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89Zr-Labeled N-suc-desferrioxamine-conjugated anti-carbonic anhydrase IX chimeric monoclonal antibody cG250-F(ab’)2 fragments

[89Zr]-cG250-F(ab')2
, PhD
National Center for Biotechnology Information, NLM, NIH, Bethesda, MD 20894

Created: ; Last Update: October 14, 2010.

Chemical name:89Zr-Labeled N-suc-desferrioxamine-conjugated anti-carbonic anhydrase IX chimeric monoclonal antibody cG250-F(ab’)2 fragments
Abbreviated name:[89Zr]-cG250-F(ab')2
Synonym:
Agent Category:Antibody
Target:Carbonic anhydrase IX
Target Category:Enzyme
Method of detection:Positron emission tomography (PET)
Source of signal / contrast:89Zr
Activation:No
Studies:
  • Checkbox In vitro
  • Checkbox Rodents
Structure not available in PubChem.

Background

[PubMed]

Hypoxic tumors are often resistant to radio- and chemotherapy, have a high metastatic potential, and predict a poor outcome for the cancer patient (1). Although several methods (invasive and noninvasive) are available for the detection of hypoxia in tumors, including the use of radiolabeled small molecules, but these methods are not completely reliable because they either yield variable diagnoses, have functional limitations, show incomplete penetration of tumors, do not detect hypoxia in all tumor types, etc (1, 2). A common feature of most solid cancerous tumor types is the presence of hypoxic conditions (3) and the overexpression of carbonic anhydrase IX (CA IX), a transmembrane cell surface enzyme that is known to regulate the pH and adhesion of tumor cells (4). Therefore, as an alternative to the currently available agents used to detect hypoxic tumors, targeting the CA IX as a hypoxia biomarker is of great interest to investigators (1-3, 5). As an alternative to small molecules, the chimeric monoclonal antibody (cMab) cG250, directed against the CA IX, was developed, labeled with 131I, and evaluated as an immunotherapeutic agent for the detection and treatment of renal cell carcinoma (RCC) (6). This cMab was subsequently labeled with other nuclides (such as 89Zr, 177Lu, 90Y, etc.) and used in preclinical studies in rats (7) and in clinical trials for the radioimmunotherapy of RCC (6). However, only minor responses were observed in the clinical investigations and dose escalation studies are ongoing (6).

The ability of radiolabeled antibodies (Abs) used to detect or treat cancer is limited because these agents show only a peripheral penetration of solid tumors (due to large size, ~150 kDa) and leave many malignant cells untreated (8). In addition, Abs have a long blood circulation time and present a high radiation-absorbed dose to the bone marrow (9). It has been shown that, compared to intact Abs, the smaller monovalent Fab (~50kDa) or the divalent F(ab’)2 (~100 kDa) fragments derived from the parent Ab have a better tumor penetration and a shorter circulating half-life and are therefore likely to yield better results if used to detect or treat solid malignant tumors (8). With these observations in mind, a divalent F(ab’)2 fragment of cG250 was developed, labeled with 131I, and compared with the intact 131I-cG250 Ab for its pharmacokinetic behavior and tumor-targeting ability in mice and RCC patients (9). However, the investigators concluded that the intact Ab was superior to the divalent fragment for targeting the RCC tumors. Recently, the cG250-F(ab’)2 fragments were labeled with 89Zr (to produce [89Zr]-cG250-F(ab’)2) and evaluated for the visualization of tumor hypoxia in mice bearing SCCNij3 cell xenograft tumors (of human head and neck squamous cell carcinoma origin) with positron emission tomography (PET) (5).

Other Sources of Information

Clinical trials on carbonic anhydrase IX inhibitors

Human carbonic anhydrase IX in Entrez Gene (Gene ID 768)

Protein and mRNA sequence of human carbonic anhydrase IX

Crystal structure of the human carbonic anhydrase IX catalytic domain

Human carbonic anhydrase IX in Online Mendelian Inheritance in Man (OMIM) database

Hypoxia response in National Cancer Institute-Nature Pathways Interaction Database

Synthesis

[PubMed]

The production and labeling of the cG250-F(ab’)2 fragment with 89Zr (using N-suc-desferrioxamine as the chelating agent) was described in detail by Hoeben et al. (5). Labeling efficiency of the labeling reaction was reported to be 60% as determined with instant thin-layer chromatography (ITLC). Based on an ITLC analysis, the radiochemical purity of [89Zr]-cG250-F(ab’)2 was >95% with a specific activity of 0.41 MBq/μg (11.07 μCi/μg). Stability of the labeled Ab fragments was not reported.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

The immunoreactive fraction of [89Zr]-cG250-F(ab’)2 was determined to be 74% using SK-RC-52 tumor cells (derived from human metastatic RCC (10)) under in vitro conditions (5).

