Distinct patterns of E-cadherin CpG island methylation in papillary, follicular, Hurthle's cell, and poorly differentiated human thyroid carcinoma.

Expression of the invasion/metastasis suppressor, E-cadherin, is diminished or lost in thyroid carcinomas. Yet, mutational inactivation of E-cadherin is rare. Herein, we show that this loss is associated with hypermethylation of the E-cadherin 5' CpG island in a panel of human thyroid cancer cell lines. This aberrant methylation is evident in 83% of papillary thyroid carcinoma, 11% of follicular thyroid carcinoma, 40% of Hurthle's cell carcinoma, and 21% of poorly differentiated thyroid carcinomas. Contrary to previous reports, the majority of these poorly differentiated thyroid carcinomas express E-cadherin, but often within the cytoplasm rather than at the cell surface. Together, our data indicate that the invasion/metastasis suppressor function of E-cadherin is frequently compromised in human papillary, Hurthle's cell, and poorly differentiated thyroid carcinoma by epigenetic and biochemical events.

Effect of estrogen on the synthesis and secretion of thyroxine-binding globulin by a human hepatoma cell line, Hep G2.

Hyperestrogenemia in humans increases both the concentration of serum T4-binding globulin (TBG) by 2- to 3-fold and the proportion having anodal mobility on isoelectric focusing (IEF). As TBG is synthesized in the liver, we studied the effect of estrogen on TBG synthesis, secretion, and degradation by cultured human hepatocarcinoma cells (Hep G2). beta-Estradiol in concentrations in the range found in pregnancy (10(-7) M) had no effect on the accumulation of immunoreactive TBG in medium over 4 days. The absence of fetal calf serum or phenol red did not alter these findings. The amount of [35S]TBG accumulated 6 h after addition of [35S]methionine was not influenced by exposure to estrogen or to serum obtained from pregnant women. However, 10(-5) M beta-estradiol suppressed TBG more severely than albumin synthesis (34% vs. 9%). The lack of an estrogen effect on TBG synthesis and secretion was supported by experiments showing no effect of estrogen on the disappearance of TBG added to the medium or the accumulation of cytoplasmic TBG mRNA. The same cultures responded to estrogen by a 10-fold increase in nuclear estrogen receptor binding sites and a 2-fold increase in apolipoprotein CII. As TBG in serum, the rate of heat denaturation was not altered in TBG synthesized by Hep G2 cells in the presence of estrogen. In contrast to the effect on TBG in serum, in Hep G2 cells estrogen did not produce an anodal shift on IEF, or increased its proportion not bound to Concanavalin A, nor reduced its clearance rate when injected into rats. However, even untreated Hep G2 cells synthesized TBG with a larger number of anodal IEF bands and proportion of Concanavalin A excluded material than TBG in pregnancy serum. Results support our hypothesis, based on analysis of TBG in pregnancy, that estrogen-induced serum TBG elevation may not be mediated through an increase in synthesis. The failure to observe estrogen induced changes in oligosaccharide structure does not exclude estrogen responsivity of Hep G2 cells. Such effect could be masked by the marked constitutive increase in number of oligosaccharide chain antennae typical in this and other neoplastic tissues.

American Thyroid Association guidelines for detection of thyroid dysfunction.

OBJECTIVE: To define the optimal approach to identify patients with thyroid dysfunction. PARTICIPANTS: The 8-member Standards of Care Committee of the American Thyroid Association prepared a draft, which was reviewed by the association's 780 members, 50 of whom responded with suggested revisions. EVIDENCE: Relevant published studies were identified through MEDLINE and the association membership's personal resources. CONSENSUS PROCESS: Consensus was reached at group meetings. The first draft was prepared by a single author (P.W.L.) after group discussion. Suggested revisions were incorporated after consideration by the committee. CONCLUSIONS: The American Thyroid Association recommends that adults be screened for thyroid dysfunction by measurement of the serum thyrotropin concentration, beginning at age 35 years and every 5 years thereafter. The indication for screening is particularly compelling in women, but it can also be justified in men as a relatively cost-effective measure in the context of the periodic health examination. Individuals with symptoms and signs potentially attributable to thyroid dysfunction and those with risk factors for its development may require more frequent serum thyrotropin testing.

