3 main lines (1) Radiochemistry. Physical characteristics of the radionuclides Basics of radiolabeling DTPA- and DOTApeptides

Transcription

1 utlines Radiochemistry Imaging of receptor-mediated processes in-vivo with radiolabeled peptides EMI, Dublin, eptember Basics of radiolabeling DTA- or DTPA-peptides with radiometals, such as 90 Y, 111 In and Basics of radiolabeling DTA-peptides with Ga (separate section) Physical characteristic and production of Ga, 90 Y, 111 In and Wouter A.P. Breeman Department of uclear Medicine Erasmus MC Rotterdam The etherlands pecific of these radionuclides pecific of radiolabeled peptides and the effect on biodistribution, pharmacokinetics, pharmacodynamics and imaging of receptor-mediated processes utlines utlines Choice of the chelator Choice of the peptide Radiochemical limitations of Ga-, 90 Y-, 111 In- and -labeled DTA- or DTPA-peptides, especially for imaging and therapy of receptor-mediated processes andling DTA- and DTPA-peptides, including storage, quality control, peptide purity and peptide content Quality control, radiolysis and shelf life of radiolabeled DTAand DTPA-peptides Definitions of pecific Activity (.A.) including definitions, confusions on.a., also as f(t) Production of via n, route from enriched 176 Points of view: production site () vs. applications.a. Quality Control eed of high.a. for PRRT (Peptide Receptor Radionuclide Therapy) Waste management radioactive (solid and liquid) legal aspects in EU and elsewhere 3 main lines (1) Physical characteristics of the radionuclides Basics of radiolabeling DTPA- and DTApeptides Quality control, RadioChemical Purity (RCP) and quenchers Production of Ge 90 Y 111 In Ge Ga 70 Ge 69 Ga Zn extra measuring.a. of

2 Production routes for possible routes reactor-produced from natural or enriched m f f f 176m f 176 f 176 m f f f Φ= n.cm -2.s -1 Calculations based on actual amount target (= ), flux, time, etc Calculation? nb epi-thermic neutrons are not included in the calculations 3 main lines (2) Physical characteristics of the radionuclides Basics of radiolabeling DTPA- and DTApeptides Quality control, Radiochemical Purity (RCP) and quenchers

3 [DTPA 0 ]ctreotide C C C C C C C C DTPA-DPhe-Cys-Phe-DTrp-Lys-Thr-Cys-Thr(ol) DTPA-DF-C-F-DW-K-T-C-T(ol) DTPA-DPhe-Cys-Phe-DTrp-Lys-Thr-Cys-Thr(ol) DTPA-DF-C-F-DW-K-T-C-T(ol) 2 2 DTAtate C C [DTPA 0 ]ctreotide C C C C C 2 2 DTPA-DPhe-Cys-Phe-DTrp-Lys-Thr-Cys-Thr(ol) DTPA-DF-C-F-DW-K-T-C-T(ol) DTA-DPhe-Cys-Tyr-DTrp-Lys-Thr-Cys-Thr DTA-DF-C-Y-DW-K-T-C-T Fig 1 FIG! FIG 1 DTA-MG11 p 0 p op the reaction? p 14 C C C C 3 DTA-DGlu-Ala-Tyr-Gly-Trp-Met-Asp-Phe- 2 DTA-E-A-Y-W-M-D-F- 2 2 The rates of formation of the complexes of radiometal-dta were found to be inversely proportional to the [ + ] olubility of In 3+, Ga 3+, Y 3+, and lanthanides as p + [ ]? FIG 2 zilagyi E, et al Moerlein M et al solubility curve Formation of Colloid 111 In, 90 Y and p = 1/3 log K s -1/3 log [M 3+ ] - log K w K s In-111 K s Y-90 K s - 1,30E-37 6,00E-24 1,00E-25 p p 8,4 8,2 8,0 7,8 7,6 7,4 MBq/ ml solubility curve 90 Y 8,6 8,4 8,2 8,0 7,8 7,6 MBq/ ml

