99mTc-Labeled murine IgM monoclonal antibody, fanolesomab, that targets the CD15 glycoprotein antigen

99mTc-Fanolesomab

Chopra A.

Publication Details

Image

Table

In vitro Humans

Background

[PubMed]

Infection is one of the most common causes of morbidity in patients, especially for those who have undergone invasive surgery, and nuclear tracers such as 67Ga-citrate and autologous leukocytes labeled with 111In-oxine- or 99mTc-hexamethylpropyleneamine oxime (111In- or 99mTc-HMPAO) are often used to localize the infection to initiate treatment (1, 2). However, these tracers have limitations. 67Ga-Citrate produces low-resolution images with a gamma camera and shows high accumulation in the liver, colon, genitals, and the urinary tract (2). To use 111In- or 99mTc-labeled HMPAO, leukocytes from the patient have to be isolated, labeled ex vivo with the radiolabeled compounds, and reinfused into the patient to detect the infection with scintigraphy. This procedure has to be performed by a skilled technician, may not always be convenient, is time-consuming, and can be risky for the technician as well as the patient (2). Investigators have also used 111In- or 99mTc-labeled human immunoglobulin or 123I-interleukin to detect infections, but these agents tend to accumulate in the interstitial spaces, have low uptake at the site of infection, and are not always able to distinguish between infections and inflammation (1). Radionuclide-labeled peptides such as ubiquicidin or its derivatives (3), P483H-99mTc (4) and 99mTc-P1827DS (5), have been used to detect infections under preclinical or clinical conditions, but none of these agents are approved by the United States Food and Drug Administration (FDA) as agents to detect infection.

Researchers have also evaluated several radiolabeled murine monoclonal antibodies (mAb) that target either the cluster of differentiation 67 (CD67) antigen or the CD15 antigen (e.g., anti–stage-specific embryonic antigen-1 (anti-SSEA-1), also known as fanolesomab) on neutrophils to detect infections in humans (for details, see Shanthly et al. (1)). However, the anti-CD67 mAbs had a low affinity for the target (Kd = 10−8 M) and required 24 h imaging to obtain proper diagnosis, and a high percentage of the patients developed antibodies against the mouse antibodies. 99mTc-Sulesomab (LeukoScan), the Fab’ fragment of IMMU-MN3, a murine mAb, is approved in some European countries for the detection of infection and inflammation in the bones of patients with osteomyelitis (6). In 2004, 99mTc-labeled fanolesomab (now known as NeutroSpec®) was approved by the FDA for use in the United States as a diagnostic radiopharmaceutical for scintigraphic imaging of patients (>5 years of age) exhibiting signs and symptoms of appendicitis (the clinical trial information is not available on the www.clinicaltrial.gov web site). This mAb binds to the 3-lactose-N-fucopentose carbohydrate that defines the CD15 antigen that is expressed on the surface of human neutrophils (1). In 2005, the marketing of 99mTc-fanolesomab was suspended in the United States because, postmarketing, two patients died and an additional 15 patients experienced life-threatening cardiopulmonary events immediately after receiving this radiochemical. The current marketing status of 99mTc-fanolesomab is not clear.

Synthesis

[PubMed]

The preparation of 99mTc-fanolesomab from a freeze-dried kit that was commercially available in the United States has been described by Tronco et al. (7). Briefly, the mAb was supplied in a vial containing maltose monohydrate, sodium potassium tartrate tetrahydrate, succinic acid, stannous tartrate, glycine, and disodium edetate dihydrate. Before use, the contents of the vial were reconstituted with 0.2–0.35 mL 99mTc-pertechnetate (740–1480 MBq (~2–4 mCi)), incubated at 37°C for 30 min, and the final volume was brought up to 1 mL with 500 mg/mL ascorbic acid. The final product had a radiochemical purity of >90% as determined with thin-layer chromatography. The specific activity of the labeled mAb was 5.5–7.4 GBq/0.66 μmol (135–200 mCi/μmol). The radiolabeled mAb was used within 6 h after synthesis.

