Effects of SGLT2 Inhibitors and GLP-1 Receptor Agonists on Renin-Angiotensin-Aldosterone System.

Sodium-glucose cotransporters inhibitors (SGLT2-i) and GLP-1 receptor agonists (GLP1-RA) are glucose-lowering drugs that are proved to reduce the cardiovascular (CV) risk in type 2 diabetes mellitus (T2DM). In this process, the renin-angiotensin-aldosterone system (RAAS) is assumed to play a role. The inhibition of SGLT2 improves hyperglycemia hampering urinary reabsorption of glucose and inducing glycosuria. This "hybrid" diuretic effect, which couples natriuresis with osmotic diuresis, potentially leads to systemic RAAS activation. However, the association between SGLT2-i and systemic RAAS activation is not straightforward. Available data indicate that SGLT2-i cause plasma renin activity (PRA) increase in the early phase of treatment, while PRA and aldosterone levels remain unchanged in chronic treated patients. Furthermore, emerging studies provide evidence that SGLT2-i might have an interfering effect on aldosterone/renin ratio (ARR) in patients with T2DM, due to their diuretic and sympathoinhibition effects. The cardio- and reno-protective effects of GLP-1-RA are at least in part related to the interaction with RAAS. In particular, GLP1-RA counteract the action of angiotensin II (ANG II) inhibiting its synthesis, increasing the inactivation of its circulating form and contrasting its action on target tissue like glomerular endothelial cells and cardiomyocytes. Furthermore, GLP1-RA stimulate natriuresis inhibiting Na+/H+ exchanger NHE-3, which is conversely activated by ANG II. Moreover, GLP1 infusion acutely reduces circulating aldosterone, but this effect does not seem to be chronically maintained in patients treated with GLP1-RA. In conclusion, both SGLT2-i and GLP1-RA seem to have several effects on RAAS, though additional studies are needed to clarify this relationship.

Malignant struma ovarii: next-generation sequencing of six cases revealed Nras, Braf, and Jak3 mutations.

PURPOSE: Struma ovarii (SO) is a highly specialized ovarian teratoma, consisting of thyroid tissue. Rarely, carcinomas histologically identical to their thyroid counterparts may occur, and are comprehensively defined as malignant struma ovarii (MSO). Their optimal management is controversial, and the molecular profile of the malignant counterpart in the ovary is incompletely known. In this study, the clinicopathological and molecular features of six MSO from different Italian Institutions were analysed, to explore genetic profiles of potential therapeutic interest. METHODS: The histopathological features and immunoprofile (according to the known markers Galectin-3, HBME1, cytokeratin 19 and CD56) were reviewed. In addition, all cases underwent genetic analysis with a next-generation sequencing (NGS) hot spot cancer panel detecting mutations in 50 genes involved in cancerogenesis. RET/PTC rearrangements and TERT promoter alterations were also evaluated. RESULTS: Papillary carcinoma in all similar to its thyroid counterpart was found in five of six cases, including classical (two tumors) and follicular variant (three tumors) types. The last case was a poorly differentiated carcinoma. An activating gene mutation, was detected in five of six cases, including two NRAS, two BRAF, and one JAK3 oncogene mutations. No alterations were found in the other panel genes, nor in TERT promoter, or in RET chromosomal regions. CONCLUSIONS: MSO is a rare condition. Papillary carcinoma is the predominant malignant type, sharing both histomorphological and molecular features of its thyroid counterpart. Interestingly, the single case of poorly differentiated carcinoma displayed a JAK3 mutation. The presence of such driving mutation could be of potential interest in guiding postoperative treatment.

Correction to: Malignant struma ovarii: next-generation sequencing of six cases revealed Nras, Braf, and Jak3 mutations.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Histone deacetylase inhibition modulates E-cadherin expression and suppresses migration and invasion of anaplastic thyroid cancer cells.

