Detection of Thrombophlebitis with Inin-Labeled

Home , Thrombophlebitis

[CANCER RESEARCH (SUPPL.) 50. 958s-961s. February I. 1990] Detection of Thrombophlebitis with In In-labeled Anti-Fibrin Antibody: Preliminary Results1

Abass Alavi,2 Naresh Gupta, Harold I. Palevsky, Mark A. Kelley, Allison D. Jatlow, Ann A. Byar, and Harvey J. Berger Division of Nuclear Medicine. Department of Radiology [A. A., N. G., A. D. J., A. A. B., H. J. B.J and Cardiovascular-Pulmonary Division, Department of Medicine Â¡H. I. P., M. A. K.], Hospital of the University of Pennsylvania, Philadelphia, Pennsylvania 19104, and Research and Development Division, Centocor, Inc., Malvern. Pennsylvania 19355 [H. J. BJ

Abstract Assessment of Current Techniques

Deep venous thrombosis remains a major medical problem, affecting The method of choice in the clinical evaluation of deep venous a large segment of the population and resulting in significant mortality thrombosis is ascending contrast venography. Considered the and morbidity. "gold standard" for clot detection, venography is capable of Current techniques available for detecting deep venous thrombosis visualizing 95% of all clots (5, 6). Able to give a detailed view present limitations that may mitigate their potential benefit to the patient. Invasive techniques, such as ascending contrast venography, carry risks of the deep venous system, venography virtually assures detec tion of life-threatening clots in the extremity under study. to the patient with regard to complications such as an allergic reaction to an iodine dye, adverse effects to renal function, and clot formation in The limitations of venography are therefore not related so a normal vein. Noninvasive techniques, such as Doppler ultrasound and much to accuracy as to the materials used in the procedure, in impedance plethysmography, evaluate only a limited segment of the which large quantities of contrast dye, typically between 100 venous bed. The need remains for a diagnostic technique that is safe, and 150 ml, are injected into the patient (7). Patients with accurate, and widely accessible. cardiac or renal dysfunction are not considered to be safe A readily available noninvasive scintigraphic technique utilizing radi- candidates for venography (6, 8). Patients may develop a reac olabeled monoclonal anti-fibrin antibody may overcome some of these tion to the contrast agent which may contain as much as 20% shortcomings. This imaging examination is quite effective in detecting clots in the lower extremities. Compared to contrast venography, '"In- iodine (9). Between 7.5 and 24% of venography patients expe labeled anti-fibrin antibody imaging appears to be as sensitive in identi rience discomfort including pain, swelling, tenderness, or ery fying acute venous thrombosis. In addition, the preliminary data indicate thema (7). One study found that 6.4% of patients had positive that anticoagulation with heparin may interfere with adequate visualiza repeat venograms after an initial negative study (9). tion of the clots with this technique. In addition, because of the technical expertise required to perform the venogram, this study is not accessible to all pa tients. First, a suitable vein must be found; failure to cannulate Introduction a vein prevents a study from being conducted. Inadequate DVT' remains a major medical problem, affecting an esti mixing of contrast materials with the unopacified blood, poor mated 2.5 million patients/year and resulting in significant injection technique, muscle tensing, use of a tourniquet, and morbidity and mortality (1). Often insidious in that they are varicosities can confound interpretations of the study. Further, frequently asymptomatic, DVT and pulmonary embolisms aris in order to avoid incomplete mixing of contrast medium and ing from DVT can elude clinical detection. Autopsy studies blood, large amounts of contrast medium should be used (10). suggest that up to 84% of patients with pulmonary emboli were Since a large quantity of contrast medium can cause complica undiagnosed prior to death (2-4). tions, it would appear that this by itself may introduce new problems. Current invasive and noninvasive diagnostic techniques are In addition to contrast venography, IPG, the '25I-labeled inadequate in visualizing thrombosis in the entire venous system fibrinogen study, Doppler ultrasound, real-time B-mode ultra- in patients suspected of thromboembolism. In addition, invasive studies such as contrast venography carry considerable risks to sonography, and MRI are other techniques for detecting the patient. Because of these shortcomings, the search for a thrombi. These methods are noninvasive and are therefore of technique that is safe, accurate, and widely accessible has con less discomfort and risk to the patient. While each method tinued for a decade. provides helpful information for the clinician regarding specific In this communication, a new noninvasive scintigraphic tech regions of the venous system, each, in turn, has its limitations nique, utilizing radiolabeled monoclonal anti-fibrin antibody, which restrict its usefulness in detecting a wide spectrum of will be discussed for its potential application as an alternative DVT. IPG is a sensitive test for clots in the thigh. Through meas diagnostic tool. After assessment of current diagnostic tech uring changes in the electrical resistance or "impedance" pro niques for the diagnosis of deep venous thrombosis, monoclonal antibody immunoscintigraphy and its advantages over estab portional to changes in blood flow, thrombi may be located lished diagnostic methods will be presented. Preliminary data (11). One disadvantage of this method is its poor sensitivity to obtained from the '"In-labeled anti-fibrin monoclonal antibody- clots in the calf (12). False positive results which can be caused study, conducted at the Hospital of the University of Pennsyl by muscle tension, arterial insufficiency, increased venous pres vania, will then be briefly discussed. sure secondary to congestive heart failure, or venoconstriction due to shock, account for between 7 and 22% of all studies (13). 1Presented at the "Second Conference on Radioimmunodetection and Ra- dioimmunotherapy of Cancer." September 8-10, 1988, Princeton. NJ. Up to 21% of the studies are false negatives, possibly due to 2To whom requests for reprints should be addressed, at Division of Nuclear nonocclusive thrombi which do not completely block blood Medicine. Department of Radiology. 117 Donner Building. Hospital of the flow, collateral circulation, and skin stretching caused by the University of Pennsylvania. .1400 Spruce St., Philadelphia, PA 19104. 3The abbreviations used are: DVT. deep venous thrombosis; IPG. impedance placement of a thigh cuff (13, 14). Dopper ultrasound techniques are based on the Doppler plethysmography: MRI. magnetic resonance imaging: DPTA. diethylenetriam- "shift" or the change in the frequency of a light or sound wave inepentaacetic acid. 958s

