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Anticancer activity, DNA-binding and DNA-denaturing aptitude of palladium(II) dithiocarbamates
Accepted Manuscript Review article Anticancer activity, DNA-binding and DNA-denaturing aptitude of palladium(II) dithiocarbamates Muhammad Kashif Amir, Shahan Zeb Khan, Faisal Hayat, Abbas Hassan Khan, Ian S. Butler, Zia-ur- Rehman PII: DOI: Reference:
S0020-1693(16)30346-2 http://dx.doi.org/10.1016/j.ica.2016.06.036 ICA 17127
To appear in:
Inorganica Chimica Acta
Received Date: Revised Date: Accepted Date:
10 March 2016 19 May 2016 26 June 2016
Please cite this article as: M.K. Amir, S.Z. Khan, F. Hayat, A.H. Khan, I.S. Butler, Z-u. Rehman, Anticancer activity, DNA-binding and DNA-denaturing aptitude of palladium(II) dithiocarbamates, Inorganica Chimica Acta (2016), doi: http://dx.doi.org/10.1016/j.ica.2016.06.036
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Anticancer activity, DNA-binding and DNA-denaturing aptitude of palladium(II) dithiocarbamates Muhammad Kashif Amira, Shahan Zeb Khana,b, Faisal Hayata, Abbas Hassan Khana, Ian S. Butlerc and Zia-ur-Rehmana* a
Department of Chemistry, Quaid-i-Azam University, Islamabad-45320, Pakistan.
b
Department of Chemistry, University of Science & Technology, Bannu-28100, KPK, Pakistan c
Department of Chemistry, McGill University, 801 Sherbrooke St. West, Montreal, Quebec, Canada H3A2K6 *
Corresponding author
Email: [email protected]/[email protected] Tel: 92-(051)90642245 Fax: 92-(051)90642241
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Abstract Although it is well known that cisplatin and its analogues are effective anticancer agents, but their clinical use is restricted by some serious side effects. Palladium complexes are emerged as altenative metallo-anticancer drugs merited by their structural similarity to platinum(II) complexes, more labile nature, minimal chemoresistance and often water solubility. However, due to exceptional high reactivity of palladium complexes than their platinum counterparts, they are not only obstructed by the sulfur containing molecules to reach their pharmocolgical targets but also significantly enhance their affinity to convert into inactive trans isomers. This hitch can be overcome by the use of appropriate ligands that can have the potential to turn down the negative lability to positive inertness. This review provides a summary of the anticancer potential, DNA-binding and DNA-denaturing aptitude of various palladium(II) dithiocarbamates in which the presence of the dithiocarbamate moiety significantly improves the anticancer action and, at the same time, reduces the possibility of any damaging side effects.
Keywords: Palladium(II) dithiocarbamates; Anticancer drugs; DNA-binding and DNAdenaturing
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1. Introduction Cancer is undoubtedly one of the world’s major health issues and several million people died from this disease every year. Following the accidental discovery of Rosenberg and his colleagues in 1965 of the eventual utility of cis-Pt(NH3)2Cl2 in cancer treatment [1], a wide variety of platinum(II)-based complexes have been investigated and now several other drugs with excellent anticancer activity are commercially available, e.g., carboplatin, oxaliplatin, nedaplatin, lobaplatin, and heptaplatin (Fig 1) [2]. The anticancer properties of the platinum(II) drugs can be attributed to the formation of intrastrand crosslinks between the guanine nitrogen bases of DNA, causing DNA bending, which subsequently interferes with DNA replication, transcription and other nuclear functions, and finally inhibits cancer cell proliferation and tumor growth. The clinical use of cisplatin and related complexes is, however, limited because of some unwelcome side effects that accompany treatment, including nephrotoxicity, ototoxicity, gastrointestinal toxicity, vomiting and developing resistance, which may be either primary (intrinsic) or secondary (acquired) [3-5]. It has been suggested that the nephrotoxicity and gastrointestinal toxicity of cisplatin may be the result of platinum complexation and inactivation of thiolcontaining enzymes [6, 7]. Platinum drug resistance is known to be a multifactorial process, which can occur through several mechanisms, including decreased platinum drug uptake due to resistance at the cell membrane, increased drug efflux, drug inactivation by glutathione and metallothioneins, evasion of apoptosis and enhancement of DNA repair [8, 9]. To overcome platinum drug resistance and toxicity, one of many strategies employed utilizes a variety of sulfur (thio)-containing ligands as detoxicant agents [10]. The small bite-angle and soft nature of the dithiocarbamato moiety as a stabilizing chelating ligand renders the platinum(II) complexes unreactive under a variety of conditions [11, 12]. In particular, dithiocarbamates have been 3
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established as excellent inhibitors of cisplatin-induced nephrotoxicity [13]. The strong binding of platinum(II) to the dithiocarbamate moiety limits any ligand exchange reactions with other sulfur-containing renal proteins and thiol-containing enzymes, thus protecting the normal tissues without undermining the anticancer action [14]. Fig. 1 Palladium complexes can be considered as substitute to platinum drugs for the treatment of different types of cancers owing to structural analogy [15-18]. However, their 105 times more reactivity than platinum complexes [19], make them more susceptible to fast hydrolysis accompanied by isomerization [19- 21]. The latter property is responsible for the formation of inactive trans isomers that lowers down anticancer activity [19]. However, this negative reactivity can be tunned to positive inertness by the right choice of the carrier ligands. It has been well notted that the bulky carrier ligands facilitate movement of the intact complex towards the target through steric shelter and encourage entrance into the cell through lipophilicity [19]. The reactivity can also be lowered down using ligands with soft donor sites i.e sulfur or phosphorous. In this context, detoxicant dithiocarbamate ligands received utmost attention due to the presence of strongly donating sulfur atoms and their chelating nature. Consequently, there is a growing interest in the designing of palladium(II) dithiocarbamates followed by their anticancer screening against different cancer cell lines, detailed DNA-binding and DNA-denaturing studies [22-28]. The detoxication effect of dithiocarbamates stems from their strong binding to palladium(II) that quenches ligand exchange reactions with sulfhydryl groups [29-31]. In this review, recent advances in the cytotoxic activities of palladium(II) dithiocarbamates against different cell lines, including sensitive 2008, A431, U2OScells and resistant C13*, A431/Pt, U2OS/Pt cells, are described. In addition, the current research on these palladium(II) complexes with respect to 4
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DNA synthesis inhibition, interstrand cross-linking, DNA–protein cross-linking, micronuclei (MN) detection, DNA-binding and denaturing is highlighted.
2. Anticancer activity of the palladium(II) dithiocarbamates The use of palladium(II) complexes containing N- and S-donor ligands in an effort to reduce the problems associated with the toxicity of cisplatin and its analogues is an important approach in anticancer studies. The palladium(II) complexes shown in Fig. 2 (1-4), which contain the 2ethoxy-2-oxoethyl(methyl)carbamodithioate ligand, have been examined for their cytotoxicity by performing dye exclusion tests on human leukemic HL-60 cells and by studying their inhibitory effects on cell proliferation in human adenocarcinoma HeLa cells [32]. Complex 1 is more active than is cisplatin against human leukemic HL-60 and adenocarcinoma HeLa cells (Table1). Notably, the neutral complexes, 1 and 2, which have a chloride ligand directly bonded to palladium, have lower IC50 values than do the ionic complexes 3 and 4, where chloride is an outer sphere ligand [32]. The higher cytotoxicities of complexes 1 and 2 may be the result of the ease of formation of covalent drug-DNA adducts. Fig. 2 Building on this notion, other neutral palladium(II) dithiocarbamates with two chiral aminoalcohols
4-(1-hydroxy-2-(methylamino)ethyl)phenol
(synephryne)
5
and
3-(2-amino-1-
hydroxyethyl)phenol (norphenylephrine) 6 (Fig. 3) have been prepared and subsequently investigated for their cytotoxicity against a panel of human cancer cell lines, including HeLa, HL60, sensitive 2008 and resistant C13* utilizing the MTT (tetrazolium salt reduction) test [33]. The cytotoxicities of complexes 5 and 6 proved to be concentration dependent against the HL60 and HeLa cell lines. An anticancer screening study of complexes 5 and 6 against the cisplatin5
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sensitive 2008 human cancer cell line and cisplatin-resistant variant C13* cells showed that they were only effective against the latter cell line [33]. An examination of the effect on DNA synthesis in Ehrlich cells (short-term effect) and on clonal growth capacity of 2008 cells (longterm effect) indicated that complexes 5 and 6 have significant antiproliferative activity similar to that of the reference drug for which the corresponding platinum complexes are completely inactive [33]. Fig. 3 Inspired by the highly interesting cytotoxic properties of the 2-picoline-based platinum(II) complex 7 (Fig 4), which were subjected to phase-I clinical trials [34-38], the heteroleptic palladium(II) complexes 8 and 9, which contain dithiocarbamate and picoline ligands (Fig. 4), were evaluated for their cytotoxicity, DNA synthesis inhibition, interstrand, DNA-protein crosslinking, and micronuclei (MN) detection properties [39]. These complexes were less active than is cisplatin against human squamous cervical adenocarcinoma (HeLa) and leukemic promyelocyte (HL60) cell. Furthermore, the 2-picoline complex 8 is more active than is the 3picoline complex 9. The high activity of the former complex may be linked with the more stable 2-picoline-palladium adduct formation due to the presence of electron releasing methyl closer to the donor site (Table 1). The efficacy of complex 8 against a panel of six human tumor cell lines, Fig. 4 and Table 1 including the sensitive (2008, A431, U2OS) and resistant(C13*, A431/pt, U2OS/pt) cell lines with reference to cisplatin was also studied utilizing the MTT assay approach [40]. The inhibitory effect of this complex on cell proliferation was concentration dependent for both the sensitive and resistant cell lines. Complex 8 proved to be slightly less active against the sensitive 2008 cells and A431 cell line but, interestingly, it was more active against the resistant C13* 6
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cells and A431/pt cell line (Table 2). This complex was also more active than was the reference cisplatin against U2OS cells at lower concentrations, but less active at higher concentrations. The resistant factors (RF) obtained for complex 8 and cisplatin showed that the former was able to overcome cisplatin resistance. In addition, HL60 cells were incubated with various concentrations of complex 8 and the mixtures were investigated with respect to DNA synthesis inhibition in terms of radioactive 3H-thymidine incorporation. This complex inhibits DNA synthesis marginally less than does cisplatin, but the analogous platinum complex has significant inhibition of DNA synthesis by comparison to that of cisplatin, even at low concentrations. Interstrand DNA cross-links (ISC) are unique, but pivotal for cytotoxicity, as they are among the critical molecular lesions that are able to inactivate DNA as a template for replication. The detection of interstrand crosslinks in 2008 cells was performed by incubating DNA with complex 8 for 6 to 12 h and subsequently exposing the cells to γ-rays, followed by alkaline solution elution [41]. The DNA ISC index was calculated by the formula (1-r0/1-r)1/2-1, where r and r0 are the fractions of 14C-thymidine-labelled DNA for the treated cells vs. the control cells remaining on the filter, respectively [41]. The DNA ISC index was greater for complex 8 than for cisplatin after 12 h exposure. The DNA-protein cross-links (DPC) were also measured as the number of DPC induced per million of nucleotides in 2008 cells. It was observed that cisplatin caused a Table 2 significant retention of DNA on the filter indicating the formation of a remarkable amount of DPC, while complex 8 proved to be totally unable to induce DPC [39]. These observations are interesting as DPCs are supposed to induce clastogenic effects in mammalian cells [42]. The genotoxic potential of complex 8 and cisplatin via micronuclei (MN) detection was evaluated on human lymphocytes. Complex 8 has a lower potential for genotoxicity than does
