Publications by authors named "Roberto Dal Zuffo"

12 Publications

  • Page 1 of 1

Discovery of Reversible Inhibitors of KDM1A Efficacious in Acute Myeloid Leukemia Models.

ACS Med Chem Lett 2020 May 13;11(5):754-759. Epub 2020 Feb 13.

Department of Experimental Oncology, Academic Drug Discovery, European Institute of Oncology IRCCS, Via Adamello 16, 20139 Milan, Italy.

Lysine-specific demethylase 1 (LSD1 or KDM1A) is a FAD-dependent enzyme that acts as a transcription corepressor or coactivator by regulating the methylation status of histone H3 lysines K4 and K9, respectively. KDM1A represents an attractive target for cancer therapy. While, in the past, the main medicinal chemistry strategy toward KDM1A inhibition was based on the optimization of ligands that irreversibly bind the FAD cofactor within the enzyme catalytic site, we and others have also identified reversible inhibitors. Herein we reported the discovery of 5-imidazolylthieno[3,2-]pyrroles, a new series of KDM1A inhibitors endowed with picomolar inhibitory potency, active in cells and efficacious after oral administration in murine leukemia models.
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http://dx.doi.org/10.1021/acsmedchemlett.9b00604DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7236255PMC
May 2020

Targeting the scaffolding role of LSD1 (KDM1A) poises acute myeloid leukemia cells for retinoic acid-induced differentiation.

Sci Adv 2020 04 8;6(15):eaax2746. Epub 2020 Apr 8.

Department of Experimental Oncology, European Institute of Oncology (IEO), IRCCS, Via Adamello 16, Milan 20139, Italy.

The histone demethylase LSD1 is deregulated in several tumors, including leukemias, providing the rationale for the clinical use of LSD1 inhibitors. In acute promyelocytic leukemia (APL), pharmacological doses of retinoic acid (RA) induce differentiation of APL cells, triggering degradation of the PML-RAR oncogene. APL cells are resistant to LSD1 inhibition or knockout, but targeting LSD1 sensitizes them to physiological doses of RA without altering of PML-RAR levels, and extends survival of leukemic mice upon RA treatment. The combination of RA with LSD1 inhibition (or knockout) is also effective in other non-APL, acute myeloid leukemia (AML) cells. Nonenzymatic activities of LSD1 are essential to block differentiation, while RA with targeting of LSD1 releases a differentiation gene expression program, not strictly dependent on changes in histone H3K4 methylation. Integration of proteomic/epigenomic/mutational studies showed that LSD1 inhibitors alter the recruitment of LSD1-containing complexes to chromatin, inhibiting the interaction between LSD1 and the transcription factor GFI1.
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http://dx.doi.org/10.1126/sciadv.aax2746DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7141832PMC
April 2020

Novel potent inhibitors of the histone demethylase KDM1A (LSD1), orally active in a murine promyelocitic leukemia model.

Future Med Chem 2017 07 19;9(11):1161-1174. Epub 2017 Jul 19.

Department of Experimental Oncology, Academic Drug Discovery, European Institute of Oncology, Ifom-IEO-Campus, via Adamello 16, 20139 Milan, Italy.

Background: Histone lysine demethylases (KDMs) are well-recognized targets in oncology drug discovery. They function at the post-translation level controlling chromatin conformation and gene transcription. KDM1A is a flavin adenine dinucleotide-dependent amine oxidase, overexpressed in several tumor types, including acute myeloid leukemia, neuroblastoma and non-small-cell lung cancer. Among the many known monoamine oxidase inhibitors screened for KDM1A inhibition, tranylcypromine emerged as a moderately active hit, which irreversibly binds to the flavin adenine dinucleotide cofactor.

Material & Methods: The KDM1A inhibitors 5a-w were synthesized and tested in vitro and in vivo. The biochemical potency was determined, modulation of target in cells was demonstrated on KDM1A-dependent genes and the anti-clonogenic activity was performed in murine acute promyelocytic Leukemia (APL) blasts. An in vivo efficacy experiment was conducted using an established murine promyelocytic leukemia model.

Results: We report a new series of tranylcypromine derivatives substituted on the cyclopropyl moiety, endowed with high potency in both biochemical and cellular assays.

