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Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2013.

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Molecular Imaging and Contrast Agent Database (MICAD) [Internet].

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68Ga-Trastuzumab F(ab’) fragment

68Ga-DCHF

Created: ; Last Update: June 25, 2007.

Chemical name:68Ga-Trastuzumab F(ab’) fragment
Abbreviated name:
Synonym:68Ga-DCHF
Agent Category:Trastuzumab F(ab’) fragment
Target:Human epidermal growth factor receptor
Target Category:Binding
Method of detection:PET
Source of signal:68Ga
Activation:No activation required
Studies:
  • Checkbox In vitro
  • Checkbox Rodents
View human EGFR protein and nucleotide sequence information.

Background

[PubMed]

The epidermal growth factor (EGF) and its receptor (EGFR) are known to play a role in the oncogenic transformation of cells (1). The EGFR is a tyrosine kinase receptor that includes four members, EGFR and three other human EGF receptors (HERs) designated as HER2/ErbB2, HER3/ErbB3, and HER4/ErbB4. The four receptors constitute the HER-kinase axis and regulate cellular responses through complex receptor-ligand interactions. The various signal transduction pathways used by the EGFRs to mediate cell responses have been described by Brandt et al. (2). The various EGFRs are known to be amplified, mutated, or overexpressed in several cancers and are a target for the development of pharmaceutical agents that inhibit the receptor or the signal transduction pathway (3). For this inhibition, a variety of small molecules and antibodies against the EGFRs have been developed (4). However, to achieve therapeutic effects the receptor or signal transduction pathway must be involved in maintenance of the malignant phenotype because HER inhibitor therapy does not necessarily result in treatment of a cancer (5).

The use of antibodies alone against EGFR (e.g., Erbitux; cetuximab) and HER2 (Herceptin; trastuzumab) for the treatment of cancer gave only moderate results; antibodies are most useful when used in combination with chemotherapy (6, 7). Geldanamycin, an ansamycin antibiotic, has a unique mechanism for inhibition of HER2 activity by inducing protesomal degradation of the receptor. The antibiotic binds to heat shock protein 90 (Hsp90), a chaperone protein that is responsible for the maturation and stability of a variety of proteins including HER2 (8). The binding of geldanamycin to Hsp90 inhibits the maturation and stability of HER2 and leads to degradation of the receptor. The geldanamycin derivative 17-N-allylamino-17-demethoxy geldanamycin (17-AAG) was shown to exhibit antitumor activity, sensitize tumors to taxanes (drugs that inhibit cell proliferation by binding to microtubules), induce degradation of HER2, and inhibit expression of the receptor (9). This drug is being studied in several clinical trials for the treatment of various cancers (www.clinicaltrials.gov).

Because 17-AAG inhibits HER2 expression, this characteristic activity of the drug has been used to image the pharmacodynamic effects of the drug with the F(ab´) fragment of an antibody against EGFR. For this, a gallium (68Ga)-labeled F(ab´) fragment of trastuzumab, which is a commercially available antibody against EGFR, was used to determine the alterations of HER2 expression in response to 17-AAG (3). The 68Ga-labeled fragment was selected for this work because it had a 3-h half-life in serum and showed no decrease in signal even 24 h after administration.

Synthesis

[PubMed]

The F(ab´) fragment of trastuzumab was generated by the digestion of trastuzumab with pepsin immobilized on agarose beads (3). Briefly, trastuzumab was diluted with sodium acetate buffer (pH 4.5) and concentrated in a centrifugal concentrator with a 60-kDa cut-off limit. This process was repeated several times and the protein was gently shaken overnight with the pepsin agarose beads. The protein fragments were obtained by removing the agarose beads after centrifugation, concentrated as before, and purified by high-performance liquid chromatography on a Superdex 200 column.

The fragments were modified with 1,4,7,10-tetraazacyclododecane-N,N’,N’’,N’’’-tetraacetic acid (DOTA) by binding one of the four carboxylic acid groups of DOTA to the primary amines in the protein structure as described by Smith-Jones et al. (10). Excess DOTA and other reactants were removed from the DOTA-F(ab´) fragment by repeated washing with sodium acetate buffer using the centrifugal concentration technique. The DOTA-conjugated trastuzumab fragment (DCHF) contained an average of 6.3 DOTA moieties per F(ab´) and were 81% immunoreactive. The fragments were labeled either with radioactive indium (111In) for biodistribution studies or with 68Ga to obtain 68Ga-DCHF for positron emission tomography (PET) investigations.