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cisplatin, as indicated by the lower levels of MN induction. Moreover, the relationship between MN induction and cytotoxicity is linear for complex 8 and cisplatin, but the fitting lines show different slopes. Different amounts of micronucleated cells were induced compared to cisplatin at the same survival rate [39]. It means the complex (8) tempt anticancer activity with reduced chromosomal damage. The homoleptic palladium(II) dithiocarbamates 10-12 have been evaluated for cytotoxicity against a panel of seven human tumor cell lines (Fig. 5) using the microculture sulforhodamine B (SRB) test [43] with doxorubicin, cisplatin, 5-fluorouracil, methotrexate, etoposide, and taxol as reference drugs. Complexes 10-12 have very low cytotoxic activities compared to those for the reference drugs and the heteroleptic complexes (which possess one dithiocarbamate moiety) [43]. The low cytotoxicity of complexes 10-12 may be due to the absence of a metal-attached, labile chloride ligand, which ultimately limits the interaction of the drug with DNA or other cellular components. Fig. 5 In an intriguing study, complexes with a dithiocarbamate moiety (13 and 14) and an amino acetate moiety (15 and 16) were synthesized with the aim of exploring the significance of the dithiocarbamate moiety in anticancer action (Fig. 6) [44]. An examination of the growth inhibitory effect of complexes 13-16 against human chronic myelogenous leukemia cell lines proved that the dithiocarbamate-based palladium(II) complexes (13 and 14) were more active than were the related complexes containing O-and N-donor ligands (complexes 15 and 16) [44]. The presence of the dithiocarbamate moiety in the structure of the Pd(II) complexes has a marked effect upon the growth suppression activity of K562 cells. Fluorescence spectroscopy is a useful technique to study the structure, dynamics and binding properties of protein molecules in solution. The effect of the Pd(II) complexes 13-16 on the 8
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structure of the two variants of carrier protein, β-lactoglobulins (BLG-A and -B), were investigated using intrinsic fluorescence measurements. The intrinsic fluorescence emissions of the proteins in the presence of 20 µM of complexes 13 and 14 with dithiocarbamate were nearly two times less than were the emissions of the proteins under the influence of 20 µM concentrations of complexes 15 and 16. This means that dithiocarbamate-based complexes, 13 and 14, have less effect on the tertiary structure of proteins at lower concentration than do the two other complexes (15 and 16) [44]. Circular dichroism (CD) spectroscopy revealed that the content of regular secondary structures of BLG-A and -B upon interaction with complexes 13-16 did not show any significant changes. Fig. 6 The lack of water solubility of some of the palladium(II) antitumor complexes explains their limited bioavailability and inferior in vivo activity [45]. The water-soluble complex 13 (Fig. 6) proved to be more active than is cisplatin against the human cancer cell line K562 (Table 3) [46]. Interestingly, changes in concentration had a little impact upon the activity. The watersoluble complex 17 (Fig. 7), with a short hydrocarbon chain ligand (ethylcarbamodithioate), was more active than is cisplatin against the human cancer cell line K562 and was observed to produce a dose-response suppression on the growth of the K562 leukemia cell line [47]. The anticancer activity jumped up markedly upon increasing the chain length associated with the N Fig. 7. attached alkyl group of dithiocarbamate ligand. Complex 18 with an octyl group (Fig. 8) was two and nearly eight times more active than were the analogous complexes with butyl (13) and ethyl(17) substituents, respectively. Complex 18 was 22 times more active than is cisplatin against the human cancer cell line K562 (Table 3) [22]. To explore the role of the auxiliary 9
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ligands on the anticancer properties, the 2,2ʹ-bipyridine ligand in complexes 13, 17 and 18 was replaced by 1,10-phenanthroline affording complexes 19, 20 and 21 (Fig. 9). The activity against the human cancer cell line K562 was increased for complexes 19 and 20 compared to the parent complexes 13 and 17 presumably due to the more planar nature of the 1,10-phenanthroline ligand [24-26]. The activity for complex 18 was greater than for the complex 21, showing the importance of both dithiocarbamate and auxiliary ligands. Figs. 8 and 9 Other
water-soluble
complexes
containing
morpholine-
(22)
and
piperidine-based
dithiocarbamate (23) ligands (Fig. 10) have been shown to inhibit the in vitro human cancer cell line K562 growth more efficiently than does cisplatin [27, 28](Table 3). The cytotoxicity of dithiocarbamate complex containing piperidine skeleton (23) was found greater than morpholine one ( 22). Complexes 22 and 23 are less active than are the analogous platinum(II) complexes at lower concentrations, but are more active at higher concentrations (45 µM). Table 3 Changing from an ethyl-based amine to morpholine or piperidine and a butyl-based amine to piperidine in the structure of the 2,2ʹ-pyridine dithiocarbamatopalladium(II) complexes results in increased cytotoxicity [27, 28, 47]. Complexes 18 and 23 had nearly equal cytotoxicities (Table 3). Fig. 10 Other
water
soluble
palladium(II)
complexes
containing
same
cationic
spheres
(pyrrolidinedithiocarbamate and 1, 10-phenanthroline) but different anions {24 (nitrate) and 25 (bromide)} exhibited more cytotoxic effect than cisplatin against chronic myelocytic leukemia
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K562 cell line (Fig. 11, table 3), however, the comparable activity of both complexes excluded the significant role of anion in anticancer activity [48,49].
Fig. 11
Dinuclear water soluble palladium(II) complexes 26-28 (Fig. 12) were also inspected for cytotoxicity against human gastric carcinoma (AGS), esophageal squamous cell carcinoma (Kyse-30), and hepatocellular carcinoma (HepG2) cell lines by MTT assay [50]. These complexes were examined to have greater cytotoxicity than cisplatin as shown from their IC50 values against these three cell lines (table 4). The results indicated that shorter alkyl chain length between two palladium nuclei enhances cytotoxicity in this class of compounds. Fig. 12 and Table 4
Mixed-ligand dithiocarbamate palladium(II) complexes having different amines or diamines in their structures have good cytotoxic activities. In order to investigate the effect of tertiary phosphine ligands, the mixed-ligand dithiocarbamate palladium(II) complexes (29-34) were synthesized (Fig. 13) and tested for their antitumor activity against cisplatin-resistant DU145 human prostate carcinoma (HTB-81) cells [51, 52]. It was observed that the complexes have a greater antitumor activity at lower concentrations (Table 5), nevertheless, except 33 the activity of 29-34 was found concentration dependent. Complexes 33 and 34 were the most active with IC50 values of 1.33 and 1.55 µM respectively [51]. The highest IC50 value was observed for complex 29 (6.94 µM). The high cytotoxicity of complex 33 can be attributed to the electron-
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donating tricyclohexylphosphine ligand resulting in a stronger Pd-P bond and so there is better possibility of reaching intact complex to the target DNA [51]. In comparison, complex 29 has a slightly less electron-donating triphenylphosphine ligand resulting in a marginally weaker Pd-P bond and the complex presumably has a greater tendency to dissociate. The movement of this dissociated complex towards DNA may be interrupted by other groups in the cell, such as glutathione, cysteines and methionines [53]. The high activity of complex 34 can be attributed to the presence of the 2-pyridyl moiety in the organophosphine ligand, which has the potential to form a hydrogen bond with DNA bases [51]. Fig. 13 and Table 5 Khan et al. [54] have reported six heteroleptic Pd(II) dithiocarbamate complexes 35-40 (Fig. 14) and studied their in vitro antitumor activity against cisplatin-resistant DU145 human prostate carcinoma (HTB-81) cells [54]. These complexes have good antitumor activity at low concentrations (Table 5) with complex 40 being the most active and complex 39 the least active. The IC50 values determined for the six complexes are lower than are the literature IC50 values for the standard drug cisplatin [54, 55]. The higher antitumor activity of complex 40 has been attributed to the presence of oxygenated species in the dithiocarbamate ligand, which have a higher potential to form hydrogen bonds with the DNA bases. The weak donating ability of phosphorous due to the presence of the electron-withdrawing chloride and phenyl groups in the organophosphine are presumably responsible for the lower cytotoxicity of complex 39. The second lowest activity was observed for complex 37 and this was attributed to the presence of the bulky t-butyl group, which hinders movement of the complex towards the target DNA. Fig. 14
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Two other piperazine based heteroleptic Pd(II) dithiocarbamates 41-42 have been recently synthesized and evaluated for their anticancer activity against Hela cell line at different concentrations [56]. The IC50 values of the complexes show that these complexes have greater cytotoxicity as compared to cisplatin against Hela cell line (Table 5). The results of DNA fragmentation in vitro by using DNA ladder assay show that the cytotoxicity of these complexes might be due to strong interaction with DNA leading to DNA damage and apoptosis [56]. Two
new
potent
anticancer
heteroleptic
palladium(II)
complexes
of
4-(2-
methoxyphenyl)piperazine-1-carbodithioate ligand with two different organophosphines namely diphenyl-p-tolylphosphine (43) and tri-p-tolylphosphine (44) were screened against five different cell lines [57]. For both complexes the activity varied in the sequence MCF7 ˃ Hepa-c1c7 ˃ PC3 ˃ LU ˃ MDA-MB-231. Furthermore, the more active nature of 44 than 43 was attributed to its extra stablility as was confirmed by the X-ray single crystal analysis (shorter Pd-S bond distances) and DFT calculation.
Fig. 15 The complexes (45-53) having expected structures (Fig. 16) have also been tested for cytotoxic activity against human cancer cell lines (2008 & A431) by MTT assay [58]. The results indicated that neutral binuclear complex (47) and ionic polynuclear complex (49) have cytotoxicity close to cisplatin (Table 6). Neutral binuclear complexes (45, 48) were less active. While neutral binuclear complex (46) and other ionic complexes (50-53) were proved to be ineffective against 2008 and A431 cancer cell lines. In order to assess cross-resistance with cisplatin the most active complexes (47, 49) were also examined against cisplatin-resistant cell line C13*. The RF values
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calculated for the complexes (47, 49) were about eight times lower than that of cisplatin declaring different cross-resistance profile from that of cisplatin [58]. Fig. 16 and Table 6
3. DNA-binding and DNA-denaturing studies The anticancer properties of metallodrugs are due to the binding and denaturing of DNA, which result in interference with DNA replication, transcription and other nuclear functions. Some metallodrugs having replaceable chloride such as cisplatin, bind covalently to the nitrogen basis of DNA. The complexes 1, 2, 5-9, 29-44 also have replaceable chloride and chances to have covalent type interaction with DNA. Some metallodrugs interact with DNA non-covalently by the formation of hydrogen bonds, electrostatic and/or hydrophobic interactions in the minor groove regions of DNA. Some metallodrugs interact with DNA non-covalently by insertion of planar moiety (planer heterocyclic groups, aromatic ring or fused aromatic rings) between adjacent base pairs of the double helix. This is interclative mode of interaction. The complexes 13-23 have interclative mode of interaction with DNA as described in the respective literature [22, 24-28, 46, 47]. The DNA-binding and DNA-denaturing behavior of complexes 13-23 has also been investigated using standard biochemical methods. In a procedure using UV-visible spectroscopy, the absorbance of the fixed concentration of the compound (drug), “A1”, is first measured. Then, the absorbance of mixed solutions of the compound with increasing concentrations of DNA, “A2”, are measured while keeping the volume and concentration of the compound constant. The change in absorbance, “ΔA” is calculated from ΔA = A1−A2. The
