_:b19902447 _:b19902466 . _:b511093511 "2"^^ . _:b19902447 _:b19902467 . _:b19902447 _:b19902468 . _:b19902447 _:b19902469 . _:b19902424 . _:b19902414 _:b19902428 . _:b511093513 . . _:b19902447 _:b19902470 . _:b511093510 "2"^^ . _:b19902414 _:b19902429 . _:b19902447 _:b19902471 . _:b19902414 _:b19902430 . _:b19902414 _:b19902431 . _:b511093512 . _:b19902425 . _:b19902414 _:b19902424 . _:b19902414 _:b19902425 . _:b511093505 "3"^^ . _:b19902414 _:b19902426 . _:b19902414 _:b19902427 . . _:b19902426 . _:b19902414 _:b19902420 . _:b511093515 . _:b19902414 _:b19902421 . _:b19902452 . _:b511093504 "3"^^ . _:b19902414 _:b19902422 . _:b19902414 _:b19902423 . _:b511093514 . _:b19902427 . _:b19902414 _:b19902416 . _:b19902414 _:b19902417 . _:b511093519 . _:b19902414 _:b19902418 . _:b511093507 "3"^^ . _:b19902414 _:b19902419 . _:b511093525 . _:b19902436 . _:b19902428 "No cellular function beyond NER has been observed for XPA and competitive inhibition of the XPA interaction with peptide fragments is effective at disrupting NER [11], [>>12<<]. Furthermore, clinically, patients that have been shown to have low expression levels of either XPA or ERCC1 demonstrate higher sensitivity to cisplatin treatment, and people deficient for XPA (or other XP proteins) are hypersensitive to" . _:b511093506 "3"^^ . _:b19902421 "demonstrated that NER is the major DNA repair mechanism that removes cisplatin-induced DNA damage, and that resistance to platinum-based therapy correlates with high expression of ERCC1, a major element of the NER machinery [4], [5], [>>6<<], [7]. In this context, one way to increase the efficacy of platinum therapy and decrease drug resistance is to regulate NER by inhibiting the activity of ERCC1 and interacting proteins using novel therapeutic compounds [8]." . _:b19902414 _:b19902415 . . _:b19902437 . _:b511093524 . _:b511093517 "2"^^ . _:b19902422 "that NER is the major DNA repair mechanism that removes cisplatin-induced DNA damage, and that resistance to platinum-based therapy correlates with high expression of ERCC1, a major element of the NER machinery [4], [5], [6], [>>7<<]. In this context, one way to increase the efficacy of platinum therapy and decrease drug resistance is to regulate NER by inhibiting the activity of ERCC1 and interacting proteins using novel therapeutic compounds [8]." . _:b19902457 "the first problem, a number of studies suggested extracting the structures at larger intervals of the MD simulation (e.g. every 5 ns or so), [24] condensing the structural ensemble generated from MD simulations using QR factorization, [>>25<<] or clustering the MD trajectory using root-mean-square-deviation (RMSD) conformational clustering, [26], [27] On the other hand, to rank the screened compounds and suggest a final set of top hits, some studies used only docking" . _:b511093527 . _:b19902438 . _:b511093516 "2"^^ . _:b19902456 . _:b511093512 . _:b511093526 . _:b19902439 . _:b511093519 "2"^^ . _:b19902456 "For the first problem, a number of studies suggested extracting the structures at larger intervals of the MD simulation (e.g. every 5 ns or so), [>>24<<] condensing the structural ensemble generated from MD simulations using QR factorization, [25] or clustering the MD trajectory using root-mean-square-deviation (RMSD) conformational clustering, [26], [27] On the other hand, to rank the" . . . _:b511093521 . _:b19902432 . _:b511093518 "2"^^ . _:b511093525 . _:b511093520 . _:b511093513 "2"^^ . _:b19902433 . _:b511093523 . _:b19902434 . _:b511093536 . _:b511093512 "2"^^ . _:b19902460 . . . _:b19902432 . _:b511093522 . _:b19902435 . _:b511093515 "2"^^ . _:b511093533 . _:b19902444 . _:b511093514 "2"^^ . . . _:b511093532 . _:b19902445 . _:b511093525 "2"^^ . . _:b511093527 . _:b511093535 . _:b19902446 . _:b19902437 "All docking simulations employed the software AutoDock, version 4.0 [>>46<<]. The docking method and parameters were similar to the ones used in our previous work [15]." . . _:b511093524 "2"^^ . _:b19902414 _:b19902432 . _:b511093534 . _:b511093527 "2"^^ . _:b19902447 . _:b511093529 . _:b19902440 . _:b511093526 "2"^^ . _:b511093528 . _:b19902441 . _:b511093521 "2"^^ . . _:b511093531 . _:b19902442 . . . _:b511093520 "2"^^ . _:b19902416 "However, in cancer cells exposed to DNA damaging agents that distort the DNA helix or form bulky injuries to the genome, NER comes into play and removes the damage, thus protecting cancer cells from death [>>1<<], [2]. A striking example of this mechanism is represented by the use of platinum compounds such as cisplatin, the backbone for many treatments of solid tumors including testicular, bladder, ovarian, head and neck, cervical, lung and" . _:b511093530 . _:b19902443 . _:b511093523 "2"^^ . _:b19902442 "The AMBER99SB force field [>>50<<] was used for protein parameterization, while the generalized AMBER force field (GAFF) provided parameters for ligands [51]." . _:b511093541 . _:b19902468 . _:b19902452 . _:b511093522 "2"^^ . . _:b19902441 "In brief, for each docking simulation a modified version of the PTRAJ module of AMBER [>>48<<] clustered the docking trials." . _:b511093540 . _:b19902453 . _:b511093533 "2"^^ . _:b511093511 . _:b511093518 . _:b19902426 . _:b511093543 . _:b19902454 . _:b511093532 "2"^^ . _:b511093542 . _:b19902455 . _:b511093535 "2"^^ . _:b19902438 "The docking method and parameters were similar to the ones used in our previous work [>>15<<]. The screening method adopted two filtering phases with the same docking parameters. First, we screened the entire CN library against a single target model followed by applying the relaxed complex scheme (RCS) [47] through docking of the" . _:b511093537 . _:b19902448 . _:b511093534 "2"^^ . _:b511093493 . _:b19902464 "Based on our previous investigations, [>>15<<], the important residues that mostly contribute to its interaction with ligands are Gly109, Pro111, Asn110, Asp 129, Phe140, Tyr145, and Arg156 (Figure 2)." . _:b19902443 "The AMBER99SB force field [50] was used for protein parameterization, while the generalized AMBER force field (GAFF) provided parameters for ligands [>>51<<]. For each ligand, partial charges were calculated with the AM1-BCC method using the Antechamber module of AMBER 10. Protonation states of all ionizable residues were calculated using the program PDB2PQR. All simulations were performed at" . _:b511093536 . _:b19902449 . _:b511093529 "2"^^ . _:b511093539 . _:b19902450 . _:b511093528 "2"^^ . _:b511093507 . _:b511093538 . _:b19902451 . . _:b19902458 . _:b511093531 "2"^^ . _:b19902460 . _:b19902460 "the MD trajectory using root-mean-square-deviation (RMSD) conformational clustering, [26], [27] On the other hand, to rank the screened compounds and suggest a final set of top hits, some studies used only docking predictions, [>>24<<], [25], [26]while others suggested (as in this thesis) using a more accurate scoring method (e.g. MM/PBSA (Molecular Mechanics/Poisson Boltzmann Surface Area)) to refine the final selected hits." . _:b511093530 "2"^^ . . _:b19902444 . _:b19902415 "In normal cells, NER removes many types of DNA lesions, protecting cell integrity [>>1<<]. However, in cancer cells exposed to DNA damaging agents that distort the DNA helix or form bulky injuries to the genome, NER comes into play and removes the damage, thus protecting cancer cells from death [1], [2]. A striking example of" . _:b19902467 "The average RMSD for the two compounds was around 6 \u00C5, which is consistent with values obtained in similar studies [>>31<<]. The RMSD for NERI01 (Figure 3-A) was more fluctuating than that of the other compound (Figure 3-B), indicating higher flexibility. This was evident in the atomic fluctuatation analysis. Many parts of NERI01 are flexible (Figure 3-C)" . _:b19902461 . _:b511093541 "2"^^ . . _:b19902462 . . _:b511093540 "2"^^ . . _:b19902445 . _:b19902463 . _:b511093513 . _:b19902462 . _:b511093543 "2"^^ . _:b511093535 . _:b19902456 . . _:b511093542 "2"^^ . _:b19902433 _:b19902434 . . _:b19902433 _:b19902435 . _:b19902433 _:b19902436 . _:b19902457 . _:b19902433 _:b19902437 . _:b19902463 . _:b511093537 "2"^^ . _:b511093523 . _:b19902433 _:b19902438 . _:b19902461 . _:b511093493 . _:b19902433 _:b19902439 . _:b511093538 . _:b511093494 . _:b19902433 _:b19902440 . _:b19902458 . _:b511093495 . _:b19902433 _:b19902441 . _:b511093536 "2"^^ . _:b19902433 _:b19902442 . _:b19902439 . _:b19902429 "patients that have been shown to have low expression levels of either XPA or ERCC1 demonstrate higher sensitivity to cisplatin treatment, and people deficient for XPA (or other XP proteins) are hypersensitive to UV radiations [>>13<<], [14]. Hence, here we continue our earlier efforts aimed at the identification and characterization of novel inhibitors of the interaction between ERCC1 and XPA [15], in order to regulate the NER pathway and offer new alternatives to be" . _:b19902433 _:b19902443 . _:b19902433 _:b19902444 . _:b19902459 . _:b19902433 _:b19902445 . _:b511093539 "2"^^ . _:b511093500 . _:b19902433 _:b19902446 . _:b511093501 . _:b511093502 . _:b511093503 . . _:b511093496 . _:b19902416 . _:b511093538 "2"^^ . _:b511093497 . _:b511093498 . _:b511093499 . _:b19902431 "Hence, here we continue our earlier efforts aimed at the identification and characterization of novel inhibitors of the interaction between ERCC1 and XPA [>>15<<], in order to regulate the NER pathway and offer new alternatives to be added to the current NER and cell cycle inhibitor UCN-01(7-hydroxystaurosporine) [16]." . _:b19902457 . . _:b19902454 . _:b19902447 _:b19902456 . _:b19902473 . _:b19902447 _:b19902457 . _:b19902447 _:b19902458 . _:b19902447 _:b19902459 . _:b19902447 _:b19902460 . _:b19902444 "This study utilized the molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) technique to rescore the preliminary ranked docking hits [>>32<<]. It combines molecular mechanics with continuum solvation models." . _:b19902447 _:b19902461 . _:b511093524 . _:b19902447 _:b19902462 . _:b19902453 . _:b511093525 . _:b19902447 _:b19902463 . _:b511093526 . _:b19902447 _:b19902448 . _:b511093527 . _:b19902447 _:b19902449 . _:b511093520 . _:b19902447 _:b19902450 . _:b511093521 . _:b19902447 _:b19902451 . . _:b511093522 . _:b19902447 _:b19902452 . _:b511093523 . _:b19902447 _:b19902453 . _:b511093532 . . _:b19902443 . _:b511093533 . _:b19902447 _:b19902454 . _:b511093493 . _:b511093534 . _:b19902447 _:b19902455 . . _:b511093535 . _:b19902447 "results and discussion" . _:b511093528 . _:b511093529 . _:b19902451 . _:b511093530 . _:b511093531 . _:b511093508 . _:b511093509 . _:b511093495 . . _:b511093510 . _:b511093511 . _:b511093504 . _:b19902450 . _:b511093534 . _:b511093505 . _:b511093494 . _:b511093506 . . _:b511093529 . _:b511093507 . _:b511093516 . _:b511093517 . _:b511093518 . _:b511093519 . _:b511093512 . _:b511093513 . _:b511093514 . _:b511093515 . _:b19902435 "The binding site was characterized in our previous work (see Figure 2) [>>15<<]. In this model, the central domain of ERCC1 (residues 99\u2013214) is bound to a fragment of XPA (residues 67\u201377)." . _:b19902467 . . _:b19902432 "characterization of novel inhibitors of the interaction between ERCC1 and XPA [15], in order to regulate the NER pathway and offer new alternatives to be added to the current NER and cell cycle inhibitor UCN-01(7-hydroxystaurosporine) [>>16<<]. The present work introduces a promising lead compound NERI01 (NER inhibitor 01) that targets the ERCC1-XPA interaction and sensitizes cancer cells to ultraviolet irradiation induced damage." . . . _:b19902471 "It was also validated as a VS refining tool and revealed excellent results in predicting the actual binding affinities and in discriminating true binders from inactive (decoy) compounds [33], [34], [>>35<<]. Its main advantages are the lack of adjustable parameters and the option of using a single MD simulation for the complete system to determine all energy values." . _:b511093501 . _:b511093504 . _:b511093500 . _:b19902466 . _:b511093540 . . _:b19902425 "ERCC1-XPF is recruited to the damage site through a direct interaction between the centeral