Publications by authors named "Federico Riccardi Sirtori"

17 Publications

  • Page 1 of 1

Discovery of Stereospecific PARP-1 Inhibitor Isoindolinone NMS-P515.

ACS Med Chem Lett 2019 Apr 13;10(4):534-538. Epub 2019 Mar 13.

Oncology, Nerviano Medical Sciences S.r.l., Viale Pasteur 10, 20014 Nerviano, Milan, Italy.

Poly(ADP-ribose) polymerase-1 (PARP-1) is an enzyme involved in signaling and repair of DNA single strand breaks. PARP-1 employs NAD to modify substrate proteins via the attachment of poly(ADP-ribose) chains. PARP-1 is a well established target in oncology, as testified by the number of marketed drugs (e.g., Lynparza, Rubraca, Zejula, and Talzenna) used for the treatment of ovarian, breast, and prostate tumors. Efforts in investigating an uncharted region of the previously identified isoindolinone carboxamide series delivered ()- (NMS-P515), a potent inhibitor of PARP-1 both in biochemical (: 0.016 μM) and cellular (IC: 0.027 μM) assays. Cocrystal structure allowed explaining NMS-P515 stereospecific inhibition of the target. After having ruled out potential loss of enantiopurity in vitro and in vivo, NMS-P515 was synthesized in an asymmetric fashion. NMS-P515 ADME profile and its antitumor activity in a mouse xenograft cancer model render the compound eligible for further optimization.
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http://dx.doi.org/10.1021/acsmedchemlett.8b00569DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6466814PMC
April 2019

MS methods to study macromolecule-ligand interaction: Applications in drug discovery.

Methods 2018 07 22;144:152-174. Epub 2018 Jun 22.

Department of Pharmaceutical Sciences, Università degli Studi di Milano, via Mangiagalli 25, 20133 Milan, Italy. Electronic address:

The interaction of small compounds (i.e. ligands) with macromolecules or macromolecule assemblies (i.e. targets) is the mechanism of action of most of the drugs available today. Mass spectrometry is a popular technique for the interrogation of macromolecule-ligand interactions and therefore is also widely used in drug discovery and development. Thanks to its versatility, mass spectrometry is used for multiple purposes such as biomarker screening, identification of the mechanism of action, ligand structure optimization or toxicity assessment. The evolution and automation of the instruments now allows the development of high throughput methods with high sensitivity and a minimized false discovery rate. Herein, all these approaches are described with a focus on the methods for studying macromolecule-ligand interaction aimed at defining the structure-activity relationships of drug candidates, along with their mechanism of action, metabolism and toxicity.
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http://dx.doi.org/10.1016/j.ymeth.2018.06.005DOI Listing
July 2018

Virtual Cross-Linking of the Active Nemorubicin Metabolite PNU-159682 to Double-Stranded DNA.

Chem Res Toxicol 2017 02 25;30(2):614-624. Epub 2017 Jan 25.

Department of Pharmaceutical and Pharmacological Sciences, University of Padova , Via Marzolo, 5, 35131 Padova, Italy.

The DNA alkylating mechanism of PNU-159682 (PNU), a highly potent metabolite of the anthracycline nemorubicin, was investigated by gel-electrophoretic, HPLC-UV, and micro-HPLC/mass spectrometry (MS) measurements. PNU quickly reacted with double-stranded oligonucleotides, but not with single-stranded sequences, to form covalent adducts which were detectable by denaturing polyacrylamide gel electrophoresis (DPAGE). Ion-pair reverse-phase HPLC-UV analysis on CG rich duplex sequences having a 5'-CCCGGG-3' central core showed the formation of two types of adducts with PNU, which were stable and could be characterized by micro-HPLC/MS. The first type contained one alkylated species (and possibly one reversibly bound species), and the second contained two alkylated species per duplex DNA. The covalent adducts were found to produce effective bridging of DNA complementary strands through the formation of virtual cross-links reminiscent of those produced by classical anthracyclines in the presence of formaldehyde. Furthermore, the absence of reactivity of PNU with CG-rich sequence containing a TA core (CGTACG), and the minor reactivity between PNU and CGC sequences (TACGCG·CGCGTA) pointed out the importance of guanine sequence context in modulating DNA alkylation.
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http://dx.doi.org/10.1021/acs.chemrestox.6b00362DOI Listing
February 2017

Nemorubicin and doxorubicin bind the G-quadruplex sequences of the human telomeres and of the c-MYC promoter element Pu22.

