Preclinical validation of putative targets in cardiovascular and metabolic disease

Cardiovascular disease is the number one cause of death worldwide. The most important risk factor for developing this disease is high cholesterol levels in the blood. Other risk factors contributing to cardiovascular disease can develop in individuals which are overweight. The clinical consequences of being overweight are clustered in the medical term: metabolic syndrome. Included in the metabolic syndrome are high blood pressure, dyslipidemia and glucose intolerance. At present, most cardiovascular disease patients are treated with statins which lower blood cholesterol levels. However, this treatment is not as effective in all patients and can cause some adverse drug reactions. Therefore, it is essential that novel therapeutic targets for the treatment of cardiovascular disease are identified. In this thesis, potential novel therapeutic targets in cardiovascular disease and metabolic syndrome are validated. In total, three potential targets were investigated: proteoglycan 4, protein arginine methyltransferase 3 and stabilin 1. Our studies showed the involvement of two of these targets in the development of cardiovascular disease and metabolic syndrome. Moreover, our results stress (1) that cardiovascular disease and metabolic syndrome are complex, multifactorial diseases with overlapping mechanisms and (2) that integration of research into both diseases can benefit therapeutic target identification and validation. Hartstichting, KNMP, Greiner Bio-OneBiopharmaceutic

Total body proteoglycan 4 (Prg4) deficiency increases atherosclerosis susceptibility in apolipoprotein E knockout and low-density lipoprotein receptor knockout mice

Proteoglycan 4 (Prg4) is a mediator in inflammatory processes, either directly through interactions with CD44 [1] or toll-like receptors [2], or indirectly as a growth factor for hematopoietic cell lineages [3]. In line with this, in our article “Proteoglycan 4 regulates macrophage function without altering atherosclerotic lesion formation in a murine bone marrow-specific deletion model” published in Atherosclerosis, we showed that Prg4 deficiency attenuated the LPS-induced pro-inflammatory response in macrophages [4].Biopharmaceutic

Inhibition of PRMT3 activity selectively prevents LXR-driven transcription of hepatic lipogenic genes in vivo

Agonists for the liver X receptor (LXR) are considered promising therapeutic moieties in cholesterol-driven diseases by promoting cellular cholesterol efflux. However, current clinical application of these agents is hampered by the concomitant LXR-induced activation of a lipogenic transcriptional network, leading to hepatic steatosis. Recent studies have suggested that protein arginine methyltransferase 3 (PRMT3) may act as a selective co-activator of LXR activity. Here we verified the hypothesis that PRMT3 inhibition selectively disrupts the ability of LXR to stimulate lipogenesis, while maintaining the capacity of LXR to modulate cholesterol homeostasis.A combination of the LXR agonist T0901317 and palm oil was administered to C57BL/6 mice to maximally stimulate LXR and PRMT3 activity, in absence and presence of the allosteric PRMT3 inhibitor SGC707.Treatment with the PRMT3 inhibitor SGC707 did not negatively influence the T0901317/palm oil induced upregulation of the cholesterol efflux genes ABCA1 and ABCG1 in peritoneal cells. In contrast, SGC707 treatment was associated with a significant decrease in the hepatic expression of the lipogenic gene FAS (-64%; pobstruction of lipogenic gene transcription coincided with a significant 2.3-fold (ptriglyceride content as compared to the T0901317 and palm oil treated control group.Inhibition of PRMT3 activity by SGC707 treatment selectively impairs LXR-driven transcription of hepatic lipogenic genes, while the positive effect of LXR stimulation on cholesterol efflux pathways is maintained.Biopharmaceutic

Sr-Bi deficiency uncouples the development of hypertriglyceridemia and atherosclerosis from hepatic steatosis aand glucose intolerance in obese mice

Scavenger receptor BI (SR-BI) plays a quantitative role  in  triglyceride  metabolism.  However,  the  relevance  of  SR-BI  in  pa- thologies  associated  with  disturbances in  triglyceride  homeostasis,  i.e. obesity  and  fatty  liver  disease,  remains  largely  unknown.  We  therefore investigated the metabolic phenotype of SR-BI knockout mice and wild- type controls after a high-fat diet challenge.Mice were fed a high-fat diet for 12 weeks.SR-BI knockout mice gained significantly more weight than wild- type  control  mice  after  12  weeks  of  high-fat  diet  feeding  (13±1  grams versus 9±1 grams, respectively; Pdeficiency was associated with higher plasma levels of free cholesterol (+111%; Plesteryl  esters  (+89%;  Pcontrast, liver triglyceride levels were lower (-29%; Prelatively high hepatic expression levels of lipogenic  genes  ACC  (+54%;  P(+199%;  Pwild-type controls (+20%; Pglucose much better than wild-type mice as judged from  the  45%  lower  (P curve  of  the  oral  glucose tolerance  test.  Notably,  atherosclerotic  lesions  could  be  detected  in  the aortic root of all SR-BI knockout mice, while none were present in wild- type mice after the high-fat diet challenge.SR-BI deficiency disassociates hypertriglyceridemia, obesity, and  atherosclerosis  development  from  hepatic  steatosis  and  glucose intolerance in high-fat diet-fed mice. Our studies provide additional prooffor the biological relevance of SR-BI in total body triglyceride metabolism.Biopharmaceutic

Inhibition of protein arginine methyltransferase 3 activity selectively impairs liver X receptor-driven transcription of hepatic lipogenic genes in vivo

