Molecular Imaging of CXCR4 Receptors

Transcription

1 Molecular Imaging of CXCR4 Receptors.J. Wester uklearmedizinische Klinik und Poliklinik, Klinikum rechts der Isar and Institut für Radiochemie TUM Campus Garching

2 Importance of CXCR4 and CXCL12 ypoxic Areas of Tumors: CXCL12 expression by fibroblasts CXCR4 expression on tumor cells tumor cell motility invasiveness CXCL12: promotes tumor cell growth by stimulating via CXCR4. induces recruitment of progenitors, which allow for tumor angiogenesis CXCR4: activation of CXCR4 leads to targeted metastasis to the marrow or other sites of high CXCL12 expression. ("hijacking" of circulating tumor cells)

3 AIM: to develop suitable ligands for CXCR4 for diagnosis, staging, therapy monitoring

4 Topochemical exploration of potent compounds using retro-enantiomer libraries of cyclic pentapeptides. T T140 FC131 Tamamura. rg Biomol Chem 2004, 2:

5 Radioligand-Binding Studies (II) Competitive binding curves of CPCR4 with 125 I-CPCR4 and 125 I-SDF-1α to Jurkat and CMS5/CXCR4 cells: I-CPCR4 125 I-SDF-1α bound tracer [%] CMS-5/CXCR4 Jurkat Comparable IC 50 values were determined for CPCR4 at both cell lines with both radioligands I-CPCR4 shows very low non-specific binding log [CPCR4]

6 Biodistribution Analysis of 125 I-CPCR4 in Mice min 60min 120min 25 %ID/g Blood Serum Lung Liver Pancreas Spleen Intestine Adrenals Kidneys igh tracer accumulation in the CXCR4-expressing tumor Low background accumulation igher tracer accumulation only in the metabolic and excreting organs (liver, intestine and kidneys) Muscle Bone CMS5-Tumor CMS5/CXCR4 Tumor

7 Advanced Multimodal CXCR4-Tumor Imaging (mirror image) Analysis of CXCR4/Luc expression and GFP control tumors: Bioluminescence Photo MRI µ-pet GFP- Fluorescence Control (GFP) CXCR4/ Luc Control (GFP) CXCR4/ Luc Control (GFP) CXCR4/ Luc CPCR4 in vivo PET and Bioluminescence Imaging: In vivo investigations of CXCR4 receptor status on tumors

8 Imaging CXCR4 Receptor Expression During Metastases ex-vivo µ-autoradiography: lung of a mouse 1h p.i. of nca. CPCR4 with a human SCLC (-1), primary tumor on the shoulder

9 Imaging of CXCR4 expression in lung metastases µ-autoradiography of lungs 1 h p.i. of 124 I-CPCR4 thickness: 1mm

10 Quantification of CPCR4-Binding ex-vivo MicroImager analysis of the lung of mice with s.c. growing metastasized SCLC tumor at 1h p.i.. Quantitative activity profile along an arbitrary selected direction

11 CXCR4 expression in lung metastases: blockade µ-autoradiography of lungs 1 h p.i. of 124 I-CPCR4 + xs CPCR4 124 I-CPCR4 + xs CPCR4 R = 6/1

12 Biodistribution 125 I-CPCR4 2h p.i. Biodistribution of 125 I-CPCR4 in mice with -1 tumors 2 h p.i. 125 I-CPCR4 2 h p.i uptake (%ID/g) 0 Blood eart Lungs Muscle Pancreas Spleen Stomach Intestine Kidney Liver Tumor

13 Acylated rn-analogs potential ligands for 18 F-labelling- Derivatized cyclic pentapeptides: First candidate for 18 F-Radiolabeling Determination of logp value: 1.07 igh lipophilicity of [ 18 F]-FB-D25 will lead to high uptake in liver and intestine Unfavourable biodistribution restricts use for future tumor imaging; further modification necessary.