4 solubility curve Experiments in small volumes (typically µl) were performed in MoBiTec tubes originated for RT-PCR thermocycler (max vol 125 µl) p MBq/mL ote: Ci per mg contains 80 % 176, conc*5 p Ga- In-111 Y-90 - Ga-67 crew cap & plunjer Double sealed Tungsten heating block p curve of 100 µl 0.1 M Cl and 1.25 M a-actetate p added volume a-acetate (µl) : at 80 C +: at 100 C, : 90 Y at 80 C : 90 Y at 100 C, : 111 In at 80 C : 111 In at 100 C Figure 5A Test model Labeling conditions, e.g. vials and stoppers Include positive control of labeling at high specific Controlled addition of ingredients, e.g. buffer Quality control ITLC PLC UPLC M, LC-M Figure 6 Figure X

5 stock solution for PRRT Cl 3 is 44 GBq per ml Incubation of fixed 6 µl of Cl 3 with x µl kit solution in MoBiTec cup, 20 min at 80 C % Incorporation undiluted total volume 6-9 µl 10* diluted total volume 50 µl µl DTAtate kit (0.4 µg per µl) p in kinetics? Confirmed under stochiometrc conditions with other metals, like Cu, Fe, Zn, i, Y Figure 7 Ga-labeling DTA-peptides Figure 8 Cyclotron-produced positron-emitter Ga p,2n EC + 69 Ga Ge Ga Zn Practical Aspects of the Ge/ Ga-generator handbook for radiolabeling DTA-peptides WAP Breeman t½ : 69 Ga : stable t½ : Ge : 270 d t½ : Ga : min* t½ : Zn : stable Ge Ga 70 Ge 69 Ga Erasmus MC Rotterdam Dept of uclear Medicine The etherlands *MeV [%] beta + Zn w.a.p.breeman@erasmusmc.nl

6 subjects Gallium chemistry history since seventies General aspects i.e. RA-quantification of Ge/ Ga, reaction kinetics (p, time, T) handling small amounts, shielding rerouting ionic Ga 3+ in vivo 0,10 0,01 Ingrowth Ga from Ge- Ga Generator [MBq per MBq] ,00 min In theory: 50 % at min 1 (63 %) at 95 min 2 (86 % at 190 min 4 (99 %) at 380 min Act( Ge ) (t0) Act Ga(t) ( Ga)* *( e ( ( Ga)- ( Ge)) ( Ge)t - - e ) ( Ga)t history on Ge generator since seventies n 2 column M Cl or EDTA, 8-hydroxy-quinoline or oxine contaminants? evaporation, pyrolysis, column chromatography, and/or purification etc more recently labeling peptides at low.a. (<20 MBq per µg) and still purification needed ew generators, other carriers (organic and oxides), knowledge, breakthrough, safety nets, PE and C 18 columns, Monographs in preparation, GMP status, Marketing Authorisation, etc Robotics Gallium chemistry: formation of colloid/gallate etc! Ga() Ga() p = 1/3 log K s - 1/3 log [M 3+ ] - log K w K sin : 1.30E-37 K sy :6.00E-24 K s : 1.00E-25 K sga : 7.10E-36 1 mci Ga 3.6*10-13 mol 1 mci 111 In 2.15*10-11 mol solubility Ga p 5,7 5,5 5,3 5,1 4,9 MBq/ ml p solubility curve MBq/mL Ga- In-111 Y-90 - Ga-67 p = 1/3 log K s - 1/3 log [M 3+ ] - log K w K sin E-37 K sy E-24 K s E-25 Ga() Ga() at p 7.4 and 25 C total solubility is 1 µm, 98.4 % as Ga() - 4, minimum at p 5.2

