Final Report Summary - FBMC2010MDG (Lead Optimisation of Novel Androgen Receptor Small Molecule Modulators - Improving Treatment of Prostate Cancer)
Executive summary
March 2011 was the starting date of the FBMC2010MDG project entitled 'Lead Optimization of Novel Androgen Receptor Small Molecule Modulators - Improving Treatment of Prostate Cancer' managed by Dr. Fernando Blanco at the Molecular Design Group (Trinity College Dublin), supervised by Pr. David Lloyd and supported by the European Commission (Marie-Curie Grant FP7 People, Project Reference: 274988). After two years of research the project has achieved most of its objectives and technical goals for the period with relatively minor deviations.
Summary description of the project objectives
One of the traditional approaches for the treatment of prostate cancer is the administration of drugs which bind to the Androgen Receptor (AR) and stop the cancer growing employing concurrent LBP directed intervention ('LBP' antiandrogens). These therapeutics bind to AR in the LBP, mimicking androgen's mode of action, but blocking the receptor's dynamic formation of the AF-2 co-regulator region through their incorporation of specific bulky substituents that prevent the 'closure' of helix 12. A major drawback of this therapy is the development of drug resistance. Castration Resistant Prostate Cancer (CRPC) is frequently metastatic and almost invariably fatal. Current treatments require high dosing and show significant side effects. The aim of this project was to develop drugs that combat prostate cancer in an alternative way.
At the beginning of this research, MDG group had computationally analysed two other potential AR binding sites termed AF-2 and BF3 and were in the process of developing small molecules that interact with the novel A-F2 site. 2 novel molecules, MDG15 and MDG173, had been identified as potential AR AF-2 ligands so that it was necessary to perform to perform a complete plan of 'Lead Optimisation'
In this scenario the objectives for this project were:
-Development of small molecules that modulate AR activity through the AF-2 site by using Structure Based Drug Design (SBDD), Ligand Based Drug Design (LBDD) and Structure Activity Relationship (SAR) protocols.
-Study and characterization of the chemical features of the new family of compounds by High Level computational methods (ab initio)
-Purchase and Biological Evaluation of new candidates through FP, FRET and WST-1 cell proliferation assays.
-Synthesis of new derivatives according to SAR criteria.
-Medicinal chemistry hit-to-lead progression followed by lead optimisation around identified hit scaffolds targeting the AR AF-2 domain.
-Establishing a strong theoretical basis of the potential 'binding mode' in the AF-2 region for the future development of new analogues.
Work performed since the beginning of the project
-Identification and full characterization by virtual screening and biological assays of diarylhydrazides as a new promising scaffold of non-LBP antiandrogens. Starting from two initial hits, we obtained up to 15 new ligands with proved activity as true antiandrogens displacing AR-coactivator interaction and having a full antagonistic profile on AR (both wt and T877A), partial antagonistic profile for PR, and selectivity for the other members of the NR-3 family (GR, ER-α, and ER-β). The small molecules non-LBP true AR modulators provided by this study will be used to further characterize the AR-coactivator interface, to understand the basis of selectivity, and to further guide rational drug design in the search of other novel scaffolds directed at this interface.
-Chemical and Theoretical characterization of the compounds under study. The preferred molecular conformations for diarylhydrazides were modeled using DFT theoretical calculations and experimentally determined through solution NMR and crystallographic di?raction analysis. Their isomerism and conformational space were fully determined. We demonstrated that the E-isomer is formed under the described synthetic conditions for this family of compounds. The Z-isomer is formed when the steric hindrance of the second imine substituent is su?ciently high, in this case as an equal mixture with the corresponding E-isomer. This was in full agreement with the theoretical predictions.
-Structure Activity Relationship study to establish the main parameters that determine the effectiveness of our compounds as AR antagonists. A new series of 25 diarylhydrazides were synthesized and evaluated for AR antagonist activity. Eight of these novel compounds showed antiandrogen activity by a proven non-LBP mechanism of action, consolidating the diarylhydrazide class of molecules as a promising alternative to traditional therapeutics for the treatment of Castration Resistant Prostate Cancer.