The radiochemical yield and specific activity of the radiochemical were not provided by the investigators; the investigators reported a linear correlation between radioactive uptake, as quantified by direct assessment of resected tumors by gamma counter, and radionuclide uptake, as estimated by PET imaging (3).

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

No publications are currently available.

Animal Studies

Rodents

[PubMed]

The biodistribution of 68Ga-DCHF was used to monitor the in vivo effects of 17-AAG on HER2 expression in tumor-bearing mice using PET imaging (3). The level of HER2 expression was determined in control mice (without 17-AAG treatment) and 17-AAG–treated mice bearing BT-474 human breast cancer xenografts. The control mice showed a constant or slightly increased HER2 expression by PET imaging over a 24-h period. In contrast, animals treated with 17-AAG showed an 80% reduction in HER2 expression 24 h after treatment, but HER2 increased to 50% of the initial expression up to 7 days after treatment. During the same period the control animals showed a 20% increase in HER2 expression up to 7 days after treatment compared to the pretreatment levels.

A decrease in 68Ga-DCHF uptake by the tumors was observed in response to the 17-AAG treatment, although no change in distribution of the radiochemical was observed in other organs (heart, lung, liver, spleen, etc.). When the radionuclide uptake was adjusted to account for the nonspecific background, the accumulation of radioactivity by the tumors correlated well with the data obtained by immunoblotting and microPET region-of-interest analysis.

The possible use of this technique to image pharmacodynamic effects of antireceptor antibodies and tyrosine kinase and Hsp90 inhibitors was suggested (3).

Other Non-Primate Mammals

[PubMed]

No publications are currently available.

Non-Human Primates

[PubMed]

No publications are currently available.

Human Studies

[PubMed]

No publications are currently available.

References

1.
Gross M.E., Shazer R.L., Agus D.B. Targeting the HER-kinase axis in cancer Semin Oncol 2004. 31 Suppl 3 1:9–20. [PubMed: 15052539]
2.
Brandt B., Meyer-Staeckling S., Schmidt H., Agelopoulos K., Buerger H. Mechanisms of egfr gene transcription modulation: relationship to cancer risk and therapy response. Clin Cancer Res. 2006;12(24):7252–60. [PubMed: 17189396]
3.
Smith-Jones P.M., Solit D.B., Akhurst T., Afroze F., Rosen N., Larson S.M. Imaging the pharmacodynamics of HER2 degradation in response to Hsp90 inhibitors. Nat Biotechnol. 2004;22(6):701–6. [PubMed: 15133471]
4.
Dancey J.E. Recent advances of molecular targeted agents: opportunities for imaging. Cancer Biol Ther. 2003;2(6):601–9. [PubMed: 14688462]
5.
Bates S.E., Fojo T. Epidermal growth factor receptor inhibitors: a moving target? Clin Cancer Res. 2005;11(20):7203–5. [PubMed: 16243788]
6.
Khalil M.Y., Grandis J.R., Shin D.M. Targeting epidermal growth factor receptor: novel therapeutics in the management of cancer. Expert Rev Anticancer Ther. 2003;3(3):367–80. [PubMed: 12820779]
7.
Sledge G.W. Gemcitabine combined with paclitaxel or paclitaxel/trastuzumab in metastatic breast cancer Semin Oncol 2003. 30 Suppl 3 2:19–21. [PubMed: 12722021]
8.
Neckers L. Hsp90 inhibitors as novel cancer chemotherapeutic agents Trends Mol Med 2002. 8 4 Suppl S55–61. [PubMed: 11927289]
9.
Solit D.B., Basso A.D., Olshen A.B., Scher H.I., Rosen N. Inhibition of heat shock protein 90 function down-regulates Akt kinase and sensitizes tumors to Taxol. Cancer Res. 2003;63(9):2139–44. [PubMed: 12727831]
10.
Smith-Jones P.M., Vallabahajosula S., Goldsmith S.J., Navarro V., Hunter C.J., Bastidas D., Bander N.H. In vitro characterization of radiolabeled monoclonal antibodies specific for the extracellular domain of prostate-specific membrane antigen. Cancer Res. 2000;60(18):5237–43. [PubMed: 11016653]
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