise in blood pressure (BP), renal dysfunction, and proteinuria associated with activation of the endothelin system. In the current study we investigated the effects of sunitinib on rat renal histology, including the resemblance with preeclampsia, as well as the roles of endothelin 1, decreased nitric oxide (NO) bioavailability, and increased oxidative stress in the development of sunitinib-induced hypertension and renal toxicity. In rats on sunitinib, light and electron microscopic examination revealed marked glomerular endotheliosis, a characteristic histological feature of preeclampsia, which was partly reversible after sunitinib discontinuation. The histological abnormalities were accompanied by an increase in urinary excretion of endothelin 1 and diminished NO metabolite excretion. In rats on sunitinib alone, BP increased (ΔBP: 31.6±0.9 mm Hg). This rise could largely be prevented with the endothelin receptor antagonist macitentan (ΔBP: 12.3±1.5 mm Hg) and only mildly with Tempol, a superoxide dismutase mimetic (ΔBP: 25.9±2.3 mm Hg). Both compounds could not prevent the sunitinib-induced rise in serum creatinine or renal histological abnormalities and had no effect on urine nitrates but decreased proteinuria and urinary endothelin 1 excretion. Our findings indicate that both the endothelin system and oxidative stress play important roles in the development of sunitinib-induced proteinuria and that the endothelin system rather than oxidative stress is important for the development of sunitinib-induced hypertension.

[New insights into the pathogenesis of pre-eclampsia: the role of angiogenesis-inhibiting factors]. / Nieuwe inzichten in pathogenese van pre-eclampsie. De rol van angiogeneseremmende factoren.

The pathogenesis of pre-eclampsia is biphasic. The first phase is characterised by insufficient placentation and the second phase by an increased placental release of 2 anti-angiogenic factors, namely, soluble fms-like tyrosine kinase 1 (sFlt-1) and soluble endoglin (sEng). Within maternal circulation, sFlt-1 and sEng inhibit the effects of vascular endothelial growth factor (VEGF) and transforming growth factor ß (TGFß). This results in endothelial cell activation and inflammation, and eventually leads to the clinical syndrome of pre-eclampsia. The rise in plasma concentrations of sFlt-1 and sEng precedes the development of pre-eclampsia with 6-8 weeks. Whether elevations in the plasma concentrations of sFlt and sEng, combined with a decrease in placental growth factor concentrations, can be utilised as a predictor for pre-eclampsia is currently under investigation.

Hypertension induced by the tyrosine kinase inhibitor sunitinib is associated with increased circulating endothelin-1 levels.

Angiogenesis inhibition with sunitinib, a multitarget tyrosine kinase inhibitor of the vascular endothelial growth factor receptor, is associated with hypertension and cardiac toxicity, of which the underlying pathophysiological mechanism is unknown. We investigated the effects of sunitinib on blood pressure (BP), its circadian rhythm, and potential mechanisms involved, including the endothelin-1 system, in 15 patients with metastatic renal cell carcinoma or gastrointestinal stromal tumors. In addition, we investigated in rats the effect of sunitinib on BP, serum endothelin-1 levels, coronary microvascular function, cardiac structure, and cardiac mitochondrial function. In patients, BP increased by ≈15 mm Hg, whereas heart rate decreased after 4 weeks of treatment. Furthermore, the nocturnal dipping of BP diminished. Plasma endothelin-1 concentration increased 2-fold (P<0.05) and plasma renin decreased (P<0.05), whereas plasma catecholamines and renal function remained unchanged. In rats, 8 days of sunitinib administration induced an ≈30-mm Hg rise in BP, an attenuation of the circadian BP rhythm, and a 3-fold rise in serum endothelin-1 and creatinine, of which all but the rise in creatinine reversed after sunitinib withdrawal. Coronary microvascular function studies after 8 days of sunitinib administration showed decreased responses to bradykinin, angiotensin II, and sodium nitroprusside, all normalizing after sunitinib withdrawal. Cardiac structure and cardiac mitochondrial function did not change. In conclusion, sunitinib induces a reversible rise in BP in patients and in rats associated with activation of the endothelin-1 system, suppression of the renin-angiotensin system, and generalized microvascular dysfunction.

Light-induced vs. bradykinin-induced relaxation of coronary arteries: do S-nitrosothiols act as endothelium-derived hyperpolarizing factors?

BACKGROUND: Light-induced relaxation depends on S-nitrosothiols. S-Nitrosothiols may also serve as endothelium-derived hyperpolarizing factors, mediating the relaxant response of porcine coronary arteries (PCAs) to bradykinin. Here we compared the mechanism of light-induced and bradykinin-induced PCA relaxation. METHODS: PCAs were mounted in organ baths in the dark, preconstricted and exposed to polychromatic light (5 min) or 100 nmol/l bradykinin. RESULTS: Light relaxed PCAs by maximally 71 +/- 1%. S-Nitrosothiol depletion abolished this relaxation. Relaxations diminished following repetitive light exposures, particularly if the dark periods between the light exposures were less than 10 min, and increased following endothelium removal or nitric oxide synthase blockade with N(omega)-nitro-L-arginine methyl ester (L-NAME), despite the prevention of guanosine-3',5'-cyclic monophosphate generation by the latter two procedures. Thus, reloading of the storage pools occurs in the dark, endothelial nitric oxide inhibits this process and photorelaxation does not depend on guanosine-3',5'-cyclic monophosphate. Bradykinin relaxed PCAs by 69 +/- 3%. The nitric oxide scavenger hydroxocobalamin and the Na+-K+ ATPase inhibitor ouabain abolished the responses to bradykinin and light. The guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one abolished the response to light, and, like L-NAME, blocked the response to bradykinin by more than 50%. On top of L-NAME, intermediate and small conductance Ca2+-dependent K+ channel (IKCa/SKCa) blockade further reduced the response to bradykinin and enhanced photorelaxation. CONCLUSION: Photorelaxation depends on stored S-nitrosothiols and their release/synthesis is negatively affected by endothelial nitric oxide and IKCa/SKCa. S-Nitrosothiols activate endothelial IKCa/SKCa and, via guanylyl cyclase, smooth muscle Na+-K+ ATPase. Thus, they possess all properties of a bradykinin-induced endothelium-derived hyperpolarizing factor.