domain of ERCC1 and XPA, an indispensible element of the NER pathways [>>9<<], [10]. No cellular function beyond NER has been observed for XPA and competitive inhibition of the XPA interaction with peptide fragments is effective at disrupting NER [11], [12]. Furthermore, clinically, patients that have been shown to" . _:b511093541 . _:b19902414 . _:b511093503 . _:b511093542 . _:b511093543 . . _:b511093536 . _:b511093537 . _:b511093538 . _:b511093502 . _:b511093539 . _:b19902415 . _:b19902434 "Our next step relied on Tsodikov\u2019s NMR crystal structure of XPA bound to ERCC1 (PDB entry 2JNW) [>>12<<]. The NMR ensemble included 10 different conformations for the proteins; all of them were used in this study." . _:b511093497 . _:b19902427 "No cellular function beyond NER has been observed for XPA and competitive inhibition of the XPA interaction with peptide fragments is effective at disrupting NER [>>11<<], [12]. Furthermore, clinically, patients that have been shown to have low expression levels of either XPA or ERCC1 demonstrate higher sensitivity to cisplatin treatment, and people deficient for XPA (or other XP proteins) are" . . _:b511093537 . _:b511093496 . . _:b511093509 . _:b19902474 . _:b511093499 . _:b511093501 . _:b511093498 . _:b511093524 . . _:b19902459 . . . _:b19902433 . . _:b19902437 . _:b19902447 . _:b511093530 . _:b19902455 . _:b19902417 . _:b511093543 . . _:b19902475 "XP is a direct consequence of lacking one out of several NER proteins such as XPA [41]\u2013[>>43<<]. A major syndrome of XP is the hypersensitivity to UV radiation and, consequently, a high susceptibility to produce skin cancer. As the role of XPA within the NER mechanism is to interact with ERCC1 and recruit the XPF-ERCC1 endonuclease" . . _:b511093495 . . _:b19902453 "Other successful examples include the identification of novel inhibitors of the acetylcholine binding protein [24], RNA-editing ligase 1 [>>25<<], the influenza protein neuraminidase [26] and Trypanosoma brucei uridine diphosphate galactose 4\u2032-epimerase [27]." . . _:b19902414 "introduction" . . . _:b19902459 "(e.g. every 5 ns or so), [24] condensing the structural ensemble generated from MD simulations using QR factorization, [25] or clustering the MD trajectory using root-mean-square-deviation (RMSD) conformational clustering, [26], [>>27<<] On the other hand, to rank the screened compounds and suggest a final set of top hits, some studies used only docking predictions, [24], [25], [26]while others suggested (as in this thesis) using a more accurate scoring method (e.g." . _:b19902422 . _:b19902451 "An excellent example is that of an HIV inhibitor, raltegravir which became the first FDA approved drug targeting HIV integrase [>>22<<], [23]. Other successful examples include the identification of novel inhibitors of the acetylcholine binding protein [24], RNA-editing ligase 1 [25], the influenza protein neuraminidase [26] and Trypanosoma brucei uridine diphosphate" . _:b19902469 "It was also validated as a VS refining tool and revealed excellent results in predicting the actual binding affinities and in discriminating true binders from inactive (decoy) compounds [>>33<<], [34], [35]." . _:b19902424 . . _:b511093528 . _:b19902428 . . _:b19902434 . _:b511093520 . _:b19902442 . . _:b19902427 . _:b19902462 "MD trajectory using root-mean-square-deviation (RMSD) conformational clustering, [26], [27] On the other hand, to rank the screened compounds and suggest a final set of top hits, some studies used only docking predictions, [24], [25], [>>26<<]while others suggested (as in this thesis) using a more accurate scoring method (e.g. MM/PBSA (Molecular Mechanics/Poisson Boltzmann Surface Area)) to refine the final selected hits." . _:b19902474 "XP is a direct consequence of lacking one out of several NER proteins such as XPA [>>41<<]\u2013[43]. A major syndrome of XP is the hypersensitivity to UV radiation and, consequently, a high susceptibility to produce skin cancer. As the role of XPA within the NER mechanism is