Relationship of oligosaccharide modification to the cause of serum thyroxine-binding globulin excess.

T4-binding globulin (TBG), a glycoprotein with four N-glycosyl complex oligosaccharide chains, exhibits sialic acid-dependent microheterogeneity on isoelectric focusing (IEF). Increasing the sialic acid content of TBG increases its anodal IEF mobility and decreases its rate of in vivo degradation, a mechanism for the elevation of serum TBG levels in pregnancy. In this study, the structure of oligosaccharides in TBG from subjects with various conditions associated with TBG excess was determined by measuring the proportion that bound to Concanavalin-A (Con-A). Since oligosaccharides with three or more branches (antennae) attached to the trimannosyl core are excluded from binding to Con-A, the percentage of serum TBG not bound to Con-A (% peak A) represented the portion of TBG molecules with three or more antennae in all oligosaccharide chains interacting with Con-A. Peak A contained the most anodal IEF bands, while the Con-A-bound TBG (peak B) contained the cathodal bands. Serum samples from 10 normal men and 10 premenopausal women did not significantly differ in terms of TBG levels, % peak A, or IEF mobility and were combined as a single group (normal). Eight subjects with elevated serum TBG levels due to inherited TBG excess [62.0 +/- 10.1 (+/- SD) mg/L] or 2 receiving 5-fluorouracil treatment (26.2 and 31.3 mg/L) compared to 20 normal (14.7 +/- 3.3 mg/L) had % peak A values and IEF mobility similar to those in normal subjects. On the other hand, high serum TBG levels in 8 women during the third trimester of pregnancy (39.2 +/- 5.3 mg/L), 2 women taking oral contraceptives (25.7 and 27.0 mg/L), and 3 women with acute hepatitis (34.8 +/- 4.8 mg/L) were associated with significant elevations of % peak A values (5.68 +/- 1.73%, 3.31% and 2.41%, and 3.25 +/- 0.78%, respectively) compared to those in normal subjects (1.33 +/- 0.40%), as well as increased anodal mobility on IEF. Treatment of a man for 3 days with ethinyl estradiol produced similar changes. Using data from densitometry measurements of IEF bands of TBG, the degree of anodal shift was quantitated (anodal index). This index correlated with the % peak A (r = 0.92) in all study subjects. We conclude that increased sites for sialylation, resulting from the increased proportions of triantennary oligosaccharide chains, account for the increased anodal mobility of TBG in hyperestrogenemia and hepatitis. Thus, in these two conditions, a reduced TBG degradation rate resulting from oligosaccharide modification is the likely mechanism of increased serum TBG levels.(ABSTRACT TRUNCATED AT 400 WORDS)

Somatostatin analogs affect proliferation of human thyroid carcinoma cell lines in vitro.

Antiproliferative effects of somatostatin (SRIH) analogs were investigated in human thyroid carcinoma cell lines. Membrane preparations from six human thyroid cell lines, including follicular (RO 87-M-1 and RO 82-W-1), papillary (NPA87), and anaplastic (RO 90-D-1) carcinomas, a follicular adenoma, as well as benign and malignant human thyroid tissues, contained high affinity SRIH-binding sites. Ligand-binding studies ([125I]Tyr11-SRIH) demonstrated mean dissociation constants ranging from 114-224 pmol/L, and 20-154 fmol/mg membrane protein mean receptor sites, in cell lines. Four cell lines were grown for 3 days in monolayers with SRIH analogs (octreotide or MK0678) at concentrations from 0.05-100 nmol/L in serum-free medium to assess changes in cell numbers. At the highest dose, MK0678 produced dose-dependent inhibition of growth in RO 87-M-1 and NPA87, each to about 70% of that in control cells. Octreotide produced dose-dependent stimulation of growth in RO 87-M-1 cells, but caused growth inhibition in NPA87 cells, with loss of effect at the highest dose. RO 82-W-1 did not respond to MK0678, yet caused biphasic inhibition with octreotide (75% and 45% of control cell numbers at 0.05 and 100 nmol/L doses, respectively). Anaplastic cells, RO 90-D-1, did not respond to either analog despite similar ligand binding. Addition of epidermal growth factor (100 micrograms/L) or TSH (200 mU/L) increased the sensitivity of RO 87-M-1 cells to growth inhibition by the lowest dose of MK0678, producing biphasic dose-response curves. In conclusion, the present data demonstrate specific SRIH binding to membranes of thyroid carcinoma cells and tissues as well as discordant growth effects of different SRIH analogs on the same cell lines. This may be a result of differential stimulation and regulation of distinct SRIH receptor subtypes.

