medicinal applications of metal complexes as therapeutic drugs have a more than 5000-year 
 history .
since the discovery of the 
 anticancer activity of cisplatin by shimizu and rosenberg 35 years ago , there has been 
 a rapid expansion in research to find new , 
 more effective metal - based anticancer drugs .
the major classes of 
 metal - based anticancer drugs include platinum ( ii ) , gold ( i ) and gold ( iii ) , 
 metalloporphyrin , ruthenium ( ii ) and ruthenium ( iii ) , bismuth ( iii ) , rhenium ( i ) , 
 and copper ( ii )   compounds .
cisplatin represents one of the most potent drugs available in the cancer chemotherapy 
 for several solid tumors , such as testicular , ovarian , bladder , and neck 
 cancers .
it is generally believed 
 that cisplatin exhibited its anticancer effects through preferentially binding 
 to quinine n-7 of dna , and then cause dna damage specifically in cancer cells , subsequently 
 leading to cell death .
after successes achieved 
 with platinum complexes , there is a tremendous increase in the search for 
 platinum complexes with different ligands that might produce more specific 
 anticancer effects . some of these platinum - based drugs have been approved by 
 the food and drug administration ( fda ) , including carboplatin for the treatment 
 of ovarian cancer [ 5 , 6 ] , oxaliplatin for metastatic 
 colorectal cancer [ 7 , 8 ] , satraplatin for 
 hormone - refractory prostate cancer , and picoplatin for 
 small - cell lung cancer .
transplatinum compounds follow different patterns of cell killing in comparison to 
 cisplatinum , thus giving a reason for optimism in their development as a new 
 class of platinum - based anticancer drugs .
the initial report of 
 anticancer properties of a dinuclear platinum complex in 1988 started a new 
 paradigm in platinum - based chemotherapy .
several multinuclear platinum 
 complexes have entered clinical trials in recent years , with varying results [ 12 , 13 ] .
the major limitations of cisplatin 
 and other platinum anticancer drugs are related to drug resistance and their 
 side effects , including nephrotoxicity , neurotoxicity , and emetogensis .
resistance to cisplatin is 
 multifactorial , most cases consist of mechanisms limiting the formation of dna 
 adducts or operating downstream of the cisplatin - dna interaction to promote 
 cell survival . gold ( i )
complexes had been used for the treatment of arthritis some decades ago , but most of them 
 disappeared from the drug market because of intolerable side effects , such as 
 gastrointestinal adverse reactions , nephrotoxicity , and haematological 
 reactions .
however , the design and testing of gold complexes , especially gold ( iii ) 
 complexes with anticancer activity begin to be intensively pursued in the past 
 few years .
the potential use of gold ( iii ) complexes as anticancer drugs were 
 based on three rationales [ 1618 ] : ( a ) analogies between square 
 planar complexes of both platinum ( ii ) and gold ( iii ) are d ions ; 
 ( b ) analogy to the immunomodulatory effects of gold ( i ) antiarthritic agents ; 
 and ( c ) complexation of gold ( i ) and gold ( iii ) with known anticancer agents to 
 form new compounds with enhanced activity .
buckley et al . first reported some organogold ( iii ) complexes endowed with 
 significant cytotoxic and anticancer properties . during the past decades , 
 various gold ( iii ) complexes with sufficient stability in the physiological 
 environment have been synthesized and evaluated for in vitro anticancer properties .
some of these gold ( iii ) 
 complexes turned out to exhibit relevant cytotoxic effects in vitro and were the subject of 
 further biochemical and pharmacological investigations [ 2033 ] .
our previous findings showed 
 that gold ( iii ) mesotetraarylporphyrin 
 1a was stable against demetallation in physiological conditions and exhibited 
 higher cytotoxicity than cisplatin against a panel of human cancer cell lines [ 3437 ] .
the major limitation of gold 
 ( iii ) complexes is that few exhibit good stability under physiological 
 conditions , due to the reduction of gold ( iii ) to gold ( i ) .
however , low cisplatin cross 
 resistance has been observed in gold complexes . there is therefore 
 considerable interest in the development of tumor - selective and stable gold 
 anticancer drugs .
metalloporphyrin drugs are new class of antioxidant enzyme mimetics with novel structure : a 
 metal in the center of porphyrin ligand .