7 Ge-generator + shielding Cassette-based Robotics Ge/ Ga-generator, bninsk, Russia Ti 2 ; 0.1 M Cl Ga: MeV (178 %), 1.8 MeV (88 %) beta + E&Z cintomics martmedix and others sample in plastic shielding in dosecalibrator ( 18 F-mode, abundance corrected <5%contribution from beta s) Ge 511 KeV in gamma counter (from Ga in (RA) equilibrium with Ge) after decay of Ga; 1 MBq Ga 1Bq Ga after 24 h hielding Pb 8 cm Tungsten VL ± 0.5 cm: k 65 mci Ge in shielding 1 µci unshielded: < 1 µv.h -1 at 10 cm Combined anion and cation column chromatography GaCl 3 + Cl Ga(Cl) G.Meyer et al., EJMMI 2003 ew acl-based technique D. MÜller et al., Bioconjugate Chemistry 2012 ew purification and concentration techniques are still being developed 3 main lines (3) Physical characteristics of the radionuclides Basics of radiolabeling DTPA- and DTApeptides Quality control, RadioChemical Purity (RCP) and quenchers extra measuring.a. of utlines Quality Control of radiolabeled DTPA- or DTAregulatory peptides for PR and PRRT RCP and focused on receptor-mediated processes EMI Dublin, eptember Wouter A.P. Breeman Department of uclear Medicine Erasmus MC Rotterdam DTPA- and DTA-Met-containing peptides 111 In and ow to quantify RCP Comparable on different systems: data interchangeable? RCP vs Gy? Model developed Parameter: PLC of the radiopeptides (chromatographic in r f intact radiopeptide vs non-intact radiopeptide) Quenchers, which and why, and effectiveness on RCP Requirements for PLC QC of the peptides, peptide purity, peptide content incl molecular extinction coefficient λ 278nm, 1300 for Y and 5600 for W The etherlands w.a.p.breeman@erasmusmc.nl

8 Radiolysis of radiopeptides measurable? Model developed RCP vs Gy 111 In and Met-containing peptides Effectiveness of quenchers + DTA-peptide -DTA-peptide -DTA-peptide, 30 min, no quenchers, 23 MBq in 200 µl baseline correction? software? -DTA-peptide, 20 h, no quenchers, 23 MBq in 200 µl -DTA-peptide, 20 h, no quenchers, 23 MBq in 200 µl Q1: how to quantify? Q2: comparison? without quenchers +10 % (v/v) ethanol + L-methionine 3 main lines (3) + ascorbic acid + ascorbic acid + ascorbic acid +10 % (v/v) ethanol +10 % (v/v) ethanol 111 In-DTA-MG11 + L-methionine Physical characteristics of the radionuclides Basics of radiolabeling DTPA- and DTApeptides Quality control, RadioChemical Purity (RCP) and quenchers 111 In-DTA-MG11 sulfoxide T= 24h extra measuring.a.

9 utlines Definitions of pecific Activity (.A.) Measuring pecific Activity of by PLC An alternative method Radiolabeling -DTA-peptides for PRRT Production of via n, route from enriched 176 Calculations of Wouter A.P. Breeman and Erik de Blois Department of uclear Medicine Calculations of.a. and Quantification of.a. by titration and PLC Erasmus MC Rotterdam The etherlands pecific mass uppose All atoms are radioactive atom % =100, 51.4 pmoles per mci 19.5 mci per nmol 720 MBq per nmol 110 Ci per µg etc pecific mass Physicists+IUPAC pecific mass pecific mm m m mass( ( ) ) pecific mm m m mass( ( f f ) ) pecific mass ( ( DTPA ordta peptide ) ) Datasheet of Vendors uclear Physicists Radiochemists Radiochemists Biochemists and in in uclear Medicine A B C D E.A. = of per mass of all present- f that is the mass of ( m++m) - f There are several methods to quantify total mass of and f, also in the hot liquid Alternative methods: Calculation? pecific (.A.) Waste ( m - or Ge-containing) management In EU, radioactive waste with half-life > 100 days is considered radioactive > 10 Bq per g All EU-members are allowed to implement stricter levels, e.g. Germany: > 1 Bq per g Bakker et al. in QJM 2006: m in -containing waste requires attention don t mix de Blois et al. in J Radioanal and uclear Chemistry 2011: concentrating and solidification of Ge-containing liquid waste