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binding constant “K” can be calculated from the slope-to-intercept ratio of the plot of A1/ΔA versus 1/[DNA] according to the Benesi–Hildebrand equation [57]. The results indicated that the complexes containing the more planar 1,10-phenanthroline ligand and the butyl or octyl chain in the dithiocarbamates have greater binding constants (Table 7). The 1/ΔA ratio is plotted versus 1/[DNA] and the value for the change in absorbance when all the binding sites on DNA are occupied by the compound “ΔAmax” is determined from the intercept on the ordinate of the linear plot [47]. Next, the absorbance of a fixed concentration of DNA “A3” is measured. The absorbance of mixed solutions of DNA with increasing concentrations of the compound “A4” is measured while keeping the volume and concentration of DNA constant [47]. Similarly, the changes in absorbance “ΔΆ” are calculated from ΔΆ = A3 −A4. The concentration of the compound bound to DNA, [L]b, is determined from the following equation [47] [L]b = ΔΆ[L]t/ΔAmax, where [L]t is the total concentration of the compound added. The concentration of free compound [L]f is obtained from the following equation [47][L]f = [L]t−[L]b. The ratio of the concentration of bound compound to total DNA concentration “ῡ” is calculated from ῡ = [L]b/ [DNA]t. Next, the Scatchard plots for these complexes are obtained by plotting ῡ/[L]f versus ῡ [23]; these proved to be curvilinear concave downwards suggesting cooperative binding. The Kolts plot is produced by plotting ῡ versus Ln[L]f [47], while the Hill plot is obtained by plotting Ln ῡ/1−ῡ versus Ln[L]f [47]. The value of binding parameter “n” the socalled Hill coefficient (which is taken as a criterion of cooperativity) can be measured from the slope of the straight line in the plot. The value of Hill coefficient, “n”, can also be measured from the slope of the Kolts plot as n = 4 × (value of slope of Kolts plot). The values of Hill coefficients indicated that these complexes have cooperative binding (Table 7), while complexes 17, 18, 21, 22 and 23 have greater cooperativity than do the other complexes. The number of 15
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binding sites per 1000 nucleotides “g” can be calculated by substituting the values of ῡ, K, n and [L]f in the Hill equation [47] ῡ = g(K[L]f)n/(1 + (K[L]f)n). The results indicated that the complex 17 has a greater number of binding sites per 1000 nucleotides (Table 7). The molar Gibbs free energy of binding “ΔG˚b” for the compound-DNA interaction at a particular temperature can be calculated from the following equation [47]ΔG˚b = −RTLnK, where R is the gas constant. Then, the molar enthalpy of binding “ΔH˚b” for the compound-DNA interactions at two different temperatures can be calculated from the following equation [47] LnKT1 /KT2 = −ΔH˚b/R(1/T2−1/T1). Finally, the molar entropy of binding “ΔS˚b” for the compound-DNA interaction at a particular temperature can be evaluated from the following equation [47] ΔG˚b =ΔH˚b−TΔS˚b. In order to determine the DNA-denaturing parameters, the UV absorption spectrum of a DNA solution at 258 nm is recorded keeping the absorbance reading below one. Then, a fixed amount of the compound is added stepwise and at each step an absorbance reading is taken. This procedure is performed until there is no further change in absorbance. The changes in the absorbance of DNA solutions are plotted against different concentrations of the compound. From this plot, the concentration of the compound at the midpoint of transition “[L]1/2” can be determined [47]. The concentrations of the compounds at the midpoint of transition calculated for these complexes are given in the Table 8. Using this DNA denaturation plot and the Pace method, the unfolding equilibrium constant K*, can be calculated from the following equation [46, 47] K* = AN−Aobs/Aobs−AD, where AN is the absorbance of natural conformation of DNA, AD is the absorbance of denatured conformation of DNA and Aobs is the absorbance in the transition region. Then, the unfolding free energy of DNA “ΔG˚” can be calculated from the
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following equation [46, 47]ΔG˚ = −RT LnK*. When these “ΔG˚” values are plotted versus the concentrations of the compound in the transition region, a straight line is obtained. The equation for this straight line can be written as [46]ΔG˚ = ΔG˚( ) − m[compound]. From the slope of straight line, the measure of DNA denaturing ability “m” can be determined. The results indicated that the complex 20 has the highest DNA denaturing ability (Table 8). From the intercept of the straight line, the conformational stability of the free DNA “ΔG˚( ) ” is determined [47]. The molar enthalpy of DNA denaturation by the compound “ΔH˚denaturation” in the range of the two temperatures (T1and T2) can then be calculated using the following Gibbs Helmholtz equation [47]ΔH˚denaturation = ΔG˚(T1)/T1−ΔG˚(T2)/T2÷ 1/T1− 1/T2. The molar enthalpy of DNA denaturation in the absence of the compound “ΔH˚( ) ” is determined by interpolation of a plot of ΔH˚denaturation against the concentration of the compound [47]. Finally, the molar entropy of DNA denaturation in the absence of the compound “ΔS˚( ) ” is obtained from the following equation ΔG˚( ) = ΔH˚( ) −TΔS˚( ). The DNA-binding and DNAdenaturing parameters thus obtained are shown in Tables 7 and 8. The results indicate that the complexes containing long-chain dithiocarbamate ligands and more planar auxiliary ligands have greater DNA-binding and DNA-denaturing abilities. These results are in agreement with the cytotoxicity results Tables 7 and 8 4. Conclusions In this review, the anticancer potential of the palladium(II) dithiocarbamates has been examined in terms of their cytotoxic activities against different cancer cell lines, and by DNA synthesis
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inhibition, detection of interstrand cross-links, DNA–protein cross-links, micronuclei detection, DNA-binding and denaturing studies. Most of these complexes (1, 13, 14, 17-44), including the water-soluble ones, have greater cytotoxic activities than does cisplatin. Moreover, they have less cross-resistance towards cisplatin-resistant human tumor cell lines. Heteroleptic palladium(II) dithiocarbamates containing organophosphines as ancillary ligands demonstrated greater cytotoxic effect than the complexes containing N-donor ligands. The mechanism of action seems to be similar to that of cisplatin itself because they strongly and irreversibly bind to DNA, and can denature DNA, even at low concentrations. Contrary to this, the complexes containing ligands with two nitrogen donor sites (bpy and phen) adopt intercalative mode of interaction with DNA. The DNA binding and, as a consequence the anticancer action of these complexes depends on the lipophilicity (carbon chain length), planarity and number of aromatic rings in the auxiliary ligands, and the presence of more labile chloride group. It will be worthwhile entering several of the water-soluble palladium(II) dithiocarbamates into clinical trials in the future. Acknowledgement We thank the Higher Education Commission of Pakistan for financial support. References [1] B. Rosenberg, L. Van Camp, T. Krigas, Nature 205 (1965) 698. [2] T. Boulikas, A. Pantos, E. Bellis, P. Christofis, Cancer Ther. 5 (2007) 537. [3] A. Eastman, Cancer cells (Cold Spring Harbor, NY) 2 (1989) 275. [4] K. WoŸniak, J. Błasiak, Acta Biochim. Pol. 49 (2002) 583.
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[5] N.P. Johnson, J.L. Butour, G. Villani, F.L. Wimmer, M. Defais, V. Pierson, V. Brabec, Metal antitumor compounds: the mechanism of action of platinum complexes. in: Ruthenium and Other Non-Platinum Metal Complexes in Cancer Chemotherapy. Springer, 1 (1989). [6] (a) P. J. O'Dwyer, J. P. Stevenson, S. W. Johnson, Cisplatin: chemistry and biochemistry of a leading anticancer drug, Wiley-VCH, (1999) 29. ISBN 3-906390-20-9; (b) B. Lippert (Ed.), Chemistry and Biochemistry of a Leading Anticancer Drug, Wiley-VCH, New York (1999); (c) T. Glaser, G.F. Von Mollard, D. Anselmetti, Inorg. Chim. Acta xxx (2016) xxx–xxx. (http://dx.doi.org/10.1016/j.ica.2016.02.013) [7] H.M. Pinedo, H.H. Schornagel, J.H. Schornagel, Platinum and Other Metal Coordination Compounds in Cancer Chemotherapy 2. Springer, 1996. [8] P.J. Morin, Drug Resist. Update. 6 (2003) 169. [9] Z.H. Siddik, Oncogene 22 (2003) 7265. [10] D. Fregona, L. Giovagnini, L. Ronconi, C. Marzano, A. Trevisan, S. Sitran, B. Biondi, F. Bordin, J. Inorg. Biochem. 93 (2003) 181. [11] K.S. Siddiqi, S.A. Nami, Y. Chebude, J. Braz. Chem. Soc. 17 (2006) 107. [12] A. Rai, S.K. Sengupta, O.P. Pandey, Spectrochim. Acta. Part A 64 (2006) 789. [13] M.S. Murthy, L.N. Rao, J.D. Khandekar, E.F. Scanlon, Cancer Res. 47 (1987) 774. [14] R. Mital, N. Jain, T. Srivastava, Inorg. Chim. Acta 166 (1989) 135. [15] H. Sigel, Metal Ions in Biological Systems: Volume 42: Metal Complexes in Tumor Diagnosis and as Anticancer Agents, CRC Press, (2004). [16] W. Micklitz, W.S. Sheldrick, B. Lippert, Inorg. Chem. 29 (1990) 211. [17] K.J. Barnham, C.J. Bauer, M.I. Djuran, M.A. Mazid, T. Rau, P.J. Sadler, Inorg. Chem. 34 (1995) 2826.
19
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[18] W.-Z. Shen, D. Gupta, B. Lippert, Inorg. Chem. 44 (2005) 8249. [19] M. Fanelli, M. Formica, V. Fusi, L. Giorgi, M. Micheloni, P. Paoli, Coord. Chem. Rev. 310 (2016) 41. [20] L. Khassanova, P. Collery, Z. Chassanova, I. Maymard, J.-C. Étienne, Metal Ions in Biology and Medicine, John Libbey Eurotext, (1990). [21] A. Furlani, V. Scarcia, G. Faraglia, L. Sindellari, L. Trincia, M. Nicolini, Eur. J. Med. Chem. 21 (1986) 261. [22] H. Mansouri-Torshizi, M. Saeidifar, F. Khosravi, A. Divsalar, A.A. Saboury, F. Hassani, Bioinorg. Chem. Appl. (2011), article ID 394506. [23] (a) A. Saboury, J. Iran. Chem. Soc. 3 (2006) 1; (b) G. Scatchard, Ann. N.Y. Acad. Sci. 51 (1949) 660. [24] H. Mansouri-Torshizi, M. Saeidifar, A. Divsalar, A. Saboury, S. Shahraki, Bull. Korean Chem. Soc. 31 (2010) 435. [25] H. Mansouri-Torshizi, M. Saeidifar, A. Divsalar, A.A. Saboury, Spectrochim. Acta Part A 77 (2010) 312. [26] H. Mansouri-Torshizi, M. Saeidifar, A. Divsalar, A. Saboury, Nucleosides, Nucleotides Nucleic Acids 30 (2011) 405. [27] H. Mansouri-Torshizi, M. Moghaddam, A. Divsalar, A. Saboury, J. Biomol. Struct. Dyn. 26 (2009) 575. [28] H. Mansouri-Torshizi, M. Eslami-Moghadam, A. Divsalar, A.-A. Saboury, Acta Chim Slov. 58 (2011) 811. [29] Q. Zhang, W. Zhong, B. Xing, W. Tang, Y. Chen, J. Inorg. Biochem. 72 (1998) 195. [30] M.-T. Hassan, I.-M. Mahbobe, S.A. Akbar, Acta Bioch. Bioph. Sin. 35 (2003) 886.
20
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[31] G. Zhao, H. Sun, H. Lin, S. Zhu, X. Su, Y. Chen, J. Inorg. Biochem. 72 (1998) 173. [32] (a) G. Faraglia, D. Fregona, S. Sitran, L. Giovagnini, C. Marzano, F. Baccichetti, U. Casellato, R. Graziani, J. Inorg. Biochem. 83 (2001) 31; (b) W.J. Durkin, V.K. Ghanta, C.M. Balch, D.W. Davis, R.N. Hiramoto, Cancer Res. 39 (1979) 402. [33] (a) V. Alverdi, L. Giovagnini, C. Marzano, R. Seraglia, F. Bettio, S. Sitran, R. Graziani, D. Fregona, J. Inorg. Biochem. 98 (2004) 1117; (b) M.C. Alley, D.A. Scudiero, A. Monks, M.L. Hursey, M.J. Czerwinski, D.L. Fine, B.J. Abbott, J.G. Mayo, R.H. Shoemaker, M.R. Boyd, Cancer Res. 48 (1988) 589; (c) D.T. Vistica, P. Skehan, D. Scudiero, A. Monks, A. Pittman, M.R. Boyd, Cancer Res. 51 (1991) 2515. [34] L.R. Kelland, S.Y. Sharp, C.F. O’Neill, F.I. Raynaud, P.J. Beale, I.R. Judson, J. Inorg. Biochem. 77 (1999) 111. [35] Y. Chen, Z. Guo, S. Parsons, P.J. Sadler, Chem. Eur. J. 4 (1998) 672. [36] J. Parkinson, P. Sadler, J. Chem. Soc. Dalton Trans. 21 (1998) 3577. [37] J. Holford, S. Sharp, B. Murrer, M. Abrams, L. Kelland, Br. J. Cancer 77 (1998) 366. [38] F.I. Raynaud, F.E. Boxall, P.M. Goddard, M. Valenti, M. Jones, B.A. Murrer, M. Abrams, L.R. Kelland, Clin. Cancer Res. 3 (1997) 2063. [39] L. Giovagnini, C. Marzano, F. Bettio, D. Fregona, J. Inorg. Biochem. 99 (2005) 2139. [40] M.C. Alley, D.A. Scudiero, A. Monks, M.L. Hursey, M.J. Czerwinski, D.L. Fine, B.J. Abbott, J.G. Mayo, R.H. Shoemaker, M.R. Boyd, Cancer Res. 48 (1988) 589. [41] K.W. Kohn, Pharmacol. Therapeut. 49 (1991) 55.
21
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22
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[51] H. Khan, A. Badshah, G. Murtaz, M. Said, C. Neuhausen, M. Todorova, B.J. Jean-Claude, I.S. Butler, Eur. J. Med. Chem. 46 (2011) 4071. [52] R. Cattaneo-Pangrazzi, H. Schott, R. Schwendener, Prostate 45 (2000) 8. [53] Y. Kasherman, S. Sturup, D. Gibson, J. Med. Chem. 52 (2009) 4319. [54] H. Khan, A. Badshah, M. Said, G. Murtaza, J. Ahmad, B.J. Jean‐Claude, M. Todorova, I.S. Butler, Appl. Organomet. Chem. 27 (2013) 387. [55] A. Amantana, C.A. London, P.L. Iversen, G.R. Devi, Mol. Cancer Ther. 3 (2004) 699. [56] S. Khan, M.K. Amir, M.M. Naseer, R. Abbasi, K. Mazhar, M.N. Tahir, I. Z. Awan, Zia-urRehman, J. Coord. Chem. 68 (2015) 2539. [57] S. Z. Khan, M. K. Amir, I. Ullah, A. Aamir, J. M. Pezzuto, T. Kondratyuk, F. BélangerGariepy, A. Ali, Zia-ur-Rehman, App. Organomet. Chem. 30 (2016) 392. [58] D. Montagner, C. Marzano, V. Gandin, Inorg. Chim. Acta 376 (2011) 574.
