Thyroglobulin Carole A

18 Thyroglobulin Carole A. Spencer and Shireen Fatemi Serum thyroglobulin (Tg) measurements are a follows that when interpreting a serum Tg value, cornerstone for managing patients with differ- it is important to weigh patient-specific pathol- entiated thyroid carcinomas (DTC). The clinical ogy and treatment together with the technical utility of serum Tg testing is predicated on the limitations of the method. This chapter will tissue-specific origin of circulating Tg protein focus on the technical strengths and pitfalls of (thyroid follicular cells). However, since the current Tg methods and the clinical utility maturation and secretion of a mature Tg mole- of using serum Tg measurement as a tumor- cule is complex, it is not surprising that circu- marker for DTC. lating Tg is heterogeneous, and that as tumors dedifferentiate they can lose their capability to synthesize, iodinate, and secrete conformation- ally normal Tg protein [1,2]. Consequently, Technical Strengths and current Tg immunometric assays (IMA), based Pitfalls of Serum Tg on monoclonal antibodies with restricted epi- tope specificity, may only recognize a limited Measurement population of Tg isoforms secreted by a neo- plasm [3,4]. It follows that specificity differences Over the last decade, IMA methodology has largely explain the wide method-to-method largely replaced radioimmunoassays (RIA) variability that precludes changing assays when for measuring serum Tg concentrations. IMA serially monitoring DTC patients. Serum Tg methods are favored by laboratories because concentrations should be interpreted relative to: they can be automated and require a shorter (1) the mass of thyroid tissue present (normal incubation to achieve maximal sensitivity, as remnant and/or tumor); (2) any inflammation compared with RIA [9,10]. Current Tg assays of, or injury to, thyroid tissue, such caused by suffer from a number of technical limitations fine-needle aspiration biopsy (FNAB), thyroid that negatively impact their clinical utility. surgery, radioactive iodine (RAI) therapy, These include (a) large between-method biases or thyroiditis; and (3) the thyroid-stimulating that preclude the use of different methods hormone (TSH) status of the patient, since the for serial monitoring of DTC patients; (b) sub- stimulation of TSH receptors by either endoge- optimal between-run precision across the long nous or recombinant human TSH (rhTSH), intervals generally used to monitor DTC the high human chorionic gonatropin (hCG) patients (~12 months) that can mask clinically of pregnancy, or the thyroid-stimulating im- important changes; (c) inadequate sensitivity munoglobulins present in Graves’ disease, that compromises the early detection of re- elevate the serum Tg concentration [5–8]. It currence; and (d) interferences by TgAb and 211 212 Practical Management of Thyroid Cancer Figure 18.1 Serum Tg concentrations measured in 88 TgAb-negative normal euthyroid volunteers (TSH 0.5–2.5mIU/L) using different methods.Method #1 = University of Southern California RIA,Los Angeles,CA,USA;method #2 = DSL RIA,Websster,Texas,USA; method #3 = Nichols Advantage ICMA, Nichols Institute Diagnostics ICMA, San Juan Capistrano, CA, USA; method #4 = CIS; method #5 = Access ICMA, Beckman-Coulter, Fullerton, CA, USA; method #6 = Immulite ICMA, Diagnostic Products Corporation, Los Angeles, CA, USA; method #7 = Brahms Kryptor, Berlin, Germany.The shaded area approximates the functional sensitivity. Median values are indicated. heterophilic antibodies (HAMA) that can lead (Figure 18.1A). The new guidelines consider a to the reporting of falsely low or high serum Tg change in serum Tg during thyroid hormone values [7,9,11–13]. suppression therapy (THST) in excess of 1.5mg/L to be clinically significant [10]. Current Between-Method Biases between-method biases alone could easily produce this magnitude of change, since current Unfortunately, a change in Tg method during biases exceed the within-person variability of the course of monitoring a DTC patient can serum Tg concentrations (10–15%) [10,14]. produce a shift in the serial Tg pattern that can Thus, significant assay biases preclude chang- cause clinical confusion. Figure 18.1 shows a ing methods during long-term monitoring of representative comparison of Tg measurements patients. For example, a change from a Tg made by two RIA and five IMA methods in sera method with a positive bias to one with a nega- from normal euthyroid subjects with TSH in the tive bias has the potential to mask recurrent 0.5–2.5mIU/L range, with or without detectable disease, whereas a change from a method with TgAb (Figures 18.1A and 18.1B, respectively). a negative bias to one with a positive bias would When TgAb was absent, the methods displayed cause concern for recurrence that could poten- a threefold difference in absolute Tg values tially lead to unnecessary imaging or RAI treat- Thyroglobulin 213 ment. It is now recommended that laboratories Suboptimal Sensitivity consult their physician-users before implement- ing any change in Tg method, and when the bias TSH