14 Synthesis of cyclic pentapeptides with spacer at rn For 18 F-labeling via aminoreactive prosthetic groups and simultaneous increase of hydrophilicity (e.g. PA-linker): 2 Ahx IC 50 : ~ 60 nm PA IC 50 : ~ 80 nm 2 2

15 Improved CPCR4 probe: 68 Ga-DTA-CPCR4-2.1 uptake (%ID/g) Biodistribution in mice with -1 tumors 2 h Ga Blood eart Lungs Muscle Pancreas Spleen Stomach Intestine Kidney Liver Tumor

16 68 Ga-CPCR4-2.1: imaging in -1 tumor bearing mice max. intensity in kidney-slices max. intensity in tumor/bladder-slices max. intensity in kidney/bladder-slices Animal 1: -1 SCLC, Injected with Ga-68-CPCRx (scale max: 177) max. intensity in kidney-slices max. intensity in tumor-slices max. intensity in kidney/bladder-slices Animal 2: -1 SCLC, Injected with Ga-68-CPCRx and xs cold peptide (scale: max 126)

17 Ga-68-labeled Ligands R 2 R: DTA n DTA 33 DTA DTA 36 DTA DTA: free: a M M: In 3+ b Ga 3+ c DTA 37 IC 50 [nm] a compd n a b c ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± ± > ± ± ±

18 Selection of tumor models to prepare first clinical studies SKBR3 DU145 MCF7 MDAMB A431 T29 MDAMB 435

19 3D Surface Plot of Immunostained Section for CXCR4 rabbit UMB-2 monoclonal antibody has been analyzed using I Image Software. Distribution of the UMB-2 antibody around the membrane of positive -1 tumour cells. The intensity of the signal has been then plotted to create a 3D Surface Plot Image of the section 400x

20 Primary -1 Tumor Stained with UMB-2 Monoclonal Antibody

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22 Dimeric DTA labeled peptides FC131 4 Peptide P: R/L R: X 2 X: ; active a X: ; inactive i 2 Gua 2 Linker L: P 1 P 2 n m m Ac Gma Demmer. et al. (submitted)

23 Dimeric DTA labeled peptides 2 2 Peptide L P 1 P 2 R m n IC 50 [nm] a ± 1 a a ± 4 a a ± 2 a a ± 1 a a ± 1 a a ± 1 a a ± 7 a a

24 Dimeric DTA labeled peptides 2 2 X X n comp X n M IC 50 [nm] ± In ± ± In ± 9 25 Gly 0-38 ± 2 26 Gly 0 In ± 2 27 Gly 1-12 ± 2 28 Gly 1 In ± 5 Ga Demmer. et al. (submitted)

25 Dimeric DTA Complexes 2-68 Ga nM Brohl F. et al. (in progress)

26 A Triazacyclononane-Based Bifunctional Phosphinate Ligand for the Preparation of Multimeric 68Ga Tracers for Positron Emission Tomography otni J et al Chem. Eur. J. 2010

27 A Triazacyclononane-Based Bifunctional Phosphinate Ligand for the Preparation of Multimeric 68Ga Tracers for Positron Emission Tomography otni J et al Chem. Eur. J. 2010

28 Immunoimaging of CXCR4 expression in brain tumor xenografts using SPECT/CT (immagadda et al, J ucl Med, 2009) 37 MBq 125 I-12G5 SPECT/CT of CXCR4 expression levels in experimental brain tumors using 125 I- labeled anti-cxcr4 mabs, hcxcr4 (12G5) and the control IgG 2A MAb 37 MBq 125 I-IgG2A SPECT/CT of CXCR4 in U87 xenografts and MIP-image of 125 I-12G5 injected mouse at 48 h after injection. The feasibility of the RID of CXCR4 expression imaging of the tumor microenvironment with a MAb is possible. Biodistribution in U87 tumor-bearing SCID mice. 74 kbq of 125 I-12G5 or 125 I-IgG2A at 24, 48, and 72 h

29 U87-stb- CXCR4 mice (90 min p.i.) immagadda S, Cancer Res (2010)

30 MDA-MB-231 (solid arrow) and DU4475 mice (open arrow)

31 Flourescent Imaging of bladder cancer ishizawa et al., Int.J Cancer 2010 (Ac-Arg-Arg-al-Cys-Tyr-Cit-Arg-D-Lys*-Pro-Tyr-Arg-Cit-Cys-Arg- 2 ; D-Lys* indicates the carboxyfluorescein-labeled D-Lys IC 50 : 11nM

32 A CXCR4 antagonist CTCE-9908 inhibits primary tumor growth and metastasis of breast cancer (uang et al, Journal of Surgical Research, 2009) (25 mg/kg, injected subcutaneously 5 d/wk KGVSLSYR K-2 KGVSLSYR CTCE-9980 structure Inhibition of CXCR4 reduced the primary tumor growth of MDA-MB-231 cells implanted into the inguinal mammary fat pad. (6 weeks) In this model, MDA-231-BSC12 cells were injected into the left cardiac ventricle to produce bone metastases. provide an sensitive method to follow the progression of the primary and metastases over time