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99mTc-Labeled rituximab, a chimeric murine/human anti-CD20 monoclonal antibody

[99mTc]Rituximab
, PhD
National Center for Biotechnology Information, NLM, Bethesda, MD 20894

Created: ; Last Update: October 25, 2012.

Chemical name:99mTc-Labeled rituximab, a chimeric murine/human anti-CD20 monoclonal antibody
Abbreviated name:[99mTc]Rituximab
Synonym:MabThera®; Rituxan®
Agent Category:Antibody
Target:Cell differentiation antigen 20 (CD20)
Target Category:Antigen
Method of detection:Single-photon emission tomography (SPECT); gamma planar imaging
Source of signal / contrast:99mTc
Activation:No
Studies:
  • Checkbox In vitro
  • Checkbox Humans
Structure not available in PubChem.

Background

[PubMed]

Rituximab is a chimeric murine/human monoclonal antibody (mAb) that targets the CD20 antigen (also known as membrane-spanning 4-domains, subfamily A, member 1; or MS4A1), which is expressed on 95% of the transformed B-cells that participate in the pathogenesis of non-Hodgkin lymphomas (NHL; diffused large-B cell, low-grade, or the follicular types), a hematological malignancy, and autoimmune diseases such as rheumatoid arthritis (RA), granulomatosis with polyangiitis (Wegener's granulomatosis), and microscopic polyangiitis. The CD20 antigen is not expressed on any other hematopoietic cells and is not shed for circulation into the plasma or internalized by the cells (1). Therefore, the CD20 antigen is considered to be an excellent target for the treatment of the various B-lymphocyte, cell-based diseases such as those mentioned above. The United States Food and Drug Administration has approved the use of rituximab for the treatment of NHL and the other diseases mentioned above. The exact mechanism of action of rituximab has been discussed by Maloney (2). Although much is known about the pathogenesis of the different forms of NHL at the molecular level, the prognosis for an individual suffering from this disease is based on the morphological and histological information obtained with invasive methods used on the patients (3). As an alternative to the invasive methods, noninvasive procedures, such as imaging with radiolabeled agents using single-photon emission computed tomography (SPECT) or positron emission tomography, are attractive options to screen for patients who can benefit the most from an anti-cancer treatment, and the same imaging technique(s) can be used to monitor and assess the efficacy of a treatment (4).

Rituximab was labeled with 99mTc ([99mTc]rituximab) and used with SPECT for the imaging of NHL (1) and several inflammatory autoimmune diseases such as RA, Behcet’s disease, sarcoidosis, and others, in humans (5).

Synthesis

[PubMed]

Rituximab was purified from a commercial formulation of the mAb and labeled with 99mTc using a photo-activation procedure as described by Stopar et al. (6). The radiolabeling yield of the reaction was >95% as determined with thin-layer chromatography (TLC) and size-exclusion high performance liquid chromatography. The radiochemical purity (RCP) and specific activity of [99mTc]rituximab were not reported.

In another study, [99mTc]rituximab was prepared with a direct labeling method for use in humans (5). The labeling efficiency of the reaction was reported to be >98%, and the preparation contained <2% colloids as determined with instant TLC. The labeled mAb was used in humans without further purification. The specific activity of [99mTc]rituximab was 3.5–3.7 GBq/6.66 nmol (94.5–100 mCi/6.66 nmol).

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

After storage at room temperature for 24 h, the RCP of [99mTc]rituximab was reported to be 95%–97.2% as determined with TLC (6). The tracer had an RCP of 89% after storage in serum (source not mentioned) for 24 h at 37°C (6).

The [99mTc]rituximab preparation was reported to have an immunoreactive fraction of 93.5% as determined in a binding assay with Ramos cells (6). The dissociation constant of [99mTc]rituximab was 2.9 nM, and the cells had a maximum binding capacity of 0.98 pmol labeled mAb per 106 cells, indicating that ~6 × 105 molecules of the tracer could bind to each cell (6). The binding of [99mTc]rituximab to Ramos cells increased from 4.7% of the total applied radioactivity to 23.9% at 4 h at 37o C and approximately 5% of the total added radioactivity was internalized by the cells (incubated with cell growth medium) (6). By comparison, the binding on the control cells was only 12.6% of the total added tracer at 4 h with almost no internalization (control cells were incubated with [99mTc]rituximab on ice for the same duration as the Ramos cells, in phosphate-buffered saline (pH 7.0) containing 1% human serum albumin) (6).

Animal Studies

Rodents

[PubMed]

No publication is currently available.