23
Revised
Fig. 1:Structures of currently approved platinum-based anticancer drugs.
Fig. 2: Mixed ligand palladium(II) 2-ethoxy-2-oxoethyl(methyl)carbamodithioates (1-4).
Fig. 3: Structure of complexes 5 and 6. 25
Revised
Fig. 4: The heteroleptic platinum and palladium(II) complexes of 2-ethoxy-2oxoethyl(methyl)carbamo- dithioateandpicoline.
Fig. 5:The homoleptic palladium(II) complexes (10-12) with dithiocarbamate ligands.
Fig. 6: Water soluble complexes with and without dithiocarbamate moiety.
26
Revised
Fig. 7: The water-soluble complex containing ethyldithiocarbamate ligand.
Fig. 8: The water-soluble complex containing octyldithiocarbamate ligand.
27
Revised
Fig. 9: The palladium complexes obtained by replacing 2,2-bipyridine with 1,10-phenanthroline from the ethyl, butyl and octyldithiocarbamate complexes.
Fig. 10:The water-soluble complexes with morpholinedithiocarbamate and piperidinedithiocarbamate ligands.
28
Revised
Fig. 11: The water-soluble complexes containing nitrate and bromide ion in the outer sphere.
Fig. 12: Dinuclear water-soluble complexes.
29
Revised
Fig. 13: The mixed-ligand palladium dithiocarbamate complexes containing organophosphine ligands.
Fig. 14:The mixed-ligand dithiocarbamate palladium complexes containing phosphine ligands.
30
Revised
Fig. 15:The piperazine based mixed-ligand dithiocarbamate palladium complexes containing phosphine ligands.
Fig. 16: Expected structures for dithiocarbamate palladium(II) complexes (45-53).
31
Revised
Table 1: In vitro cytotoxic activity results (IC50, µM) of palladium(II) dithiocarbamates (complexes 1-4, 8 and 9) against different cell lines. Complex
IC50( µM) against cancer cell lines
Refs.
HL-60
HeLa
1
3.11
5.61
[32]
2
5.83
7.50
[32]
3
27.44
77.0
[32]
4
24.27
59.99
[32]
8
59.62
69.54
[39]
9
˃100
˃100
[39]
cisplatin
3.40
6.33
[39]
Table 2: Cross-resistance profiles of the investigated complex (8) toward some established cisplatin-sensitive and resistant human tumor cell lines. Complex
IC50 (µM)
R.F.
Refs.
2008 cells
C13* cells
8
60.0
128.4
2.14
[39]
Cisplatin
42
616.1
14.68
[39]
A431 cells
A431/Pt cells
8
81.2
161.5
1.99
[39]
Cisplatin
76.4
297.1
3.89
[39]
U2OS cells
U2OS/Pt cells
8
54.3
108.3
2.02
[39]
Cisplatin
30.2
191.2
6.33
[39]
R.F. = Resistance factor = IC50 resistant cell line/ IC50 sensitive cell line.
32
Revised
Table 3:The 50% cytotoxic concentrations of the palladium (II) dithiocarbamate complexes. Complex
Cc50/IC50 (µM)
Refs
K562 13
14
[44]
14
36
[44]
15
251
[44]
16
320
[44]
13
18
[46]
17
55
[47]
18
7
[22]
19
10
[24]
20
10
[25]
21
15.6
[26]
22
26
[27]
23
10.5
[28]
24
46
[48]
25
45
[49]
Cisplatin
154
[46]
Table 4: IC50 (µg) values of the dinuclear palladium (II) complexes against AGS, Kyse-30 and HepG2 cell lines Complex 26 27 28 Cisplatin
AGS 0.68 0.78 1.20 4.08
IC50 (µg) Kyse-30 0.88 0.98 1.06 1.90
Refs HepG2 1.02 1.17 1.19 2.08
[50] [50] [50] [50]
33
Revised
Table 5: The 50% inhibitory concentration (IC50) of the mixed ligand palladium(II) dithiocarbamates. Complex
IC50 (µM) HTB-81 cells
Refs. Hela cells
29
6.94
[51]
30
-
[51]
31
5.71
[51]
32
3.10
[51]
33
1.33
[51]
34
1.55
[51]
35
3.67
[54]
36
-
[54]
37
9.52
[54]
38
4.57
[54]
39
21.7
[54]
40
2.12
[54]
41
22
[56]
42
26
[56]
Cisplatin
78
[56]
34
Revised
Table 6: Cytotoxic activity (IC50, µM) of palladium dithiocarbamate complexes.
Complex
Formula
2008
A431
C13*
RF
Refences
45
[Pd2Cl2(PyDT)2(dah)]
52.9
60.1
-
-
[58]
46
[Pd2Cl2(PyDT)2(dab)]
˃100
˃100
-
-
[58]
47
[Pd2Cl2(ESDT)2(dah)]
16.4
14.3
14.5
0.9
[58]
48
[Pd2Cl2(ESDT)2(dab)]
42.5
36.5
-
49
[Pd(ESDT)(dab)]nCln
18.4
21.5
17.6
0.9
[58]
50
[Pd(PyDT)(en)]Cl
˃100
˃100
-
-
[58]
51
[Pd(PyDT)(dap)]Cl
˃100
˃100
-
-
[58]
52
[Pd(PyDT)(en)][PdCl2(PyDT)]
˃100
˃100
-
-
[58]
53
Bu4N[PdCl2(DMDT)]
˃100
˃100
-
-
[58]
Cisplatin
Pt (NH3)2Cl2
12.2
19.5
95.4
7.8
[58]
[58]
PyDT = pyrrolidine dithiocarbamate, ESDT = sarcosine ethyl ester dithiocarbamate, DMDT = dimethyl dithiocarbamate, en = ethylenediamine, dab = 1, 4-diaminobutane, dah = 1, 7-diaminoheptane, dap = 1, 3diaminopropane
35
Revised
Table 7: Values of ∆Amax and binding parameters in the Hill equation for interaction between Pd(II) complexes and DNA in 10 mmol/L Tris–HCl buffer and pH 7.0.
Complex
Temp(℃)
∆Amaxa
gb
Kc×104 M-1
nd
Errore
Refs.
13
27
0.047
8
0.317
1.48
0.005
[46]
37
0.033
8
0.401
1.79
0.008
[46]
27
0.014
9
0.180
4.77
0.03
[47]
37
0.011
9
0.270
4.79
0.06
[47]
27
0.035
6
5.28
5.92
0.13
[22]
37
0.058
5.86
6.18
0.14
[22]
27
0.246
5
6.693
1.972
0.0007
[24]
37
0.167
5
4.360
1.604
0.0004
[24]
27
0.213
6
0.1902
1.432
0.00004
[25]
37
0.101
6
0.1772
1.239
0.0006
[25]
27
0.038
3
8.13
4.95
0.0043
[26]
37
0.040
3
11.78
3.079
0.0041
[26]
27
0.07
6
0.018
3.5
0.06
[27]
37
0.08
6
0.024
4.74
0.09
[27]
27
0.098
6
0.020
3.91
0.039
[28]
37
0.048
6
0.033
3.8
0.063
[28]
17
18
19
20
21
22
23
a
Change in the absorbance when all the binding sites on DNA were occupied by a metal complex. The number of binding sites per 1000 nucleotides. c The apparent binding constant. d The Hill coefficient (as a criterion of cooperativity). e Maximum error between theoretical and experimental values of ῡ. b
36
Revised
Table 8: Thermodynamic parameters of DNA denaturation by palladium complexes.
Complex
Temp (℃)
M
∆G˚(H2O)
∆H˚(H2O)
∆S˚(H2O)
[L]1/2
(kJmol-1K)
(mM/L)
0.073
0.093
[46]
0.085
[46]
0.121
[47]
0.105
[47]
0.093
[22]
0.086
[22]
0.055
0.0134
[24]
0.030
0.0127
[24]
0.065
0.0101
[25]
0.065
0.0099
[25]
~0
0.012
[26]
~0
0.011
[26]
0.08
0.15
[27]
0.14
[27]
~0
0.128
[28]
~0
0.125
[28]
Refs.
-1
13
17
18
19
20
21
22
23
(kJmol-1)(mM)-1
(kJmol-1)
27
72.12
7.47
37
89.9
6.74
27
159.03
18.81
37
158.4
17.81
27
195
13.44
37
218
12.93
27
589.6
9.846
37
710.1
19.610
27
1119.6
11.54
37
1019.2
10.88
27
544.05
7.877
37
515.77
6.845
27
159.4
23.7
37
115.9
15.7
27
80.1
20.33
37
86.6
19.00
(kJmol )
29.36
48.90
28.09
37.58
31.14
5.154
262.8
20.5
0.10
0.05
37
Revised
Graphical Abstract (synopsis) This review provides a summary of the anticancer potential, DNA-binding and DNA-denaturing aptitude of palladium(II) dithiocarbamates.
39
Revised
Graphical Abstract (pictogram)
40
Revised
Muhammad Kashif Amir has completed his M.Phil Inorganic Chemistry from Bahauddin Zakariya University Multan, pakistan. Currently he is PhD scholar in department of Chemistry, Quaid-i-Azam University Islamabad, Pakistan under the supervision of Dr. Zia-ur-Rehman. His research is focused on the synthesis, characterization and medicinal applications of metal based compounds.
Shahan Zeb Khan is a lecturer in University of Science and Technology Bannu Khyber Pakhtunkhwa, Pakistan. He did his M.Sc and M.Phil from Quaid-I-Azam University Islamabad and currently he is pursuing his Ph.D. at QAU under the supervision of Dr. Zia-ur-Rehman. His research is focused on the synthesis and characterization of metal based compounds and their Biological applications.
41
Revised
Professor Ian S. Butler was educated at the University of Bristol in the U.K. and, following postdoctoral work at Indiana and Northwestern Universities in the U.S.A., he began his academic career at McGill University in Montreal, Quebec, Canada in 1966, where he is still teaching and doing research in a wide range of applications of molecular spectroscopy. Well over 50 M.Sc. and PhD. students have obtained their degrees under his direction and ~500 articles and ~400 national and international conference presentations have emanated from their research work to date. He has been a Visiting Professor in the U.K., France, China, Brazil, Hungary and Australia. His teaching and research efforts have been recognized by the Gerhard Herzberg Award from the Spectroscopy Society of Canada and the David Thomson Award from McGill University. He has co-authored several textbooks, including Relevant Problems for Chemical Principles (with Dr. A.E. Grosser) and Inorganic Chemistry: Principles and Applications (with Dr. J.F. Harrod). He has been married to his wife , Pamela, a former dancer with American Ballet Theatre in New York CIty and now a retired Professor of Political Science, for 48 years.
42
Revised
Dr. Abbas Hassan was educated at The University of Texas at Austin. Currently, he is working as Assistant Professor in Department of Chemistry Quaid-i-Azam University Islamabad Pakistan. His field of research is catalysic organic synthesis.
Dr. Zia-ur-Rehman was educated at the Quaid-i-Azam University Islamabad Pakistan and McGill University Canada in 2009. He began his academic career at QAU in 2009, where he is still teaching and doing research in metallo-drug, electrochemistry, supramolecular chemistry, environmental and drug delivery applications of novel surfactants, and nanotechnology. He published 84 research articles in various journals of international repute, and a co-author of a book entitled “DNA Binding and DNA Extraction: Methods, Applications and Limitations”. In his short academic carrier, 11 M.Phil students obtained their degrees in his direction. His teaching and research efforts have been recognized by the Dr. Abus Salam Award from TWAS. He has been married to his wife Kausar for 4 years and together they have a son Muhammad Ahmmad and a daughter Musfira.