Thakur et al. isolated anti-SSEA-1 mAb from a hybridoma cell line (MCA-480) and labeled it with 99mTc using two different methods (8). For the first procedure, the mAb was conjugated with diethylenetriamine pentaacetic acid (DTPA) and then labeled with 99mTc (8). In the second procedure, the anti-SSEA-1 mAb was labeled directly with the radionuclide. The radiochemical yields with the two procedures were reported to be 94.8 ± 1.4% and 95.8 ± 3.5%, respectively, and the amount of colloids in both preparations was <5%. The radiochemical purity and specific activities of the mAb labeled with the two procedures were not reported. The final product from both procedures was formulated in 0.9% sodium chloride containing 3.0% human serum albumin and sterilized by passing through a 0.22-micron filter. The Rf value of the labeled mAb on instant thin-layer chromatography was reported to be 1.0, and the Rf of the colloid fraction was 0.0 (8).

In another study (9), the anti-SSEA-1 mAb was labeled with 99mTc after conjugation with DTPA (the DTPA:mAb ratio was determined to be 1), and the 99mTc-labeled mAb had a radiochemical purity of at least 80% as determined with high-performance liquid chromatography. The radiochemical yield and specific activity of the final product were not reported.

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

In an effort to develop an animal model to study the biological characteristics of 99mTc-anti-SSEA-1, human, rat, guinea pig, dog, rabbit, sheep, and pig polymorph nuclear leukocytes (PMN) were exposed to the labeled mAb under in vitro conditions (9). Only the human PMNs were reported to bind the radiolabeled mAb (80 ± 5%), and PMNs from the other animal species bound only ~3% of the tracer. This indicated that the mAb had a high binding specificity for the human cells (9). On the basis of this observation, the investigators concluded that this mAb could be used only in humans.

In another study, 99mTc-anti-SSEA-1 mAb was shown to have a Kd of 1.6 × 10−11 M for human neutrophils and could bind to 5.1 × 105 antigenic sites on these cells (9).

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]

Twelve patients with clinically confirmed inflammation were intravenously injected with 99mTc-anti-SSEA-1 mAb, and serial anterior and posterior spot-views and whole-body single-photon emission computed tomography (SPECT) scans were acquired for all patients up to 3 h postinjection (p.i.) (8). Infections (perforated appendicitis, osteomyelitis, abscess, and inflammation) were visible in all the patients within 3 h p.i., and 11 of these infections were confirmed with other modalities. Biodistribution of the tracer in the patients was determined as described elsewhere (8). Radioactivity accumulated in each organ was presented as a percentage of the geometric mean of the total body counts. The highest uptake of radioactivity was observed in the liver (49.9 ± 3.2%), followed by the spleen (7.7 ± 0.1%), kidneys (2.4 ± 0.03%), and the bladder (1.3 ± 0.4%). At 3 h p.i., the amount of label bound to the PMNs of the patients varied from 15% to 51%, and the amount of label in the whole blood was <25% at this time point.

Tronco et al. reported that SPECT imaging with 99mTc-fanolesomab at 18–30 h p.i. can be used to detect prosthetic vascular graft infection with 95% accuracy (7).

A recent report describes the use of 99mTc-fanolesomab with SPECT for the imaging of five patients exhibiting renal allograft complications (10). It is pertinent to mention that it is assumed this study is published now, but it was performed before 99mTc-fanolesomab was withdrawn for marketing in the United States. Two patients had very high uptake in the transplant kidney compared to the lower lumbar spine (used as control tissue for this study), and the patients were determined to have pyelonephritis. One individual who had acute renal failure due to secondary humoral rejection showed a similar accumulation of the tracer in the transplant kidney and the lower lumbar spine. One patient showed a high level of radioactivity in the pelvis of the transplant kidney, indicating normal excretion of the free 99mTc-pertechnetate by the organ. Another patient, who was off immunosuppressive therapy and on maintenance hemodialysis for 4 months, had an uptake of the label that was comparable to the lumbar spine, indicating there was chronic rejection of the transplant kidney in this individual. From this study, the investigators concluded that SPECT scans with 99mTc-fanolesomab can probably be used to distinguish a normal functioning allograft kidney from an infected or a rejected transplant kidney. However, they caution that this procedure will have to be evaluated in a large number of patients, including a large control group, before it can be used as a diagnostic technique in the clinics.