Reduced clearance rate of thyroxine-binding globulin (TBG) with increased sialylation: a mechanism for estrogen-induced elevation of serum TBG concentration.

Hyperestrogenemic states, including pregnancy, cause an increase in serum T4-binding globulin (TBG) concentrations and an increase in the proportion of TBG molecules with greater anodal mobility on isoelectric focusing, indicating greater sialic acid content. The possible causal relationship between the degree of sialylation and accumulation of TBG in serum was explored by measuring the in vivo half-lives (t1/2) of TBGs with different isoelectric points. TBG in unfractionated serum and its major peaks, isolated by chromatofocusing and defined by their isoelectric points on isoelectric focusing were each injected iv into rats. The resulting TBG concentrations, measured by specific RIA in serum samples obtained at intervals after injection, were used for the calculation of the t1/2. TBG in serum from a pregnant woman had a significantly longer t1/2 of 17.2 +/- 1.2 h (mean +/- SD) compared to those of 13.3 +/- 1.5 and 12.9 +/- 0.9 h for TBG in serum from a man and a nonpregnant woman, respectively. TBG peaks II, III, IV, and V, with increasing anodal mobility, had progressively longer t1/2 values of 11, 13, 15, and 33 h, respectively. However, TBG peaks of the same mobility on IEF isolated from serum of pregnant or nonpregnant subjects had similar t1/2 values. Neither the TBG concentration nor estrogen had a direct effect on the rate of TBG clearance. Indeed, the t1/2 of TBG from a subject with inherited TBG excess was not different from that of TBG from a nonpregnant woman or a man. Chronic treatment of rats with estradiol did not alter the rate of clearance of injected human TBG. Finally, the more heavily sialylated anodal bands of purified but unfractionated serum TBG, analyzed by Western blots, survived longer in the circulation of a rat. These results indicate that the rate of in vivo metabolism of TBG is dependent on its sialic acid content. The increased proportion of TBG molecules with higher sialic acid content thus contributes to the increase in the serum TBG concentration in hyperestrogenemic states.

Antineoplastic activity of taxol against human anaplastic thyroid carcinoma cell lines in vitro and in vivo.

Anaplastic thyroid carcinoma is a rapidly fatal neoplasm that fails to adequately respond to any known chemotherapeutic regimen. We tested taxol (paclitaxel) against six human anaplastic thyroid carcinoma cell lines (DRO-90, ARO-81, KAT-4, KAT-18, SW-1736, and BHT-101). Each cell line monolayer culture, in log phase growth, was treated with taxol concentrations ranging from 0.001-5.0 mumol/L. Cell numbers, after 24-, 48-, and 72-h growth periods in separate experiments, were expressed as percentages of control cell numbers without taxol. All cell lines showed maximal inhibition with 0.05 mumol/L taxol at 3-28% of control cell numbers. Greater inhibition was seen with longer growth periods. Three cell lines (DRO-90, ARO-81, and KAT-4) were grown as sc xenograft tumors in nude mice for 18-26 days. Treatment groups received sc taxol injections in sites distant from the tumors, whereas control mice received vehicle. All taxol-treated xenografts were inhibited to near-starting volume or disappeared, whereas control xenograft volumes increased 9- to 59-fold. These results suggest that taxol may have beneficial clinical effects in anaplastic thyroid carcinoma patients.