43
Revised
Highlights Potential anticancer palladium(II) dithiocarbamates DNA-binding and DNA-denaturing studies Structure-activity relationship
45
S0020-1693(16)30346-2 http://dx.doi.org/10.1016/j.ica.2016.06.036 ICA 17127
To appear in:
Inorganica Chimica Acta
Received Date: Revised Date: Accepted Date:
10 March 2016 19 May 2016 26 June 2016
Please cite this article as: M.K. Amir, S.Z. Khan, F. Hayat, A.H. Khan, I.S. Butler, Z-u. Rehman, Anticancer activity, DNA-binding and DNA-denaturing aptitude of palladium(II) dithiocarbamates, Inorganica Chimica Acta (2016), doi: http://dx.doi.org/10.1016/j.ica.2016.06.036
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Revised
Anticancer activity, DNA-binding and DNA-denaturing aptitude of palladium(II) dithiocarbamates Muhammad Kashif Amira, Shahan Zeb Khana,b, Faisal Hayata, Abbas Hassan Khana, Ian S. Butlerc and Zia-ur-Rehmana* a
Department of Chemistry, Quaid-i-Azam University, Islamabad-45320, Pakistan.
b
Department of Chemistry, University of Science & Technology, Bannu-28100, KPK, Pakistan c
Department of Chemistry, McGill University, 801 Sherbrooke St. West, Montreal, Quebec, Canada H3A2K6 *
Corresponding author
Email: [email protected]/[email protected] Tel: 92-(051)90642245 Fax: 92-(051)90642241
1
Revised
Abstract Although it is well known that cisplatin and its analogues are effective anticancer agents, but their clinical use is restricted by some serious side effects. Palladium complexes are emerged as altenative metallo-anticancer drugs merited by their structural similarity to platinum(II) complexes, more labile nature, minimal chemoresistance and often water solubility. However, due to exceptional high reactivity of palladium complexes than their platinum counterparts, they are not only obstructed by the sulfur containing molecules to reach their pharmocolgical targets but also significantly enhance their affinity to convert into inactive trans isomers. This hitch can be overcome by the use of appropriate ligands that can have the potential to turn down the negative lability to positive inertness. This review provides a summary of the anticancer potential, DNA-binding and DNA-denaturing aptitude of various palladium(II) dithiocarbamates in which the presence of the dithiocarbamate moiety significantly improves the anticancer action and, at the same time, reduces the possibility of any damaging side effects.
Keywords: Palladium(II) dithiocarbamates; Anticancer drugs; DNA-binding and DNAdenaturing
2
Revised
1. Introduction Cancer is undoubtedly one of the world’s major health issues and several million people died from this disease every year. Following the accidental discovery of Rosenberg and his colleagues in 1965 of the eventual utility of cis-Pt(NH3)2Cl2 in cancer treatment [1], a wide variety of platinum(II)-based complexes have been investigated and now several other drugs with excellent anticancer activity are commercially available, e.g., carboplatin, oxaliplatin, nedaplatin, lobaplatin, and heptaplatin (Fig 1) [2]. The anticancer properties of the platinum(II) drugs can be attributed to the formation of intrastrand crosslinks between the guanine nitrogen bases of DNA, causing DNA bending, which subsequently interferes with DNA replication, transcription and other nuclear functions, and finally inhibits cancer cell proliferation and tumor growth. The clinical use of cisplatin and related complexes is, however, limited because of some unwelcome side effects that accompany treatment, including nephrotoxicity, ototoxicity, gastrointestinal toxicity, vomiting and developing resistance, which may be either primary (intrinsic) or secondary (acquired) [3-5]. It has been suggested that the nephrotoxicity and gastrointestinal toxicity of cisplatin may be the result of platinum complexation and inactivation of thiolcontaining enzymes [6, 7]. Platinum drug resistance is known to be a multifactorial process, which can occur through several mechanisms, including decreased platinum drug uptake due to resistance at the cell membrane, increased drug efflux, drug inactivation by glutathione and metallothioneins, evasion of apoptosis and enhancement of DNA repair [8, 9]. To overcome platinum drug resistance and toxicity, one of many strategies employed utilizes a variety of sulfur (thio)-containing ligands as detoxicant agents [10]. The small bite-angle and soft nature of the dithiocarbamato moiety as a stabilizing chelating ligand renders the platinum(II) complexes unreactive under a variety of conditions [11, 12]. In particular, dithiocarbamates have been 3
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established as excellent inhibitors of cisplatin-induced nephrotoxicity [13]. The strong binding of platinum(II) to the dithiocarbamate moiety limits any ligand exchange reactions with other sulfur-containing renal proteins and thiol-containing enzymes, thus protecting the normal tissues without undermining the anticancer action [14]. Fig. 1 Palladium complexes can be considered as substitute to platinum drugs for the treatment of different types of cancers owing to structural analogy [15-18]. However, their 105 times more reactivity than platinum complexes [19], make them more susceptible to fast hydrolysis accompanied by isomerization [19- 21]. The latter property is responsible for the formation of inactive trans isomers that lowers down anticancer activity [19]. However, this negative reactivity can be tunned to positive inertness by the right choice of the carrier ligands. It has been well notted that the bulky carrier ligands facilitate movement of the intact complex towards the target through steric shelter and encourage entrance into the cell through lipophilicity [19]. The reactivity can also be lowered down using ligands with soft donor sites i.e sulfur or phosphorous. In this context, detoxicant dithiocarbamate ligands received utmost attention due to the presence of strongly donating sulfur atoms and their chelating nature. Consequently, there is a growing interest in the designing of palladium(II) dithiocarbamates followed by their anticancer screening against different cancer cell lines, detailed DNA-binding and DNA-denaturing studies [22-28]. The detoxication effect of dithiocarbamates stems from their strong binding to palladium(II) that quenches ligand exchange reactions with sulfhydryl groups [29-31]. In this review, recent advances in the cytotoxic activities of palladium(II) dithiocarbamates against different cell lines, including sensitive 2008, A431, U2OScells and resistant C13*, A431/Pt, U2OS/Pt cells, are described. In addition, the current research on these palladium(II) complexes with respect to 4
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DNA synthesis inhibition, interstrand cross-linking, DNA–protein cross-linking, micronuclei (MN) detection, DNA-binding and denaturing is highlighted.
2. Anticancer activity of the palladium(II) dithiocarbamates The use of palladium(II) complexes containing N- and S-donor ligands in an effort to reduce the problems associated with the toxicity of cisplatin and its analogues is an important approach in anticancer studies. The palladium(II) complexes shown in Fig. 2 (1-4), which contain the 2ethoxy-2-oxoethyl(methyl)carbamodithioate ligand, have been examined for their cytotoxicity by performing dye exclusion tests on human leukemic HL-60 cells and by studying their inhibitory effects on cell proliferation in human adenocarcinoma HeLa cells [32]. Complex 1 is more active than is cisplatin against human leukemic HL-60 and adenocarcinoma HeLa cells (Table1). Notably, the neutral complexes, 1 and 2, which have a chloride ligand directly bonded to palladium, have lower IC50 values than do the ionic complexes 3 and 4, where chloride is an outer sphere ligand [32]. The higher cytotoxicities of complexes 1 and 2 may be the result of the ease of formation of covalent drug-DNA adducts. Fig. 2 Building on this notion, other neutral palladium(II) dithiocarbamates with two chiral aminoalcohols
4-(1-hydroxy-2-(methylamino)ethyl)phenol
(synephryne)
5
and
3-(2-amino-1-
hydroxyethyl)phenol (norphenylephrine) 6 (Fig. 3) have been prepared and subsequently investigated for their cytotoxicity against a panel of human cancer cell lines, including HeLa, HL60, sensitive 2008 and resistant C13* utilizing the MTT (tetrazolium salt reduction) test [33]. The cytotoxicities of complexes 5 and 6 proved to be concentration dependent against the HL60 and HeLa cell lines. An anticancer screening study of complexes 5 and 6 against the cisplatin5
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sensitive 2008 human cancer cell line and cisplatin-resistant variant C13* cells showed that they were only effective against the latter cell line [33]. An examination of the effect on DNA synthesis in Ehrlich cells (short-term effect) and on clonal growth capacity of 2008 cells (longterm effect) indicated that complexes 5 and 6 have significant antiproliferative activity similar to that of the reference drug for which the corresponding platinum complexes are completely inactive [33]. Fig. 3 Inspired by the highly interesting cytotoxic properties of the 2-picoline-based platinum(II) complex 7 (Fig 4), which were subjected to phase-I clinical trials [34-38], the heteroleptic palladium(II) complexes 8 and 9, which contain dithiocarbamate and picoline ligands (Fig. 4), were evaluated for their cytotoxicity, DNA synthesis inhibition, interstrand, DNA-protein crosslinking, and micronuclei (MN) detection properties [39]. These complexes were less active than is cisplatin against human squamous cervical adenocarcinoma (HeLa) and leukemic promyelocyte (HL60) cell. Furthermore, the 2-picoline complex 8 is more active than is the 3picoline complex 9. The high activity of the former complex may be linked with the more stable 2-picoline-palladium adduct formation due to the presence of electron releasing methyl closer to the donor site (Table 1). The efficacy of complex 8 against a panel of six human tumor cell lines, Fig. 4 and Table 1 including the sensitive (2008, A431, U2OS) and resistant(C13*, A431/pt, U2OS/pt) cell lines with reference to cisplatin was also studied utilizing the MTT assay approach [40]. The inhibitory effect of this complex on cell proliferation was concentration dependent for both the sensitive and resistant cell lines. Complex 8 proved to be slightly less active against the sensitive 2008 cells and A431 cell line but, interestingly, it was more active against the resistant C13* 6
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cells and A431/pt cell line (Table 2). This complex was also more active than was the reference cisplatin against U2OS cells at lower concentrations, but less active at higher concentrations. The resistant factors (RF) obtained for complex 8 and cisplatin showed that the former was able to overcome cisplatin resistance. In addition, HL60 cells were incubated with various concentrations of complex 8 and the mixtures were investigated with respect to DNA synthesis inhibition in terms of radioactive 3H-thymidine incorporation. This complex inhibits DNA synthesis marginally less than does cisplatin, but the analogous platinum complex has significant inhibition of DNA synthesis by comparison to that of cisplatin, even at low concentrations. Interstrand DNA cross-links (ISC) are unique, but pivotal for cytotoxicity, as they are among the critical molecular lesions that are able to inactivate DNA as a template for replication. The detection of interstrand crosslinks in 2008 cells was performed by incubating DNA with complex 8 for 6 to 12 h and subsequently exposing the cells to γ-rays, followed by alkaline solution elution [41]. The DNA ISC index was calculated by the formula (1-r0/1-r)1/2-1, where r and r0 are the fractions of 14C-thymidine-labelled DNA for the treated cells vs. the control cells remaining on the filter, respectively [41]. The DNA ISC index was greater for complex 8 than for cisplatin after 12 h exposure. The DNA-protein cross-links (DPC) were also measured as the number of DPC induced per million of nucleotides in 2008 cells. It was observed that cisplatin caused a Table 2 significant retention of DNA on the filter indicating the formation of a remarkable amount of DPC, while complex 8 proved to be totally unable to induce DPC [39]. These observations are interesting as DPCs are supposed to induce clastogenic effects in mammalian cells [42]. The genotoxic potential of complex 8 and cisplatin via micronuclei (MN) detection was evaluated on human lymphocytes. Complex 8 has a lower potential for genotoxicity than does
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cisplatin, as indicated by the lower levels of MN induction. Moreover, the relationship between MN induction and cytotoxicity is linear for complex 8 and cisplatin, but the fitting lines show different slopes. Different amounts of micronucleated cells were induced compared to cisplatin at the same survival rate [39]. It means the complex (8) tempt anticancer activity with reduced chromosomal damage. The homoleptic palladium(II) dithiocarbamates 10-12 have been evaluated for cytotoxicity against a panel of seven human tumor cell lines (Fig. 5) using the microculture sulforhodamine B (SRB) test [43] with doxorubicin, cisplatin, 5-fluorouracil, methotrexate, etoposide, and taxol as reference drugs. Complexes 10-12 have very low cytotoxic activities compared to those for the reference drugs and the heteroleptic complexes (which possess one dithiocarbamate moiety) [43]. The low cytotoxicity of complexes 10-12 may be due to the absence of a metal-attached, labile chloride ligand, which ultimately limits the interaction of the drug with DNA or other cellular components. Fig. 5 In an intriguing study, complexes with a dithiocarbamate moiety (13 and 14) and an amino acetate moiety (15 and 16) were synthesized with the aim of exploring the significance of the dithiocarbamate moiety in anticancer action (Fig. 6) [44]. An examination of the growth inhibitory effect of complexes 13-16 against human chronic myelogenous leukemia cell lines proved that the dithiocarbamate-based palladium(II) complexes (13 and 14) were more active than were the related complexes containing O-and N-donor ligands (complexes 15 and 16) [44]. The presence of the dithiocarbamate moiety in the structure of the Pd(II) complexes has a marked effect upon the growth suppression activity of K562 cells. Fluorescence spectroscopy is a useful technique to study the structure, dynamics and binding properties of protein molecules in solution. The effect of the Pd(II) complexes 13-16 on the 8
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structure of the two variants of carrier protein, β-lactoglobulins (BLG-A and -B), were investigated using intrinsic fluorescence measurements. The intrinsic fluorescence emissions of the proteins in the presence of 20 µM of complexes 13 and 14 with dithiocarbamate were nearly two times less than were the emissions of the proteins under the influence of 20 µM concentrations of complexes 15 and 16. This means that dithiocarbamate-based complexes, 13 and 14, have less effect on the tertiary structure of proteins at lower concentration than do the two other complexes (15 and 16) [44]. Circular dichroism (CD) spectroscopy revealed that the content of regular secondary structures of BLG-A and -B upon interaction with complexes 13-16 did not show any significant changes. Fig. 6 The lack of water solubility of some of the palladium(II) antitumor complexes explains their limited bioavailability and inferior in vivo activity [45]. The water-soluble complex 13 (Fig. 6) proved to be more active than is cisplatin against the human cancer cell line K562 (Table 3) [46]. Interestingly, changes in concentration had a little impact upon the activity. The watersoluble complex 17 (Fig. 7), with a short hydrocarbon chain ligand (ethylcarbamodithioate), was more active than is cisplatin against the human cancer cell line K562 and was observed to produce a dose-response suppression on the growth of the K562 leukemia cell line [47]. The anticancer activity jumped up markedly upon increasing the chain length associated with the N Fig. 7. attached alkyl group of dithiocarbamate ligand. Complex 18 with an octyl group (Fig. 8) was two and nearly eight times more active than were the analogous complexes with butyl (13) and ethyl(17) substituents, respectively. Complex 18 was 22 times more active than is cisplatin against the human cancer cell line K562 (Table 3) [22]. To explore the role of the auxiliary 9
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ligands on the anticancer properties, the 2,2ʹ-bipyridine ligand in complexes 13, 17 and 18 was replaced by 1,10-phenanthroline affording complexes 19, 20 and 21 (Fig. 9). The activity against the human cancer cell line K562 was increased for complexes 19 and 20 compared to the parent complexes 13 and 17 presumably due to the more planar nature of the 1,10-phenanthroline ligand [24-26]. The activity for complex 18 was greater than for the complex 21, showing the importance of both dithiocarbamate and auxiliary ligands. Figs. 8 and 9 Other
water-soluble
complexes
containing
morpholine-
(22)
and
piperidine-based
dithiocarbamate (23) ligands (Fig. 10) have been shown to inhibit the in vitro human cancer cell line K562 growth more efficiently than does cisplatin [27, 28](Table 3). The cytotoxicity of dithiocarbamate complex containing piperidine skeleton (23) was found greater than morpholine one ( 22). Complexes 22 and 23 are less active than are the analogous platinum(II) complexes at lower concentrations, but are more active at higher concentrations (45 µM). Table 3 Changing from an ethyl-based amine to morpholine or piperidine and a butyl-based amine to piperidine in the structure of the 2,2ʹ-pyridine dithiocarbamatopalladium(II) complexes results in increased cytotoxicity [27, 28, 47]. Complexes 18 and 23 had nearly equal cytotoxicities (Table 3). Fig. 10 Other
water
soluble
palladium(II)
complexes
containing
same
cationic
spheres
(pyrrolidinedithiocarbamate and 1, 10-phenanthroline) but different anions {24 (nitrate) and 25 (bromide)} exhibited more cytotoxic effect than cisplatin against chronic myelocytic leukemia
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K562 cell line (Fig. 11, table 3), however, the comparable activity of both complexes excluded the significant role of anion in anticancer activity [48,49].
