Biological effects of insulin and its analogs on cancer cells with different insulin family receptor expression.

Hyperinsulinemia is a likely cause of the increased cancer incidence and mortality in diabetic patients, but its role is difficult to define in vivo. Previous in vitro studies testing the mitogenic potential of insulin and its analogs provided incomplete and sometimes contradictory results. To better evaluate cancer cell responsiveness to insulin, to its analogs and to IGF-I, we measured under identical experimental conditions cell proliferation, invasiveness, and foci formation in six cancer cell lines with different insulin receptor family expression levels. The cancer cells studied have a different expression of insulin receptor (IR), its isoforms (IR-A and IR-B), and of the IGF-I receptor. The data indicate that insulin stimulates proliferation in all cancer cell lines, invasiveness in some, and foci formation in none. Cancer cell responses to insulin (and IGF-I) are not related to receptor expression levels; moreover, hormone-stimulated proliferation and invasiveness are not correlated. IGF-I is a more potent stimulator than insulin in most but not all cancer cell lines. Insulin analogs including M1 and M2 Glargine metabolites stimulate cancer cells similar to insulin. However, exceptions occur for specific analogs in particular cancer cells. In conclusion, in vitro insulin is an effective growth factor for all cancer cells but the biological response to insulin cannot be predicted on the basis of receptor expression levels. In the clinical setting, these observations should be taken in account when deciding treatment for diabetic patients who are at risk of undiscovered cancer or survivors of oncological diseases.

Thyroid cancer in thyroglossal duct cysts requires a specific approach due to its unpredictable extension.

CONTEXT: Differentiated thyroid cancer (DTC) in thyroglossal duct cysts is uncommon. The requirement of total thyroidectomy and lymph node dissection is still controversial. SETTING: The study was performed in a referral thyroid cancer center at an academic hospital. PATIENTS: We conducted a single center retrospective study of a consecutive series of 26 patients with DTC in thyroglossal duct cyst, all having undergone cyst resection and total thyroidectomy. MAIN OUTCOME MEASURES: Diagnostic modalities, surgical treatment, histopathological features, and clinical outcome were included in the study. RESULTS: Thyroglossal duct cyst cancer histotype was papillary in 23 of 26 patients (88.5%) and follicular-Hurthle in 3 of 26 cases (11.5%). A concomitant papillary DTC in the thyroid gland was found in 16 of 26 cases (61.5%), and it was multifocal in 8 of 16 cases (50%). At presentation, the patients with cancer in both the thyroglossal duct cyst and the thyroid were older than the patients who only had cancer in the thyroglossal duct cyst (44.9 ± 7.6 vs 32.0 ± 12.7; P = .006). Lymph node dissection, performed in 17 of 26 patients (65.4%), indicated that the central compartment was involved in 6 patients (35.3%, all having cancer also in the thyroid), the laterocervical compartments in 10 patients (58.8%), and the submental in 4 (23.5%). Six patients (23.1%) had persistent disease at 6-year median follow-up. CONCLUSIONS: DTC in thyroglossal duct cysts occurs at a younger age and with more aggressive features at presentation. Concomitant cancer in the thyroid and lymph node metastases is present in most cases. Lymph node compartment involvement is different from that of cancers in the thyroid gland. Therefore, surgical treatment should include both thyroglossal duct cyst resection and total thyroidectomy, with individualized surgical nodal dissection. Subsequent management should follow current DTC guidelines.

Update on thyroid cancer treatment.

Surgery and radioiodine therapy are usually effective for most patients with differentiated thyroid cancer. However, poorly differentiated and anaplastic thyroid carcinomas represent a challenge to physicians on the basis of the current cancer treatment modalities. These cancer subtypes are often lethal and refractory to radioiodine therapy as well as most of the common chemotherapy drugs. Several kinase inhibitors are promising targeted therapies for these malignancies; however, clinical trials involving these drugs have provided controversial results and their clinical use is still under debate. Advanced medullary thyroid carcinomas may also be refractory to conventional therapies and novel kinase inhibitors may also be useful to control tumor progression in certain patients. Novel evidence is emerging that thyroid cancer is a stem cell disease, thereby implying that the driving force of thyroid cancers is a subset of undifferentiated cells (thyroid cancer stem cells) with unlimited growth potential and resistance to conventional therapeutic regimens. Thyroid cancer stem cells have been proposed as responsible for tumor invasiveness, metastasis, relapse and differentiation. Therefore, drugs that selectively target these cells could serve as a cornerstone in the treatment of poorly differentiated thyroid cancer.