Somatostatin receptor subtype expression in human thyroid and thyroid carcinoma cell lines.

Somatostatin (SRIH) analogs can suppress the proliferation of human differentiated thyroid carcinoma cell lines that express SRIH receptors (SSTRs) demonstrated by radioligand binding analysis. Five distinct human SSTR subtypes (hSSTR1-5) that bind native SRIH exhibit diverse affinities to a wide range of SRIH analogs. Reverse transcriptase-PCR amplification of ribonucleic acids (RNAs) obtained from normal thyroid tissues and nine human thyroid carcinoma cell lines, grown as monolayer cultures and xenograft tumors in nude mice, were used to discriminate expression of SSTR subtype messenger RNAs (mRNAs). The cell lines were derived from a follicular adenoma (KAK-1), two follicular carcinomas (MRO-87 and WRO-82), two papillary carcinomas (NPA87 and KAT-10), and four anaplastic thyroid carcinomas (DRO-90, ARO-81, KAT-4, and KAT-18). Most thyroid cancer cell line monolayers and xenografts expressed SSTR3 and SSTR5 mRNAs. SSTR1 expression was more varied between monolayers and xenografts, whereas SSTR2 mRNA was only faintly detectable at the most extreme resolution. SSTR4 mRNA was faintly positive in only one anaplastic carcinoma xenograft. Normal thyroid also expressed SSTR3 and SSTR5 mRNAs, with only faint expression of SSTR1 and SSTR2 mRNAs (in one of five and three of five samples, respectively). SSTR mRNA expression was dependent upon in vitro culture conditions, as xenograft SSTR mRNA expression tended to decrease compared to that in each respective monolayer culture. Characterization of SSTR subtype expression in human thyroid carcinomas may permit targeting of specific SRIH analogs to inhibit proliferation of differentiated and anaplastic thyroid carcinomas in patients.

Restoration of iodide uptake in dedifferentiated thyroid carcinoma: relationship to human Na+/I-symporter gene methylation status.

Disseminated dedifferentiated thyroid epithelial carcinoma, which cannot sufficiently concentrate therapeutic radioiodide, is a terminal disease without any effective systemic treatment or chemotherapy. This is a likely consequence of loss of human sodium-iodide symporter (hNIS) function. We hypothesized that hNIS transcriptional failure in thyroid carcinoma could be consequent to methylation of DNA in critical regulatory regions and could be reversed with chemical demethylation treatment. Analysis of hNIS messenger ribonucleic acid (mRNA) expression in 23 tumor samples revealed that although loss of this expression corresponded to loss of clinical radioiodide uptake, some thyroid carcinomas with hNIS mRNA expression did not concentrate iodide, suggesting additional posttranscriptional mechanisms for loss of hNIS function. In addition, analysis of DNA methylation in CpG-rich regions of the hNIS promoter extending to the first intron failed to define specific methylation patterns associated with transcriptional failure in human thyroid tumor samples. In seven human thyroid carcinoma cell lines lacking hNIS mRNA, treatment with 5-azacytidine or sodium butyrate was able to restore hNIS mRNA expression in four cell lines and iodide transport in two cell lines. Investigation of methylation patterns in these cell lines revealed that successful restoration of hNIS transcription was associated with demethylation of hNIS DNA in the untranslated region within the first exon. This was also associated with restoration of expression of thyroid transcription factor-1. These results suggest a role for DNA methylation in loss of hNIS expression in thyroid carcinomas as well as a potential application for chemical demethylation therapy in restoring responsiveness to therapeutic radioiodide.

Serum thyrotropin and prolactin in the syndrome of generalized resistance to thyroid hormone: responses to thyrotropin-releasing hormone stimulation and short term triiodothyronine suppression.

