Toxic Desserts for Cancer Cells

by Lindsay Kinyon

Background

Beginning in the early 1980s, researchers began examining the interactions of octahedrally coordinated complexes of copper, nickel, zinc, iron, cobalt, chromium, rhodium, osmium, and, especially, ruthenium with DNA. Octahedrally coordinated organometallic compounds (see Figure 1) are molecules that contain a metal center with six binding sites occupied by small organic molecules (ligands).

The generic compound formulation [M(N-N)3]n+ will be used to represent molecules of this type. These compounds have demonstrated site-specific DNA interaction (intercalation, groove-binding or electrostatic) and some have been shown to engage in photolytic strand cleavage. Notably absent from this collection of transition metal DNA probes were the octahedrally coordinated platinum family analogs, [Pt(N-N)3]4+ & [Pd(N-N)3]4+, most likely due to the difficulty of solvating the high charge density of the platinum (4+) & palladium (4+) ions.

Early in the summer of 1996, Dr. Robert Granger developed a new, versatile scheme for the synthesis of homoleptic (only one type of ligand) water-soluble octahedrally-coordinated platinum(IV) complexes with intercalative ligands^,. Dr. Granger then obtained a three-year grant from the Jeffress Memorial Trust to study the synthesis and characterization of these new Pt(IV) and Pd(IV) diimene complexes. That research focused primarily on the synthesis of Pt(IV) and Pd(IV) complexes of 1,10-phenanthroline and 2,2’-dipyridine (see Figure 2).

Figure 2

Although many metal complexes are used as DNA markers, only compounds of the platinum family are widely used medically as chemotherapeutic agents. Cisplatin (cis-diamminedichloroplatinum, a square-planar Pt(II) compound) is widely used alone or in combination with other chemotherapeutic drugs in the treatment of several aggressive cancers, including ovarian, lung, testicular and bladder carcinomas. Unfortunately, resistance to cisplatin can develop and the drug itself is toxic to the patient, with the kidneys, gastrointestinal tract, bone marrow and nervous system all experiencing distress resulting from treatment. Furthermore, not all tumors respond to cisplatin. So, there is the hope that new drugs might be found which would be effective against cisplatin-resistant tumors.

To date, none of the other octahedrally coordinated metal center complexes have shown to have anti-cancer activity, but because of the importance of platinum in other forms as anti-cancer drugs, Dr. Granger wished to explore the bioactivity of his new platinum(IV) and palladium(IV) complexes against malignant cells. In the summer of 1997, Dr. Robin Davies began preliminary cell culture experiments using Dr. Granger’s novel platinum (IV) and palladium (IV) complexes. To their surprise, these new platinum (IV) and palladium (IV) complexes were extremely active against malignant cell lines (see Figure 3).

Figure 3: Cell count vs. Time for human leukemia cells exposed to drug.

These results serve to indicate that further investigation of the cytotoxic effects of these novel Pt(IV) and Pd(IV) diimene complexes is warranted and to suggest that the proposed investigations are likely to prove fruitful. These investigations are currently underway in Dr. Davies research group.

In addition, Dr. Granger has also completed studies on the binding mode of [Pt(phen)₃]⁴⁺ with DNA and has been able to demonstrate that the (4+) charge on these complexes bends the DNA strand upon binding (see Figure 4).

Figure 4: The localized charge on the metal center "pulls" the anionic phosphates towards the metal.As the charge on the metal center increases, intercal-kinking increases.

Late in the summer of 1999, Dr.Granger began to focus his synthetic efforts on modifications to the intercalating ligand. Specifically, Dr. Granger has begun developing a series of extended ligand systems that will be able to insert deeper into the DNA backbone (see Scheme 1 for additional possible ligand systems). Use of these extended ligands will hold the metals (4+) charge farther away from the DNA’s backbone, thus promoting an intercalative binding mode.

Also, Dr. Granger has developed a synthetic scheme that will allow for the exact placement of individual ligands. This is the portion of the project upon which I have focused my summer research efforts.

Lindsay Kinyon's Summer Research 2000

The goal for my summer research was to synthesize a series of ligands similar to the ligand dppz (see Scheme 1). The first thing I had to do was to synthesize the necessary template molecule 5,6-dione-1,10-phenanthroline (see Scheme 2).

I then selected three commercially available diamene compounds for use in synthesizing three new dppz-like ligands (see Scheme 3).

Each of these syntheses involved refluxing (boiling) the 5,6-dione-1,10-phenanthroline ligand with the requisite diamine. A classic condensation reaction between the dione oxygen and the amine hydrogens occurs yielding water and our desired compound. The products of the three condensation reactions were confirmed by GC-Mass spectrometry (GC-MS). This technique allows for the determination of the exact mass of the molecule in addition to identifying fragmentation products (see Figures 5 & 6)

 

Figure 5: GC-MS of bdppz. The parent peak at 355 represents the ethyl ester of the expected product. The two peaks at 281 and 73 represent the two fragmentation products dppz and -CO₂CH₂CH₃.

Figure 6: GC-MS of dpnpz. The parent peak at 415 shows the ethyl ester of the expected product. The peak at 327 shows the expected product.

A surprise we encountered during this project occurred in the analysis of the product bdppz. The GC-MS did not find the expected product bdppz but instead we found the ethyl ester of bdppz. In retrospect, this is not surprising. By the same condensation reaction used to make dppz-like ligands, carboxylic acids will condense with alcohols to form esters. Since the reaction was conducted in ethanol, this is not a surprising result (see Scheme 4).

The discovery of the ethyl ester of bdppz led us to make some exciting conjectures. I then tried to repeat the bdppz synthesis in a non-alcohol solvent, which

would verify the stability of bdppz. However, I could not find a non-alcohol solvent that my starting materials were both soluble in. Once I found a suitable solvent, my next step would be to attempt to place a metabolite such as a simple sugar onto the bdppz ligand. Since our goal is to make cancer drugs, it only makes sense to exploit the exaggerated metabolism of cancer cells in order to actively transport our DNA drugs into the cancer cells (see Scheme 5).

 

Once this scheme is (Scheme 5) successful, we will want to place this ligand onto a metal center and create our DNA drugs (see Figure 8).

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This page updated Decmber 20, 2001