Severe Graves' ophthalmopathy after percutaneous ethanol injection in a nontoxic thyroid nodule.

BACKGROUND: Percutaneous ethanol injection (PEI) is used to treat cystic or mixed benign thyroid nodules. This treatment can result in rare complications, and three cases of Graves' disease (GD) without Graves' ophthalmopathy (GO) have been reported after PEI treatment of toxic thyroid adenomas. Here we present a 55-year-old male patient who developed GD and severe GO after PEI treatment of a mixed cystic-solid, nontoxic thyroid nodule. PATIENT FINDINGS: Six months after PEI, the nodule volume had decreased from 8.9 to 3.0 mL, but we observed severe hyperthyroidism with elevated serum free triiodothyronine, free thyroxine, and thyrotropin receptor antibody levels. We also observed ophthalmopathy with symmetrical orbit and soft tissue involvement (grade b/c) and a clinical activity score of 4/7. The diagnosis of GO was confirmed by bilateral corneal damage, increased intraocular pressure on upgaze, and inconstant diplopia. A computed tomography scan showed that the inferior, medial, and superior extraocular muscles were bilaterally enlarged, the perineural space at the orbital cone was slightly reduced and the ophthalmic vein was congested. SUMMARY: A cause-effect relationship between PEI and GD/GO was likely in this patient because of the temporal sequence. Although the mechanism was unknown, we speculated that the thyroid tissue damage caused by PEI released a large amount of antigenic materials from follicular thyroid cells, including thyrotropin receptor protein, which triggered the autoimmune inflammatory response against the thyroid itself and the orbital soft tissues. The patient did not have any risk factors for either GD or GO. CONCLUSIONS: This observation raises the concern, therefore, that unpredictable and severe complications, such as GD and GO, may occur in a few patients treated with PEI.

A diffuse sclerosing variant of papillary thyroid carcinoma: clinical and pathologic features and outcomes of 34 consecutive cases.

BACKGROUND: The diffuse sclerosing variant of papillary thyroid carcinoma (DSPC) is a relatively rare variant of papillary thyroid cancer. Large studies of patients with DSPC have been infrequently performed, and controversy still exists concerning some DSPC features and outcomes. The aim of the present study was to retrospectively evaluate the clinicopathologic features and outcomes in a series of 34 consecutive patients with DSPC and to compare them with a larger group of 245 consecutive patients with the classic variant of papillary thyroid carcinoma (cPTC) that were evaluated in the same period. PATIENTS AND METHODS: Clinical and histological features (sex, age, tumor size,multifocality, bilaterality, extra thyroid extension, and local and distant metastases) were recorded in all patients, as well as any persistent or recurrent disease and the patients' disease status at last observation. Patients with cPTC were classified as either low (122) or high risk (123). DSPC and high-risk patients were all treated with the same protocol, including ¹³¹I treatment. All patients were included in a Cox regression model analysis to investigate the effect of each variable on the hazard ratio. RESULTS: As expected, multifocality, bilaterality, and extra thyroid extension were more frequently noted at presentation, and the pT1 category of TNM classification was less frequently noted in DSPC and high-risk patients with cPTC compared with low-risk patients with cPTC. No significant difference was found between patients with DSPC and those with high-risk cPTC, except that extra thyroid extension was found more frequently in the patients with DSPC. Using multivariate analysis, diffuse sclerosing variant was an independent variable for predicting a high risk of persistent and recurrent disease during initial follow-up. However, at a later time, and after further treatment, the disease status was not different between patients with DSPC and those with high-risk cPTC, and only the presence of distant metastases affected the final outcome. CONCLUSIONS: DSPC is a thyroid papillary carcinoma variant characterized by high aggressiveness. In patients with DSPC, the outcome is worse than in patients with low-risk cPTC; and, at presentation, characteristics are somewhat worse than for patients with high-risk cPTC.At medium term, the outcome is similar to that observed in patients with high-risk cPTC, provided aggressive treatment is used (additional surgical intervention, when required, and/or ¹³¹I radiotherapy).