Fig. 11
Dinuclear water soluble palladium(II) complexes 26-28 (Fig. 12) were also inspected for cytotoxicity against human gastric carcinoma (AGS), esophageal squamous cell carcinoma (Kyse-30), and hepatocellular carcinoma (HepG2) cell lines by MTT assay [50]. These complexes were examined to have greater cytotoxicity than cisplatin as shown from their IC50 values against these three cell lines (table 4). The results indicated that shorter alkyl chain length between two palladium nuclei enhances cytotoxicity in this class of compounds. Fig. 12 and Table 4
Mixed-ligand dithiocarbamate palladium(II) complexes having different amines or diamines in their structures have good cytotoxic activities. In order to investigate the effect of tertiary phosphine ligands, the mixed-ligand dithiocarbamate palladium(II) complexes (29-34) were synthesized (Fig. 13) and tested for their antitumor activity against cisplatin-resistant DU145 human prostate carcinoma (HTB-81) cells [51, 52]. It was observed that the complexes have a greater antitumor activity at lower concentrations (Table 5), nevertheless, except 33 the activity of 29-34 was found concentration dependent. Complexes 33 and 34 were the most active with IC50 values of 1.33 and 1.55 µM respectively [51]. The highest IC50 value was observed for complex 29 (6.94 µM). The high cytotoxicity of complex 33 can be attributed to the electron-
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donating tricyclohexylphosphine ligand resulting in a stronger Pd-P bond and so there is better possibility of reaching intact complex to the target DNA [51]. In comparison, complex 29 has a slightly less electron-donating triphenylphosphine ligand resulting in a marginally weaker Pd-P bond and the complex presumably has a greater tendency to dissociate. The movement of this dissociated complex towards DNA may be interrupted by other groups in the cell, such as glutathione, cysteines and methionines [53]. The high activity of complex 34 can be attributed to the presence of the 2-pyridyl moiety in the organophosphine ligand, which has the potential to form a hydrogen bond with DNA bases [51]. Fig. 13 and Table 5 Khan et al. [54] have reported six heteroleptic Pd(II) dithiocarbamate complexes 35-40 (Fig. 14) and studied their in vitro antitumor activity against cisplatin-resistant DU145 human prostate carcinoma (HTB-81) cells [54]. These complexes have good antitumor activity at low concentrations (Table 5) with complex 40 being the most active and complex 39 the least active. The IC50 values determined for the six complexes are lower than are the literature IC50 values for the standard drug cisplatin [54, 55]. The higher antitumor activity of complex 40 has been attributed to the presence of oxygenated species in the dithiocarbamate ligand, which have a higher potential to form hydrogen bonds with the DNA bases. The weak donating ability of phosphorous due to the presence of the electron-withdrawing chloride and phenyl groups in the organophosphine are presumably responsible for the lower cytotoxicity of complex 39. The second lowest activity was observed for complex 37 and this was attributed to the presence of the bulky t-butyl group, which hinders movement of the complex towards the target DNA. Fig. 14
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Two other piperazine based heteroleptic Pd(II) dithiocarbamates 41-42 have been recently synthesized and evaluated for their anticancer activity against Hela cell line at different concentrations [56]. The IC50 values of the complexes show that these complexes have greater cytotoxicity as compared to cisplatin against Hela cell line (Table 5). The results of DNA fragmentation in vitro by using DNA ladder assay show that the cytotoxicity of these complexes might be due to strong interaction with DNA leading to DNA damage and apoptosis [56]. Two
new
potent
anticancer
heteroleptic
palladium(II)
complexes
of
4-(2-
methoxyphenyl)piperazine-1-carbodithioate ligand with two different organophosphines namely diphenyl-p-tolylphosphine (43) and tri-p-tolylphosphine (44) were screened against five different cell lines [57]. For both complexes the activity varied in the sequence MCF7 ˃ Hepa-c1c7 ˃ PC3 ˃ LU ˃ MDA-MB-231. Furthermore, the more active nature of 44 than 43 was attributed to its extra stablility as was confirmed by the X-ray single crystal analysis (shorter Pd-S bond distances) and DFT calculation.
Fig. 15 The complexes (45-53) having expected structures (Fig. 16) have also been tested for cytotoxic activity against human cancer cell lines (2008 & A431) by MTT assay [58]. The results indicated that neutral binuclear complex (47) and ionic polynuclear complex (49) have cytotoxicity close to cisplatin (Table 6). Neutral binuclear complexes (45, 48) were less active. While neutral binuclear complex (46) and other ionic complexes (50-53) were proved to be ineffective against 2008 and A431 cancer cell lines. In order to assess cross-resistance with cisplatin the most active complexes (47, 49) were also examined against cisplatin-resistant cell line C13*. The RF values
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calculated for the complexes (47, 49) were about eight times lower than that of cisplatin declaring different cross-resistance profile from that of cisplatin [58]. Fig. 16 and Table 6
3. DNA-binding and DNA-denaturing studies The anticancer properties of metallodrugs are due to the binding and denaturing of DNA, which result in interference with DNA replication, transcription and other nuclear functions. Some metallodrugs having replaceable chloride such as cisplatin, bind covalently to the nitrogen basis of DNA. The complexes 1, 2, 5-9, 29-44 also have replaceable chloride and chances to have covalent type interaction with DNA. Some metallodrugs interact with DNA non-covalently by the formation of hydrogen bonds, electrostatic and/or hydrophobic interactions in the minor groove regions of DNA. Some metallodrugs interact with DNA non-covalently by insertion of planar moiety (planer heterocyclic groups, aromatic ring or fused aromatic rings) between adjacent base pairs of the double helix. This is interclative mode of interaction. The complexes 13-23 have interclative mode of interaction with DNA as described in the respective literature [22, 24-28, 46, 47]. The DNA-binding and DNA-denaturing behavior of complexes 13-23 has also been investigated using standard biochemical methods. In a procedure using UV-visible spectroscopy, the absorbance of the fixed concentration of the compound (drug), “A1”, is first measured. Then, the absorbance of mixed solutions of the compound with increasing concentrations of DNA, “A2”, are measured while keeping the volume and concentration of the compound constant. The change in absorbance, “ΔA” is calculated from ΔA = A1−A2. The
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binding constant “K” can be calculated from the slope-to-intercept ratio of the plot of A1/ΔA versus 1/[DNA] according to the Benesi–Hildebrand equation [57]. The results indicated that the complexes containing the more planar 1,10-phenanthroline ligand and the butyl or octyl chain in the dithiocarbamates have greater binding constants (Table 7). The 1/ΔA ratio is plotted versus 1/[DNA] and the value for the change in absorbance when all the binding sites on DNA are occupied by the compound “ΔAmax” is determined from the intercept on the ordinate of the linear plot [47]. Next, the absorbance of a fixed concentration of DNA “A3” is measured. The absorbance of mixed solutions of DNA with increasing concentrations of the compound “A4” is measured while keeping the volume and concentration of DNA constant [47]. Similarly, the changes in absorbance “ΔΆ” are calculated from ΔΆ = A3 −A4. The concentration of the compound bound to DNA, [L]b, is determined from the following equation [47] [L]b = ΔΆ[L]t/ΔAmax, where [L]t is the total concentration of the compound added. The concentration of free compound [L]f is obtained from the following equation [47][L]f = [L]t−[L]b. The ratio of the concentration of bound compound to total DNA concentration “ῡ” is calculated from ῡ = [L]b/ [DNA]t. Next, the Scatchard plots for these complexes are obtained by plotting ῡ/[L]f versus ῡ [23]; these proved to be curvilinear concave downwards suggesting cooperative binding. The Kolts plot is produced by plotting ῡ versus Ln[L]f [47], while the Hill plot is obtained by plotting Ln ῡ/1−ῡ versus Ln[L]f [47]. The value of binding parameter “n” the socalled Hill coefficient (which is taken as a criterion of cooperativity) can be measured from the slope of the straight line in the plot. The value of Hill coefficient, “n”, can also be measured from the slope of the Kolts plot as n = 4 × (value of slope of Kolts plot). The values of Hill coefficients indicated that these complexes have cooperative binding (Table 7), while complexes 17, 18, 21, 22 and 23 have greater cooperativity than do the other complexes. The number of 15
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binding sites per 1000 nucleotides “g” can be calculated by substituting the values of ῡ, K, n and [L]f in the Hill equation [47] ῡ = g(K[L]f)n/(1 + (K[L]f)n). The results indicated that the complex 17 has a greater number of binding sites per 1000 nucleotides (Table 7). The molar Gibbs free energy of binding “ΔG˚b” for the compound-DNA interaction at a particular temperature can be calculated from the following equation [47]ΔG˚b = −RTLnK, where R is the gas constant. Then, the molar enthalpy of binding “ΔH˚b” for the compound-DNA interactions at two different temperatures can be calculated from the following equation [47] LnKT1 /KT2 = −ΔH˚b/R(1/T2−1/T1). Finally, the molar entropy of binding “ΔS˚b” for the compound-DNA interaction at a particular temperature can be evaluated from the following equation [47] ΔG˚b =ΔH˚b−TΔS˚b. In order to determine the DNA-denaturing parameters, the UV absorption spectrum of a DNA solution at 258 nm is recorded keeping the absorbance reading below one. Then, a fixed amount of the compound is added stepwise and at each step an absorbance reading is taken. This procedure is performed until there is no further change in absorbance. The changes in the absorbance of DNA solutions are plotted against different concentrations of the compound. From this plot, the concentration of the compound at the midpoint of transition “[L]1/2” can be determined [47]. The concentrations of the compounds at the midpoint of transition calculated for these complexes are given in the Table 8. Using this DNA denaturation plot and the Pace method, the unfolding equilibrium constant K*, can be calculated from the following equation [46, 47] K* = AN−Aobs/Aobs−AD, where AN is the absorbance of natural conformation of DNA, AD is the absorbance of denatured conformation of DNA and Aobs is the absorbance in the transition region. Then, the unfolding free energy of DNA “ΔG˚” can be calculated from the
16
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following equation [46, 47]ΔG˚ = −RT LnK*. When these “ΔG˚” values are plotted versus the concentrations of the compound in the transition region, a straight line is obtained. The equation for this straight line can be written as [46]ΔG˚ = ΔG˚( ) − m[compound]. From the slope of straight line, the measure of DNA denaturing ability “m” can be determined. The results indicated that the complex 20 has the highest DNA denaturing ability (Table 8). From the intercept of the straight line, the conformational stability of the free DNA “ΔG˚( ) ” is determined [47]. The molar enthalpy of DNA denaturation by the compound “ΔH˚denaturation” in the range of the two temperatures (T1and T2) can then be calculated using the following Gibbs Helmholtz equation [47]ΔH˚denaturation = ΔG˚(T1)/T1−ΔG˚(T2)/T2÷ 1/T1− 1/T2. The molar enthalpy of DNA denaturation in the absence of the compound “ΔH˚( ) ” is determined by interpolation of a plot of ΔH˚denaturation against the concentration of the compound [47]. Finally, the molar entropy of DNA denaturation in the absence of the compound “ΔS˚( ) ” is obtained from the following equation ΔG˚( ) = ΔH˚( ) −TΔS˚( ). The DNA-binding and DNAdenaturing parameters thus obtained are shown in Tables 7 and 8. The results indicate that the complexes containing long-chain dithiocarbamate ligands and more planar auxiliary ligands have greater DNA-binding and DNA-denaturing abilities. These results are in agreement with the cytotoxicity results Tables 7 and 8 4. Conclusions In this review, the anticancer potential of the palladium(II) dithiocarbamates has been examined in terms of their cytotoxic activities against different cancer cell lines, and by DNA synthesis
17
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inhibition, detection of interstrand cross-links, DNA–protein cross-links, micronuclei detection, DNA-binding and denaturing studies. Most of these complexes (1, 13, 14, 17-44), including the water-soluble ones, have greater cytotoxic activities than does cisplatin. Moreover, they have less cross-resistance towards cisplatin-resistant human tumor cell lines. Heteroleptic palladium(II) dithiocarbamates containing organophosphines as ancillary ligands demonstrated greater cytotoxic effect than the complexes containing N-donor ligands. The mechanism of action seems to be similar to that of cisplatin itself because they strongly and irreversibly bind to DNA, and can denature DNA, even at low concentrations. Contrary to this, the complexes containing ligands with two nitrogen donor sites (bpy and phen) adopt intercalative mode of interaction with DNA. The DNA binding and, as a consequence the anticancer action of these complexes depends on the lipophilicity (carbon chain length), planarity and number of aromatic rings in the auxiliary ligands, and the presence of more labile chloride group. It will be worthwhile entering several of the water-soluble palladium(II) dithiocarbamates into clinical trials in the future. Acknowledgement We thank the Higher Education Commission of Pakistan for financial support. References [1] B. Rosenberg, L. Van Camp, T. Krigas, Nature 205 (1965) 698. [2] T. Boulikas, A. Pantos, E. Bellis, P. Christofis, Cancer Ther. 5 (2007) 537. [3] A. Eastman, Cancer cells (Cold Spring Harbor, NY) 2 (1989) 275. [4] K. WoŸniak, J. Błasiak, Acta Biochim. Pol. 49 (2002) 583.