Biochim Biophys Acta 2016 Jun 23;1860(6):1129-38. Epub 2016 Feb 23.

Department of Food, Environmental and Nutritional Sciences, Division of Chemistry and Molecular Biology, University of Milan, Via Celoria 2, 20133 Milano, Italy. Electronic address:

Background: Intra-molecular G-quadruplex structures are present in the guanine rich regions of human telomeres and were found to be prevalent in gene promoters. More recently, the targeting of c-MYC transcriptional control has been suggested, because the over expression of the c-MYC oncogene is one of the most common aberration found in a wide range of human tumors.

Methods: The interaction of nemorubicin and doxorubicin with DNA G-quadruplex structures has been studied by NMR, ESI-MS and molecular modelling, in order to obtain further information about the complex and the multiple mechanisms of action of these drugs.

Results And Conclusions: Nemorubicin intercalates between A3 and G4 of d(TTAGGGT)4 and form cap-complex at the G6pT7 site. The presence of the adenine in this sequence is important for the stabilization of the complex, as was shown by the interaction with d(TTGGGTT)4 and d(TTTGGGT)4, which form only a 1:1 complex. The interaction of doxorubicin with d(TTAGGGT)4 is similar, but the complex appears less stable. Nemorubicin also binds with high efficiency the c-MYC G-quadruplex sequence Pu22, to form a very well defined complex. Two nemorubicin molecules bind to the 3'-end and to the 5'-end, forming an additional plane of stacking over each external G-tetrad. The wild type c-MYCPu22 sequence forms with nemorubicin the same complex.

General Significance: Nemorubicin and doxorubicin, not only intercalate into the duplex DNA, but also result in significant ligands for G-quadruplex DNA segments, stabilizing their structure; this may in part explain the multiple mechanisms of action of their antitumor activity.
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http://dx.doi.org/10.1016/j.bbagen.2016.02.011DOI Listing
June 2016

Discovery of 2-[1-(4,4-Difluorocyclohexyl)piperidin-4-yl]-6-fluoro-3-oxo-2,3-dihydro-1H-isoindole-4-carboxamide (NMS-P118): A Potent, Orally Available, and Highly Selective PARP-1 Inhibitor for Cancer Therapy.

J Med Chem 2015 Sep 26;58(17):6875-98. Epub 2015 Aug 26.

Oncology, Nerviano Medical Sciences Srl , Viale Pasteur 10, 20014 Nerviano, Milan, Italy.

The nuclear protein poly(ADP-ribose) polymerase-1 (PARP-1) has a well-established role in the signaling and repair of DNA and is a prominent target in oncology, as testified by the number of candidates in clinical testing that unselectively target both PARP-1 and its closest isoform PARP-2. The goal of our program was to find a PARP-1 selective inhibitor that would potentially mitigate toxicities arising from cross-inhibition of PARP-2. Thus, an HTS campaign on the proprietary Nerviano Medical Sciences (NMS) chemical collection, followed by SAR optimization, allowed us to discover 2-[1-(4,4-difluorocyclohexyl)piperidin-4-yl]-6-fluoro-3-oxo-2,3-dihydro-1H-isoindole-4-carboxamide (NMS-P118, 20by). NMS-P118 proved to be a potent, orally available, and highly selective PARP-1 inhibitor endowed with excellent ADME and pharmacokinetic profiles and high efficacy in vivo both as a single agent and in combination with Temozolomide in MDA-MB-436 and Capan-1 xenograft models, respectively. Cocrystal structures of 20by with both PARP-1 and PARP-2 catalytic domain proteins allowed rationalization of the observed selectivity.
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http://dx.doi.org/10.1021/acs.jmedchem.5b00680DOI Listing
September 2015

Establish an automated flow injection ESI-MS method for the screening of fragment based libraries: Application to Hsp90.