The role of insulin receptors and IGF-I receptors in cancer and other diseases.

There is evidence, both in vitro and in vivo, that receptor tyrosine kinases play a key role in the formation and progression of human cancer. In particular, the insulin-like growth factor receptor (IGF-IR), a tyrosine kinase receptor for IGF-I and IGF-II, has been well documented in cell culture, animal studies, and humans to play a role in malignant transformation, progression, protection from apoptosis, and metastasis. In addition, the hormone insulin (which is very closely related to the IGFs) and its tyrosine kinase receptor (the IR, which is very closely related to the IGR-IR) have been documented both in vitro and in vivo to play a key role in cancer biology. Indeed, several epidemiological studies have shown that insulin resistance status, characterized by hyperinsulinaemia, is associated with an increased risk for a number of malignancies, including carcinomas of the breast, prostate, colon and kidney. Recent data have elucidated some molecular mechanisms by which IR is involved in cancer. IR is over-expressed in several human malignancies. Interestingly, one of the two IR isoform (IR-A) is especially over-expressed in cancer. IR-A is the IR foetal isoform and has the peculiar characteristic to bind not only insulin but also IGF-II. In addition, the IR contributes to formation of hybrid receptors with the IGF-IR (HR). By binding to hybrid receptors, insulin may stimulate specific IGF-IR signalling pathways. Over-expression of IR-A is, therefore, a major mechanism of IGF system over-activation in cancer. In this respect, IR-A isoform and hybrid receptors should be regarded as potential molecular targets, in addition to IGF-IR, for novel anti-cancer therapy. These findings may have important implications for both the prevention and treatment of common human malignancies. They underline the concept that hyperinsulinaemia, associated with insulin resistance and obesity, should be treated by changes in life style and/or pharmacological approaches to avoid an increased risk for cancer. Moreover, native insulin and insulin analogue administration should be carefully evaluated in terms of the possible increase in cancer risk.

TAp73 alpha increases p53 tumor suppressor activity in thyroid cancer cells via the inhibition of Mdm2-mediated degradation.

p53 family proteins include p53 tumor suppressor, p63, and p73. Despite the high similarity in structure and function with p53, p63, and p73 function in tumor suppression is still controversial. Here, we show that TAp73alpha, a transcriptionally active p73 isoform, is able to synergize p53 tumor suppressor function in thyroid cancer cells. Indeed, depletion of p73 by small interfering RNA in thyroid cancer cells resulted in a reduced transcriptional activity of p53. Ectopic coexpression of both p53 and TAp73alpha in thyroid cancer cells resulted in increased transcription and tumor suppressor function compared with p53 or TAp73alpha alone, as well as in increased p53 protein levels. The enhancing effect of TAp73alpha on p53 activity is Mdm2 dependent because it is prevented by Mdm2 depletion by small interfering RNA. At least two mechanisms may explain the interference of TAp73alpha with p53 function. First, in thyroid cancer cells, TAp73alpha inhibits the effect of p53 on Mdm2 induction by antagonizing p53 at the Mdm2 promoter level. Second, a TAp73alpha mutant (G264W), which is devoid of DNA binding capability, is still able to increase p53 protein levels by competing with p53 for Mdm2 protein binding. Taken together, these results indicate that in thyroid cancer cells, TAp73alpha is able to increase p53 protein level and function by interfering with Mdm2-mediated p53 degradation. These results may be useful for designing gene therapies aimed at restoring a normal p53 function in thyroid cancer cells.

Effects of either LT4 monotherapy or LT4/LT3 combined therapy in patients totally thyroidectomized for thyroid cancer.