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[5] N.P. Johnson, J.L. Butour, G. Villani, F.L. Wimmer, M. Defais, V. Pierson, V. Brabec, Metal antitumor compounds: the mechanism of action of platinum complexes. in: Ruthenium and Other Non-Platinum Metal Complexes in Cancer Chemotherapy. Springer, 1 (1989). [6] (a) P. J. O'Dwyer, J. P. Stevenson, S. W. Johnson, Cisplatin: chemistry and biochemistry of a leading anticancer drug, Wiley-VCH, (1999) 29. ISBN 3-906390-20-9; (b) B. Lippert (Ed.), Chemistry and Biochemistry of a Leading Anticancer Drug, Wiley-VCH, New York (1999); (c) T. Glaser, G.F. Von Mollard, D. Anselmetti, Inorg. Chim. Acta xxx (2016) xxx–xxx. (http://dx.doi.org/10.1016/j.ica.2016.02.013) [7] H.M. Pinedo, H.H. Schornagel, J.H. Schornagel, Platinum and Other Metal Coordination Compounds in Cancer Chemotherapy 2. Springer, 1996. [8] P.J. Morin, Drug Resist. Update. 6 (2003) 169. [9] Z.H. Siddik, Oncogene 22 (2003) 7265. [10] D. Fregona, L. Giovagnini, L. Ronconi, C. Marzano, A. Trevisan, S. Sitran, B. Biondi, F. Bordin, J. Inorg. Biochem. 93 (2003) 181. [11] K.S. Siddiqi, S.A. Nami, Y. Chebude, J. Braz. Chem. Soc. 17 (2006) 107. [12] A. Rai, S.K. Sengupta, O.P. Pandey, Spectrochim. Acta. Part A 64 (2006) 789. [13] M.S. Murthy, L.N. Rao, J.D. Khandekar, E.F. Scanlon, Cancer Res. 47 (1987) 774. [14] R. Mital, N. Jain, T. Srivastava, Inorg. Chim. Acta 166 (1989) 135. [15] H. Sigel, Metal Ions in Biological Systems: Volume 42: Metal Complexes in Tumor Diagnosis and as Anticancer Agents, CRC Press, (2004). [16] W. Micklitz, W.S. Sheldrick, B. Lippert, Inorg. Chem. 29 (1990) 211. [17] K.J. Barnham, C.J. Bauer, M.I. Djuran, M.A. Mazid, T. Rau, P.J. Sadler, Inorg. Chem. 34 (1995) 2826.
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[18] W.-Z. Shen, D. Gupta, B. Lippert, Inorg. Chem. 44 (2005) 8249. [19] M. Fanelli, M. Formica, V. Fusi, L. Giorgi, M. Micheloni, P. Paoli, Coord. Chem. Rev. 310 (2016) 41. [20] L. Khassanova, P. Collery, Z. Chassanova, I. Maymard, J.-C. Étienne, Metal Ions in Biology and Medicine, John Libbey Eurotext, (1990). [21] A. Furlani, V. Scarcia, G. Faraglia, L. Sindellari, L. Trincia, M. Nicolini, Eur. J. Med. Chem. 21 (1986) 261. [22] H. Mansouri-Torshizi, M. Saeidifar, F. Khosravi, A. Divsalar, A.A. Saboury, F. Hassani, Bioinorg. Chem. Appl. (2011), article ID 394506. [23] (a) A. Saboury, J. Iran. Chem. Soc. 3 (2006) 1; (b) G. Scatchard, Ann. N.Y. Acad. Sci. 51 (1949) 660. [24] H. Mansouri-Torshizi, M. Saeidifar, A. Divsalar, A. Saboury, S. Shahraki, Bull. Korean Chem. Soc. 31 (2010) 435. [25] H. Mansouri-Torshizi, M. Saeidifar, A. Divsalar, A.A. Saboury, Spectrochim. Acta Part A 77 (2010) 312. [26] H. Mansouri-Torshizi, M. Saeidifar, A. Divsalar, A. Saboury, Nucleosides, Nucleotides Nucleic Acids 30 (2011) 405. [27] H. Mansouri-Torshizi, M. Moghaddam, A. Divsalar, A. Saboury, J. Biomol. Struct. Dyn. 26 (2009) 575. [28] H. Mansouri-Torshizi, M. Eslami-Moghadam, A. Divsalar, A.-A. Saboury, Acta Chim Slov. 58 (2011) 811. [29] Q. Zhang, W. Zhong, B. Xing, W. Tang, Y. Chen, J. Inorg. Biochem. 72 (1998) 195. [30] M.-T. Hassan, I.-M. Mahbobe, S.A. Akbar, Acta Bioch. Bioph. Sin. 35 (2003) 886.
20
Revised
[31] G. Zhao, H. Sun, H. Lin, S. Zhu, X. Su, Y. Chen, J. Inorg. Biochem. 72 (1998) 173. [32] (a) G. Faraglia, D. Fregona, S. Sitran, L. Giovagnini, C. Marzano, F. Baccichetti, U. Casellato, R. Graziani, J. Inorg. Biochem. 83 (2001) 31; (b) W.J. Durkin, V.K. Ghanta, C.M. Balch, D.W. Davis, R.N. Hiramoto, Cancer Res. 39 (1979) 402. [33] (a) V. Alverdi, L. Giovagnini, C. Marzano, R. Seraglia, F. Bettio, S. Sitran, R. Graziani, D. Fregona, J. Inorg. Biochem. 98 (2004) 1117; (b) M.C. Alley, D.A. Scudiero, A. Monks, M.L. Hursey, M.J. Czerwinski, D.L. Fine, B.J. Abbott, J.G. Mayo, R.H. Shoemaker, M.R. Boyd, Cancer Res. 48 (1988) 589; (c) D.T. Vistica, P. Skehan, D. Scudiero, A. Monks, A. Pittman, M.R. Boyd, Cancer Res. 51 (1991) 2515. [34] L.R. Kelland, S.Y. Sharp, C.F. O’Neill, F.I. Raynaud, P.J. Beale, I.R. Judson, J. Inorg. Biochem. 77 (1999) 111. [35] Y. Chen, Z. Guo, S. Parsons, P.J. Sadler, Chem. Eur. J. 4 (1998) 672. [36] J. Parkinson, P. Sadler, J. Chem. Soc. Dalton Trans. 21 (1998) 3577. [37] J. Holford, S. Sharp, B. Murrer, M. Abrams, L. Kelland, Br. J. Cancer 77 (1998) 366. [38] F.I. Raynaud, F.E. Boxall, P.M. Goddard, M. Valenti, M. Jones, B.A. Murrer, M. Abrams, L.R. Kelland, Clin. Cancer Res. 3 (1997) 2063. [39] L. Giovagnini, C. Marzano, F. Bettio, D. Fregona, J. Inorg. Biochem. 99 (2005) 2139. [40] M.C. Alley, D.A. Scudiero, A. Monks, M.L. Hursey, M.J. Czerwinski, D.L. Fine, B.J. Abbott, J.G. Mayo, R.H. Shoemaker, M.R. Boyd, Cancer Res. 48 (1988) 589. [41] K.W. Kohn, Pharmacol. Therapeut. 49 (1991) 55.
21
Revised
[42] C. Marzano, F. Baccichetti, F. Carlassare, A. Chilin, S. Lora, F. Bordin, Photochem. Photobiol. 71 (2000) 263. [43] (a) F. Shaheen, A. Badshah, M. Gielen, M. Dusek, K. Fejfarova, D. de Vos, B. Mirza, J. Organomet. Chem. 692 (2007) 3019; (b) Y.P. Keepers, P.E. Pizao, G.J. Peters, J. van Ark-Otte, B. Winograd, H.M. Pinedo, Eur. J. Cancer Clin. 27 (1991) 897. [44] A. Divsalar, A. Saboury, R. Yousefi, A. Moosavi-Movahedi, H. Mansoori-Torshizi, Int. J. Biol. Macromol. 40 (2007) 381. [45] N. Farrell, Met. Ions Biol. Syst. 32 (1996) 603. [46] (a) H. Mansouri-Torshizi, I. Mahboube, A. Divsalar, A.-A. Saboury, Bioorg. Med. Chem. 16 (2008) 9616; (b) A. King, B. Nicholson, Biochem. J. 114 (1969) 679. [47] (a) M. Islami-Moghaddam, H. Mansouri-Torshizi, A. Divsalar, A. Saboury, J. Iran. Chem. Soc. 6 (2009) 552; (b) R. Mitai, K.S. Ray, T. Srivastava, R. Bhattacharya, J. Inorg. Biochem. 27 (1986) 133. [48] S. Shahraki, H. Mansouri-Torshizi, A. Heydari, A. Ghahghaei, A. Divsalar, A. A. Saboury, H. Ghaemi, M. Doostkami and S. Zareian, Iran. J. Sci. Tech. 39A2 (2015) 187. [49] H. Mansouri-Torshizi, S. Shahraki, Z.S. Nezami, A. Ghahghaei, S. Najmedini, A. Divsalar, H. Ghaemi and A.A. Saboury, Complex Met. 1 (2014) 23. [50] S. Hadizadeh, N. Najafzadeh, M. Mazani, M. Amani, H.Mansouri-Torshizi and A. Niapour, Biochem. Res. Int., (2014). http://dx.doi.org/10.1155/2014/813457.