Eur J Pharm Sci 2015 Aug 4;76:83-94. Epub 2015 May 4.

Oncology Business Unit, Nerviano Medical Sciences, Viale Pasteur 10, 20014 Nerviano (MI), Italy.

ESI-MS is a well established technique for the study of biopolymers (nucleic acids, proteins) and their non covalent adducts, due to its capacity to detect ligand-target complexes in the gas phase and allows inference of ligand-target binding in solution. In this article we used this approach to investigate the interaction of ligands to the Heat Shock Protein 90 (Hsp90). This enzyme is a molecular chaperone involved in the folding and maturation of several proteins which has been subjected in the last years to intensive drug discovery efforts due to its key role in cancer. In particular, reference compounds, with a broad range of dissociation constants from 40pM to 100μM, were tested to assess the reliability of ESI-MS for the study of protein-ligand complexes. A good agreement was found between the values measured with a fluorescence polarization displacement assay and those determined by mass spectrometry. After this validation step we describe the setup of a medium throughput screening method, based on ESI-MS, suitable to explore interactions of therapeutic relevance biopolymers with chemical libraries. Our approach is based on an automated flow injection ESI-MS method (AFI-MS) and has been applied to screen the Nerviano Medical Sciences proprietary fragment library of about 2000 fragments against Hsp90. In order to discard false positive hits and to discriminate those of them interacting with the N-terminal ATP binding site, competition experiments were performed using a reference inhibitor. Gratifyingly, this group of hits matches with the ligands previously identified by NMR FAXS techniques and confirmed by X-ray co-crystallization experiments. These results support the use of AFI-MS for the screening of medium size libraries, including libraries of small molecules with low affinity typically used in fragment based drug discovery. AFI-MS is a valid alternative to other techniques with the additional opportunities to identify compounds interacting with unpredicted or allosteric sites, without the need of any binding probes.
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http://dx.doi.org/10.1016/j.ejps.2015.05.001DOI Listing
August 2015

Effective immuno-targeting of the IDH1 mutation R132H in a murine model of intracranial glioma.

Acta Neuropathol Commun 2015 Jan 21;3. Epub 2015 Jan 21.

The R132H mutation of cytosolic isocitrate dehydrogenase (IDH1) is present in the majority of low grade gliomas.Immunotherapy in these tumors has an interesting, still unexploited, therapeutic potential, as they are less immunosuppressive than glioblastomas. Using site-directed mutagenesis we introduced the R132H mutation into the murine glioma cell line GL261,creating mIDH1-GL261. Presence of the mutation was confirmed by immunoblotting and production of the oncometabolite 2-hydroxyglutarate (2HG), demonstrated by mass spectrometry (LC-MS/MS) performed on cell supernatant. In vitro mIDH1-GL261 had different morphology but similar growth rate than parental GL261 (p-GL261). After intracranial injection, MRI suggested that the initial growth rate was slower in mIDH1-GL261 than p-GL261 gliomas but overall survival was similar. mIDH1-GL261 gliomas showed evidence of R132H expression and of intratumoral 2HG production (evaluated by MRS and LC-MS/MS). Immunizations were performed nine days after intracranial implantation of mIDH1- or p-GL261 cells by three subcutaneous injections of five different peptides encompassing the IDH1 mutation site, all emulsified with Montanide ISA-51, in association with GM-CSF. Control mice were injected with four ovalbumin peptides or vehicle. Mice with mIDH1-GL261 but not p-GL261 gliomas treated with mIDH1 peptides survived longer than controls; 25% of them were cured. Immunized mice showed higher amounts of peripheral CD8+ T cells, higher production of IFN-γ, and evidence of anti-mIDH1 antibodies.Immunizations led to intratumoral up-regulation of IFN-γ, granzyme-b and perforin-1 and down-regulation of TGF-β2 and IL-10. These results support the translational potential of immunotherapeutic targeting of gliomas carrying IDH1 mutations.
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http://dx.doi.org/10.1186/s40478-014-0180-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4359524PMC
January 2015