-Theoretical study (docking and molecular dynamics) of the potential binding mode of our systems within the AR AF-2. In the absence of a ligand-cocrystal structure, a combined study of docking and molecular dynamics was performed, affording new and relevant insights to the potential binding mode of our active ligands to the AF-2 site. The results strongly support the main conclusions deduced from the SAR analysis and provided enough information to hypothesize a bioactive conformation as a basis for the design of future new candidates.
Description of the main results achieved
Prostate cancer (PC) is one of the most prevalent cancers in the male population. Each year, approximately 400,000 patients are diagnosed with the disease in the seven major markets. Patients diagnosed with early-stage prostate cancer have excellent prognosis and can survive for many years after surgery through a chemotherapeutic antihormonal treatment regimen. However, prolonged use of antihormonal drugs often results in the disease becoming hormone-refractory, where prognosis deteriorates dramatically due to the lack of effective treatment. The existing antihormonal treatment for prostate cancer, androgen ablation therapy (AAT), focuses on preventing binding of the endogenous androgen receptor (AR) ligand, 5alpha-dihydrotestosterone (DHT), by competitively binding to the ligand-binding pocket (LBP) of the AR. Existing chemotherapeutics following that strategy have high dosing requirements and so exhibit significant side-effects and are prone to hormone-refraction resistance on continued administration.
In the present project, it has been conducted a comprehensive research in the field of rational design of new non-LBP androgen receptor antagonists with potential activity in the AF-2 region as a new alternative to known PC-therapeutics. The protocol followed has allowed the identification of 15 new hits by combining virtual screening, similarity, and docking techniques. All this compounds have shown proved non-LBP antiandrogen activity evaluated by TR-FRET and FP assays. The new family of compounds (diarylhydrazides) has been analyzed from a chemical perspective determining its most relevant structural features. Furthermore, it has been conducted a SAR study whose results allow a better understanding and a 'control' over substitution effects in the systems analyzed. Additionally, a Molecular Dynamics theoretical study has been performed, affording new and relevant insights to the potential binding mode of our active ligands to the AF-2 site. These results also strongly support the main conclusions deduced from the SAR analysis and have provided enough information to hypothesize a bioactive conformation as a basis for the design of future new candidates.
the results obtained at the end of this project are of great interest in the field of design of new therapeutics for the treatment of Castration Resistant Prostate Cancer.
March 2011 was the starting date of the FBMC2010MDG project entitled 'Lead Optimization of Novel Androgen Receptor Small Molecule Modulators - Improving Treatment of Prostate Cancer' managed by Dr. Fernando Blanco at the Molecular Design Group (Trinity College Dublin), supervised by Pr. David Lloyd and supported by the European Commission (Marie-Curie Grant FP7 People, Project Reference: 274988). After two years of research the project has achieved most of its objectives and technical goals for the period with relatively minor deviations.
Summary description of the project objectives
One of the traditional approaches for the treatment of prostate cancer is the administration of drugs which bind to the Androgen Receptor (AR) and stop the cancer growing employing concurrent LBP directed intervention ('LBP' antiandrogens). These therapeutics bind to AR in the LBP, mimicking androgen's mode of action, but blocking the receptor's dynamic formation of the AF-2 co-regulator region through their incorporation of specific bulky substituents that prevent the 'closure' of helix 12. A major drawback of this therapy is the development of drug resistance. Castration Resistant Prostate Cancer (CRPC) is frequently metastatic and almost invariably fatal. Current treatments require high dosing and show significant side effects. The aim of this project was to develop drugs that combat prostate cancer in an alternative way.
At the beginning of this research, MDG group had computationally analysed two other potential AR binding sites termed AF-2 and BF3 and were in the process of developing small molecules that interact with the novel A-F2 site. 2 novel molecules, MDG15 and MDG173, had been identified as potential AR AF-2 ligands so that it was necessary to perform to perform a complete plan of 'Lead Optimisation'
In this scenario the objectives for this project were:
-Development of small molecules that modulate AR activity through the AF-2 site by using Structure Based Drug Design (SBDD), Ligand Based Drug Design (LBDD) and Structure Activity Relationship (SAR) protocols.
-Study and characterization of the chemical features of the new family of compounds by High Level computational methods (ab initio)
-Purchase and Biological Evaluation of new candidates through FP, FRET and WST-1 cell proliferation assays.
