Six most potent compound. Compound 8 with the less

Six lipophilic and one hydrophilic derivatives of
hydroxyurea (table 1) were synthesized in 24-72% yield based on method that
shown in scheme 1. All of compounds characterized by TLC followed by IR and
proton NMR.

The cytostatic activity of compounds 2-9 were determined
using an in-vitro assay against Hella and Panc-1 cell lines and the results at
24 and 48 hours are summarized in tables 2-4 and figure 2. Due to great
difference of molecular weight of Hydroxyurea and compounds 2-9, the IC50
values were reported in micrograms and micromole scales. Based on the IC50
value (table 4) all the designed and prepared compounds are more potent than hydroxyurea
at 24 and 48 hours on both cell lines, that cytostatic activity at 48 hour is
more than of 24 hour. Lipophilicity (log P) strongly affect the cytostatic
activity of prepared ligands and compounds 7 and 5 with high amount of logp,
4.88 and 2.79 respectively, showed the high cytostatic activity that compound 7
with the highest logp was the most potent compound. Compound 8 with the less
amount of logp was the less cytostatic agent in this series.

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 Comparison of
compounds 4, 5 and 6 with close amounts of lipophilicity which having 2 phenyl
ring with different structure, orientation and volumes reveals in addition to
the logp, size and volume of substituent play very important role in activity. So
compound 4 with logp around 3.05 due to its more blocker structure is less
active than compounds 5 and 6 with logp 2.79 and 2.54 respectively.  Also compound 9 with high lipophilicity, due
to it geometrical shape, imine moiety, showed weak cytostatic activity. Comparison
of compound 8 with hydroxyurea that have close amounts of logp (-0.83 and -.082
respectively) and different amounts of molecular weight, reveals heavier
compound 8 is more active than hydroxyurea. Based on our docking studies and
in-vitro evaluation results, it is suggested that size, shape and hydrophobic
character of substituents strongly affect the pharmacodynamics and
pharmacokinetic of these ligands.

Finally, overall cytostatic activity (table 4) show that  all the synthetized compounds are very active
specially compounds 7, 5 and 3  can be
acted as a new lead compounds to design and finding novel and too potent
cytostatic agents.  


   The docked
conformations of compound 7 as the most potent ligand were ranked into
clusters based on the binding energy and the top ranked conformations were
visually analyzed. Drug-receptor interaction profile was analyzed using
AutoDock Tools (version 1.50) and Ligplot 16.   As it was reported already the inhibitor
must be ready to destroy the Tyrosyl radical and thus the inhibitors such as compound
7 should be bonded at the active site with Tyrosine 176 that is a
radical harboring, to reduce the radical activity and inhibit cell division

The docking energies of compound 7 and hydroxyurea as
reference drug, are noted as -5.49 and -4.11 kcal/mol, respectively that based
on the lowest docking energy compound 7 shows higher affinity than
hydroxyurea to human RNR protein. The LIGPLOT diagrams of compound 7 and
human RNR docking interactions are shown in figure 1.

Compound 7 is easily occupied at the active site of
receptor via two strong hydrogen bonds using the residue of Ser173. In
addition, some hydrophobic interactions by the residues Tyr176, Ala135, Ser177,
Arg190, Ala132, Leu193 and Ile180 also help the drug for its conformational
stability in the active site. Dichlorophenyl moiety of ligand was incorporated
into the hydrophobic pocket that created by Arg190, Ala132, Leu193 and Ile180. Hydroxyl
moiety of ligand has good interactions with surrounding residues, Ser173, Tyr176
and Ala135.



   All studied
compounds have more cytotoxic effect against Hella and Punc cell line than
hydroxyurea at 24 and 48 hours.  Compound
7 is the most active against both cell lines. It was concluded that lipophilicity,
size and shape of substituents strongly affect the cytostatic activity and
ligands with high lipophilicity and small size and the compact shape has good
pharmacokinetic (cell permeability and stability) and pharmacodynamics.