Mass spectrometric strategies for the identification and characterization of human serum albumin covalently adducted by amoxicillin: ex vivo studies.

Chem Res Toxicol 2014 Sep 13;27(9):1566-74. Epub 2014 Aug 13.

Department of Pharmaceutical Sciences, Università degli Studi di Milano , via Mangiagalli 25, 20133, Milan, Italy.

This study addresses the detection and characterization of the modification of human serum albumin (HSA) by amoxicillin (AX) in ex vivo samples from healthy subjects under oral amoxicillin administration (acute intake of 1 g every 8 h for 48 h). To reach this goal, we used an analytical strategy based on targeted and untargeted mass spectrometric approaches. Plasma samples withdrawn before AX oral intake represented the negative control samples to test the method selectivity, whereas HSA incubated in vitro with AX was the positive control. Different MS strategies were developed, particularly (1) multiple reaction monitoring (MRM) and precursor ion scan (PIS) using a HPLC system coupled to a triple quadrupole MS analyzer and (2) a dedicated data-dependent scan and a customized targeted MS/MS analysis carried out using a nano-LC system coupled to a high-resolution MS system (LTQ Orbitrap XL). Lys 190 was identified as the only modification site of HSA in the ex vivo samples. The AX adduct was identified and fully characterized by complementary targeted approaches based on triple quadrupole (MRM mode) and orbitrap (SIC mode) mass analyzers. The SIC mode also permitted the relative amount of AX-adducted HSA to be measured, ranging from 1 to 2% (6-12 μM) at 24 and 48 h after the oral intake. No adduct in any ex vivo sample was identified by the untargeted methods (PIS and data-dependent scan mode analysis). The results on one hand indicate that MS, in particular high-resolution MS, analysis represents a suitable analytical tool for the identification/characterization of covalently modified proteins/peptides; on the other hand, they give deeper insight into AX-induced protein haptenation, which is required to better understand the mechanisms involved in AX-elicited allergic reactions.
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http://dx.doi.org/10.1021/tx500210eDOI Listing
September 2014

Insights into PARP Inhibitors' Selectivity Using Fluorescence Polarization and Surface Plasmon Resonance Binding Assays.

J Biomol Screen 2014 Sep 10;19(8):1212-9. Epub 2014 Jun 10.

Nerviano Medical Sciences S.r.l., Nerviano, Italy

PARP inhibitors are an exciting new class of antineoplastic drugs that have been proven to be efficacious as single agents in cancer settings with inherent DNA repair defects, as well as in combination with DNA-damaging chemotherapeutics. Currently, they are designed to target the catalytic domain of PARP-1, the most studied member of the family, with a key role in the DNA-damage repair process. Because PARP inhibitors are substrate (NAD(+)) competitors, there is a need for a deeper understanding of their cross-reactivity. This is particularly relevant for PARP-2, the PARP-1 closest homologue, for which an embryonic lethal phenotype has been observed in double knockout mice. In this study, we describe the development and validation of binding assays based on fluorescence polarization (FP) and surface plasmon resonance (SPR) techniques. PARP-1, PARP-2, PARP-3, and TNKS-1 FP displacement assays are set up by employing ad hoc synthesized probes. These assays are suitable for high-throughput screening (HTS) and selectivity profiling, thus allowing the identification of NAD(+)binding site selective inhibitors. The PARP-1 and PARP-2 complementary SPR binding assays confirm displacement data and the in-depth inhibitor characterization. Moreover, these formats have the potential to be broadly applicable to other members of the PARP family.
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http://dx.doi.org/10.1177/1087057114538319DOI Listing
September 2014

Alkylsulfanyl-1,2,4-triazoles, a new class of allosteric valosine containing protein inhibitors. Synthesis and structure-activity relationships.