Downloaded from cancerres.aacrjournals.org on September 24, 2021. © 1990 American Association for Cancer Research. "In-LABELED ANTI-FIBRIN ANTIBODY DETECTION OF THROMBOPHLEBITIS when reflected from a moving source. A blood clot causes an Every technique mentioned has its limitations which reduce abnormal response due to decreased venous blood flow velocity its effectiveness in the ability to unequivocably locate deep in the area of the clot (13). Like IPG, Doppler ultrasound venous thrombi. Several of the methods described are adequate techniques are excellent in detecting thigh clots (popliteal, in the detection of DVT for specific regions of the body but are femoral, and iliac veins); reported accuracies in Doppler ultra inadequate (i.e., high false positive, false negative percentages) sound for this application are greater than 90% but have a for other regions. For example, impedance plethymography sensitivity of less than 50% in the region of the calf (13, 15). In and Doppler ultrasound are adequate for examining the proxi addition, as with IPG, nonocclusive clots can cause false nega mal venous system but inadequate for the detection of thrombi tive results. in the calves, while i:5I-labeled fibrinogen is sensitive to clots Unlike other noninvasive techniques, 125Ifibrinogen is highly in the calves but not in the thighs. Contrast venography is sensitive (up to 92%) for clots in the region of the calf (16). usually applied in examining only one extremity to reduce This method has its disadvantages as well: dependence on active contrast load and discomfort. thrombus formation in order to detect the presence of a clot; As an invasive technique, contrast venography carries with it interference from surrounding structures such as bladder activ the risk of complications from the procedure including an ity, which results in poor sensitivity to proximal venous throm allergic reaction to the iodine-containing contrast medium, bosis; a waiting period of at least 24 to 48 h between injection while certain medical conditions, such as cardiac and renal and detectable evidence of a clot; and results based on relative dysfunction, render participation in this procedure unsafe. In counting rates from a nonimaging probe, not from -y-scintilla Doppler ultrasound, nonocclusive thrombi can cause false neg tion imaging. ative results, whereas in MRI, the extent of the thrombus is Real-time B-mode ultrasonography produces a two-dimen exaggerated by an area of slow blood flow resembling a clot. sional picture of the structure being examined. A transducer emits and detects sound waves in rapid succession, permitting a "moving picture" to appear on the viewing screen. The time Immunoscintigraphy: A New Noninvasive Technique of echo return and the direction in which the ultrasound beam Considering the limitations of both invasive and noninvasive is reflected are used to construct the two-dimensional image. techniques currently applied by clinicians for the detection of Solid mass areas are characterized by more echoes (hyperech- DVT, it should be evident that a better imaging modality is ogenicity) than fluid-filled areas. Studies using B-mode ultra needed. Immunoscintigraphy, or the imaging of pathological sound to detect deep venous thrombosis have a sensitivity of conditions using radiolabeled antibody and antibody fragments, 100% in the detection of the femoral and iliac thrombi; this is a promising new technology that may help to address the sensitivity drops to 62.5% for calf thrombi (17-19). diagnostic inadequacies encountered in other techniques. False positive results in real-time B-mode ultrasonography The production of monoclonal antibodies originated with can result when the patient has a hematoma, a popliteal cyst, Kohler and Milstein (22) in 1975. A single hybrid cell, stimu lower limb ischemia, cardiac or respiratory insufficiency, and lated to produce a specific antibody in response to a selected obesity (17-19). In addition, this method cannot conclusively antigen, is the parent cell for a colony of genetically identical distinguish between acute and chronic thrombi. Given these daughter cells. This colony or clone is referred to as a "hybrid- limitations, real-time B-mode ultrasonography is often used in oma"; the antibodies generated by the hybridoma are therefore conjunction with other noninvasive studies such as IPG and monoclonal. Monoclonal antibodies are biochemically identical Doppler ultrasound. to one another, since they are produced by genetically identical MRI is an alternative noninvasive technique. MRI is based daughter cells from a single parent cell. on the observed phenomenon that a hydrogen atom will reso The selection of an antigen is a critical component in radioim- nate (produce a signal) at a frequency which is directly related munoimaging study. Fibrin, the biomolecule that forms the to the strength of the magnetic field to which it is exposed. The structural matrix of the blood clot and the biochemical end strength of the signal will also depend on the properties of the product in the process of blood coagulation, was chosen as the material (i.e., the density of hydrogen atoms in a given tissue). antigen in this study. When clot formation is initiated, fibrin is The intensity of the energy emitted, where the timing and produced in relatively large quantities (mg amounts). It is strength of the magnetic field is known at any given point relatively accessible to the antibody, since the clot is produced within the space being scanned, can provide important anatom within the vascular space making it an ideal candidate as a ical information. monoclonal antibody target. Platelets have been used as the The ability of MRI to detect DVT has been investigated; it selected antigen in some studies, but it has been found that has been determined that MRI is an excellent technique from monoclonal anti-platelet antibodies tend to aggregate at only which the resolution of a thrombi can be monitored; however, fresh thrombi sites (23-25). slow blood flow or stasis may create problems in scan interpre Monoclonal anti-fibrin antibodies can be further processed tation (20). Thrombi can have magnetic properties that may from whole antibodies into antibody fragments, which have the differ from other stationary tissue and from flowing blood. The advantage of being less likely to stimulate a harmful immune effects on signal intensity of slow blood flow in proximity to response in the patient. This response is usually referred to as the thrombus may not be measurably different (i.e., below a producing human anti-murine antibody. Some studies have threshold) from that of the thrombus itself, exaggerating its been conducted using whole antibodies (26). In this study, a size. Also, contrast quality is compromised when adjustments Fab fragment (59D8) anti-fibrin antibody was used, in order to are made to decrease this problem. Limited flip angle, gradient- avoid the potential problem of an allergic reaction developing refocused MRI demonstrates some improvements over tradi in the patient. tional spin-echo MRI; this method uses pulse sequences with This antibody (59D8) is fibrin specific and does not react short repetition times and flip angles of less than 90 degrees with fibrinogen (the precursor of fibrin which is biochemically (21). The age of the thrombus may also contribute to the almost identical and therefore potentially confusing to the variability in the MRI image obtained. antibody). This antibody was developed by Hui et al. (27) and 959s