Serum TSH and PRL levels and their response to TRH were measured in 11 patients with generalized resistance to thyroid hormone (GRTH), 6 euthyroid subjects, and 6 patients with primary hypothyroidism. TSH and PRL levels and their response to TRH were also measured after the consecutive administration of 50, 100, and 200 micrograms T3 daily, each for a period of 3 days. Using a sensitive TSH assay, all GRTH patients had TSH values that were elevated or within the normal range. On the basis of a normal or elevated TSH level, GRTH patients were classified as GRTH-N1 TSH (5 patients) or GRTH-Hi TSH (6 patients), respectively. Only GRTH patients with previous thyroid ablative therapy had basal TSH values greater than 20 mU/L. TSH responses, in terms of percent increment above baseline, were appropriate for the basal TSH level in all subjects. No GRTH patient had an elevated basal PRL level. PRL responses to TRH were significantly increased only in the hypothyroid controls compared to values in all other groups. On 50 micrograms T3, 7 of 12 (58%) nonresistant (euthyroid and hypothyroid) and 1 of 11 (9%) resistant subjects had a greater than 75% suppression of the TSH response to TRH. On the same T3 dose, 2 of 12 (17%) nonresistant and 4 of 11 (36%) resistant subjects had a greater than 50% suppression of the PRL response to TRH. On 200 micrograms T3, all subjects, except for 1 with GRTH, had a greater than 75% suppression of the TSH response to TRH. On the same T3 dose, while 11 of 12 (92%) nonresistant subjects had a greater than 50% reduction of the PRL response to TRH, only 3 of 10 (30%) resistant patients showed this degree of suppression (P less than 0.005). Without previous ablative therapy, serum TSH in patients with GRTH is usually normal or mildly elevated. The TSH response to TRH is proportional to the basal TSH level and is suppressed by exogenous T3. However, on 200 micrograms T3 basal TSH was not detectable (less than 0.1 mU/L) in all euthyroid subjects, but it was measurable in three of four GRTH patients with normal TSH levels before T3 treatment. PRL levels in GRTH are normal even when TSH is elevated. The PRL response to TRH is not increased in GRTH. In all subjects, exogenous T3 suppresses the PRL response to TRH to a lesser degree than the TSH response, but this difference is much greater in patients with GRTH.

Lithium as a potential adjuvant to 131I therapy of metastatic, well differentiated thyroid carcinoma.

As lithium inhibits the release of iodine from the thyroid but does not change iodine uptake, it may potentiate 131I therapy of thyroid cancer. The effects of lithium on the accumulation and retention of 131I in metastatic lesions and thyroid remnants were evaluated in 15 patients with differentiated thyroid carcinoma. Two 131I turnover studies were performed while the patients were hypothyroid. One was performed while the patient received lithium; the second served as a control study. From a series of gamma-camera images, it was found that lithium increased 131I retention in 24 of 31 metastatic lesions and in 6 of 7 thyroid remnants. A comparison of 131I retention during lithium with that during the control period showed that the mean increase in the biological or retention half-life was 50% in tumors and 90% in remnants. This increase occurred in at least 1 lesion in each patient and was proportionally greater in lesions with poor 131I retention. When the control biological half life was less than 3 days, lithium prolonged the effective half-life, which combines both biological turnover and isotope decay, in responding metastases by more than 50%. More 131I also accumulated during lithium therapy, probably as a consequence of its effect on iodine release. The increase in the accumulated 131I and the lengthening of the effective half-life combined to increase the estimated 131I radiation dose in metastatic tumor by 2.29 +/- 0.58 (mean +/- SEM) times. These studies suggest that lithium may be a useful adjuvant for 131I therapy of thyroid cancer, augmenting both the accumulation and retention of 131I in lesions.

Effects of p53-expressing adenovirus on the chemosensitivity and differentiation of anaplastic thyroid cancer cells.