CONTEXT: Anaplastic thyroid cancer cells are characterized by a mesenchymal phenotype, as revealed by spindle-shaped cells and absent or reduced levels of E-cadherin. Epigenetic silencing is considered one of the leading mechanisms of E-cadherin impairment, which causes the acquisition of the invasive and metastatic phenotype of anaplastic thyroid cancer. OBJECTIVES: In this study we investigated the effects of histone deacetylase inhibition on E-cadherin expression, cell motility, and invasion in anaplastic thyroid cancer cell cultures. DESIGN: Three stabilized cell lines and primary cultures of anaplastic thyroid cancer were treated with various histone deacetylase inhibitors. After treatment, we evaluated histone acetylation by Western blotting and E-cadherin expression by RT-real time PCR. The proper localization of E-cadherin/ß-catenin complex was assessed by immunofluorescence and Western blot. Transcription activity of ß-catenin was measured by luciferase reporter gene and cyclin D1 expression. The effect on cell motility and invasion was studied both in vitro using scratch-wound and transwell invasion assays and in anaplastic thyroid carcinomas tumor xenografts in mice in vivo. RESULTS: Histone deacetylase inhibition induced the E-cadherin expression and the proper membrane localization of the E-cadherin/ß-catenin complex, leading to reduced cancer cell migration and invasion. CONCLUSIONS: We here demonstrate an additional molecular mechanism for the anticancer effect of histone deacetylase inhibition. The antiinvasive effect in addition to the cytotoxic activity of histone deacetylase inhibitors opens up therapeutic perspectives for the anaplastic thyroid tumor that does not respond to conventional therapy.

Cytotoxic activity of the histone deacetylase inhibitor panobinostat (LBH589) in anaplastic thyroid cancer in vitro and in vivo.

Anaplastic thyroid carcinoma (ATC) has a rapidly fatal clinical course, being resistant to multimodal treatments. Microtubules, α/ß tubulin heterodimers, are crucial in cell signaling, division and mitosis and are among the most successful targets for anticancer therapy. Panobinostat (LBH589) is a potent deacetylase inhibitor acting both on histones and nonhistonic proteins, including α-tubulin. In vitro LBH589, evaluated in three ATC cell lines (BHT-101, CAL-62 and 8305C), resulted in impairment of cell viability, inhibition of colony formation, cell cycle arrest and apoptosis induction. Mechanistically, we showed that LBH589 not only affected the expression of p21 and cyclin D1, but markedly determined microtubule stabilization as evidenced by tubulin acetylation and increased tubulin polymerization. In a SCID xenograft model implanted with CAL-62 cells, the cytotoxic properties of LBH589 were confirmed. The drug at the dose of 20 mg/kg significantly impaired tumor growth (final tumor volume 2.5-fold smaller than in untreated animals); at this dose, no relevant side effects were observed. In tumors of treated animals, a significant reduction of Ki67, which was negatively correlated with tubulin acetylation, was observed. Moreover, acetyl-tubulin levels negatively correlated with tumor volume at sacrifice, reinforcing the opinion that tubulin acetylation has a role in the inhibition of tumor growth. In conclusion, LBH589, acting on both histones and nonhistonic proteins in anaplastic thyroid cancer, appears to be a promising therapeutic agent for the treatment of this kind of cancer which is known not to respond to conventional therapy.

Emerging molecular therapies of advanced thyroid cancer.

Advanced thyroid cancer refers to thyroid tumors which are resistant to conventional therapies and do not respond to radioiodine and comprises metastatic or recurrent differentiated cancers, poorly differentiated and anaplastic tumors. Progress in the knowledge of genetic/epigenetic alterations in thyroid cancer cells is rapidly offering several opportunities to develop new drugs directed to specific targets. Drugs currently proposed for molecular therapy include: (a) monoclonal antibodies; (b) kinase inhibitors; (c) anti-angiogenetic drugs; (d) proteasome inhibitors; (e) retinoic acid and PPAR-gamma ligands; (f) radionuclide therapy; (g) epigenetic drugs (deacetylase inhibitors and demethylating agents). The results of several phase II trials using molecular drugs look promising. None of the treated patients, however, had a complete response, and only a minority of them had a partial response. The review will focus especially on epigenetic therapy, whose goal is to target the chromatin in rapidly dividing tumor cells and potentially restore normal cell functions. Deacetylases inhibitors modulate both epigenetic and multiple non-epigenetic mechanisms; they are, thus, viewed as a promising class of anticancer drugs. Experimental data show that deacetylase inhibitors are effective against advanced thyroid cancer. However, since multiple pathways need to be inhibited in order to substantially affect thyroid cancer growth, it is likely that a significant increase in the response rate to treatment of advanced thyroid cancer will be achieved through combinatorial drug therapies. Actually, many pre-clinical and clinical studies evaluate the combination of either two epigenetic drugs or a non-epigenetic chemotherapeutic and an epigenetic drug, in the effort to increase response rates.

Effects of the histone deacetylase inhibitor valproic acid on the sensitivity of anaplastic thyroid cancer cell lines to imatinib.