Downloaded from cancerres.aacrjournals.org on September 24, 2021. © 1990 American Association for Cancer Research. "In-LABELED ANTI-FIBRIN ANTIBODY DETECTION OF THROMBOPHLEBITIS was shown to detect clots for up to 4 days in rabbits and up to the need remains for a safe, accurate, and widely available 24 h in dogs, in one study (28). Other studies confirm the technique for diagnosing the condition. The invasive procedure efficacy of immunoscintigraphy to detect the presence of of ascending contrast venography and the noninvasive tech thrombi using the monoclonal anti-fibrin antibody T2G1 (29- niques of impedance plethysmography, Doppler ultrasound, 31). '"Â¡-labeled fibrinogen, real-time B-mode ultrasound, and MRI One potential advantage from using monoclonal antibody lack one or more of the attributes described above. Our results immunoscintigraphy over conventional techniques, such as IPG indicate that the '"In-labeled 59D8 Fab-DPTA monoclonal and Doppler ultrasound, is that it has the capability of detecting antibody is accurate in targeting thrombi and that adoption of the presence of thrombi over the entire body, as opposed to a this technique would not require major investment in either the limited region. Another advantage is that, as a noninvasive purchase of new equipment or the employment of highly skilled procedure, it does not carry the risks associated with an invasive personnel. These studies can be adequately carried out using technique such as contrast venography. The following prelimi the existing facilities of modern nuclear medicine laboratories. nary results indicate the potential usefulness of immunoscintig Further investigation of radioimmunoimaging with monoclonal raphy in detecting DVT. anti-fibrin antibodies may demonstrate that this method pos sesses similar or perhaps greater diagnostic capabilities without the liabilities of established procedures. Preliminary Results