metalloporphyrins ( e.g. , mntbap ) have previously 
 been used to inhibit age - related oxidative damage in myocardium of mice that 
 are lacking mitochondrial enzyme manganese superoxide dismutase .
afterwards , metalloporphyrin 
 drugs began to be used as photodynamic therapy agents for certain solid tumors .
photodynamic therapy is 
 based on the concept that porphyrins are known to be rapidly and preferentially 
 taken up by the tumor cells with higher intakes of lipoproteins [ 42 , 43 ] .
when such photosensitizers 
 are irradiated with an appropriate wavelength of visible or near infrared ( nir ) 
 light , the excited molecules can transfer their energy to molecular oxygen in 
 the surroundings , which is normally in its triplet ground state .
this results 
 in the formation of cytotoxic reactive oxygen species ( ross ) , particularly 
 singlet oxygen .
ross are responsible for 
 oxidizing various cellular compartments including plasma , mitochondrial , 
 lysosomal , and nuclear membranes , resulting in irreversible damage of tumor [ 34 , 44 ] .
therefore , under appropriate 
 conditions , photodynamic therapy offers the advantage of an effective and 
 selective method of destroying diseased tissues without damaging adjacent 
 healthy cells [ 42 , 43 ] . since the approval 
 of photofrin by fda for chemotherapy , porphyrin derivatives with 
 different metal in the center of the molecule have been widely used as 
 photosensitizers for photodynamic therapy in the treatment of cancer , including 
 chlorophyllin copper complex as superoxide dismutase mimics [ 46 , 47 ] , fetbap and mntbap [ 48 , 49 ] , zntbap , motexafin gadolinium ( mgd ) .
for example , mgd has been shown to inhibit heme 
 oxygenase-1 ( ho-1 ) activity that results in inactivation of the antiapoptotic properties of the 
 products of ho-1 . while fetbap and mntbap have 
 been reported to be superoxide anion scavengers .
mgd is also an active 
 inhibitor of cytochrome p450 enzymes , although with a lower potency than that 
 exhibited for inhibition of ho-1 . in recent years , other approaches in the search for new ,
metal - based anticancer agents are to 
 examine complexes that contain other transition metals . in the design of these 
 new drugs , octahedral ruthenium ( ii ) and ruthenium ( iii ) complexes
tetraammine- , 
 pentaammine- , heterocycle- , and dimethylsulfoxide - coordinated ruthenium 
 complexes have been synthesized and shown high affinity for nitrogen donor 
 ligands in vitro and as a result exhibit anticancer action in 
 vivo [ 5254 ] .
other transition metals have 
 been used as anticancer drugs , including bismuth ( iii ) labeled antibodies for 
 systemic radioimmunotherapy [ 55 , 56 ] , rhenium ( i ) complexes as 
 dna - binding agents , ( mtr)2zn complex that induces cancer cell death by binding to chromatin , and cu compound 
 chlorophyllin initiated apoptosis in human colon cancer cells through caspase-8 
 and apoptosis - inducing factor ( aif ) activation in a cytochrome c - independent 
 manner .
the proteome is defined as all expressed protein complement of a cell , organ or organism , and 
 it includes all isoforms and posttranslational variants .
proteomic technology , 
 first coined in 1995 , attempts to separate , 
 identify , and characterize a global set of proteins in an effort to provide 
 information about protein abundance , location , modification , and 
 protein - protein interaction in the proteome of a given biological system [ 59 , 60 ] .
this postgenomic technology 
 provides a direct measurement of the presence and relative abundance of 
 proteins , and reveals the consequence of protein functioning in establishing 
 the biological phenotype of organisms in different states . by studying 
 interrelationships of protein expressions and modifications in health and 
 disease or drug treatment ,
proteomics contributes important insights into 
 determining the pathophysiological basis of disease , validating drug targets , and illustrating drug action 
 , toxicity , and side effects . in the field of proteomics ,
several 
 well - established methods persist as means to resolve and analyze complex mixtures 
 of proteins derived from cells and tissues .
currently , the most commonly used 
 proteomic platforms include two - dimensional gel electrophoresis ( 2de ) and 
 protein chip arrays , isotope - coded affinity tages ( icats ) , and immobilized metal affinity 
 chromatography ( imac ) ( table 1 ) .