Supplemental Information

[Disclaimers]

No supplemental information is currently available.

NIH Support

References

1.
Shanthly N., Aruva M.R., Zhang K., Mathew B., Thakur M.L. 99mTc-Fanolesomab: affinity, pharmacokinetics and preliminary evaluation. Q J Nucl Med Mol Imaging. 2006;50(2):104–12. [PubMed: 16770300]
2.
Love C., Tronco G.G., Palestro C.J. Imaging of infection and inflammation with 99mTc-Fanolesomab. Q J Nucl Med Mol Imaging. 2006;50(2):113–20. [PubMed: 16770301]
3.
Melendez-Alafort L., Nadali A., Pasut G., Zangoni E., De Caro R., Cariolato L., Giron M.C., Castagliuolo I., Veronese F.M., Mazzi U. Detection of sites of infection in mice using 99mTc-labeled PN(2)S-PEG conjugated to UBI and 99mTc-UBI: a comparative biodistribution study. Nucl Med Biol. 2009;36(1):57–64. [PubMed: 19181269]
4.
Chopra, A., Acetyl-lys-lys-lys-lys-lys-cys-gly-cys-gly-gly-pro-leu-tyr-lys-lys-ile-ile-lys-lys-leu-leu-glu-ser-heparin-[99mTc] (P483H-99mTc). Molecular Imaging and Contrast agent Database (MICAD) [database online]. National Library of Medicine, NCBI, Bethesda, MD, USA. Available from www​.micad.nih.gov, 2004 -to current. [PubMed: 20641948]
5.
Chopra, A., 99mTc]-Acetyl-arg-arg-arg-arg-arg-cys-gly-cys-gly-gly-pro-leu-tyr-arg-arg-ile-ile-arg-arg-leu-leu-glu-ser-dermatan sulfate (99mTc-P1827DS). Molecular Imaging and Contrast agent Database (MICAD) [database online]. National Library of Medicine, NCBI, Bethesda, MD, USA. Available from www​.micad.nih.gov, 2004 -to current. [PubMed: 20641659]
6.
Quigley A.M., Gnanasegaran G., Buscombe J.R., Hilson A.J. Technetium-99m-labelled sulesomab (LeukoScan) in the evaluation of soft tissue infections. Med Princ Pract. 2008;17(6):447–52. [PubMed: 18836272]
7.
Tronco G.G., Love C., Rini J.N., Yu A.K., Bhargava K.K., Nichols K.J., Pugliese P.V., Palestro C.J. Diagnosing prosthetic vascular graft infection with the antigranulocyte antibody 99mTc-fanolesomab. Nucl Med Commun. 2007;28(4):297–300. [PubMed: 17325593]
8.
Thakur M.L., Marcus C.S., Henneman P., Butler J., Sinow R., Diggles L., Minami C., Mason G., Klein S., Rhodes B. Imaging inflammatory diseases with neutrophil-specific technetium-99m-labeled monoclonal antibody anti-SSEA-1. J Nucl Med. 1996;37(11):1789–95. [PubMed: 8917176]
9.
Thakur M.L., Richard M.D., White F.W. 3rd. Monoclonal antibodies as agents for selective radiolabeling of human neutrophils. J Nucl Med. 1988;29(11):1817–25. [PubMed: 3141596]
10.
Aygen M., Amirmokri P., Berk F., Nguyen N., Bastani B., Civelek A.C. A preliminary report on 99mTc-fanolesomab imaging of renal allograft. Nucl Med Commun. 2011;32(10):925–8. [PubMed: 21862945]