After total thyroidectomy all thyroid cancer patients require lifelong treatment with thyroid hormones; the treatment of choice is synthetic levothyroxine (LT4). The question of whether these patients might benefit from the combined LT4 and liothyronine (LT3) treatment has been addressed with conflicting conclusions. The aim of the present study was to compare the effects of combined low LT4/LT3 molar ratio therapy versus LT4 monotherapy on various target organs and tissues in patients thyroidectomized for thyroid cancer. Urine collection (24 hour), a fasting blood sample for laboratory examinations, thyroid function clinical score, and cardiovascular, neurological, and neuropsychological evaluations were obtained. Clinical parameters and peripheral markers of thyroid function were measured during the two different treatment regimens in 20 patients. Mean serum aspartate aminotransferase, alanine aminotransferase, sex hormone binding globulin, and osteocalcin values were significantly higher during the combined treatment. No significant differences in the clinical score, the systolic and diastolic performance, and the neurological and neuropsychological evaluations were observed between the two treatment regimens. Moreover, no alteration due to subclinical hyperthyroidism or to the fluctuations in serum T3 concentrations during the combined therapy was observed. In conclusion, we found no evidence that combined therapy with a low LT4/LT3 molar ratio resulted in improved well-being and cognitive function or in increased thyroid hormone action on peripheral tissues in respect to LT4 monotherapy. Until future large, blind, randomized, and controlled trials prove otherwise, LT4 should remain the standard treatment for thyroid cancer patients.

HMGA1 inhibits the function of p53 family members in thyroid cancer cells.

HMGA1 is an architectural transcription factor expressed at high levels in transformed cells and tumors. Several lines of evidence indicate that HMGA1 up-regulation is involved in the malignant transformation of thyroid epithelial cells. However, the mechanisms underlying the effect of HMGA1 on thyroid cancer cell phenotype are not fully understood. We now show that in thyroid cancer cells, HMGA1 down-regulation by small interfering RNA and antisense techniques results in enhanced transcriptional activity of p53, TAp63alpha, TAp73alpha, and, consequently, increased apoptosis. Coimmunoprecipitation and pull-down experiments with deletion mutants showed that the COOH-terminal oligomerization domain of p53 family members is required for direct interaction with HMGA1. Moreover, inhibition of HMGA1 expression in thyroid cancer cells resulted in increased p53 oligomerization in response to the DNA-damaging agent doxorubicin. Finally, electrophoretic mobility shift assay experiments showed that the p53-HMGA1 interaction results in reduced DNA-binding activity. These results indicate a new function of HMGA1 in the regulation of p53 family members, thus providing new mechanistic insights in tumor progression.

High prevalence of differentiated thyroid carcinoma in acromegaly.

OBJECTIVE: Acromegaly is a chronic disease caused by increased GH secretion and associated with a greater risk of developing both benign and malignant tumours. In the present study we evaluated the prevalence of thyroid disorders and thyroid malignancies in a series of acromegalic subjects. DESIGN AND PATIENTS: We studied, retrospectively, a continuous series of 125 acromegalic patients referred to the Endocrinology Centres at the University Hospitals of Catania and Ferrara, Italy, over a 22-year period. MEASUREMENTS: Diagnosis and management of acromegaly were based on standardized criteria. In all patients thyroid function and morphology were evaluated by serum free T4, free T3, TSH measurements and ultrasound scanning, respectively. Fine-needle aspiration biopsy (FNAB) was performed in all solid-mixed thyroid nodules of diameter greater than 1 cm. IGF-1 expression was assessed by semiquantitative immunohistochemical analysis in some patients with differentiated thyroid cancer. RESULTS: Abnormal thyroid function was found in eight cases (6%). A diffuse or nodular goitre was present in 102 cases (82%). Thyroidectomy was performed in 17 patients. Histological examination revealed a differentiated thyroid cancer in seven of the patients. No significant association of thyroid cancer with GH or IGF-1 levels was found. Semiquantitative assessment of IGF-1 expression by immunohistochemistry revealed a more intense staining in thyroid cancer from acromegalic subjects than in papillary thyroid cancer from nonacromegalic subjects. CONCLUSIONS: The frequency of thyroid disorders in our series of acromegalic subjects was similar to that previously observed in these patients. However, the prevalence of thyroid cancer was not only strikingly elevated (5.6%) in comparison to the estimated prevalence in the general population (0.093%), but it was even higher than that reported for acromegalic subjects. Sustained exposure to high serum IGF-1 levels is likely to play a role in the development of thyroid cancer in this disease. An additive role for the autocrine/paracrine action of locally produced IGF-1 is also possible. Our results suggest that thyroid function and morphology should be carefully monitored in all acromegalic patients.