and Tg assay development share many between the old and proposed new method analogies concerning their quests for improved exceeds 10% the re-baselining of patients is rec- sensitivity. The clinical utility of TSH measure- ommended [10]. Practically, the relative bias ment has been dramatically enhanced over between methods can be assessed from a com- the last decade by the optimization of IMA parison between the mean reference range value methodology, leading to a 100-fold improve- of the methods in question. ment in TSH assay functional sensitivity. Unfor- There are a number of reasons for persis- tunately, the development of more sensitive Tg tent biases between different Tg methods. The assays is still in its infancy. Currently, Tg assay biosynthesis of a mature Tg molecule within the functional sensitivity ranges between 0.3 and follicular cell is a complex vectoral process that 2.5mg/L (Figure 18.1). As shown in Figure 18.2, is orchestrated by different molecular chaper- a 100-fold improvement in Tg assay functional one proteins [15]. Given the complexity of this, sensitivity (~0.01mg/L) would dramatically im- it is not surprising that Tg molecular processing prove the diagnostic sensitivity of measuring Tg may become unregulated in thyroid tumors, during THST.Specifically,Figure 18.2 shows that leading to heterogeneity in tumor-derived Tg there is a striking linear relationship between protein in the circulation.Different Tg assay for- the serum Tg nadir measured without TSH mulations employ a variety of Tg monoclonal stimulation in the first postoperative year and antibody reagents that vary with respect to their long-term (median 8-year) recurrence risk [16]. specificity for different epitopes on tumor- This suggests that less than 1% of patients derived Tg. This can result in the measurement with serum Tg below 0.1mg/L during their first of different Tg isoforms in the specimen and postoperative year would suffer a recurrence give rise to differences in serum Tg measure- [16,17]. ments made by assays [4]. Compounding these As Tg assay sensitivity becomes a marketing specificity differences are differences in assay issue, there will be increasing commercial pres- standardization and the use of non-serum cali- sure for the diagnostic kit manufacturers to make brator matrices that behave differently from unrealistic claims for the sensitivity of their tests. human sera [9]. The guidelines state that the functional sensitivity of a Tg assay should be determined from the lowest Tg value that can be measured with Between-Run Precision 20% between-run coefficient of variation, using TgAb-negative human sera measured across a Current Tg assays have suboptimal between- 6- to 12-month period and using different lots run precision across the long follow-up interval of reagent [10]. It is critical that Tg method (6–12 months) typical of monitoring patients comparisons are made on the basis of functional with DTC. It is well known that the between- sensitivity and that descriptive terms such as run precision of a biochemical test erodes over “ultrasensitive” and “supersensitive” are not used time as a consequence of changing reagent lots, for marketing purposes [10]. instrument calibrations and a myriad of other Unfortunately, even with Tg assay functional less well-defined factors. The loss of assay pre- sensitivity determined according to the stan- cision over the time between clinical evaluations dardized protocol, the absolute (mg/L) func- negatively impacts the ability to detect small tional sensitivity of different methods cannot be clinically significant changes, especially at the compared because of method biases (Figure extremes of the Tg measurement range. The 18.1A). The ability to detect small amounts of guidelines suggest that laboratories should tumor is related to the degree of discrimination archive specimens remaining after serum Tg between the assay lower reference limit for testing to allow concurrent measurement of the normal euthyroid subjects and its functional past and current specimens from the patient in sensitivity. Currently, there is very little dis- the same run, thereby eliminating between-run crimination between the lower reference limit errors and improving the clinical sensitivity of for normal euthyroid subjects and the func- the test [7,10]. tional sensitivity limits of current methods, as 214 Practical Management of Thyroid Cancer Figure 18.2 The relationship between cumulative percent recurrence in a cohort of 278 papillary thyroid cancer patients followed over a median of 8 years and the median group serum Tg nadir value reported (in the absence of TSH stimulation) during the first year fol- lowing thyroidectomy. G1 had serum Tg nadirs between 1.0 and 1.9mg/L; G2 had serum Tg nadirs between 2.0 and 4.9mg/L;G3 had serum Tg nadirs between 5 and 10mg/L; G4 had serum Tg nadirs above 10mg/L [16]. shown in Figure 18.1A. In fact, serum Tg was studies can be used to supply reassurance that paradoxically undetectable for some subjects by disease is absent.

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