Paolo Vigneri

Diabetes and cancer

Diabetes and cancer are two heterogeneous, multifactorial, severe, and chronic diseases. Because of their frequency, reciprocal influences - even minor influences - may have a major impact. Epidemiological studies clearly indicate that the risk of several types of cancer (including pancreas, liver, breast, colorectal, urinary tract, and female reproductive organs) is increased in diabetic patients. Mortality is also moderately increased. Several confounding factors, having general or site-specific relevance, make it difficult to accurately assess cancer risk in diabetic patients. These factors include diabetes duration, varying levels of metabolic control, different drugs used for therapy, and the possible presence of chronic complications. Hyperinsulinemia most likely favors cancer in diabetic patients as insulin is a growth factor with pre-eminent metabolic but also mitogenic effects, and its action in malignant cells is favored by mechanisms acting at both the receptor and post-receptor level. Obesity, hyperglycemia, and increased oxidative stress may also contribute to increased cancer risk in diabetes. While anti-diabetic drugs have a minor influence on cancer risk (except perhaps the biguanide metformin that apparently reduces the risk), drugs used to treat cancer may either cause diabetes or worsen a pre-existing diabetes. In addition to the well-known diabetogenic effect of glucocorticoids and anti-androgens, an increasing number of targeted anti-cancer molecules may interfere with glucose metabolism acting at different levels on the signaling substrates shared by IGF-I and insulin receptors. In conclusion, diabetes and cancer have a complex relationship that requires more clinical attention and better-designed studies.

Targeting autophagy potentiates tyrosine kinase inhibitor-induced cell death in Philadelphia chromosome-positive cells, including primary CML stem cells.

Imatinib mesylate (IM), a potent inhibitor of the BCR/ABL tyrosine kinase, has become standard first-line therapy for patients with chronic myeloid leukemia (CML), but the frequency of resistance increases in advancing stages of disease. Elimination of BCR/ABL-dependent intracellular signals triggers apoptosis, but it is unclear whether this activates additional cell survival and/or death pathways. We have shown here that IM induces autophagy in CML blast crisis cell lines, CML primary cells, and p210BCR/ABL-expressing myeloid precursor cells. IM-induced autophagy did not involve c-Abl or Bcl-2 activity but was associated with ER stress and was suppressed by depletion of intracellular Ca2+, suggesting it is mechanistically nonoverlapping with IM-induced apoptosis. We further demonstrated that suppression of autophagy using either pharmacological inhibitors or RNA interference of essential autophagy genes enhanced cell death induced by IM in cell lines and primary CML cells. Critically, the combination of a tyrosine kinase inhibitor (TKI), i.e., IM, nilotinib, or dasatinib, with inhibitors of autophagy resulted in near complete elimination of phenotypically and functionally defined CML stem cells. Together, these findings suggest that autophagy inhibitors may enhance the therapeutic effects of TKIs in the treatment of CML.

Expression of Angiogenesis Stimulators and Inhibitors in Human Thyroid Tumors and Correlation with Clinical Pathological Features

Experimental evidence has shown, both in vitro and in animal models, that neoplastic growth and subsequent metastasis formation depend on the tumors ability to induce an angiogenic switch. This requires a change in the balance of angiogenic stimulators and inhibitors. To assess the potential role of angiogenesis factors in human thyroid tumor growth and spread, we analyzed their expression by semiquantitative RT-PCR and immunohistochemistry in normal thyroid tissues, benign lesions, and different thyroid carcinomas. Compared to normal tissues, in thyroid neoplasias we observed a consistent increase in vascular endothelial growth factor (VEGF), VEGF-C, and angiopoietin-2 and in their tyrosine kinase receptors KDR, Flt-4, and Tek. In particular, we report the overexpression of angiopoietin-2 and VEGF in thyroid tumor progression from a prevascular to a vascular phase. In fact, we found a strong association between tumor size and high levels of VEGF and angiopoietin-2. Furthermore, our results show an increased expression of VEGF-C in lymph node invasive thyroid tumors and, on the other hand, a decrease of thrombospondin-1, an angioinhibitory factor, in thyroid malignancies capable of hematic spread. These results suggest that, in human thyroid tumors, angiogenesis factors seem involved in neoplastic growth and aggressiveness. Moreover, our findings are in keeping with a recent hypothesis that in the presence of VEGF, angiopoietin-2 may collaborate at the front of invading vascular sprouts, serving as an initial angiogenic signal that accompanies tumor growth.