these technological platforms are most often incorporated 
 with matrix - assisted laser desorption / ionization time of flight ( maldi - tof ) , surface enhanced laser 
 desorption ionization time of flight ( seldi - tof ) , electrospray ionization 
 ( esi ) , and/or tandem mass 
 spectrometry ( ms / ms ) .
in addition , inductively coupled plasma ( icp ) mass spectrometry 
 has also been applied in proteomic - based research of drug discovery .
ever since the initial discovery of the anticancer activity of cisplatin , major efforts have 
 been devoted to elucidate the biochemical mechanisms of anticancer activities 
 of metal - based drugs to facilitate rational design of novel metal - based drugs 
 with better pharmacological profiles .
a comprehensive understanding of the 
 molecular action mechanisms , which are triggered by metal - based drugs to kill 
 cancer cells can lead to the design of more effective anticancer drugs , as well 
 as to provide new therapeutic strategies based on the molecular activity of 
 metal - based drug activity .
target discovery , which involves the identification and early validation of disease - associated 
 targets , is an essential first step in the drug discovery pipeline .
indeed , the drive to 
 determine protein function has been stimulated , both in industry and academia , 
 by the human genome and proteome projects in progress .
proteomics , the study of 
 cellular protein expression , is an evolving technology platform that has the 
 potential to identify novel proteins involved in key biological processes in 
 cells .
it must 
 be pointed out , however , that numerous drug - targeted proteins are 
 membrane - bound proteins , for example , receptors and ion channels .
these 
 proteins may not be amenable for study by proteomics due to their poor 
 solubility and low abundance , and thus they are disproportionally represented 
 in proteome profiles .
up to date , only a fraction 
 of putative drug targets has been identified by proteomic approaches , including 
 the volume - sensitive organic osmolyte / anion channel as key elements of tumor 
 development , migration , and invasiveness , and integrin alpha-4 as a 
 molecular target of oxidative stress .
studying protein expression profiling of drug - treatment leads to the identification of a number 
 of drug - specific targets both in vivo and in vitro . using 
 hplc - maldi - tof ms , hasinoff et al .
have identified topoisomerase ii contained at least five free cysteins ( 170 , 
 216 , 300 , 392 , and 405 ) and two disulfide - bonded cyteine pairs ( 427 - 455 and 
 997 - 1008 ) .
cisplatin was found to antagonize 
 the formation of a fluorescence adduct between topoisomeraser ii and the 
 sulfhydryl - reactive maleimide reagent 
 10-(2,5-dihydro-2,5-dioxo-1h - pyrrol-1-yl)-9-methoxy-    3-oxo-3h - naphtho[2,1-b]pyran-2-carboxylic 
 acid methyl   ester ( thioglo-1 ) .
based on these results , the authors suggested 
 that topoisomeraser ii cysteines may be possible sites responsible for the 
 inhibition of the catalytic activity of topoisomeraser ii observed in the 
 presence of cisplatin , and topoisomeraser ii cysteins and dna as targets 
 responsible for cisplatin - induced inhibition of topoisomeraser ii . besides , mitochondrial 
 proteins , especially atp synthase - beta subunit , have been reported to be key 
 proteins that serve as primary target of manganese porphyrin [ mntnhex-2-pyp(5 + ) ] 
 treatment during renal ischemia / reperfusion injury by proteomic study .
icp ms coupled with capillary electrophoresis ( ce ) has been used in the identification , 
 characterization , and determination of different chemical species of an element 
 in complex biological systems , that is , the impetus of biochemical speciation 
 analysis [ 87 , 88 ] .
polec - pawlak et al . have used this approach to 
 study the platinum group metallodrug - protein binding aiming to characterize the 
 interactions between cancer - inhibiting metal complexes and serum transport 
 proteins .
such binding does not only 
 regulate the uptake and accumulation of the drug in tumor tissue but also 
 determines its overall distribution and exertion and differences in efficacy , activity , and 
 toxicity [ 90 , 91 ] .
their study provides clear 
 evidence that a ruthenium ( iii ) complex 
 [ trans - tetrachlorobis(1h - indazole)ruthenate ( iii ) ] ( kp1019 ) preferentially 
 binds toward albumin whose adduct is a dominating protein - bound species of ruthenium 
 .
insights into toxic responses are an asset for the interpretation of adverse drug effects and 
 contribution to accurate risk assessment for humans
. proteomics in combination 
 with combinatorial chemistry and high - throughput screening can help to bring 
 forward validating toxicity and resistance of an unprecedented number of 
 potential lead compounds .