-Synthesis of new derivatives according to SAR criteria.
-Medicinal chemistry hit-to-lead progression followed by lead optimisation around identified hit scaffolds targeting the AR AF-2 domain.
-Establishing a strong theoretical basis of the potential 'binding mode' in the AF-2 region for the future development of new analogues.
Work performed since the beginning of the project
-Identification and full characterization by virtual screening and biological assays of diarylhydrazides as a new promising scaffold of non-LBP antiandrogens. Starting from two initial hits, we obtained up to 15 new ligands with proved activity as true antiandrogens displacing AR-coactivator interaction and having a full antagonistic profile on AR (both wt and T877A), partial antagonistic profile for PR, and selectivity for the other members of the NR-3 family (GR, ER-α, and ER-β). The small molecules non-LBP true AR modulators provided by this study will be used to further characterize the AR-coactivator interface, to understand the basis of selectivity, and to further guide rational drug design in the search of other novel scaffolds directed at this interface.
-Chemical and Theoretical characterization of the compounds under study. The preferred molecular conformations for diarylhydrazides were modeled using DFT theoretical calculations and experimentally determined through solution NMR and crystallographic di?raction analysis. Their isomerism and conformational space were fully determined. We demonstrated that the E-isomer is formed under the described synthetic conditions for this family of compounds. The Z-isomer is formed when the steric hindrance of the second imine substituent is su?ciently high, in this case as an equal mixture with the corresponding E-isomer. This was in full agreement with the theoretical predictions.
-Structure Activity Relationship study to establish the main parameters that determine the effectiveness of our compounds as AR antagonists. A new series of 25 diarylhydrazides were synthesized and evaluated for AR antagonist activity. Eight of these novel compounds showed antiandrogen activity by a proven non-LBP mechanism of action, consolidating the diarylhydrazide class of molecules as a promising alternative to traditional therapeutics for the treatment of Castration Resistant Prostate Cancer.
-Theoretical study (docking and molecular dynamics) of the potential binding mode of our systems within the AR AF-2. In the absence of a ligand-cocrystal structure, a combined study of docking and molecular dynamics was performed, affording new and relevant insights to the potential binding mode of our active ligands to the AF-2 site. The results strongly support the main conclusions deduced from the SAR analysis and provided enough information to hypothesize a bioactive conformation as a basis for the design of future new candidates.
Description of the main results achieved
Prostate cancer (PC) is one of the most prevalent cancers in the male population. Each year, approximately 400,000 patients are diagnosed with the disease in the seven major markets. Patients diagnosed with early-stage prostate cancer have excellent prognosis and can survive for many years after surgery through a chemotherapeutic antihormonal treatment regimen. However, prolonged use of antihormonal drugs often results in the disease becoming hormone-refractory, where prognosis deteriorates dramatically due to the lack of effective treatment. The existing antihormonal treatment for prostate cancer, androgen ablation therapy (AAT), focuses on preventing binding of the endogenous androgen receptor (AR) ligand, 5alpha-dihydrotestosterone (DHT), by competitively binding to the ligand-binding pocket (LBP) of the AR. Existing chemotherapeutics following that strategy have high dosing requirements and so exhibit significant side-effects and are prone to hormone-refraction resistance on continued administration.
In the present project, it has been conducted a comprehensive research in the field of rational design of new non-LBP androgen receptor antagonists with potential activity in the AF-2 region as a new alternative to known PC-therapeutics. The protocol followed has allowed the identification of 15 new hits by combining virtual screening, similarity, and docking techniques. All this compounds have shown proved non-LBP antiandrogen activity evaluated by TR-FRET and FP assays. The new family of compounds (diarylhydrazides) has been analyzed from a chemical perspective determining its most relevant structural features. Furthermore, it has been conducted a SAR study whose results allow a better understanding and a 'control' over substitution effects in the systems analyzed. Additionally, a Molecular Dynamics theoretical study has been performed, affording new and relevant insights to the potential binding mode of our active ligands to the AF-2 site. These results also strongly support the main conclusions deduced from the SAR analysis and have provided enough information to hypothesize a bioactive conformation as a basis for the design of future new candidates.
the results obtained at the end of this project are of great interest in the field of design of new therapeutics for the treatment of Castration Resistant Prostate Cancer.