to interact with ERCC1 and recruit the XPF-ERCC1" . . . _:b19902470 "It was also validated as a VS refining tool and revealed excellent results in predicting the actual binding affinities and in discriminating true binders from inactive (decoy) compounds [33], [>>34<<], [35]. Its main advantages are the lack of adjustable parameters and the option of using a single MD simulation for the complete system to determine all energy values." . . . _:b19902441 . _:b19902446 "Media with or without potential inhibitors was added and cells were incubated for another 72 hours before living cells were quantified with methylthiazoletetrazolium (MTT) assay as previously described [>>57<<]." . . _:b511093515 . _:b19902471 . . . _:b19902466 "This adaptive approach was tested on other targets and led to successful outcomes [28], [>>29<<], [30]. For MD simulations, starting from the optimal binding mode is the most efficient route to reach equilibrium. Therefore, by running the clustering protocol on each ligand and filtering the hits in terms of the population of the" . _:b19902475 . _:b19902425 . _:b19902454 "Other successful examples include the identification of novel inhibitors of the acetylcholine binding protein [24], RNA-editing ligase 1 [25], the influenza protein neuraminidase [>>26<<] and Trypanosoma brucei uridine diphosphate galactose 4\u2032-epimerase [27]." . _:b19902440 . . _:b19902449 . . . _:b19902468 "In this context, our VS protocol utilized the MM-PBSA [>>32<<] to suggest the final ranked set of top hits (see Materials and Methods)." . _:b19902417 "However, in cancer cells exposed to DNA damaging agents that distort the DNA helix or form bulky injuries to the genome, NER comes into play and removes the damage, thus protecting cancer cells from death [1], [>>2<<]. A striking example of this mechanism is represented by the use of platinum compounds such as cisplatin, the backbone for many treatments of solid tumors including testicular, bladder, ovarian, head and neck, cervical, lung and colorectal" . . _:b19902438 . . . . _:b19902435 . _:b19902418 . _:b19902472 . _:b19902436 . _:b19902463 "used only docking predictions, [24], [25], [26]while others suggested (as in this thesis) using a more accurate scoring method (e.g. MM/PBSA (Molecular Mechanics/Poisson Boltzmann Surface Area)) to refine the final selected hits. [>>21<<] All of these approaches, similar to the work presented here, were aiming at keeping the balance between significantly reducing the number of target structures and, in the meantime, retaining their capacity to describe the conformational" . _:b511093516 . _:b19902420 . _:b511093532 . _:b511093522 . . _:b19902431 . _:b19902423 "In this context, one way to increase the efficacy of platinum therapy and decrease drug resistance is to regulate NER by inhibiting the activity of ERCC1 and interacting proteins using novel therapeutic compounds [>>8<<]." . _:b19902449 "In our present work, we exploited the published ERCC1-XPA NMR structure [>>12<<]. However, as the initial screening involved an enormous number of compounds (\u223C 90,000) (see below), it was important to start the docking simulations using a representative target structure." . _:b19902469 . . _:b511093494 . _:b511093500 . . _:b19902473 "As aforementioned, NER is a major DNA repair pathway that eliminates DNA lesions induced by UV light [>>40<<]. A deficiency in NER leads to dramatic diseases characterized by hypersensitivity to UV and a prominent clinical and genetic heterogeneity." . . _:b19902436 "Prior to docking, the XPA peptide was removed, protonation states of the residues constituting the ERCC1 pocket were adjusted using the software PDB2PQR [>>45<<], and the protein structures were conformationally relaxed using the NAMD molecular dynamics software with constraints on the backbone atoms (see below)." . _:b511093498 . _:b19902458 "simulation (e.g. every 5 ns or so), [24] condensing the structural ensemble generated from MD simulations using QR factorization, [25] or clustering