Towards the development of activity-based probes for detection of lysine-specific demethylase-1 activity

The implications of lysine-specific demethylase-1 (LSD1) in tumorigenesis have urged scientists to develop diagnostic tools in order to explore the function of this enzyme. In this work, we present our efforts on the development of tranylcypromine (TCP)-based functionalized probes for activity-based protein profiling (ABPP) of LSD1 activity. Biotinylated forms of selected compounds enabled dose-dependent enzyme labeling of recombinant LSD1. However, treatment with LSD1 inhibitors did not clearly reduce the LSD1 labeling efficiency thus indicating that labeling using these probes is not activity dependent. This calls for alternative strategies to develop probes for ABPP of the enzyme LSD1

Chemical epigenetics to assess the role of HDAC1-3 inhibition in macrophage pro-inflammatory gene expression

Histone deacetylases (HDACs) have been used as pharmacological targets for the treatment of various diseases. Some non-selective HDAC inhibitors (HDACi) have been clinically-used as therapeutic agents for treatment of hematological cancers but their cytotoxic side effects are an important downside. The discovery of more selective inhibitors has certified the involvement of individual HDACs in pathological processes but the elucidation of the role of specific family members in inflammatory responses still remains a challenge. Here, we report the development of closely related, structural analogues of the clinically-used HDACi Entinostat via a chemical epigenetic approach. Three compounds were designed and synthesized in which the cap moiety of Entinostat was replaced by an azobenzene group that is either para, meta or ortho substituted. The compounds were then evaluated for selectivity towards HDACs 1-3 and their effect on pro-inflammatory gene expression in macrophages. One analogue, compound 4, lacked selectivity and demonstrated inhibition of NF-kappa B reporter gene activity and pro-inflammatory gene expression in RAW264.7 macrophages, thus indicating that there is a delicate balance between the selectivity of HDACi over specific family members and their pro-or anti-inflammatory effects

Quinoline-based p300 histone acetyltransferase inhibitors with pro-apoptotic activity in human leukemia U937 cells.

Chemical manipulations performed on 2-methyl-3-carbethoxyquinoline (1), a histone acetyltransferase inhibitor previously identified by our research group and active at the sub-millimolar/millimolar level, led to compounds bearing higher alkyl groups at the C2-quinoline or additional side chains at the C6- quinoline positions. Such compounds displayed at least threefold improved inhibitory potency toward p300 protein lysine acetyltransferase activity; some of them decreased histone H3 and H4 acetylation levels in U937 cells and induced high degrees of apoptosis (three compounds >10-fold higher than compound 1) after treatment of U937 cells

Discovery of salermide-related sirtuin inhibitors: binding mode studies and antiproliferative effects in cancer cells including cancer stem cells.

Chemical changes performed on 1a (sirtinol) led to a series of SIRT1/2 inhibitors, in some cases more potent than 1a mainly against SIRT1. Tested in human leukemia U937 cells, the benzamide and anilide derivatives 1b, 1c, 2b, and 2c as well as the 4-(2-phenylpropyl)thioanalogue 4c showed huge apoptosis induction, while some sulfinyl and sulfonyl derivatives (5b, 5c, and 6a−c) were highly efficient in granulocytic differentiation. When assayed in human leukemia MOLT4 as well as in human breast MDA-MB-231 and colon RKO cancer cell lines, the anilide 2b (salermide) and the phenylpropylthio analogue 4b emerged as the most potent antiproliferative agents. Tested on colorectal carcinoma and glioblastoma multiforme cancer stem cells (CSCs) from patients, 2b was particularly potent against colorectal carcinoma CSCs, while 4b, 6a, and the SIRT2-selective inhibitor AGK-2 showed the highest effect against glioblastoma multiforme CSCs. Such compounds will be further explored for their broad-spectrum anticancer properties

Discovery of salermide-related sirtuin inhibitors: Binding mode studies and antiproliferative effects in cancer cells including cancer stem cells

Chemical changes performed on 1a (sirtinol) led to a series of SIRT1/2 inhibitors, in some cases more potent than 1a mainly against SIRT1. Tested in human leukemia U937 cells, the benzamide and anilide derivatives 1b, 1c, 2b, and 2c as well as the 4-(2-phenylpropyl)thioanalogue 4c showed huge apoptosis induction, while some sulfinyl and sulfonyl derivatives (5b, 5c, and 6a-c) were highly efficient in granulocytic differentiation. When assayed in human leukemia MOLT4 as well as in human breast MDA-MB-231 and colon RKO cancer cell lines, the anilide 2b (salermide) and the phenylpropylthio analogue 4b emerged as the most potent antiproliferative agents. Tested on colorectal carcinoma and glioblastoma multiforme cancer stem cells (CSCs) from patients, 2b was particularly potent against colorectal carcinoma CSCs, while 4b, 6a, and the SIRT2-selective inhibitor AGK-2 showed the highest effect against glioblastoma multiforme CSCs. Such compounds will be further explored for their broad-spectrum anticancer properties. \uc2\ua9 2012 American Chemical Society

1,3,4-Oxadiazole-Containing Histone Deacetylase Inhibitors: Anticancer Activities in Cancer Cells

We describe 1,3,4-oxadiazole-containing hydroxamates (<b>2</b>) and 2-aminoanilides (<b>3</b>) as histone deacetylase inhibitors. Among them, <b>2t</b>, <b>2x</b>, and <b>3i</b> were the most potent and selective against HDAC1. In U937 leukemia cells, <b>2t</b> was more potent than SAHA in inducing apoptosis, and <b>3i</b> displayed cell differentiation with a potency similar to MS-275. In several acute myeloid leukemia (AML) cell lines, as well as in U937 cells in combination with doxorubicin, <b>3i</b> showed higher antiproliferative effects than SAHA

An Inflammatory Signature to Predict the Clinical Benefit of First-Line Cetuximab Plus Platinum-Based Chemotherapy in Recurrent/Metastatic Head and Neck Cancer

Epidermal growth factor receptor (EGFR) pathway has been shown to play a crucial role in several inflammatory conditions and host immune-inflammation status is related to tumor prognosis. This study aims to evaluate the prognostic significance of a four-gene inflammatory signature in recurrent/metastatic (R/M) head and neck squamous cell carcinoma (HNSCC) patients treated with the EGFR inhibitor cetuximab plus chemotherapy. The inflammatory signature was assessed on 123 R/M HNSCC patients, enrolled in the multicenter trial B490 receiving first-line cetuximab plus platinum-based chemotherapy. The primary endpoint of the study was progression free survival (PFS), while secondary endpoints were overall survival (OS) and objective response rate (ORR). The patient population was subdivided into 3 groups according to the signature score groups. The four-genes-signature proved a significant prognostic value, resulting in a median PFS of 9.2 months in patients with high vs. 6.2 months for intermediate vs. 3.9 months for low values (p = 0.0016). The same findings were confirmed for OS, with median time of 18.4, 13.4, and 7.5 months for high, intermediate, and low values of the score, respectively (p = 0.0001). When ORR was considered, the signature was significantly higher in responders than in non-responders (p = 0.0092), reaching an area under the curve (AUC) of 0.65 (95% CI: 0.55-0.75). Our findings highlight the role of inflammation in the response to cetuximab and chemotherapy in R/M-HNSCC and may have translational implications for improving treatment selection