Animal Studies

Rodents

[PubMed]

In an ex vivo study to determine the correlation of spatial distribution of [89Zr]-cG250-F(ab’)2 to the microscopic distribution of tumor cells expressing CA IX, investigators injected mice bearing SCCNij3 cell xenograft tumors with the tracer; 4 h later, just before euthanization, the animals were injected with the pimonidazole stain (a hypoxia marker) and the Hoechst 33342 dye (a perfusion marker) (5). The tumors were removed from the animals, and an ex vivo autoradiographic analysis (for [89Zr]-cG250-F(ab’)2 uptake) and a histochemical analysis (for CA IX staining using an anti-CA IX Ab) of the tumors showed there was a significant (P < 0.0001) correlation (r = 0.57–0.74) between the CA IX staining and radioactivity uptake at both time points. A significant (P < 0.0001) correlation (r = 0.46–0.68) between pimonidazole staining and tracer uptake of the tumor was also reported at the two time points. In addition, a significant (P = 0.0067) correlation (r = 0.93) between the tumor maximum standardized uptake value and the tumor uptake determined ex vivo was reported. A similar correlation between CA IX and pimonidazole staining of tumor sections, respectively, and radioactivity accumulated in the tumor sections was also observed.

Mice bearing SCCNij3 cell xenograft tumors (n = 4 animals/time point) were injected intravenously with [89Zr]-cG250-F(ab’)2, and small-animal PET images were acquired at 4 h and 24 h post-injection (p.i.) (5). Tumors were clearly visible in the PET images at 4 h p.i., but the tumor image intensity was reduced by 24 h p.i. The percent injected dose per gram tissue (% ID/g) values for the tumor, blood, and muscles were ~3.75% ID/g, 4.50% ID/g, and 0.50% ID/g, respectively, at 4 h p.i. These values were reduced to ~1.60% ID/g, 0.20% ID/g, and 0.25% ID/g for the tumor, blood, and muscles, respectively, at 24 h p.i.

From these studies, the investigators concluded that [89Zr]-cG250-F(ab’)2 was a suitable tracer for the detection of head and neck xenograft tumors in a murine model. In addition, the accumulation of [89Zr]-cG250-F(ab’)2 in the tumor correlated well with the expression of CA IX in these tissue (5).

Other Non-Primate Mammals

[PubMed]

No references are currently available.

Non-Human Primates

[PubMed]

No references are currently available.

Human Studies

[PubMed]

No references are currently available.

Supplemental Information

[Disclaimers]

No information is currently available.

References

1.
Michalski, M.H. and X. Chen, Molecular imaging in cancer treatment. Eur J Nucl Med Mol Imaging. 2010. [PMC free article: PMC3022114] [PubMed: 20661557]
2.
Mees G., Dierckx R., Vangestel C., Van de Wiele C. Molecular imaging of hypoxia with radiolabelled agents. Eur J Nucl Med Mol Imaging. 2009;36(10):1674–86. [PMC free article: PMC2758191] [PubMed: 19565239]
3.
Carlin S., Khan N., Ku T., Longo V.A., Larson S.M., Smith-Jones P.M. Molecular targeting of carbonic anhydrase IX in mice with hypoxic HT29 colorectal tumor xenografts. PLoS One. 2010;5(5):e10857. [PMC free article: PMC2877709] [PubMed: 20523727]
4.
De Simone G., Supuran C.T. Carbonic anhydrase IX: Biochemical and crystallographic characterization of a novel antitumor target. Biochim Biophys Acta. 2010;1804(2):404–9. [PubMed: 19679200]
5.
Hoeben B.A., Kaanders J.H., Franssen G.M., Troost E.G., Rijken P.F., Oosterwijk E., van Dongen G.A., Oyen W.J., Boerman O.C., Bussink J. PET of hypoxia with 89Zr-labeled cG250-F(ab')2 in head and neck tumors. J Nucl Med. 2010;51(7):1076–83. [PubMed: 20554724]
6.
Stillebroer A.B., Oosterwijk E., Oyen W.J., Mulders P.F., Boerman O.C. Radiolabeled antibodies in renal cell carcinoma. Cancer Imaging. 2007;7:179–88. [PMC free article: PMC2151324] [PubMed: 18055291]
7.
Brouwers A., Verel I., Van Eerd J., Visser G., Steffens M., Oosterwijk E., Corstens F., Oyen W., Van Dongen G., Boerman O. PET radioimmunoscintigraphy of renal cell cancer using 89Zr-labeled cG250 monoclonal antibody in nude rats. Cancer Biother Radiopharm. 2004;19(2):155–63. [PubMed: 15186595]
8.
Schmidt M.M., Wittrup K.D. A modeling analysis of the effects of molecular size and binding affinity on tumor targeting. Mol Cancer Ther. 2009;8(10):2861–71. [PMC free article: PMC4078872] [PubMed: 19825804]
9.
Brouwers A., Mulders P., Oosterwijk E., Buijs W., Corstens F., Boerman O., Oyen W. Pharmacokinetics and tumor targeting of 131I-labeled F(ab')2 fragments of the chimeric monoclonal antibody G250: preclinical and clinical pilot studies. Cancer Biother Radiopharm. 2004;19(4):466–77. [PubMed: 15453961]
10.
Gastl G., Ebert T., Finstad C.L., Sheinfeld J., Gomahr A., Aulitzky W., Bander N.H. Major histocompatibility complex class I and class II expression in renal cell carcinoma and modulation by interferon gamma. J Urol. 1996;155(1):361–7. [PubMed: 7490887]

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