J Med Chem 2013 Jan 4;56(2):437-50. Epub 2013 Jan 4.

Nerviano Medical Sciences S.r.l., Oncology, Viale Pasteur 10, 20014 Nerviano (MI), Italy.

Valosine containing protein (VCP), also known as p97, is a member of AAA ATPase family that is involved in several biological processes and plays a central role in the ubiquitin-mediated degradation of misfolded proteins. VCP is an ubiquitously expressed, highly abundant protein and has been found overexpressed in many tumor types, sometimes associated with poor prognosis. In this respect, VCP has recently received a great deal of attention as a potential new target for cancer therapy. In this paper, the discovery and structure-activity relationships of alkylsulfanyl-1,2,4-triazoles, a new class of potent, allosteric VCP inhibitors, are described. Medicinal chemistry manipulation of compound 1, identified via HTS, led to the discovery of potent and selective inhibitors with submicromolar activity in cells and clear mechanism of action at consistent doses. This represents a first step toward a new class of potential anticancer agents.
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http://dx.doi.org/10.1021/jm3013213DOI Listing
January 2013

The interaction of nemorubicin metabolite PNU-159682 with DNA fragments d(CGTACG)(2), d(CGATCG)(2) and d(CGCGCG)(2) shows a strong but reversible binding to G:C base pairs.

Bioorg Med Chem 2012 Dec 3;20(24):6979-88. Epub 2012 Nov 3.

DeFENS-Department of Food, Environmental and Nutritional Sciences, via Celoria 2, 20133 Milano, Italy.

The antitumor anthracycline nemorubicin is converted by human liver microsomes to a major metabolite, PNU-159682 (PNU), which was found to be much more potent than its parent drug toward cultured tumor cells and in vivo tumor models. The mechanism of action of nemorubicin appears different from other anthracyclines and until now is the object of studies. In fact PNU is deemed to play a dominant, but still unclear, role in the in vivo antitumor activity of nemorubicin. The interaction of PNU with the oligonucleotides d(CGTACG)(2), d(CGATCG)(2) and d(CGCGCG)(2) was studied with a combined use of (1)H and (31)P NMR spectroscopy and by ESI-mass experiments. The NMR studies allowed to establish that the intercalation between the base pairs of the duplex leads to very stable complexes and at the same time to exclude the formation of covalent bonds. Melting experiments monitored by NMR, allowed to observe with high accuracy the behaviour of the imine protons with temperature, and the results showed that the re-annealing occurs after melting. The formation of reversible complexes was confirmed by HPLC-tandem mass spectra, also combined with endonuclease P1digestion. The MS/MS spectra showed the loss of neutral PNU before breaking the double helix, a behaviour typical of intercalators. After digestion with the enzyme, the spectra did not show any compound with PNU bound to the bases. The evidence of a reversible process appears from both proton and phosphorus NOESY spectra of PNU bound to d(CGTACG)(2) and to d(CGATCG)(2). The dissociation rate constants (k(off)) of the slow step of the intercalation process, measured by (31)P NMR NOE-exchange experiments, showed that the kinetics of the process is slower for PNU than for doxorubicin and nemorubicin, leading to a 10- to 20-fold increase of the residence time of PNU into the intercalation sites, with respect to doxorubicin. A relevant number of NOE interactions allowed to derive a model of the complexes in solution from restrained MD calculations. The conformation of PNU bound to the oligonucleotides was also derived from the coupling constant values.
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http://dx.doi.org/10.1016/j.bmc.2012.10.033DOI Listing
December 2012

Molecular recognition of T:G mismatched base pairs in DNA as studied by electrospray ionization mass spectrometry.