Induction of apoptosis in chronic myelogenous leukemia cells through nuclear entrapment of BCR-ABL tyrosine kinase.

The chimeric BCR–ABL oncoprotein is the molecular hallmark of chronic myelogenous leukemia (CML). BCR–ABL contains nuclear import and export signals but it is localized only in the cytoplasm where it activates mitogenic and anti-apoptotic pathways. We have found that inhibition of the BCR–ABL tyrosine kinase, either by mutation or by the drug STI571, can stimulate its nuclear entry. By combining STI571 with leptomycin B (LMB) to block nuclear export, we trapped BCR–ABL in the nucleus and the nuclear BCR–ABL tyrosine kinase activates apoptosis. As a result, the combined treatment with STI571 and LMB causes the irreversible and complete killing of BCR–ABL transformed cells, whereas the effect of either drug alone is fully reversible. The combined treatment with STI571 and LMB also preferentially eliminates mouse bone marrow cells that express BCR–ABL. These results indicate that nuclear entrapment of BCR–ABL can be used as a therapeutic strategy to selectively kill chronic myelogenous leukemia cells.

Cancer Stem Cells and Chemosensitivity

Cancer lethality is mainly due to the onset of distant metastases and refractoriness to chemotherapy. Thus, the development of molecular targeted agents that can restore or increase chemosensitivity will provide valuable therapeutic options for cancer patients. Growing evidence indicates that a cellular subpopulation with stem cell–like features, commonly referred to as cancer stem cells (CSCs), is critical for tumor generation and maintenance. Recent advances in stem cell biology are revealing that this cellular fraction shares many properties with normal adult stem cells and is able to propagate the parental tumor in animal models. CSCs seem to be protected against widely used chemotherapeutic agents by means of different mechanisms, such as a marked proficiency in DNA damage repair, high expression of ATP-binding cassette drug transporters, and activation of PI3K/AKT and Wnt pathways. Moreover, microenvironmental stimuli such as those involved in the epithelial-mesenchymal transition and hypoxia indirectly contribute to chemoresistance by inducing in cancer cells a stem-like phenotype. Understanding how CSCs overcome chemotherapy-induced death stimuli, and integrating such knowledge into clinical research methodology, has become a priority in the process of identifying innovative therapeutic strategies aimed at improving the outcome of cancer patients. Clin Cancer Res; 17(15); 4942–7. ©2011 AACR.

V gamma 9V delta 2 T lymphocytes efficiently recognize and kill zoledronate-sensitized, imatinib-sensitive, and imatinib-resistant chronic myelogenous leukemia cells.

Imatinib mesylate (imatinib), a competitive inhibitor of the BCR-ABL tyrosine kinase, is highly effective against chronic myelogenous leukemia (CML) cells. However, because 20–30% of patients affected by CML display either primary or secondary resistance to imatinib, intentional activation of Vγ9Vδ2 T cells by phosphoantigens or by agents that cause their accumulation within cells, such as zoledronate, may represent a promising strategy for the design of a novel and highly innovative immunotherapy capable to overcome imatinib resistance. In this study, we show that Vγ9Vδ2 T lymphocytes recognize, trogocytose, and efficiently kill imatinib-sensitive and -resistant CML cell lines pretreated with zoledronate. Vγ9Vδ2 T cell cytotoxicity was largely dependent on the granule exocytosis- and partly on TRAIL-mediated pathways, was TCR-mediated, and required isoprenoid biosynthesis by zoledronate-treated CML cells. Importantly, Vγ9Vδ2 T cells from patients with CML can be induced by zoledronate to develop antitumor activity against autologous and allogeneic zoledronate-treated leukemia cells, both in vitro and when transferred into immunodeficient mice in vivo. We conclude that intentional activation of Vγ9Vδ2 T cells by zoledronate may substantially increase their antileukemia activities and represent a novel strategy for CML immunotherapy.

