Energetics of Complexation of Y with DOTA, a Model for Cancer Radiotherapy

1. Energetics of Complexation of Y with DOTA, a Model for Cancer Radiotherapy Yun Hee Jang, Mario Blanco, Siddharth Dasgupta, William A. Goddard, III MSC, Beckman Institute, Caltech David A. Keire, John E. Shively The Beckman Research Institute of the City of Hope

2. Chelating ligand (DOTA) 90 Antibody targeting b-emitting (2.25 MeV, t1/2=64h) Tumor cell Cancer Radioimmunotherapy: Bifunctional Chelating Agent • Therapy: 90Y3+(64h) • Diagnosis: 111In3+(2.8d) • 64Cu2+(12.8h) • MRI contrast agent: Gd3+ D. Parker, Chem. Soc. Rev. 19, 271 (1990)

3. Criteria of Good Chelating Agent DOTA (log K=24.8) DTPA (log K=22.1) Thermodynamic stability Kinetic inertness at pH 2~8w.r.t. acid- promoted dissociation <0.5% dissociated over 18 days in serum (pH 7.4, 37oC): inert Not inert leading to bone-marrow toxicity Rapid complexation x1600 slower than Y-DTPA formation Lewis, Raubitschek and Shively, Bioconjugate Chem. 5, 565 (1994)

4. fast slow Kinetics of metal binding of DOTA E.T. Clark and A.E. Martell, Inorg.Chim.Acta 190, 27 (1991) X.Y. Wang, et al. Inorg.Chem. 31, 1095 (1992) Y3+ + Y3+ or Y3+ Y3+ Type II: stable/inert Y3+ + H2(DOTA)2- Type I: labile

5. Objectives • Calculate structure/energy change occurring during complex-formation • Identify the rate-determining step: • Deprotonation or conformation change? • Design new chelating agent and predict its energetics/kinetics Calculation method • B3LYP/LACVP* // HF/LACVP* (6-31g* for C/H/O/N; Hay-Wadt ECP for Y) • Vibration analysis  ZPE / thermodynamic quantity  Gibb’s free energy • Continuum solvation calculation by solving Poisson-Boltzmann equation • Jaguar 3.5 (Schrodinger Inc.)

6. Optimized structure after sequential deprotonation -H+ -H+ YH2(DOTA)+ YH(DOTA) Y(DOTA)- Y3+ outside the cage the same as x-ray structure of final complex • Y3+ moves into the cage spontaneously with deprotonation. • RMS deviation between ring conformations < 0.5 Å. • Deprotonation is the rate-determining step.

7. How can the proton be removed? Direct attack of outside base on the ring proton? No room for it. top view side view bottom view

8. How can the proton be removed? Conformation change to the one favorable to attack? Too high cost, especially, for YH(DOTA) 4-coordinate 3-coordinate 2-coordinate YH2(DOTA)+ 16.6* (12.1)** kcal/mol 42.7* (34.5)** kcal/mol * 1.807 Å for r(Y) ** 1.673 Å for r(Y) in solvation calculation YH(DOTA) 21.6* (24.6)** kcal/mol

9. How can the proton be removed? Proton transfer from ring NH to COO (more accessible to outside base)? reactant (NH...COO) TS (N..H..COO) product (N...COOH) ***experimental DG for Eu,Gd,Ce,Ca-complexes (Inorg.Chem. 32, 4193 (1993)) • Proton transfer is easier than conformation change. • Calculated activation free energy is in agreement with experimental value.

10. Suggestion: DO3A1Pr Structural change leading to more stable TS: 6-membered ring of DO3A1Pr rather than 5-membered ring of DOTA TS (DO3A1Pr) DO3A1Pr(Pr=propionate) TS (DOTA)

11. DO3A1Pr: Protonation site Hpr(DO3A1Pr): 0.0 kcal/mol Hac(DO3A1Pr): 7.8 kcal/mol Protonation at propionate site is more stable.  6-membered ringTS

12. Summary • Deprotonation from ring nitrogen is the rate-determining step. • Deprotonation occurs by proton transfer from ring nitrogen to carboxylate. • Adding CH2 to one carboxylate arm can improve the incorporation rate. Future work • Explicitly-coordinated water molecules • How many water molecules? • Effect on structure/energetics • Introduction of amide linkage

13. Acknowledgement Caltech William A. Goddard, III Siddharth Dasgupta Mario Blanco Daniel Mainz Sungu Hwang City of Hope John E. Shively David Keire Supported by NSF