ChemMedChem 2012 Jun 4;7(6):1112-22. Epub 2012 Apr 4.

Oncology Business Unit, Nerviano Medical Sciences, Viale Pasteur 10, 20014 Nerviano (MI), Italy.

Postreplicative mismatch repair (MMR) is a cellular system involved in the recognition and correction of DNA polymerase errors that escape detection in proofreading. Of the various mismatched bases, T:G pairing in DNA is one of the more common mutations leading to the formation of tumors in humans. In addition, the absence of the MMR system can generate resistance to several chemotherapeutic agents, particularly DNA-damaging substances. The main purpose of this study was the setup and validation of an electrospray ionization (ESI) mass spectrometry method for the identification of small molecules that are able to recognize T:G mismatches in DNA targets. These findings could be useful for the discovery of new antitumor drugs. The analytical method is based on the ability of electrospray to preserve the noncovalent adducts present in solution and transfer them to the gas phase. Lexitropsin derivatives (polyimidazole compounds) have been previously described as selective for T:G mismatch binding by NMR and ITC studies. We synthesized and tested various polyimidazole derivatives, one of which in particular (NMS-057) showed a higher affinity for an oligonucleotide DNA sequence containing a T:G mismatched base pair. To rationalize these findings, molecular docking studies were performed using available NMR structures. Moreover, ESI-MS experiments, performed on an orbitrap mass spectrometer, highlighted the formation of heterodimeric complexes between DNA sequences, distamycin A, and polyimidazole compounds. Our results confirm that this ESI method could be a valuable tool for the identification of new molecules able to specifically recognize T:G mismatched base pairs.
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http://dx.doi.org/10.1002/cmdc.201100526DOI Listing
June 2012

Development of biochemical assays for the identification of eIF4E-specific inhibitors.

J Biomol Screen 2012 Jun 5;17(5):581-92. Epub 2012 Mar 5.

Biotechnology Department, Nerviano Medical Sciences, Nerviano (MI), Italy.

Control of mRNA translation plays a critical role in cell growth, proliferation, and differentiation and is tightly regulated by AKT and RAS oncogenic pathways. A key player in the regulation of this process is the mRNA 5' cap-binding protein, eukaryotic translation initiation factor 4E (eIF4E). eIF4E contributes to malignancy by selectively enabling the translation of a limited pool of mRNAs that generally encode key proteins involved in cell cycle progression, angiogenesis, and metastasis. Several data indicate that the inhibition of eIF4E in tumor cell lines and xenograft models impairs tumor growth and induces apoptosis; eIF4E, therefore, can be considered a valuable target for cancer therapy. Targeting the cap-binding pocket of eIF4E should represent a way to inhibit all the eIF4E cellular functions. We present here the development and validation of different biochemical assays based on fluorescence polarization and surface plasmon resonance techniques. These assays could support high-throughput screening, further refinement, and characterization of eIF4E inhibitors, as well as selectivity assessment against CBP80/CBP20, the other major cap-binding complex of eukaryotic cells, overall providing a robust roadmap for development of eIF4E-specific inhibitors.
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http://dx.doi.org/10.1177/1087057112438554DOI Listing
June 2012

Thieno[3,2-c]pyrazoles: a novel class of Aurora inhibitors with favorable antitumor activity.

Bioorg Med Chem 2010 Oct 25;18(19):7113-20. Epub 2010 Jul 25.

Nerviano Medical Sciences-Oncology, via Pasteur 10, 20014 Nerviano, Milan, Italy.