Left ventricular function in acute hypothyroidism: a Doppler echocardiography study.

BACKGROUND: Acute changes in cardiac parameters may occur after L-thyroxine withdrawal in patients totally thyroidectomized for thyroid cancer. The literature data regarding cardiac function in acute hypothyroidism are limited and discordant. METHODS: In order to evaluate the effects of acute hypothyroidism on cardiac function, 20 athyreotic patients (3 males, 17 females, mean age 46.4 +/- 8.6 years, range 18-58 years) underwent Doppler echocardiography during L-thyroxine therapy (euthyroid phase) and 5 weeks after hormone therapy withdrawal (hypothyroid phase). RESULTS: Significant changes in the left ventricular mass (83 +/- 12 vs 93 +/- 17 g/m2, p = 0.004) and end-diastolic volume (56 +/- 9 vs 50 +/- 9 ml/m2, p = 0.01) were found. Among systolic function parameters, the pre-ejection period/left ventricular ejection time (PEP/LVET) ratio (0.33 +/- 0.07 vs 0.40 +/- 0.08, p = 0.0002), aortic peak flow velocity corrected for heart rate (3.9 +/- 0.7 vs 3.5 +/- 0.5 cm/s, p = 0.02) and mean aortic acceleration corrected for heart rate (45 +/- 15 vs 38 +/- 9 cm/s2, p = 0.007) showed significant variations, whereas the left ventricular fractional shortening (39 +/- 5 vs 40 +/- 6%, p = NS) and ejection fraction (69 +/- 6 vs 68 +/- 7%, p = NS) did not change. Among diastolic function parameters, only the E-wave velocity decreased (73 +/- 17 vs 65 +/- 12 cm/s, p = 0.01); no significant modification was found in the A-wave velocity (62 +/- 19 vs 58 +/- 14 cm/s, p = NS), E/A ratio (1.2 +/- 0.5 vs 1.1 +/- 0.3, p = NS), isovolumic relaxation time (93 +/- 16 vs 95 +/- 37 ms, p = NS) and E-wave deceleration time (233 +/- 48 vs 235 +/- 45 ms, p = NS). The pattern of left ventricular filling remained unchanged, except in 2 patients. The Suga-Sagawa's index, a known parameter of myocardial contractility, was unchanged (5.6 +/- 2 vs 6.1 +/- 2 mmHg/ml, p = NS). The systemic vascular resistance increased (1511 +/- 599 vs 2216 +/- 408 dynes-s-cm(-5), p = 0.002), while the stroke index (39 +/- 8 vs 33 +/- 7 ml/m2, p = 0.001) and cardiac index (2.74 +/- 0.6 vs 2.07 +/- 0.5 l/min/m2, p = 0.0001) significantly decreased. CONCLUSIONS: Acute hypothyroidism was associated with left ventricular systolic dysfunction, probably due to pre- and afterload alterations rather than to an impaired myocardial contractility. The diastolic function was not significantly modified. An increase in cardiac mass was also found, possibly a consequence of early interstitial myxedema. Unlike the PEP/LVET ratio, both the fractional shortening and ejection fraction may be unreliable indicators of left ventricular systolic dysfunction in patients with acute hypothyroidism.

The diagnostic use of the rhTSH/thyroglobulin test in differentiated thyroid cancer patients with persistent disease and low thyroglobulin levels.