benefits can be expected in 
 optimized clinical trials based on the availability of biologically relevant 
 markers of drug efficacy and safety .
proteomics has demonstrated 
 proof - of - concept in toxicology as shown by a number of successful applications 
 in mechanistic toxicology and lead selection .
proteomic studies of liver 
 toxicity have been carried out with thioacetamide   and ethanol .
other researches have 
 studied nephrotoxicity of cyclosporine a by proteomic approaches [ 94 , 95 ] and reported a profound downregulation 
 of the calcium binding protein calbindin d28 responsible for cyclosporine 
 a - induced kidney toxicity .
however , this technology - driven acceleration in drug 
 discovery moves the bottlenecks in drug development to the downstream , which is 
 the improvement in the selection of patient populations for clinical trials .
proteomic approaches have been used to identify the underlying mechanisms for cisplatin 
 resistance . in this study , the authors 
 used cervix squamous cell carcinoma cell line a431 and its cisplatin - resistant 
 subline , a431/pt as model system . the identified differentially expressed 
 proteins can be classified into several groups , including molecular chaperones 
 ( e.g. , heat - shock protein hsp60 , hsp90 , and hsc71 ) , ca - binding 
 proteins ( e.g. , calmodulin and calumenin ) , proteins involved in drug 
 detoxification ( e.g. , peroxiredoxin 2 and 6 , and glutathione - s - transferase ) , 
 antiapoptotic proteins ( e.g. , 14 - 3 - 3 switched on in cisplatin - exposed cells ) 
 and ion channels ( e.g. , voltage - dependent anion channel 1 , voltage - dependent 
 anion - selective channel ) . besides this , proteome profile of cisplatin sensitive 
 ovarian cell line igrov1 and its cisplatin - resistant counterpart igrov1-r10 
 have been compared aiming to find any protein markers or to establish new 
 therapeutic strategies [ 97 , 98 ] . increased expression of 
 cytokeratin 8 and cytokeratin 18
was considered to play a role in acquired chemoresistance of 
 igrov1-r10 cancer cell line to cisplatin [ 97 , 98 ] .
cytokeratin 8 and cytokeratin 
 18 have been implicated in resistance to tnf--induced apoptosis by binding the 
 cytoplasmic domain of tumor necrosis factor receptor 1 [ 69 , 99 ] . moreover ,
human 
 nasopharyngeal carcinoma cells deficient for cytokeratin 8 were more sensitive 
 to cisplatin - induced apoptosis .
in addition , acquired and intrinsic cellular drug resistances are multifactorial processes , 
 involving induction of drug detoxifying mechanisms , quantitative and 
 qualitative modification of drug targets , cell cycle arrest , regulation of dna 
 replication or reparation mechanisms , modulation of apoptosis , and other mechanisms 
 [ 101 , 102 ] .
global examination of the glycoproteomes of 
 the cisplatin - resistant ovarian cancer cell line igrov-1/cp using shotgun 
 glycopeptide capture approach coupled with ms has been used to study cisplatin resistance . in this approach , 

glycopeptides derived from glycoproteins are enriched by selective capture onto 
 a solid support using hydrazide chemistry followed by enzymatic release of the 
 peptides and subsequent analysis by ms / ms .
this method improves solubility of 
 large membrane proteins and exposes all of the glycosylation sites to ensure 
 equal accessibility to capture reagents .
stewart et al . also used isotope - coded affinity tags ( icats ) integrated 
 with mrna expression levels to study cisplatin resistance in ovarian cancer 
 cells .
their study identified three 
 pathways in panther database ( http://www.pantherdb.org/ ) that were significantly ( p < .05 ) upregulated 
 in cisplatin - sensitive cells , including glycolysis , interleukin signaling 
 pathway , and pi 3-kinase pathway .
an understanding of protein function within the context of complex cellular networks is required 
 to facilitate the discovery of novel drug targets and , subsequently , new 
 therapies directed against them .
being the basic 
 biochemical mode of drug activities , drug action mechanism should be better 
 understood to provide valuable insights into drug modification and new drug 
 development [ 34 , 105 ] .
successful examples in drug 
 mechanism study using proteomics include the illustration of insulin - like 
 growth factor - binding protein-6-induced sublethal hydrogen peroxide stress in 
 human diploid fibroblasts cells . by using esi ms / ms , kanski et al .
