The Application of ATR-FTIR Spectroscopy and the Reversible DNA Conformation as a Sensor to Test the Effectiveness of Platinum(II) Anticancer Drugs

Abstract

Platinum(II) complexes have been found to be effective against cancer cells. Cisplatin curbs cell replication by interacting with the deoxyribonucleic acid (DNA), reducing cell proliferation and eventually leading to cell death. In order to investigate the ability of platinum complexes to affect cancer cells, two examples from the class of polyfluorophenylorganoamidoplatinum(II) complexes were synthesised and tested on isolated DNA. The two compounds trans-[N,N'-bis(2,3,5,6-tetrafluorophenyl)ethane-1,2-diaminato(1-)](2,3,4,5,6-pentafluorobenzoato)(pyridine)platinum(II) (PFB) and trans-[N,N'-bis(2,3,5,6-tetrafluorophenyl)ethane-1,2-diaminato(1-)](2,4,6-trimethylbenzoato)(pyridine)platinum(II) (TMB) were compared with cisplatin through their reaction with DNA. Attenuated Total Reflection Fourier Transform Infrared (ATR-FTIR) spectroscopy was applied to analyse the interaction of the Pt^(II) complexes with DNA in the hydrated, dehydrated and rehydrated states. These were compared with control DNA in acetone/water (PFB, TMB) and isotonic saline (cisplatin) under the same conditions. Principle Component Analysis (PCA) was applied to compare the ATR-FTIR spectra of the untreated control DNA with spectra of PFB and TMB treated DNA samples. Disruptions in the conformation of DNA treated with the Pt^(II) complexes upon rehydration were mainly observed by monitoring the position of the IR-band around 1711 cm^-1 assigned to the DNA base-stacking vibration. Furthermore, other intensity changes in the phosphodiester bands of DNA at ~1234 cm^-1 and 1225 cm^-1 and shifts in the dianionic phosphodiester vibration at 966 cm^-1 were observed. The isolated double stranded DNA (dsDNA) or single stranded DNA (ssDNA) showed different structural changes when incubated with the studied compounds. PCA confirmed PFB had the most dramatic effect by denaturing both dsDNA and ssDNA. Both compounds, along with cisplatin, induced changes in DNA bands at 1711, 1088, 1051 and 966 cm^-1 indicative of DNA conformation changes. The ability to monitor conformational change with infrared spectroscopy paves the way for a sensor to screen for new anticancer therapeutic agents.

Keywords: DNA; IR; platinum.

Figure 1

Chemical structures of PFB, TMB and cisplatin.

Figure 2

DNA interaction with Pt^II and conformation change.

Figure 3

Infrared (IR) raw spectra (left) and 2nd derivative (right) of compounds PFB (1), TMB (2) and cisplatin.

Figure 4

Raw IR spectra of the transition of ss and ds A-DNA to B-DNA upon rehydration. Green: control (DNA treated with acetone), dark blue: DNA treated with PFB, light blue: DNA treated with TMB.

Figure 5

IR Average spectra (second derivative) of dsDNA treated with acetone (control 1) PFB, TMB, saline (control 2) and cisplatin in the course of dehydration (left) colour-coded from blue (hydrated) to red (dehydrated) and in the course of rehydration (right) colour-coded from red (dehydrated) to blue (hydrated). The inserts show spectral features around 1225 and 1088/1051 cm⁻¹.

Figure 6

IR Average spectra (second derivative) of ssDNA treated with acetone (control 1) PFB, TMB, saline (control 2) and cisplatin in the course of dehydration (left) colour-coded from blue (hydrated) to red (dehydrated) and in the course of rehydration (right) colour-coded from red (dehydrated) to blue (hydrated). The inserts show spectral features around 1225 and 1088/1051 cm⁻¹.

Figure 7

(A) PC1 versus PC2 scores plot depicting ssDNA treated with the PFB and TMB and the non-treated DNA (control in acetone/water) in the dehydrated state; (B) PC1 versus PC2 scores plot for the rehydrated ssDNA with and without drug treatment; (C) PC1 versus PC2 scores plot for dehydrated dsDNA with and without drug treatment; (D) PC1 versus PC2 Scores Plot for rehydrated dsDNA with and without the drug treatment.

Figure 8

(A) PC1 loadings plot depicting ssDNA treated with PFB and TMB and the non-treated DNA (control in acetone/water) in the dehydrated state. The positive loadings are associated with the controls and the negative loadings are associated with the drug inoculated cells; (B) PC1 loadings plot for the rehydrated ssDNA with and without drug treatment. The positive loadings are associated with the negative scores. The positive loadings are associated with the controls and the negative loadings are associated with the drug inoculated cells; (C) PC1 loadings plot for dehydrated dsDNA with and without drugs. The positive loadings are associated with the drug inoculated cells and the negative loadings are associated with the control cells; (D) PC1 versus PC2 loadings plot for rehydrated dsDNA with and without the drugs. The positive loadings are associated with the drug inoculated cells and the negative loadings are associated with the control cells.