New therapeutic approaches are mandatory for anaplastic thyroid cancer. We investigated the ability of a new combined treatment using valproic acid (VPA), the only clinically available histone deacetylase inhibitor, and the tyrosine-kinase inhibitor imatinib mesylate to control the cell growth of anaplastic thyroid cancer cell lines. We showed that treatment with imatinib alone is unable to affect the cell growth of anaplastic thyroid cancer cells, whereas in ARO cells, the combined treatment resulted in a cytostatic effect, with clinically achievable doses of imatinib and VPA. The effect is mediated by G1 growth arrest, acting through p21 expression and the impairment of AKT phosphorylation.

Valproic acid enhances tubulin acetylation and apoptotic activity of paclitaxel on anaplastic thyroid cancer cell lines.

The introduction of paclitaxel into multimodal therapy for anaplastic thyroid carcinoma has failed to improve overall survival. Toxicity rules out the high doses required, especially in older patients. The search for strategies to enhance paclitaxel antineoplastic activity and reduce its side effects is thus advisable. The study aimed to determine whether the histone deacetylase (HDAC) inhibitor valproic acid (VPA) improves the anticancer action of paclitaxel and elucidate the mechanisms underlying the effects of combined treatment. We examined the effect of VPA on the sensitivity to paclitaxel of two anaplastic thyroid carcinoma cell lines (CAL-62 and ARO), and the ability of the drug to determine tubulin acetylation and enhance paclitaxel-induced acetylation. The addition of as little as 0.7 mM VPA to paclitaxel enhances both cytostatic and cytotoxic effects of paclitaxel alone. Increased apoptosis explains the enhancement of the cytotoxic effect. The mechanism underlying this effect is through inhibition of HDAC6 activity, which leads to tubulin hyperacetylation. The results suggest a mechanistic link between HDAC6 inhibition, tubulin acetylation, and the VPA-induced enhancement of paclitaxel effects, and provide the rationale for designing future combination therapies.

Valproic acid, a histone deacetylase inhibitor, enhances sensitivity to doxorubicin in anaplastic thyroid cancer cells.

Multimodality treatments (i.e. surgery, chemotherapy, and radiotherapy) are recommended for anaplastic thyroid carcinoma (ATC), an extremely lethal human cancer, but to date there is little evidence that such approaches improve survival rates. It is thus necessary to seek new therapeutic tools. Histone deacetylase (HDAC) inhibitors are a promising class of anti-neoplastic agents that induce differentiation and apoptosis. Moreover, they may enhance the cytotoxicity of drugs targeting DNA through acetylation of histones. Using two ATC cell lines (CAL-62 and ARO), we show here that valproic acid (VPA), a clinically available HDAC inhibitor, enhances the activity of doxorubicin, whose anti-tumor properties involve binding to DNA and inhibiting topoisomerase II. A meager 0.7 mM VPA, which corresponds to serum concentrations in patients treated for epilepsy, is able to increase the cytotoxicity of doxorubicin about threefold in CAL-62 cells and twofold in ARO cells. The sensitizing effect, which is through histone acetylation, involves increased apoptosis, which is also shown by the increased caspase 3 activation and the enhancement of doxorubicin-induced G2 cell cycle arrest. These results might offer a rationale for clinical studies of a new combined therapy in an effort to improve the outcome of patients with anaplastic thyroid cancer.

Valproic acid induces apoptosis and cell cycle arrest in poorly differentiated thyroid cancer cells.

Poorly differentiated thyroid carcinoma is an aggressive human cancer that is resistant to conventional therapy. Histone deacetylase inhibitors are a promising class of drugs, acting as antiproliferative agents by promoting differentiation, as well as inducing apoptosis and cell cycle arrest. Valproic acid (VPA), a class I selective histone deacetylase inhibitor widely used as an anticonvulsant, promotes differentiation in poorly differentiated thyroid cancer cells by inducing Na(+)/I(-) symporter and increasing iodine uptake. Here, we show that it is also highly effective at suppressing growth in poorly differentiated thyroid cancer cell lines (N-PA and BHT-101). Apoptosis induction and cell cycle arrest are the underlying mechanisms of VPA's effect on cell growth. It induces apoptosis by activating the intrinsic pathway; caspases 3 and 9 are activated but not caspase 8. Cell cycle is selectively arrested in G(1) and is associated with the increased expression of p21 and the reduced expression of cyclin A. Both apoptosis and cell cycle arrest are induced by treatment with 1 mm VPA, a dose that promotes cell redifferentiation and that is slightly above the serum concentration reached in patients treated for epilepsy. These multifaceted properties make VPA of clinical interest as a new approach to treating poorly differentiated thyroid cancer.