A novel series of 3-amino-1H-thieno[3,2-c]pyrazole derivatives demonstrating high potency in inhibiting Aurora kinases was developed. Here we describe the synthesis and a preliminary structure-activity relationship, which led to the discovery of a representative compound (38), which showed low nanomolar inhibitory activity in the anti-proliferation assay and was able to block the cell cycle in HCT-116 cell line. This compound demonstrated favorable pharmacokinetic properties and good efficacy in the HL-60 xenograft tumor model.
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http://dx.doi.org/10.1016/j.bmc.2010.07.048DOI Listing
October 2010

Cell division cycle 7 kinase inhibitors: 1H-pyrrolo[2,3-b]pyridines, synthesis and structure-activity relationships.

J Med Chem 2009 Jul;52(14):4380-90

Nerviano Medical Sciences, 20014 Nerviano, Milano, Italy.

Cdc7 kinase has recently emerged as an attractive target for cancer therapy and low-molecular-weight inhibitors of Cdc7 kinase have been found to be effective in the inhibition of tumor growth in animal models. In this paper, we describe synthesis and structure-activity relationships of new 1H-pyrrolo[2,3-b]pyridine derivatives identified as inhibitors of Cdc7 kinase. Progress from (Z)-2-phenyl-5-(1H-pyrrolo[2,3-b]pyridin-3-ylmethylene)-3,5-dihydro-4H-imidazol-4-one (1) to [(Z)-2-(benzylamino)-5-(1H-pyrrolo[2,3-b]pyridin-3-ylmethylene)-1,3-thiazol-4(5H)-one] (42), a potent ATP mimetic inhibitor of Cdc7 kinase with IC(50) value of 7 nM, is also reported.
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http://dx.doi.org/10.1021/jm900248gDOI Listing
July 2009

6-Substituted pyrrolo[3,4-c]pyrazoles: an improved class of CDK2 inhibitors.

ChemMedChem 2007 Jun;2(6):841-52

Oncology Business Unit, Department of Chemistry, Nerviano Medical Sciences, Viale Pasteur 10, 20014 Nerviano MI, Italy.

We have recently reported a new class of CDK2/cyclin A inhibitors based on a bicyclic tetrahydropyrrolo[3,4-c]pyrazole scaffold. The introduction of small alkyl or cycloalkyl groups in position 6 of this scaffold allowed variation at the other two diversity points. Conventional and polymer-assisted solution phase chemistry provided a way of generating compounds with improved biochemical and cellular activity. Optimization of the physical properties and pharmacokinetic profile led to a compound which exhibited good efficacy in vivo on A2780 human ovarian carcinoma.
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http://dx.doi.org/10.1002/cmdc.200600302DOI Listing
June 2007

A fully automated method for accurate mass determination using high-performance liquid chromatography with a quadrupole/orthogonal acceleration time-of-flight mass spectrometer.

Rapid Commun Mass Spectrom 2004 ;18(4):511-7

Pharmaceutical Sciences, Pharmacia Italia S.p.A., via Pasteur 10, 20014 Nerviano-Milan, Italy.

A generic LC/ESI(+)-oaTOFMS method has been developed for routine automated high accuracy mass determinations of different classes of substances. The system makes use of micro-high-performance liquid chromatography and a hybrid quadrupole/orthogonal acceleration time-of-flight (Q-oaTOF) mass spectrometer. Reproducible and accurate mass measurements were obtained using an electrospray dual sprayer with reserpine as reference compound, introduced into the mass spectrometer alternating with the samples. Experiments were performed to optimize analyte/reference response ratio, statistical algorithm correction setting, and analyte concentration. In these experiments, a clear dependence of the mass measurement error on the analyte/reference response ratio was observed. The dependence of average mass error versus different dead time correction algorithm settings (Np factors) was also explored. In the final automated procedure, verified for a statistically significant set of compounds ( approximately 550) obtained from a medicinal chemistry department, about 70% of the analyzed samples satisfied the acceptance criteria fixed at a maximum error of +/-5 ppm (mass range 150-800 Da).
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http://dx.doi.org/10.1002/rcm.1368DOI Listing
May 2004