We investigated the p53 status and the ability of exogenous wildtype (wt) p53 to affect chemosensitivity in three anaplastic thyroid carcinoma cell lines (BHT-101, SW-1736, and KAT-4). All three cell lines had nonfunctional p53. Treatment with mitomycin C or adriamycin did not result in accumulation of p53 or induction of p21WAF1/CIP1 or Mdm-2 and did not cause Rb dephosphorylation. BHT-101 and KAT-4 cells had mutant p53. SW-1736 cells were functionally mutant because of marked down-regulation of wt p53 messenger ribonucleic acid, representing a novel mechanism of p53 dysfunction. Infection with a p53-expressing adenovirus (Ad-p53) induced high levels of p21 and Mdm-2 proteins. In BHT-101 cells, induction of p21 and Mdm-2 was evident 10 h after infection. In KAT-4 cells, induction of p21 and Mdm-2 was observed 1 day after infection, and continued to increase over the ensuing 24 h. SW-1736 cells demonstrated intermediate kinetics. Sensitivity to the cytotoxic effect of Ad-p53 paralleled the kinetics of p21/Mdm-2 induction. BHT-101 cells were most sensitive to killing by Ad-p53, with an IC50 of less than 2 multiplicity of infection; SW-1736 cells were intermediate in sensitivity; KAT-4 cells were resistant. All three cell lines became more sensitive to adriamycin after wt p53 expression, with a 10-fold decrease in IC50 values. The latter observation may make a combination of wt p53 and chemotherapeutic drugs an attractive modality for treating anaplastic thyroid cancer.

Papillary thyroid carcinoma. Etiology, assessment, and therapy.

Most papillary carcinomas do not cause death or significant morbidity. Current modes of therapy when appropriately applied, appear effective for 80% to 90% of patients. The clinical challenge is to identify the minority of patients who will suffer greatly from tumors to direct appropriately the best therapeutic efforts. New appreciation of histologic subtypes of papillary carcinoma has provided fresh clues to identify cancers with poor prognosis. We look forward to innovative therapies to deal with currently untreatable disease.

A unique interaction between polyamine and multidrug resistance (P-glycoprotein) transporters in cultured Chinese hamster ovary cells transfected with mouse mdr-1 gene.

We have shown that a functional link exists between the polyamine transporter and the multi-drug resistance (MDR) efflux transporter (P-glycoprotein, P-gp) in MDR-positive cancer cells. To further explore the nature of this interaction, we have examined the effect of reduced polyamine transport activity on cellular expression and activity of P-gp acquired by either selection or transfection. Chinese hamster ovary (CHO) cells and their polyamine transport-deficient mutants (CHOMGBG) were transfected with mouse mdr-1b gene. The activity of P-gp in these cells was quantified by measuring cellular accumulation of radiolabeled taxol and etoposide in the presence and absence of the P-gp modulator SDZ PSC-833 (valspodar; a semisynthetic undecapeptide derived from cyclosporin D). The mdr-1b-transfected CHO cells accumulated 2- to 3-fold less taxol and etoposide than the controls, an accumulation defect reversed by the potent MDR modulator PSC-833. Despite expression of P-gp on the surface of mdr-1b-transfected CHOMGBG cells, this classic MDR phenotype was not observed. Similarly, CHO cells, but not CHOMGBG cells, showed MDR activity after selection with doxorubicin as determined by reduced accumulation of radiolabeled taxol. Treatment with 50 microM of reduced polymer of spermine and glutaraldehyde, a selective blocker of the polyamine transport system, reduced MDR activity in mdr-1-transfected CHO cells and restored cellular accumulation of etoposide and taxol to control levels, effects not observed in mdr-1-transfected CHOMGBG cells. Notably, mdr-1-transfected CHO cells were 4- to 16-fold more resistant to the cytotoxic effects of the P-gp substrates doxorubicin, taxol, and etoposide than were the mdr-1-transfected CHOMGBG cells. CHO cells transfected with the mdr-1 gene exhibited a 23% reduction in cellular uptake of [14C]spermidine compared with untransfected controls; spermidine accumulation in CHOMGBG cells was no different than that in untransfected controls. These data suggest that the existence of a functioning polyamine transport system may be a requirement for MDR transporter activity, while the expression of functioning P-gp appears to reduce polyamine transporter activity.