Other Non-Primate Mammals

[PubMed]

No publication is currently available.

Non-Human Primates

[PubMed]

No publication is currently available.

Human Studies

[PubMed]

The use of [99mTc]rituximab was evaluated to assess the expression of CD20 in NHL patients (n = 10 individuals); nine patients had CD20+ B-cell NHL; one patient had CD20 NHL. Among the ten patients, five individuals had follicular NHL and diffuse large cell B-lymphoma, respectively (1). The patients were intravenously administered 500–600 MBq (18.5–22.2 mCi) [99mTc]rituximab, and whole-body scintigraphic images were acquired from the individuals at 6 h and 20 h postinjection (p.i.). To study early biodistribution of the tracer, additional images were acquired from three patients at 1 h and 3–4 h p.i. At 1 h p.i. and 3 h p.i., the images showed that the radioactivity was present mainly in the blood and the liver of the patients, but at 3 h p.i. some accumulation was also visible in the kidneys and the spleen (1). At 3 h p.i., one patient with a splenic lymphoma showed high uptake of the tracer in the spleen. In three patients, the tumors were clearly visible at 20 h p.i. compared with the earlier time point. From this study, the investigators concluded that [99mTc]rituximab was probably a suitable imaging agent to assess the expression of CD20 and to visualize spread of the B-cell NHL disease in humans (1).

In another study, the use of [99mTc]rituximab was assessed for the imaging of B-lymphocyte infiltration in the affected tissues of patients with chronic inflammatory autoimmune disease (n = 20 individuals) (5). The patients were given an intravenous injection of 350–370 MBq (9.5–10 mCi; 600 pmol) [99mTc]rituximab, and whole-body SPECT images were acquired from the individuals at 6 h p.i. and 20 h p.i. The label was observed to accumulate rapidly in the spleen, and the radioactivity was excreted primarily through the urinary route in all the patients. At 6 h p.i., the inflamed joints were clearly visible in the patients with RA; the salivary and lacrimal glands showed variable uptake of radioactivity in patients with Sjögren’s syndrome; moderate lung uptake was evident in a patient with sarcoidosis; a patient with Behçet’s disease showed increased accumulation of radioactivity in the oral mucosa; moderately high uptake of label was evident in the neck cartilage of a patient with polychondritis; and a moderate to high accumulation of the tracer was observed in the skin of dermatopolymyositis patients (5). From this study, the investigators concluded that [99mTc]rituximab can probably be used to assess the infiltration of B lymphocyte in the affected organs of patients suffering from the different types of autoimmune diseases (5).

Supplemental Information

[Disclaimers]

No information is currently available.

References

1.
Gmeiner Stopar T., Fettich J., Zver S., Mlinaric-Rascan I., Hojker S., Socan A., Peitl P.K., Mather S. 99mTc-labelled rituximab, a new non-Hodgkin's lymphoma imaging agent: first clinical experience. Nucl Med Commun. 2008;29(12):1059–65. [PubMed: 18987526]
2.
Maloney D.G. Anti-CD20 antibody therapy for B-cell lymphomas. N Engl J Med. 2012;366(21):2008–16. [PubMed: 22621628]
3.
Natarajan A., Gowrishankar G., Nielsen C.H., Wang S., Iagaru A., Goris M.L., Gambhir S.S. Positron Emission Tomography of (64)Cu-DOTA-Rituximab in a Transgenic Mouse Model Expressing Human CD20 for Clinical Translation to Image NHL. Mol Imaging Biol. 2012;14(5):608–16. [PubMed: 22231277]
4.
Willmann J.K., van Bruggen N., Dinkelborg L.M., Gambhir S.S. Molecular imaging in drug development. Nat Rev Drug Discov. 2008;7(7):591–607. [PubMed: 18591980]
5.
Malviya G., Anzola K.L., Podesta E., Lagana B., Del Mastro C., Dierckx R.A., Scopinaro F., Signore A. (99m)Tc-labeled Rituximab for Imaging B Lymphocyte Infiltration in Inflammatory Autoimmune Disease Patients. Mol Imaging Biol. 2012;14(5):637–46. [PMC free article: PMC3443359] [PubMed: 22127469]
6.
Stopar T.G., Mlinaric-Rascan I., Fettich J., Hojker S., Mather S.J. (99m)Tc-rituximab radiolabelled by photo-activation: a new non-Hodgkin's lymphoma imaging agent. Eur J Nucl Med Mol Imaging. 2006;33(1):53–9. [PubMed: 16172899]

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