Conclusion: The most interesting derivative (5a) significantly improved survival rate after oral administration in a murine model of promyelocitic leukemia.
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http://dx.doi.org/10.4155/fmc-2017-0003DOI Listing
July 2017

Discovery of a Novel Inhibitor of Histone Lysine-Specific Demethylase 1A (KDM1A/LSD1) as Orally Active Antitumor Agent.

J Med Chem 2016 Feb 7;59(4):1501-17. Epub 2016 Jan 7.

Department of Experimental Oncology, Academic Drug Discovery, European Institute of Oncology , Via Adamello 16, 20139 Milan, Italy.

We report the stereoselective synthesis and biological activity of a novel series of tranylcypromine (TCPA) derivatives (14a-k, 15, 16), potent inhibitors of KDM1A. The new compounds strongly inhibit the clonogenic potential of acute leukemia cell lines. In particular three molecules (14d, 14e, and 14g) showing selectivity versus MAO A and remarkably inhibiting colony formation in THP-1 human leukemia cells, were assessed in mouse for their preliminary pharmacokinetic. 14d and 14e were further tested in vivo in a murine acute promyelocytic leukemia model, resulting 14d the most effective. Its two enantiomers were synthesized: the (1S,2R) enantiomer 15 showed higher activity than its (1R,2S) analogue 16, in both biochemical and cellular assays. Compound 15 exhibited in vivo efficacy after oral administration, determining a 62% increased survival in mouse leukemia model with evidence of KDM1A inhibition. The biological profile of compound 15 supports its further investigation as a cancer therapeutic.
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http://dx.doi.org/10.1021/acs.jmedchem.5b01209DOI Listing
February 2016

Beclin 1 restrains tumorigenesis through Mcl-1 destabilization in an autophagy-independent reciprocal manner.

Nat Commun 2014 Dec 4;5:5637. Epub 2014 Dec 4.

1] Department of Experimental Oncology, European Institute of Oncology, IEO, 20139 Milan, Italy [2] Department of Biosciences, University of Milan, 20100 Milan, Italy [3] Drug Development Program, European Institute of Oncology, IEO, 20139 Milan, Italy.

Mcl-1 is a unique Bcl-2 family member that plays crucial roles in apoptosis. Apoptosis-unrelated functions of Mcl-1 are however emerging, further justifying its tight regulation. Here we unravel a novel mechanism of Mcl-1 regulation mediated by the haplo-insufficient tumour suppressor Beclin 1. Beclin 1 negatively modulates Mcl-1 stability in a reciprocal manner whereby depletion of one leads to the stabilization of the other. This co-regulation is independent of autophagy and of their physical interaction. Both Beclin 1 and Mcl-1 are deubiquitinated and thus stabilized by binding to a common deubiquitinase, USP9X. Beclin 1 and Mcl-1 negatively modulate the proteasomal degradation of each other through competitive displacement of USP9X. The analysis of patient-derived melanoma cells and tissue samples shows that the levels of Beclin 1 decrease, while Mcl-1 levels subsequently increase during melanoma progression in a significant inter-dependent manner. The identified inverse co-regulation of Beclin 1 and Mcl-1 represents a mechanism of functional counteraction in cancer.
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http://dx.doi.org/10.1038/ncomms6637DOI Listing
December 2014

Chiral resolution and pharmacological characterization of the enantiomers of the Hsp90 inhibitor 2-amino-7-[4-fluoro-2-(3-pyridyl)phenyl]-4-methyl-7,8-dihydro-6H-quinazolin-5-one oxime.

ChemMedChem 2014 Jul 17;9(7):1574-85. Epub 2014 Apr 17.

Genextra Group, Congenia s.r.l., Via Adamello 16, 20139 Milan (Italy); Current address: Drug Discovery Program, Department of Experimental Oncology, IEO-European Institute of Oncology, Via Adamello 16, 20139 Milan (Italy).