Alzheimer's amyloid beta-peptide associated free radicals increase rat embryonic neuronal polyamine uptake and ornithine decarboxylase activity: protective effect of vitamin E.

Recent evidence indicates that alterations in brain polyamine metabolism may be critical for nerve cell survival after a free radical initiated neurodegenerative process. It has been shown previously that A beta(1-42) and A beta(25-35) are toxic to neurons through a free radical dependent oxidative mechanism. Treatment of rat embryonic hippocampal neuronal cultures with A beta-peptides increased ornithine decarboxylase (ODC) activity and spermidine uptake, suggesting that oxidative stress upregulates the polyamine mechanism for the repair of free radical damage. Pretreatment of the cells with vitamin E prior to A beta exposure decreased ODC activity and spermidine uptake to control level. This study is the first to demonstrate that A beta treated cells show an increased polyamine metabolism in response to free radical mediated oxidative stress and that the free radical scavenger vitamin E prevents these attenuations. These results are discussed with reference to Alzheimer's disease.

Effects of restricting levothyroxine dosage strength availability.

We conducted a prospective, randomized, controlled trial to assess whether hospital formulary restrictions involving limiting dosage strengths of levothyroxine affect physicians' ability to manage patients effectively and provide pharmacy cost savings in a tertiary care federal government research hospital. Thirty-three endocrinologists were randomly assigned to prescribe levothyroxine from a restrictive (dosage strengths of 25, 50, 100, 125, and 150 micrograms) or a nonrestrictive (dosage strengths of 25, 50, 75, 100, 112, 125, 150, 175, 200, and 300 micrograms) formulary through a central computer system. Their 241 respective outpatients' laboratory results and drug compliance were outcome measures. Achievement of treatment objectives was measured by thyroid function tests (free and total thyroxine, total triiodothyronine, thyrotropin), number of clinic visits, and compliance (survey method). Additional measures were drug distribution patterns, drug costs, and pharmacy inventory costs. Restriction of levothyroxine's dosage strength did not significantly alter therapeutic outcomes. However, the restricted formulary was associated with more complex dosing regimens, and resulted in no significant cost savings. It is not known whether such restriction would adversely affect the care of patients of nonspecialists. Prospective studies are required to verify presumed cost-containment measures before such measures are adopted for widespread application.

Anaplastic thyroid carcinoma: behavior, biology, and therapeutic approaches.

Anaplastic thyroid carcinoma (ATC), although exceedingly rare, is the most aggressive solid tumor known. Early studies on the effects of different therapies may be biased by the inclusion of responsive "small cell" ATC patients, which are now known to be mostly lymphoma patients. Local control of disease with surgery and/or external beam radiotherapy (XRT) is of fundamental importance to enhance survival. Ultimately, nearly all ATC patients die from their disease, which is widely metastatic. Development of effective systemic chemotherapy agents would provide the best chance for long-term survival of patients. Early preliminary data suggest that paclitaxel may be helpful, although no agent has yet been identified to result in dramatic improvements in survival. Select patients may benefit from aggressive multimodal therapy, although it is important to provide appropriate palliative care when desired.

Cloning of the human sodium-iodide symporter promoter and characterization in a differentiated human thyroid cell line, KAT-50.

Elucidation of the regulation of human sodium-iodide symporter (hNIS) gene expression is critical to understanding its effects on iodide concentration abilities of thyroid and thyroid carcinomas. To explore this issue, a 1.2-kb portion of the 5'-flanking region of the hNIS gene was isolated and characterized. Transient transfections with chimeric luciferase-reporter constructs into a differentiated human thyroid cell line, KAT-50, as well as non-thyroidal cells, defined an active promoter with tissue-specificity. Reverse-transcriptase polymerase chain reaction analysis for hNIS mRNA expression in normal human tissues was positive in thyroid, salivary gland, omentum, and gallbladder. KAT-50 cells expressed hNIS mRNA and were capable of thyrotropin-responsive iodide uptake in vitro. Despite the failure to exhibit iodide concentration in clinical anaplastic carcinoma tumors, 4 of 5 cell lines from this cancer phenotype expressed hNIS mRNA. Definition of the active promoter provides further insights and tools to uncover new approaches to use of radioiodine for therapy of thyroid carcinomas.