BACKGROUND: Serum thyroglobulin (Tg) measurement after TSH stimulation, by either thyroid hormone withdrawal or recombinant human TSH (rhTSH) administration, is the most sensitive method for early detection of patients with persistent or recurrent differentiated thyroid cancer (DTC) after total thyroidectomy and 131I ablation. The use of rhTSH is now increasing because it avoids thyroid hormone suppressive therapy (THST) withdrawal and the consequent symptoms of severe hypothyroidism. Current guidelines suggest measurement of serum Tg 4 days after starting a 2-day course of rhTSH injections, and assumes that Tg reaches maximum serum levels at that time. OBJECTIVE: The present study was carried out to evaluate the accuracy of rhTSH/thyroglobulin test in DTC patients with persistent disease and low thyroglobulin levels. PATIENTS AND MEASUREMENTS: A series of 13 DTC patients was selected because they had proven persistent disease associated with low Tg levels (< 2.0 micro g/l) under l-thyroxine treatment. In all of them, serum Tg was > 5.0 micro g/l at the last THST withdrawal. We measured serum Tg and TSH levels on days 0.5, 1, 1.5, 2, 4, 7, 10 and 15 after the first of a 2-day course of intramuscular rhTSH injections. RESULTS: Serum Tg values were variable in terms of both peak and time-course. Detectable serum Tg levels were recorded on day 4 in all patients. However, among these 13 patients, the peak Tg value was reached earlier than day 4 in three patients and later in two others. In one patient, Tg level at day 2 was higher (3.0 micro g/l) than at day 4 (1.8 micro g/l). In six of the 13 patients studied we compared Tg values after rhTSH to those subsequently obtained after THST withdrawal: in five of them Tg values were two to three times higher after the latter stimulation. Serum Tg value variability after rhTSH was partially accounted for by variability of serum TSH levels, which were inversely related to patient body surface. CONCLUSIONS: In DTC patients with persistent disease and low Tg levels, optimization of the diagnostic use of Tg measurement after rhTSH may require rhTSH dose adjustment to the patient body surface area and repeated blood sampling, in order to improve diagnostic accuracy. In these patients not even a TSH-stimulated serum Tg cut-off of 2.0 micro g/l on day 4 provides 100% accuracy, whereas a cut-off of 1.0 micro g/l seems more appropriate. Therefore, in this subset of patients, if any detectable Tg level >or= 1.0 micro g/l is found after rhTSH, re-evaluation after THST should be advised.

False-positive findings on (131)I whole-body scans because of posttraumatic superficial scabs.

UNLABELLED: Nonspecific (131)I uptake may affect both the specificity and the accuracy of whole-body scanning (WBS) performed after (131)I administration in the follow-up of thyroid carcinoma after thyroidectomy. We report a newly identified cause of false-positive WBS findings: posttraumatic superficial scabs. METHODS: Four patients who underwent thyroidectomy for differentiated thyroid carcinoma were studied after therapeutic administration of 3,700 MBq (131)I. RESULTS: WBS revealed an area of uptake in the lower limbs, in a site corresponding to a slight abrasion of the skin that had incidentally occurred between a few hours before and 24 h after radioiodine administration. In 2 patients, a radioiodine concentration in the scab was shown by the disappearance of the radioactivity in the leg after removal of the scab and by detection of radioactivity in the collected material. CONCLUSION: Posttraumatic superficial scabs may be a cause of false-positive WBS findings. High (131)I doses, although providing increased sensitivity, may also increase the number of false-positive results.

A novel autocrine loop involving IGF-II and the insulin receptor isoform-A stimulates growth of thyroid cancer.

The insulin receptor (IR) occurs in two isoforms (IR-A and IR-B) resulting from alternative splicing of exon 11 of the gene. The IR-A isoform is predominantly expressed in fetal tissues and malignant cells and binds IGF-II with high affinity. We previously observed that IRs are overexpressed in thyroid cancer cells; now we evaluated whether these cells preferentially express IR-A and produce IGF-II, which would activate a growth-promoting autocrine loop. The IR content ranged 6.0-52.6 ng/100 microg cell membrane protein in thyroid cancer primary cultures (n = 8) and permanent cell lines (n = 6) vs. 1.2-1.7 in normal thyroid cells (n = 11 primary cultures; P < 0.0001). IR-A isoform relative abundance ranged from 36-79% in cancer cells (with the highest values in undifferentiated cancers) vs. 27-39% in normal cells. Similar results were obtained in normal vs. cancer thyroid tissue specimens. IGF-II caused IR autophosphorylation with an ED(50) of 1.5-40.0 nM in cancer cells vs. more than 100 nM in normal cells; IGF-II affinity correlated with the relative abundance of IR-A (r = 0.628; P < 0.0001). IGF-II was expressed in all cancer cells, highly expressed in anaplastic cells, and less expressed in normal cells. In conclusion, malignant thyrocytes, especially when poorly differentiated, produce IGF-II and overexpress IR, predominantly as IGF-II-sensitive isoform A. A growth-promoting autocrine loop is activated, therefore, and may affect thyroid cancer biology.