Agonists for the liver X receptor (LXR) are considered promising therapeutic moieties in cholesterol-driven diseases by promoting cellular cholesterol efflux pathways. However, current clinical application of these agents is hampered by concomitant LXR-induced activation of a lipogenic transcriptional network, leading to hepatic steatosis. Recent studies have suggested that protein arginine methyltransferase 3 (PRMT3) may act as a selective co-activator of LXR activity. Here, we verified the hypothesis that PRMT3 inhibition selectively disrupts the ability of LXR to stimulate lipogenesis while maintaining its capacity to modulate macrophage cholesterol homeostasis. A combination of the LXR agonist T0901317 and palm oil was administered to C57BL/6 mice to maximally stimulate LXR and PRMT3 activity. PRMT3 activity was inhibited using the allosteric inhibitor SGC707. Treatment with SGC707 did not negatively influence the T0901317/palm oil-induced up-regulation of the cholesterol efflux ATP-binding cassette transporter genes, ABCA1 and ABCG1, in peritoneal cells. In contrast, SGC707 treatment was associated with a significant decrease in the hepatic expression of the lipogenic gene fatty acid synthase (-64%). A similar trend was observed for stearoyl-coenzyme A desaturase and acetyl CoA carboxylase expression (-43%; -56%). This obstruction of lipogenic gene transcription coincided with a significant 2.3-fold decrease in liver triglyceride content as compared with the T0901317 and palm oil-treated control group. We showed that inhibition of PRMT3 activity by SGC707 treatment selectively impairs LXR-driven transcription of hepatic lipogenic genes, while the positive effect of LXR stimulation on macrophage cholesterol efflux pathways is maintained.Biopharmaceutic

Simvastatin treatment aggravates the glucocorticoid insufficiency associated with hypocholesterolemia in mice

Statin treatment disrupts HMG-CoA reductase-mediated endogenous cholesterol synthesis and lowers plasma LDL-cholesterol levels. Although statin treatment can theoretically impair adrenal steroid hormone synthesis, thus far, no effect on glucocorticoid output has been described, as LDL-cholesterol levels usually remain within the physiological range. However, novel statin-based treatment regimens that dramatically decrease LDL-cholesterol levels are currently employed. Here, we assessed whether inhibition of cholesterol synthesis under these relatively hypocholesterolemic conditions may alter adrenal glucocorticoid output. Hypocholesterolemic apolipoprotein A1 (apoA1) knockout mice were administered high dose simvastatin twice daily for 3 days. Simvastatin treatment did not change plasma cholesterol levels or modify the adrenal expression levels of genes involved in cholesterol metabolism. However, simvastatin treatment lowered basal plasma levels of the primary glucocorticoid corticosterone (-62%; p < 0.05). Upon injection with adrenocorticotropic hormone, control-treated apoA1 knockout mice already showed only a mild increase in plasma corticosterone levels, indicative of relative glucocorticoid insufficiency. Importantly, simvastatin treatment further diminished the adrenal glucocorticoid response to adrenocorticotropic hormone exposure (two-way ANOVA p < 0.05 for treatment). Peak corticosterone levels were 49% lower (p < 0.01) upon simvastatin treatment. We have shown that simvastatin treatment aggravates the glucocorticoid insufficiency associated with hypocholesterolemia in mice. Our data suggest that (1) HMG-CoA reductase activity controls the adrenal steroidogenic capacity under hypocholesterolemic conditions and (2) imply that it might be important to monitor adrenal function in humans subjected to statin-based treatments aimed at achieving sub-physiological LDL-cholesterol levels, as these may potentially execute a negative impact on the glucocorticoid function in humans.Biopharmaceutic

Inhibition of PRMT3 activity reduces hepatic steatosis without altering atherosclerosis susceptibility in apoE knockout mice

The nuclear receptor liver X receptor (LXR) impacts on cholesterol metabolism as well as hepatic lipogenesis via transcriptional regulation. It is proposed that inhibition of the protein arginine methyltransferase 3 (PRMT3) uncouples these two transcriptional pathways in vivo by acting as a specific lipogenic coactivator of LXR. Here we validated the hypothesis that treatment with the allosteric PRMT3 inhibitor SGC707 will diminish the hepatic steatosis extent, while leaving global cholesterol metabolism, important in cholesterol-driven pathologies like atherosclerosis, untouched. For this purpose, 12-week old hyperlipidemic apolipoprotein E knockout mice were fed a Western-type diet for six weeks to induce both hepatic steatosis and atherosclerosis. The mice received 3 intraperitoneal injections with SGC707 or solvent control per week. Mice chronically treated with SGC707 developed less severe hepatic steatosis as exemplified by the 51% reduced (P < 0.05) liver triglyceride levels. In contrast, the extent of in vivo macrophage foam cell formation and aortic root atherosclerosis was not affected by SGC707 treatment. Interestingly, SGC707-treated mice gained 94% less body weight (P < 0.05), which was paralleled by changes in white adipose tissue morphology, i.e. reduction in adipocyte size and browning. In conclusion, we have shown that through PRMT3 inhibitor treatment specific functions of LXR involved in respectively the development of fatty liver disease and atherosclerosis can be uncoupled, resulting in an overall diminished hepatic steatosis extent without a negative impact on atherosclerosis susceptibility. As such, our studies highlight that PRMT3 inhibition may constitute a novel therapeutic approach to limit the development of fatty liver disease in humans.Biopharmaceutic

Quantitative Monitoring of NPM1A Mutations Can Be Used to Guide Immunotherapeutic Interventions After Allogeneic Stem Cell Transplantation for Acute Myeloid Leukemia

Immunobiology of allogeneic stem cell transplantation and immunotherapy of hematological disease

Quantitative Monitoring of NPM1A Mutations Can Be Used to Guide Immunotherapeutic Interventions After Allogeneic Stem Cell Transplantation for Acute Myeloid Leukemia

Immunobiology of allogeneic stem cell transplantation and immunotherapy of hematological disease