Heat-shock protein 90 (Hsp90) is a molecular chaperone involved in the stabilization of key oncogenic signaling proteins, and therefore, inhibition of Hsp90 represents a new strategy in cancer therapy. 2-Amino-7-[4-fluoro-2-(3-pyridyl)phenyl]-4-methyl-7,8-dihydro-6H-quinazolin-5-one oxime is a racemic Hsp90 inhibitor that targets the N-terminal adenosine triphosphatase site. We developed a method to resolve the enantiomers and evaluated their inhibitory activity on Hsp90 and the consequent antitumor effects. The (S) stereoisomer emerged as a potent Hsp90 inhibitor in biochemical and cellular assays. In addition, this enantiomer exhibited high oral bioavailability in mice and excellent antitumor activity in two different human cancer xenograft models.
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http://dx.doi.org/10.1002/cmdc.201400037DOI Listing
July 2014

Synthesis and biological characterization of spiro[2H-(1,3)-benzoxazine-2,4'-piperidine] based histone deacetylase inhibitors.

Eur J Med Chem 2013 Jun 10;64:273-84. Epub 2013 Apr 10.

Genextra Group, Congenia s.r.l., Via Adamello 16, 20139 Milan, Italy.

Histone Deacetylases (HDACs) have become important targets for the treatment of cancer and other diseases. In previous studies we described the development of novel spirocyclic HDAC inhibitors based on the combination of privileged structures with hydroxamic acid moieties as zinc binding group. Herein, we report further explorations, which resulted in the discovery of a new class of spiro[2H-(1,3)-benzoxazine-2,4'-piperidine] derivatives. Several compounds showed good potency of around 100 nM and less in the HDAC inhibition assays, submicromolar IC50 values when tested against tumour cell lines and a remarkable stability in human and mouse microsomes. Two representative examples exhibited a good pharmacokinetic profile with an oral bioavailability equal or higher than 35% and one of them studied in an HCT116 murine xenograft model showing a robust tumour growth inhibition. In addition, the two benzoxazines were found to have a minor affinity for the hERG potassium channel compared to their corresponding ketone analogues.
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http://dx.doi.org/10.1016/j.ejmech.2013.03.061DOI Listing
June 2013

A dual role for Hdac1: oncosuppressor in tumorigenesis, oncogene in tumor maintenance.

Blood 2013 Apr 25;121(17):3459-68. Epub 2013 Feb 25.

Department of Experimental Oncology at the IFOM-IEO Campus, European Institute of Oncology, via Adamello 16, Milan, Italy.

Aberrant recruitment of histone deacetylases (HDACs) by the oncogenic fusion protein PML-RAR is involved in the pathogenesis of acute promyelocytic leukemia (APL). PML-RAR, however, is not sufficient to induce disease in mice but requires additional oncogenic lesions during the preleukemic phase. Here, we show that knock-down of Hdac1 and Hdac2 dramatically accelerates leukemogenesis in transgenic preleukemic mice. These events are not restricted to APL because lymphomagenesis driven by deletion of p53 or, to a lesser extent, by c-myc overexpression, was also accelerated by Hdac1 knock-down. In the preleukemic phase of APL, Hdac1 counteracts the activity of PML-RAR in (1) blocking differentiation; (2) impairing genomic stability; and (3) increasing self-renewal in hematopoietic progenitors, as all of these events are affected by the reduction in Hdac1 levels. This led to an expansion of a subpopulation of PML-RAR-expressing cells that is the major source of leukemic stem cells in the full leukemic stage. Remarkably, short-term treatment of preleukemic mice with an HDAC inhibitor accelerated leukemogenesis. In contrast, knock-down of Hdac1 in APL mice led to enhanced survival duration of the leukemic animals. Thus, Hdac1 has a dual role in tumorigenesis: oncosuppressive in the early stages, and oncogenic in established tumor cells.
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http://dx.doi.org/10.1182/blood-2012-10-461988DOI Listing
April 2013

Spiro[chromane-2,4'-piperidine]-based histone deacetylase inhibitors with improved in vivo activity.

ChemMedChem 2012 Apr 22;7(4):709-21. Epub 2012 Feb 22.

Genextra Group, Congenia s.r.l., Via Adamello 16, 20139 Milan,

A series of spiro[chromane-2,4'-piperidine] derivatives based on a previously published lead benzyl spirocycle 1 and bearing various N-aryl and N-alkylaryl substituents on the piperidine ring were prepared as novel histone deacetylase (HDAC) inhibitors. The compounds were evaluated for their abilities to inhibit nuclear HDACs, their in vitro antiproliferative activities, and in vitro ADME profiles. Based on these activities, 4-fluorobenzyl and 2-phenylethyl spirocycles were selected for further characterization. In vivo pharmacokinetic (PK) studies showed that both compounds exhibit an overall lower clearance rate, an increased half-life, and higher AUCs after intravenous and oral administration than spiropiperidine 1 under the conditions used. The improved PK behavior of these two compounds also correlated with superior in vivo antitumor activity in an HCT-116 xenograft model.
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http://dx.doi.org/10.1002/cmdc.201200024DOI Listing
April 2012

Discovery, synthesis, and pharmacological evaluation of spiropiperidine hydroxamic acid based derivatives as structurally novel histone deacetylase (HDAC) inhibitors.