The changing epidemiology of thyroid cancer: why is incidence increasing?

Purpose of review Thyroid cancer incidence is increasing worldwide. Causes are highly debated. Recent findings Thyroid cancer increase has been associated to socioeconomic status, better access to healthcare and rising use of thyroid imaging. Therefore, the rise could be apparent because of the useless identification of a large reservoir of subclinical papillary lesions that will never affect patient health (overdiagnosis). However, not all epidemiological and clinical data support this hypothesis. The increasing number of large tumors, the increasing thyroid cancer-related mortality in spite of earlier treatment and the changes in thyroid cancer molecular profile suggest a true increase. Recently increased and thyroid-specific environmental carcinogens could be responsible, such as radiation (mostly medical radiation), increased iodine intake and chronic lymphocytic thyroiditis and environmental pollutants such as nitrates, heavy metals and other compounds largely used in the industrialized society. Possible mechanisms await further investigation. Summary The increased incidence of thyroid cancer is the likely result of two coexisting processes: increased detection (apparent increase) and increased number of cases (true increase) due to unrecognized thyroid-specific carcinogens. To identify causal factors and to differentiate stationary cancers from those that will progress are major urgent requirements for both clinical and scientific purposes.

Altered Expression of c-IAP1, Survivin, and Smac Contributes to Chemotherapy Resistance in Thyroid Cancer Cells

Resistance to chemotherapy predicts an unfavorable outcome for patients with radioiodine-insensitive thyroid cancer. To investigate the mechanisms underlying this resistance, we evaluated the expression of four different inhibitor of apoptosis proteins, and their antagonist, Smac, in thyroid cancer cells that survived 48 hours of exposure to cisplatin, doxorubicin, or taxol. We found high levels of c-IAP1 after cisplatin treatment and increased expression of survivin following exposure to doxorubicin. Cells that endured treatment with taxol showed reduced expression of Smac and released minimal amounts of this protein from the mitochondria. Down-regulation of c-IAP1 and survivin increased the cytotoxicity of cisplatin and doxorubicin, whereas overexpression of Smac improved the efficacy of taxol. Finally, thyroid cancer cells permanently resistant to doxorubicin or cisplatin showed increased expression of c-IAP1 and survivin, respectively. However, silencing of these proteins by RNA interference restored sensitivity to doxorubicin and cisplatin. Thus, in thyroid cancer cells, early resistance to chemotherapeutic agents requires high levels of c-IAP1 and survivin and low levels of Smac. Furthermore, increased expression of c-IAP1 and survivin contributes to the acquisition of permanent resistance to cytotoxic compounds.

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.

Tyrosine kinase inhibitor STI571 enhances thyroid cancer cell motile response to Hepatocyte Growth Factor

The Hepatocyte Growth Factor (HGF) and its receptor Met are physiological regulators of cell migration. HGF and Met have also been implicated in tumor progression and metastasis. We show here that the tyrosine kinase inhibitor STI571 has a stimulatory effect on HGF-induced migration and branching morphogenesis in thyroid cancer but not in primary or immortalized thyroid epithelial cells. These stimulatory effects of STI571 are observed at a concentration that is clinically relevant. The STI571-enhanced motile response can be correlated with an increase in the Met receptor tyrosine phosphorylation as well as ERK and Akt activation by HGF. Interestingly, one of the targets of STI571, namely the c-Abl tyrosine kinase, is activated by HGF and is recruited at the migrating edge of thyroid cancer cells. These data suggests that c-Abl and/or STI571-inhibited tyrosine kinases can negatively regulate the Met receptor to restrain the motile response in thyroid cancer cells.