OnT4-Thyroglobulin Assay Before rhTSH-Aided Radioiodine Ablation
OnT4-Thyroglobulin Assay Before rhTSH-Aided Radioiodine Ablation
Thyroidectomy followed by administration of large activities of 131-iodine (131I) is the
 treatment of choice for differentiated thyroid carcinoma (DTC). The serum thyroglobulin (Tg)
 measurement during hypothyroidism (offT4-Tg), just before radioiodine thyroid ablation, has
 proved to be effective for predicting persistent/recurrent disease. However, the Tg
 measurement cannot be used as a corresponding value for preablative offT4-Tg when rhTSH is
 used as stimulous before treatment. The present study was undertaken to evaluate if
 post-thyroidectomy Tg values, measured before rhTSH-stimulation and radioiodine
 administration, is of prognostic value in patients affected by DTC. We enrolled 28 patients
 with DTC and submitted to total thyroidectomy. Thyroxine (T4) treatment was started just
 after surgery to suppress TSH levels. Six to nine weeks later Tg levels were measured both
 basally (onT4-Tg) and after rhTSH (rhTSH-Tg) stimulation. Subsequently, T4 was stopped and
 serum Tg measured (offT4-Tg) just before 3700 MBq of 131I-iodide administration. A
 post-treatment whole body scan (PT-WBS) was performed and neck radioiodine uptake (RAIU)
 measured. A significant relationship was found between onT4-Tg and both rhTSH-Tg and
 offT4-Tg. The onT4-Tg levels of 0.2 ng/mL or higher predicted PT-WBS results with a 100%
 negative and 43% positive predictive values, respectively. Additionally onT4-Tg levels of 0.9
 ng/mL or more predicts 12-months recurrences with 100% negative and 60% positive predictive
 value. In comparison, 1.0 ng/mL or higher offT4-Tg values predicted PT-WBS results and
 12-months restaging with 94% and 100% negative and 45% and 27% positive predictive value,
 respectively. Basing on our data we conclude that preablative onT4-Tg may be of value as
 prognostic marker when rhTSH-aided radioiodine ablation is done. Additionally, the role of
 preblative onT4-Tg measurement as a yard-stick for radioiodine ablation should be further
 evaluate.
Introduction Total (or near-total) thyroidectomy followed by TSH-stimulated administration of
 large activities of 131-iodine (131I) is the treatment of choice for DTC [1-3]. The serum
 thyroglobulin (Tg) measurement during hypothyroidism, just before radioiodine thyroid
 ablation, has proved to be effective for predicting persistent/recurrent disease [4-8].
 Recently recombinant human TSH (rhTSH) showed to be safely employed instead of thyroxine (T4)
 withdrawal (offT4) to prepare patients for radioiodine ablation [9-10]. However, the Tg level
 are measured 48 hours after radioiodine administration when rhTSH is used as stimulation
 [11]. Consequently, due to the radioiodine-induced thyroid cells damage and Tg release, the
 Tg measurement would not have reliable predictive value in patients treated by rhTSH
 stimulation [12]. The present study was undertaken to evaluate if preablativeTg measurement
 under T4 treatment is of prognostic value and serves as surrogate marker of offT4-stimulated
 preablative Tg.

 Patients and methods

 Patients selection We retrospectively enrolled 28 consecutive patients affected by
 histologically proven DTC (23 papillary, 5 follicular) submitted to total thyroidectomy and
 central compartment lymph-node dissection. Thyroxine (T4) treatment was started immediately
 after surgery to suppress TSH levels. Six to nine weeks later Tg levels were measured both
 basally (onT4-Tg) and after rhTSH (rhTSH-Tg) stimulation as previously described [13].
 Subsequently, T4 was stopped for 4 weeks and serum Tg measured (offT4-Tg) just before 3700
 MBq of 131I-iodide administration. A post-treatment whole body scan (PT-WBS) with radioiodine
 uptake (RAIU) calculation was performed according to a previously described protocol [1]. All
 non physiologic iodine uptake areas out of the thyroid bed were considered as positive
 findings [14, 15]. Patients with positive PT-WBS underwent specific treatment and
 personalized follow-up. Patients with negative PT-WBS immediately restarted T4 suppressive
 treatment. Final restaging was performed in all patients 12 months after the last treatment
 by neck ultrasound, onT4-Tg assay and both offT4-diagnostic WBS (DgWBS) and Tg assay (4 weeks
 T4 withdrawal; required TSH>30 mUI/L). Clinical and pathological characteristics of selected
 patients were summarized in the Table 1.

 Serum Tg assay and screening for interferences Serum Tg was assayed in duplicate by a
 specific high-sensitive IRMA assay (DYNOtest® Tg-plus, BRAHMS Diagnostica GmbH, Berlin,
 Germany) according to the producer' instructions. This method provided a sensitivity of 0.05
 ng/mL and a functional sensitivity of 0.2 ng/mL [16, 17]. As previously published,
 preablative offT4 serum Tg values above 4.5 ng/mL and 3.2 ng/mL were considered as positive
 with respect to PT-WBS and 12 months restaging results, respectively [8]. Otherwise, offT4-Tg
 values higher than 0.2 ng/ml measured 12 months after thyroid ablation were considered
 positive for persisting/relapsing disease. [18, 19]. The presence of anti-thyroglobulin
 antibodies (AbTg) was screened by a specific radioimmunoassay (DYNOtest® anti-TGn, BRAHMS
 Diagnostica GmbH, Berlin, Germany) and by recovery test with a specific Tg-recovery buffer
 provided by the producer. Sera showing AbTg levels more than 60 U/mL and/or recover less than
 80% or more than 120% were excluded from the study. Quality control was ensured by assaying
 two levels of control sera in each series, by re-assessing all sera showing a coefficient of
 variation exceeding 10% and by a bimonthly partecipation in the European inter-laboratory
 control OncocheckTM.