J Med Chem 2011 Apr 1;54(8):3051-64. Epub 2011 Apr 1.

Genextra Group, DAC SRL, Milan, Italy.

New spiro[chromane-2,4'-piperidine] and spiro[benzofuran-2,4'-piperidine] hydroxamic acid derivatives as HDAC inhibitors have been identified by combining privileged structures with a hydroxamic acid moiety as zinc binding group. The compounds were evaluated for their ability to inhibit nuclear extract HDACs and for their in vitro antiproliferative activity on different tumor cell lines. This work resulted in the discovery of spirocycle 30d that shows good oral bioavailability and tumor growth inhibition in an HCT-116 murine xenograft model.
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http://dx.doi.org/10.1021/jm200146uDOI Listing
April 2011

Interplay between oncogene-induced DNA damage response and heterochromatin in senescence and cancer.

Nat Cell Biol 2011 Mar 20;13(3):292-302. Epub 2011 Feb 20.

IFOM Foundation - FIRC Institute of Molecular Oncology Foundation, Milan, Italy.

Two major mechanisms have been causally implicated in the establishment of cellular senescence: the activation of the DNA damage response (DDR) pathway and the formation of senescence-associated heterochromatic foci (SAHF). Here we show that in human fibroblasts resistant to premature p16(INK4a) induction, SAHF are preferentially formed following oncogene activation but are not detected during replicative cellular senescence or on exposure to a variety of senescence-inducing stimuli. Oncogene-induced SAHF formation depends on DNA replication and ATR (ataxia telangiectasia and Rad3-related). Inactivation of ATM (ataxia telangiectasia mutated) or p53 allows the proliferation of oncogene-expressing cells that retain increased heterochromatin induction. In human cancers, levels of heterochromatin markers are higher than in normal tissues, and are independent of the proliferative index or stage of the tumours. Pharmacological and genetic perturbation of heterochromatin in oncogene-expressing cells increase DDR signalling and lead to apoptosis. In vivo, a histone deacetylase inhibitor (HDACi) causes heterochromatin relaxation, increased DDR, apoptosis and tumour regression. These results indicate that heterochromatin induced by oncogenic stress restrains DDR and suggest that the use of chromatin-modifying drugs in cancer therapies may benefit from the study of chromatin and DDR status of tumours.
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http://dx.doi.org/10.1038/ncb2170DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3918344PMC
March 2011

Pathology tissue-chromatin immunoprecipitation, coupled with high-throughput sequencing, allows the epigenetic profiling of patient samples.

Proc Natl Acad Sci U S A 2010 Dec 24;107(50):21535-40. Epub 2010 Nov 24.

Department of Biomolecular Sciences, University of Urbino Carlo Bo, 61032 Fano, Italy.

Epigenetic alterations in the pattern of DNA and histone modifications play a crucial role in cancer development. Analysis of patient samples, however, is hampered by technical limitations in the study of chromatin structure from pathology archives that usually consist of heavily fixed, paraffin-embedded material. Here, we present a methodology [pathology tissue-ChIP (PAT-ChIP)] to extract and immunoprecipitate chromatin from paraffin-embedded patient samples up to several years old. In a pairwise comparison with canonical ChIP, PAT-ChIP showed a high reproducibility of results for several histone marks and an identical ability to detect dynamic changes in chromatin structure upon pharmacological treatment. Finally, we showed that PAT-ChIP can be coupled with high-throughput sequencing (PAT-ChIP-Seq) for the genome-wide analysis of distinct chromatin modifications. PAT-ChIP therefore represents a versatile procedure and diagnostic tool for the analysis of epigenetic alterations in cancer and potentially other diseases.
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http://dx.doi.org/10.1073/pnas.1007647107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3003125PMC
December 2010