Treatment of anaplastic thyroid carcinoma with paclitaxel: phase 2 trial using ninety-six-hour infusion. Collaborative Anaplastic Thyroid Cancer Health Intervention Trials (CATCHIT) Group.

Anaplastic thyroid carcinoma is a rare, lethal disease with no effective systemic therapies. Preclinical studies demonstrated antineoplastic activity of paclitaxel. This prompted a prospective phase 2 clinical trial to determine activity of paclitaxel against anaplastic thyroid carcinoma in patients with persistent or metastatic disease despite surgery or local radiation therapy. Twenty patients, entered through 6 of 12 study sites, were treated with 96-hour continuous infusion paclitaxel every 3 weeks for 1 to 6 cycles; the first 7 patients received 120 mg/m2 per 96 hours and the rest received 140 mg/m2 per 96 hours. Total responses to therapy were assessed using modified criteria with response durability acceptable at 2 or more weeks, due to the exceedingly rapid growth rate of this tumor. Plasma samples were obtained for pharmacokinetic analyses. Off-protocol, data showed that 9 patients were later treated with 225 mg/m2 paclitaxel as weekly 1-hour infusions. Nineteen evaluable patients demonstrated a 53% total response rate (95% confidence interval, 29%-76%) with one complete response and nine partial responses (including one off protocol). Results of historical review off-protocol showed 2 of 7 patients, with prior partial responses to the 96-hour infusion, had subsequent partial responses to weekly treatment and 1 of 2 prior nonresponders gained a partial response to weekly therapy. No toxicities greater than grade 2 were seen with 96-hour infusions, while peripheral neuropathy (up to grade 3) was most common with postprotocol weekly infusions. Paclitaxel appears to be the only agent with significant clinical systemic activity against anaplastic thyroid carcinoma; however, it is not capable of altering the lethality of this malignancy, suggesting the need for additional therapeutic innovations. Decreased time intervals between paclitaxel infusions may be more efficacious.

Thyroid hormone levels affected by time of blood sampling in thyroxine-treated patients.

Patients receiving exogenous levothyroxine are reported to have higher total and free serum thyroxine levels than euthyroid controls. This may be an artifact of the serum collection time. We explored the effect of collection time on serum levels of thyroid hormones in outpatients receiving levothyroxine for replacement therapy (26 patients) or suppression of thyrotropin (25 patients). Blood samples, obtained during regular clinic visits (random samples) and at more than 22 h from ingestion of levothyroxine (trough samples), were assayed for total and free thyroxine, triiodothyronine, and thyrotropin. Four athyreotic patients on levothyroxine therapy had serial blood sampling over 24 h. Compared to corresponding trough samples, random samples had elevated total thyroxine levels in patients receiving replacement (8.1 +/- 1.2%, mean +/- SE, p = 0.0001) and in patients undergoing suppression (8.8 +/- 1.6%, p = 0.0001). Free thyroxine was increased by 12.7 +/- 2.6% (p = 0.0003) and 14.5 +/- 2.3% (p = 0.0001), respectively, compared with trough samples. Thyrotropin levels were 18.9 +/- 6.8% (p = 0.003) lower in patients receiving replacement and triiodothyronine levels showed small or no changes. Time-course analysis showed that free and total thyroxine levels remained significantly elevated above baseline for 9 and 5 h, respectively, after a levothyroxine dose. In conclusion, there is a transient increase in thyroid hormone levels for 9 h after an oral levothyroxine dose. Accurate assessment of thyroid hormone levels in patients receiving levothyroxine therapy should take this into account. This has greatest significance in selecting minimal levothyroxine dosages for suppression of thyrotropin.