the MD trajectory using root-mean-square-deviation (RMSD) conformational clustering, [>>26<<], [27] On the other hand, to rank the screened compounds and suggest a final set of top hits, some studies used only docking predictions, [24], [25], [26]while others suggested (as in this thesis) using a more accurate scoring method (e.g." . . _:b511093497 . _:b511093505 . . . . "PMC0" . _:b511093502 . _:b19902429 . . _:b19902465 . _:b19902455 "examples include the identification of novel inhibitors of the acetylcholine binding protein [24], RNA-editing ligase 1 [25], the influenza protein neuraminidase [26] and Trypanosoma brucei uridine diphosphate galactose 4\u2032-epimerase [>>27<<]. These applications employed alternative ways to solve two main problems with the method, namely, reducing the number of extracted target conformations and deciding on how to select the final set of hits after carrying out the screening" . _:b511093539 . . _:b511093541 . _:b19902418 "example of this mechanism is represented by the use of platinum compounds such as cisplatin, the backbone for many treatments of solid tumors including testicular, bladder, ovarian, head and neck, cervical, lung and colorectal cancer [>>3<<]. It has been demonstrated that NER is the major DNA repair mechanism that removes cisplatin-induced DNA damage, and that resistance to platinum-based therapy correlates with high expression of ERCC1, a major element of the NER machinery" . . _:b19902423 . "10.1371%2Fjournal.pone.0051329" . . _:b511093503 . . . _:b511093517 . _:b19902430 "patients that have been shown to have low expression levels of either XPA or ERCC1 demonstrate higher sensitivity to cisplatin treatment, and people deficient for XPA (or other XP proteins) are hypersensitive to UV radiations [13], [>>14<<]. Hence, here we continue our earlier efforts aimed at the identification and characterization of novel inhibitors of the interaction between ERCC1 and XPA [15], in order to regulate the NER pathway and offer new alternatives to be added" . _:b19902445 "Most compounds have not been reported elsewhere, but the synthesis of AB-00005094 [>>54<<], AB-00012818 and AB-00012800 [55], [56] have previously been published." . _:b19902464 . . _:b19902448 . . _:b19902426 "ERCC1-XPF is recruited to the damage site through a direct interaction between the centeral domain of ERCC1 and XPA, an indispensible element of the NER pathways [9], [>>10<<]. No cellular function beyond NER has been observed for XPA and competitive inhibition of the XPA interaction with peptide fragments is effective at disrupting NER [11], [12]. Furthermore, clinically, patients that have been shown to have" . _:b19902440 "Clustering of the docking results followed the same adaptive procedure as the one employed in our previous study [>>15<<]. In brief, for each docking simulation a modified version of the PTRAJ module of AMBER [48] clustered the docking trials." . . _:b511093496 . . . . _:b19902468 . _:b19902452 "Other successful examples include the identification of novel inhibitors of the acetylcholine binding protein [>>24<<], RNA-editing ligase 1 [25], the influenza protein neuraminidase [26] and Trypanosoma brucei uridine diphosphate galactose 4\u2032-epimerase [27]." . _:b19902469 . _:b19902446 . _:b19902448 "For example, Philippopoulos et al. suggested NMR structures as the most effective source for protein conformations [>>20<<]. A set of 15 NMR conformations for ribonuclease HI was compared to a trajectory obtained from a 1.7 ns MD simulation. The NMR data explored the conformational space of the protein more efficiently than the conventional MD simulation. In" . _:b19902470 . _:b19902472 "Ms\u22121) through the following equation: KA\u200A=\u200AKQ \u03C40 [37], where KA is the association constant, KQ is the biomolecular rate quenching rate constant and \u03C40 is the average lifetime of the biomolecule without a quencher (\u03C40\u200A=\u200A10\u22128 s) [>>38<<]. The results obtained from this study show that the estimated