22
Revised
[51] H. Khan, A. Badshah, G. Murtaz, M. Said, C. Neuhausen, M. Todorova, B.J. Jean-Claude, I.S. Butler, Eur. J. Med. Chem. 46 (2011) 4071. [52] R. Cattaneo-Pangrazzi, H. Schott, R. Schwendener, Prostate 45 (2000) 8. [53] Y. Kasherman, S. Sturup, D. Gibson, J. Med. Chem. 52 (2009) 4319. [54] H. Khan, A. Badshah, M. Said, G. Murtaza, J. Ahmad, B.J. Jean‐Claude, M. Todorova, I.S. Butler, Appl. Organomet. Chem. 27 (2013) 387. [55] A. Amantana, C.A. London, P.L. Iversen, G.R. Devi, Mol. Cancer Ther. 3 (2004) 699. [56] S. Khan, M.K. Amir, M.M. Naseer, R. Abbasi, K. Mazhar, M.N. Tahir, I. Z. Awan, Zia-urRehman, J. Coord. Chem. 68 (2015) 2539. [57] S. Z. Khan, M. K. Amir, I. Ullah, A. Aamir, J. M. Pezzuto, T. Kondratyuk, F. BélangerGariepy, A. Ali, Zia-ur-Rehman, App. Organomet. Chem. 30 (2016) 392. [58] D. Montagner, C. Marzano, V. Gandin, Inorg. Chim. Acta 376 (2011) 574.
23
Revised
Fig. 1:Structures of currently approved platinum-based anticancer drugs.
Fig. 2: Mixed ligand palladium(II) 2-ethoxy-2-oxoethyl(methyl)carbamodithioates (1-4).
Fig. 3: Structure of complexes 5 and 6. 25
Revised
Fig. 4: The heteroleptic platinum and palladium(II) complexes of 2-ethoxy-2oxoethyl(methyl)carbamo- dithioateandpicoline.
Fig. 5:The homoleptic palladium(II) complexes (10-12) with dithiocarbamate ligands.
Fig. 6: Water soluble complexes with and without dithiocarbamate moiety.
26
Revised
Fig. 7: The water-soluble complex containing ethyldithiocarbamate ligand.
Fig. 8: The water-soluble complex containing octyldithiocarbamate ligand.
27
Revised
Fig. 9: The palladium complexes obtained by replacing 2,2-bipyridine with 1,10-phenanthroline from the ethyl, butyl and octyldithiocarbamate complexes.
Fig. 10:The water-soluble complexes with morpholinedithiocarbamate and piperidinedithiocarbamate ligands.
28
Revised
Fig. 11: The water-soluble complexes containing nitrate and bromide ion in the outer sphere.
Fig. 12: Dinuclear water-soluble complexes.
29
Revised
Fig. 13: The mixed-ligand palladium dithiocarbamate complexes containing organophosphine ligands.
Fig. 14:The mixed-ligand dithiocarbamate palladium complexes containing phosphine ligands.
30
Revised
Fig. 15:The piperazine based mixed-ligand dithiocarbamate palladium complexes containing phosphine ligands.
Fig. 16: Expected structures for dithiocarbamate palladium(II) complexes (45-53).
31
Revised
Table 1: In vitro cytotoxic activity results (IC50, µM) of palladium(II) dithiocarbamates (complexes 1-4, 8 and 9) against different cell lines. Complex
IC50( µM) against cancer cell lines
Refs.
HL-60
HeLa
1
3.11
5.61
[32]
2
5.83
7.50
[32]
3
27.44
77.0
[32]
4
24.27
59.99
[32]
8
59.62
69.54
[39]
9
˃100
˃100
[39]
cisplatin
3.40
6.33
[39]
Table 2: Cross-resistance profiles of the investigated complex (8) toward some established cisplatin-sensitive and resistant human tumor cell lines. Complex
IC50 (µM)
R.F.
Refs.
2008 cells
C13* cells
8
60.0
128.4
2.14
[39]
Cisplatin
42
616.1
14.68
[39]
A431 cells
A431/Pt cells
8
81.2
161.5
1.99
[39]
Cisplatin
76.4
297.1
3.89
[39]
U2OS cells
U2OS/Pt cells
8
54.3
108.3
2.02
[39]
Cisplatin
30.2
191.2
6.33
[39]
R.F. = Resistance factor = IC50 resistant cell line/ IC50 sensitive cell line.
32
Revised
Table 3:The 50% cytotoxic concentrations of the palladium (II) dithiocarbamate complexes. Complex
Cc50/IC50 (µM)
Refs
K562 13
14
[44]
14
36
[44]
15
251
[44]
16
320
[44]
13
18
[46]
17
55
[47]
18
7
[22]
19
10
[24]
20
10
[25]
21
15.6
[26]
22
26
[27]
23
10.5
[28]
24
46
[48]
25
45
[49]
Cisplatin
154
[46]
Table 4: IC50 (µg) values of the dinuclear palladium (II) complexes against AGS, Kyse-30 and HepG2 cell lines Complex 26 27 28 Cisplatin
AGS 0.68 0.78 1.20 4.08
IC50 (µg) Kyse-30 0.88 0.98 1.06 1.90
Refs HepG2 1.02 1.17 1.19 2.08
[50] [50] [50] [50]
33
Revised
Table 5: The 50% inhibitory concentration (IC50) of the mixed ligand palladium(II) dithiocarbamates. Complex
IC50 (µM) HTB-81 cells
Refs. Hela cells
29
6.94
[51]
30
-
[51]
31
5.71
[51]
32
3.10
[51]
33
1.33
[51]
34
1.55
[51]
35
3.67
[54]
36
-
[54]
37
9.52
[54]
38
4.57
[54]
39
21.7
[54]
40
2.12
[54]
41
22
[56]
42
26
[56]
Cisplatin
78
[56]
34
Revised
Table 6: Cytotoxic activity (IC50, µM) of palladium dithiocarbamate complexes.
Complex
Formula
2008
A431
C13*
RF
Refences
45
[Pd2Cl2(PyDT)2(dah)]
52.9
60.1
-
-
[58]
46
[Pd2Cl2(PyDT)2(dab)]
˃100
˃100
-
-
[58]
47
[Pd2Cl2(ESDT)2(dah)]
16.4
14.3
14.5
0.9
[58]
48
[Pd2Cl2(ESDT)2(dab)]
42.5
36.5
-
49
[Pd(ESDT)(dab)]nCln
18.4
21.5
17.6
0.9
[58]
50
[Pd(PyDT)(en)]Cl
˃100
˃100
-
-
[58]
51
[Pd(PyDT)(dap)]Cl
˃100
˃100
-
-
[58]
52
[Pd(PyDT)(en)][PdCl2(PyDT)]
˃100
˃100
-
-
[58]
53
Bu4N[PdCl2(DMDT)]
˃100
˃100
-
-
[58]
Cisplatin
Pt (NH3)2Cl2
12.2
19.5
95.4
7.8
[58]
[58]
PyDT = pyrrolidine dithiocarbamate, ESDT = sarcosine ethyl ester dithiocarbamate, DMDT = dimethyl dithiocarbamate, en = ethylenediamine, dab = 1, 4-diaminobutane, dah = 1, 7-diaminoheptane, dap = 1, 3diaminopropane
35
Revised
Table 7: Values of ∆Amax and binding parameters in the Hill equation for interaction between Pd(II) complexes and DNA in 10 mmol/L Tris–HCl buffer and pH 7.0.
Complex
Temp(℃)
∆Amaxa
gb
Kc×104 M-1
nd
Errore
Refs.
13
27
0.047
8
0.317
1.48
0.005
[46]
37
0.033
8
0.401
1.79
0.008
[46]
27
0.014
9
0.180
4.77
0.03
[47]
37
0.011
9
0.270
4.79
0.06
[47]
27
0.035
6
5.28
5.92
0.13
[22]
37
0.058
5.86
6.18
0.14
[22]
27
0.246
5
6.693
1.972
0.0007
[24]
37
0.167
5
4.360
1.604
0.0004
[24]
27
0.213
6
0.1902
1.432
0.00004
[25]
37
0.101
6
0.1772
1.239
0.0006
[25]
27
0.038
3
8.13
4.95
0.0043
[26]
37
0.040
3
11.78
3.079
0.0041
[26]
27
0.07
6
0.018
3.5
0.06
[27]
37
0.08
6
0.024
4.74
0.09
[27]
27
0.098
6
0.020
3.91
0.039
[28]
37
0.048
6
0.033
3.8
0.063
[28]
17
18
19
20
21
22
23
a
Change in the absorbance when all the binding sites on DNA were occupied by a metal complex. The number of binding sites per 1000 nucleotides. c The apparent binding constant. d The Hill coefficient (as a criterion of cooperativity). e Maximum error between theoretical and experimental values of ῡ. b
36
Revised
Table 8: Thermodynamic parameters of DNA denaturation by palladium complexes.
Complex
Temp (℃)
M
∆G˚(H2O)
∆H˚(H2O)
∆S˚(H2O)
[L]1/2
(kJmol-1K)
(mM/L)
0.073
0.093
[46]
0.085
[46]
0.121
[47]
0.105
[47]
0.093
[22]
0.086
[22]
0.055
0.0134
[24]
0.030
0.0127
[24]
0.065
0.0101
[25]
0.065
0.0099
[25]
~0
0.012
[26]
~0
0.011
[26]
0.08
0.15
[27]
0.14
[27]
~0
0.128
[28]
~0
0.125
[28]
Refs.
-1
13
17
18
19
20
21
22
23
(kJmol-1)(mM)-1
(kJmol-1)
27
72.12
7.47
37
89.9
6.74
27
159.03
18.81
37
158.4
17.81
27
195
13.44
37
218
12.93
27
589.6
9.846
37
710.1
19.610
27
1119.6
11.54
37
1019.2
10.88
27
544.05
7.877
37
515.77
6.845
27
159.4
23.7
37
115.9
15.7
27
80.1
20.33
37
86.6
19.00
(kJmol )
29.36
48.90
28.09
37.58
31.14
5.154
262.8
20.5
0.10
0.05
37
Revised
Graphical Abstract (synopsis) This review provides a summary of the anticancer potential, DNA-binding and DNA-denaturing aptitude of palladium(II) dithiocarbamates.
39
Revised
Graphical Abstract (pictogram)
40
Revised
Muhammad Kashif Amir has completed his M.Phil Inorganic Chemistry from Bahauddin Zakariya University Multan, pakistan. Currently he is PhD scholar in department of Chemistry, Quaid-i-Azam University Islamabad, Pakistan under the supervision of Dr. Zia-ur-Rehman. His research is focused on the synthesis, characterization and medicinal applications of metal based compounds.
Shahan Zeb Khan is a lecturer in University of Science and Technology Bannu Khyber Pakhtunkhwa, Pakistan. He did his M.Sc and M.Phil from Quaid-I-Azam University Islamabad and currently he is pursuing his Ph.D. at QAU under the supervision of Dr. Zia-ur-Rehman. His research is focused on the synthesis and characterization of metal based compounds and their Biological applications.
41
Revised
Professor Ian S. Butler was educated at the University of Bristol in the U.K. and, following postdoctoral work at Indiana and Northwestern Universities in the U.S.A., he began his academic career at McGill University in Montreal, Quebec, Canada in 1966, where he is still teaching and doing research in a wide range of applications of molecular spectroscopy. Well over 50 M.Sc. and PhD. students have obtained their degrees under his direction and ~500 articles and ~400 national and international conference presentations have emanated from their research work to date. He has been a Visiting Professor in the U.K., France, China, Brazil, Hungary and Australia. His teaching and research efforts have been recognized by the Gerhard Herzberg Award from the Spectroscopy Society of Canada and the David Thomson Award from McGill University. He has co-authored several textbooks, including Relevant Problems for Chemical Principles (with Dr. A.E. Grosser) and Inorganic Chemistry: Principles and Applications (with Dr. J.F. Harrod). He has been married to his wife , Pamela, a former dancer with American Ballet Theatre in New York CIty and now a retired Professor of Political Science, for 48 years.
42
Revised
Dr. Abbas Hassan was educated at The University of Texas at Austin. Currently, he is working as Assistant Professor in Department of Chemistry Quaid-i-Azam University Islamabad Pakistan. His field of research is catalysic organic synthesis.
Dr. Zia-ur-Rehman was educated at the Quaid-i-Azam University Islamabad Pakistan and McGill University Canada in 2009. He began his academic career at QAU in 2009, where he is still teaching and doing research in metallo-drug, electrochemistry, supramolecular chemistry, environmental and drug delivery applications of novel surfactants, and nanotechnology. He published 84 research articles in various journals of international repute, and a co-author of a book entitled “DNA Binding and DNA Extraction: Methods, Applications and Limitations”. In his short academic carrier, 11 M.Phil students obtained their degrees in his direction. His teaching and research efforts have been recognized by the Dr. Abus Salam Award from TWAS. He has been married to his wife Kausar for 4 years and together they have a son Muhammad Ahmmad and a daughter Musfira.
43
Revised
Highlights Potential anticancer palladium(II) dithiocarbamates DNA-binding and DNA-denaturing studies Structure-activity relationship
45