 Statistics Quantitative data are expressed as mean ± SD. Differences between groups were
 assessed by two-tailed unpaired t-test. The relationship between variables was assessed by
 linear regression analysis. In order to allow statistical analysis the value of undetectable
 serum Tg expressed as < 0.2 ng/nL was arbitrarily changed in 0.10 ng/mL. Statistical
 significance was defined by a p-value < 0.05.

 Ethics All diagnostic and therapeutic procedures were performed according to the regulations
 of the local ethics committee. Informed consensus was obtained from each patient.

 Results The overall results are summarized in the Table I. Relationship Between Post-surgery
 onT4-Tg, rhTSH-Tg and offT4-Tg A significant positive relationship was found between onT4-Tg
 and both rhTSH-Tg (p<.0001) and offT4-Tg (p<0.0001) (Figure 1, A-B-C) as well as between
 rhTSH and offT4-stimulated Tg (p<.0001).

 Relationship Between Post-surgery Serum Tg and Thyroid Remnant Radioiodine Uptake (RAIU)
 Among 22 patients showing no DTC metastasis on PT-WBS the serum distribution of onT4-Tg,
 rhTSH-Tg and offT4-Tg was 0.342±0.402; 0.664±0.803 and 1.195±1.485 ng/mL respectively. Both
 rhTSH and offT4-stimulated Tg levels were related to RAIU (p <.05 and <.005, respectively)
 while no significant relationship was found between RAIU and onT4-Tg levels (Figure 1,
 D-E-F).

 Relationship Between Post-surgery Serum Tg and PT-WBS results The patients with positive
 PT-WBS showed higher onT4-Tg (0.617±0.445 vs 0.341±0.402 ng/mL), rhTSH-Tg (2.150±1.249 vs
 0.664±0.803 ng/mL) and offT4-Tg (4.417±2.136 vs 1.195±1.485 ng/mL) levels as compared with
 patients with negative scan. Among 14 patients with undetectable onT4-Tg (i.e. ≤0.2 ng/mL)
 none had positive PT-WBS neither recurrences at 12-months restaging. Additionally, none of
 these patients showed stimulated Tg values more than 0.4 ng/mL and 1.0 ng/mL after rhTSH
 stimulation and T4 withdrawal, respectively. Viceversa, among 14 patients with onT4-Tg levels
 more than 0.2 ng/mL, 6 had positive PT-WBS (onT4-Tg: 0.3 to 1.4 ng/mL) and 3 showed DTC
 recurrences at 12 months restaging (onT4-Tg: 0.9 to 1.7 ng/mL). Consequently, the onT4-Tg
 levels predicts PT-WBS results with a 100% negative and 43% positive predictive values,
 respectively. Additionally onT4-Tg levels of 0.9 ng/mL or more predicts 12-months recurrences
 with 100% negative and 60% positive predictive value. In comparison, 1.0 ng/mL or higher
 offT4-Tg values predicted PT-WBS results and 12-months restaging with 94% and 100% negative
 and 45% and 27% positive predictive value, respectively.

 Discussion Many reports indicate the usefulness of Tg concentration measurement before
 radioiodine treatment to early detection of DTC relapse or metastasis [4-8, 20, 21]. Three
 factors determine Tg concentration in most clinical situations: thyroid cell mass, thyroid
 cell damage and activation of TSH receptors [22]. When Tg is measured before radioiodine
 ablation the effects of surgical damage are generally vanished and endogenous TSH levels are
 increased in all patients: consequently the thyroid remnant mass is the major determinant of
 the serum Tg concentrations [23]. However, the rhTSH-stimulated Tg cannot be used instead of
 preablative offT4-Tg when rhTSH is employed to prepare radioiodine ablation [12]. Therefore
 we evaluated the role of post-surgery onT4-Tg as surrogate prognostic marker. We choiced to
 suppress TSH levels in order to normalize the effect of TSH stimulation on thyroid remnants.
 Clearly, the TSH suppression reduced Tg levels: however, the high-sensitive Tg assays provide
 a good distinction between the lower limit of the euthyroid reference range and the
 functional sensitivity limit detecting small amounts of thyroid tissue even in the
 TSH-suppressed state (22, 23). We showed a significant positive relationship between
 post-surgery Tg measured during T4 treatment and after TSH stimulation. No relationship was
 found between RAIU (i.e. expression of remnant mass) and onT4-Tg, probably due the clustering
 of all Tg levels lower than 0.2 ng/mL (see statistics paragraph). However, undetectable
 onT4-Tg after surgery identifies patients free of metastasis at PT-WBS and without late
 recurreces during early 12-months follow-up. None of these patients showed a significant
 increase in both rhTSH and offT4-stimulated Tg before radioiodine ablation. This seems to
 indicate that the relationship between Tg expression and thyroid tissue mass is manteined
 even in TSH suppression state: Therefore undetectable serum onT4-Tg really identifies
 patients without significant thyroid tissue amount as well as stimulated Tg. All patients
 performed extracapsular total thyroidectomy in a dedicated thyroid surgery unit and the
 thyroid remnant, expressed as RAIU, was lower in our series than in others. This means that
 our data cannot directly translated to patients treated by more limited surgery. Globally,
 basing on our data we conclude that preablative onT4-Tg may be of value as prognostic marker
 when rhTSH-aided radioiodine ablation is done. Additionally, the role of preblative onT4-Tg
 measurement as a yard-stick for radioiodine ablation should be further evaluate.

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Inclusion Criteria:

 -  Hiostologically proved DTC (M0)

 Exclusion Criteria:

 -  Preoperative metastasis