values for KA are greater than the maximum scatter quenching constant of various quenchers with the biopolymers (KQ\u200A=\u200A2\u00D71010 Ms\u22121) [39] which indicates that the observed static" . . _:b19902471 . _:b19902464 . _:b19902414 . _:b19902465 "This adaptive approach was tested on other targets and led to successful outcomes [>>28<<], [29], [30]." . _:b19902465 . _:b19902415 . _:b511093508 . _:b19902466 . . _:b511093495 "4"^^ . _:b19902450 "The second stage was a more rigorous docking approach that employed the RCS methodology [>>21<<]. In the RCS approach, all-atom MD simulations (e.g., 2\u20135 ns simulation) are applied to explore the conformational space of the target, while docking is subsequently used for the fast screening of drug libraries against an ensemble of" . . _:b19902467 . _:b19902420 "been demonstrated that NER is the major DNA repair mechanism that removes cisplatin-induced DNA damage, and that resistance to platinum-based therapy correlates with high expression of ERCC1, a major element of the NER machinery [4], [>>5<<], [6], [7]. In this context, one way to increase the efficacy of platinum therapy and decrease drug resistance is to regulate NER by inhibiting the activity of ERCC1 and interacting proteins using novel therapeutic compounds [8]." . _:b19902419 . _:b511093493 "7"^^ . _:b511093494 "4"^^ . _:b19902461 "the MD trajectory using root-mean-square-deviation (RMSD) conformational clustering, [26], [27] On the other hand, to rank the screened compounds and suggest a final set of top hits, some studies used only docking predictions, [24], [>>25<<], [26]while others suggested (as in this thesis) using a more accurate scoring method (e.g. MM/PBSA (Molecular Mechanics/Poisson Boltzmann Surface Area)) to refine the final selected hits." . . . _:b511093509 . _:b19902420 . _:b511093506 . . _:b511093508 . _:b19902421 . _:b19902472 . _:b511093501 "3"^^ . _:b511093514 . _:b19902421 . _:b511093511 . _:b19902422 . . _:b19902473 . _:b511093500 "3"^^ . _:b19902470 . _:b511093521 . _:b19902424 "The protein ERCC1 forms a heterodimer with XPF. The resulting complex is an endonuclease enzyme that cleaves the 5` end of the damage whereas XPG cleaves in the 3\u2032 position (for a comprehensive review on NER, see ref. [>>2<<]). ERCC1-XPF is recruited to the damage site through a direct interaction between the centeral domain of ERCC1 and XPA, an indispensible element of the NER pathways [9], [10]." . _:b19902423 . _:b511093510 . _:b19902474 . _:b511093503 "3"^^ . _:b511093505 . _:b19902416 . _:b19902475 . _:b511093531 . _:b511093502 "3"^^ . _:b511093504 . _:b19902417 . _:b19902433 "materials and methods" . _:b511093510 . . . _:b511093533 . . _:b511093497 "4"^^ . _:b511093540 . _:b511093507 . _:b19902418 . _:b19902419 "It has been demonstrated that NER is the major DNA repair mechanism that removes cisplatin-induced DNA damage, and that resistance to platinum-based therapy correlates with high expression of ERCC1, a major element of the NER machinery [>>4<<], [5], [6], [7]. In this context, one way to increase the efficacy of platinum therapy and decrease drug resistance is to regulate NER by inhibiting the activity of ERCC1 and interacting proteins using novel therapeutic compounds [8]." . _:b511093496 "4"^^ . _:b511093526 . _:b19902419 . _:b511093506 . . _:b511093499 "3"^^ . . . _:b511093517 . _:b19902428 . _:b511093499 . _:b511093498 "3"^^ . _:b19902430 . _:b19902447 _:b19902472 . _:b19902447 _:b19902473 . _:b19902429 . _:b511093516 . . . . _:b19902447 _:b19902474 . _:b19902447 _:b19902475 . _:b511093509 "3"^^ . _:b511093519 . _:b19902430 . . _:b511093542 . _:b511093508 "3"^^ . _:b19902447 _:b19902464 . _:b19902447 _:b19902465 . _:b19902439 "First, we screened the entire CN library against a single target model followed by applying the relaxed complex scheme (RCS) [>>47<<] through docking of the top 2,000 hits from the first screen against the rest of the ten target structures (see results for more details)." . _:b19902431 . _:b511093518 .