have applied proteomic 
 analysis of protein nitration in aging skeletal muscle and identified 
 nitrotyrosine - containing sequences in vivo . in our previous study , we have used 2de - based proteomic technology to compare the protein 
 profile of human nasopharyngeal carcinoma sune1 cell line treated with gold ( iii ) 
 porphyrin 1a , and a number of differentially expressed proteins were identified 
 .
these proteins can be 
 classified into several categories based on their major biological functions , 
 including cellular structural proteins , stress - related and chaperone proteins , 
 proteins involved in ros , enzyme proteins and translation factors , proteins 
 that mediate cell death and survival signaling , and proteins that participate 
 in the internal degradation system . among these proteins ,
one of 
 the significant increased proteins is voltage - dependent anion channel 1 ( vdac 
 1 ) .
vdac 1 is a mitochondrial outer membrane channel protein , which functions 
 as the pathway for the movement of various substances in and out of the 
 mitochondria .
it is considered to be a 
 component of the permeability transition pore oligoprotein complex that plays a 
 role in the permeability transition [ 109 , 110 ] .
vdac 1 also plays an 
 essential role in bax / bak - induced apoptotic mitochondrial changes in the 
 process of mammalian cell death [ 111113 ] . in this process , the 
 proapoptotic proteins bax and bak bind to vdac 1 , and enhance its permeability 
 so that cytochrome c passes through the channel and releases to cytoplasm [ 111113 ] .
our data on vdac 1 
 upregulation   and bax overexpression   suggest that gold ( iii ) 
 porphyrin 1a may induce cell death via the mitochondria - mediated apoptosis 
 pathway .
further functional studies revealed that gold ( iii ) porphyrin 1a 
 caused depletion of mitochondrial transmembrane potential ( m ) soon 
 after uptake with suppression of bcl-2 , and activation of caspase 9 and caspase 
 3 . taken together , these 
 results suggested that mitochondria are the primary target of gold ( iii ) 
 porphyrin 1a .
quantitative proteomic analysis on other metal - based anticancer drugs has also been pursuit . 

schmidt et al . have used nano - lc coupled offline maldi - tof / tof - ms 
 to study cisplatin - induced apoptosis in jurkat t cells .
their results showed that 
 this method is more accurate than the commonly used online lc - esi - ms .
the potential value of proteomics in metal - based drug development , especially in 
 mapping drug action mechanisms , has been demonstrated in many successful 
 examples .
proteomic approaches have been recognized as promising techniques 
 that can facilitate the systematic characterization of a drug targets ' 
 physiology , thereby helping to reduce the typically high attrition rates in 
 discovery projects , and improving the overall efficiency of pharmaceutical 
 research processes . however , at present stage , the bottleneck for taking full 
 advantage of this new experimental technology is the rapidly growing volumes of 
 automatically produced biological data , and technical challenges with regards 
 to sampling , tumor heterogeneity , and lack of standardized methodologies . in 
 addition , to complement the limitation of current proteomic technology , systematic 
 biological and pharmaceutical studies should be integrated with proteomics to 
 better serve the purpose of illustrating the action mechanism of drugs and thus 
 contribute to the success in metal - based drug development .
 medicinal inorganic chemistry has been stimulating largely by the success of the anticancer drug , cisplatin . 
 various metal complexes are currently used as therapeutic agents ( e.g. , pt , au , and ru ) in the treatment of malignant diseases , including several types of cancers . understanding the mechanism of action of these metal - based drugs is for the design of more effective drugs . 
 proteomic approaches combined with other biochemical methods can provide comprehensive understanding of responses that are involved in metal - based anticancer drugs - induced cell death , including insights into cytotoxic effects of metal - based anticancer drugs , correlation of protein alterations to drug targets , and prediction of drug resistance and toxicity . 
 this information , when coupled with clinical data , can provide rational basses for the future design and modification of present used metal - based anticancer drugs . 
1. METAL-BASED ANTICANCER DRUGS
2. PROTEOMICS
3. POTENTIAL APPLICATIONS OF  PROTEOMICS IN ILLUSTRATING METAL-BASED DRUG  DEVELOPMENT AND DISCOVERY
4. CONCLUSION AND FUTURE PROSPECTS