Eighteen patients were given injections of approximately 2.0 References mCi of 59D8 anti-fibrin antibody Fab fragments labeled with '"In in conjugation with the chelating agent DPTA. The study Dalen. J. E., and Alpcrt. J. S. Natural history of pulmonary emboli. Prog. Cardiovasc. Dis., 17: 257-270. 1975. group consisted of nine men and nine women ranging in age Uhland, H., and Goldberg, L. M. Pulmonary embolism: a commonly missed from 26 to 83 years. Eleven patients had undergone total knee entity. Dis. Chest, 45: 533, 1964. Morrei, M. T.. and Dunnill, M. S. The post-mortem incidence of pulmonary or hip arthroplasty. Fifteen of these patients also underwent embolism in a hospitalized population. Br. Med. J.. 55: 347. 1968. contrast venography; all but one of the venograms were per Coon, W. W. Risk factors in pulmonary embolism. Surg. Gynecol. Obstet., formed within 48 h of antibody injection. One venogram was 143: 385-390. 1976. Grollman. J. H.. Webber. M. M.. and Gomes. A. S. 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24. Peters, A. M., Lavender, J. P., Needham, S. G., et al. Imaging thrombus Science (Wash. DC), 222: 1129-1132, 1983. with radiolabcled monoclonal antibody to platelets. Br. Med. J.. 293: 1525- 28. Knight, L. C., Maurer, A. H., Mattis, J. A., Shealy. D. J., and Ammar. 1. A. 1527,1986. Evaluation of In III labeled anti-fibrin antibody for imaging vascular 25. Oster. Z. H., Srivastava, S. C, Som, P.. et al. Thrombus radioimmunoscin- thrombi. J. NucÃ.Med.. 29: 494-502. 1988. tigraphy, an approach using monoclonal antiplatelet antibody. Proc. Nati. 29' Krudr>k-B- RohÂ°za-Au' Ahadl- N'" Â£hln.J" and ,Welbc- MfEr Specificity Acad. Sci. USA. 82:3465-3468. 1985. (.'f a mo,no^.0"o S^iSS NH2-term,nal region of fibrin. Mol. Â», â€”¿. . , â€ž¿., ~ ... .. - â€ž¿ ... , . ... . Ininumol., 21: Xv-94. 19X4. 26. Feitsma, R. I. J.. BlÃ¶k.D., Wasser M. N. J. M N,euwenhu,7.en, W and 3Q Roscbro h. s E . Kudr k

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Downloaded from cancerres.aacrjournals.org on September 24, 2021. © 1990 American Association for Cancer Research. Detection of Thrombophlebitis with 111In-labeled Anti-Fibrin Antibody: Preliminary Results

Abass Alavi, Naresh Gupta, Harold I. Palevsky, et al.

Cancer Res 1990;50:958s-961s.

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