Progress and pitfalls in the identification of cancer stem cell-targeting therapies in head and neck squamous cell carcinoma.

Head and neck squamous cell carcinoma (HNSCC) is a highly prevalent and deadly cancer that has not shown improvements in survival rates for many years. Current treatments of HNSCC include surgical resection, radiotherapy, and chemotherapy, which are relatively ineffective for the management of recurrent or metastatic tumors. Cancer stem cells (CSC) within HNSCC offer an attractive therapeutic target for improving the survival rates for such cases, as eliminating the cells responsible for tumor initiation will theoretically prevent the onset of metastasis and recurrence. Since CSCs were initially isolated from HNSCC, there have been a handful of papers published that examine the potential of certain agents to selectively inhibit HNSCC CSCs, although a review of these papers has not yet been performed. Here we review the current literature for potential compounds or particles which have been proposed to selectively target the HNSCC CSC subpopulation. The various agents that have been tested so far include RNA interference, cell-based immunotherapy, antibodies, chemicals, microRNA precursors, and lentiviral microRNA vectors. Although many of these compounds showed considerable promise, few, if any, of the studies provided comprehensive evidence showing that the proposed agents were specific to CSCs and were considerably more effective than conventional therapy (radiation, cisplatin, etc). The proposed treatments in these studies require further investigation in these two regards, especially through rigorous in vivo experimentation, before they can be considered as true potential CSC inhibitors, let alone be considered for use in clinical trials.

[Immunohistochemistry study of EGFR expression in head and neck squamous cell carcinoma]. / Estudio por inmunohistoquímica de la expresión de EGFR en carcinoma de células escamosas de cabeza y cuello.

EGFR is an important transmembrane receptor member of the family of tyrosine kinases, that translates signals from both outside and inside the cell and plays a key role in numerous proceses that affect tumour development, growth, progresion, differentiation, inhibition of apoptosis and metastasis. Immunohistochemistry studies have shown that 40-80% of head and neck squamous cell carcinomas express EGFR and it has been suggested as a potential independent prognostic parameter. The objective of this study is to evaluate by immunohistochemistry the expresi6n of EGFR in a series of Head and Neck Squamous Cell Carcinoma and correlate it to clinico-pathological features and prognostic significance. We investigated expression of EGFR in 44 samples. There was a high expression in 41% of the cases. Even if we have not found that the expression of EGFR correlated with the prognosis of these patients the presence of EGFR is very important because there are chemical agents or drugs that can inhibit its activity.

Estudio por inmunohistoquímica de la expresión de EGFR en carcinoma de células escamosas de cabeza y cuello / Immunohistochemistry study of EGFR expression in head and neck squamous cell carcinoma

EGFR es un receptor transmembrana miembro de la familia de tirosín-kinasas que traduce señales tanto del exterior como del interior de la célula y juega un papel importante en numerosos procesos que afectan a la evolución del tumor, crecimiento, progresión, diferenciación, inhibición de apoptosis y al desarrollo de metástasis. Su expresión mediante inmunohistoquímica en carcinoma de células escamosas varía entre el 40-80% y se le ha adjudicado un valor pronóstico independiente en estos carcinomas. El objetivo del presente estudio es el de analizar por inmunohistoquímica la expresión de esta proteína en una serie de carcinomas epidermoides de cabeza y cuello y correlacionarlo con parámetros clínico-patológicos y supervivencia. Se realiza inmunohistoquímica de 44 muestras en las que el 41% es positivo. A pesar de no haber encontrado relación con el pronóstico de estos pacientes, la presencia de EGFR es muy importante por el hecho de que existen fármacos y agentes químicos capaces de inhibir su actividad

EGFR is an important transmembrane receptor member of the family of tyrosine kinases, that translates signals from both outside and inside the cell and plays a key role in numerous proceses that affect tumour development, growth, progresion, differentiation, inhibition of apoptosis and metastasis. Immunohistochemistry studies have shown that 40-80% of head and neck squamous cell carcinomas express EGFR and it has been suggested as a potential independent prognostic parameter. The objective of this study is to evaluate by immunohistochemistry the expresión of EGFR in a series of Head and Neck Squamous Cell Carcinoma and correlate it to clinico-pathological features and prognostic significance. We investigated expression of EGFR in 44 samples. There was a high expression in 41% of the cases. Even if we have not found that the expression of EGFR correlated with the prognosis of these patients the presence of EGFR is very important because there are chemical agents or drugs that can inhibit its activity

A possible role of p73 on the modulation of p53 level through MDM2.

MDM2, one of the transcriptional targets of p53, can target p53 for degradation in a negative feedback loop. The p53-related protein p73, however, can bind to MDM2 but is not consequently down-regulated. Here we demonstrate that p73 could transactivate the MDM2 promoter in p53-null cell lines. In p53-null cell lines, the level of MDM2 was increased by p73 due to increases in transcription and protein stability of MDM2. In transient transfection assays, inhibition of the transcriptional activity of p73 required a higher amount of MDM2 than that of p53. This is probably due to the fact that MDM2 can target p53, but not p73, for degradation. We demonstrated further that the level of p53 could be altered by a cooperation between MDM2 and p73, but not by transcriptional inactive mutants of p73. Expression of p73 resulted in a reduction of the ectopically expressed p53 in transient transfections or of the endogenous p53 induced by Adriamycin- or UV-mediated damage. These reductions of p53 were likely to be due to an increase in MDM2-mediated proteolysis. These results suggest the possibility that different levels of p73 in the cell may act as a mechanism to modulate p53 responses after DNA damage and other stresses and that an increase rather than a decrease in p73 may play a role in tumorigenesis.

MDM2 and MDMX bind and stabilize the p53-related protein p73.

The p53 gene encodes one of the most important tumor suppressors in human cells and undergoes frequent mutational inactivation in cancers. MDM2, a transcriptional target of p53, binds p53 and can both inhibit p53-mediated transcription [1] [2] and target p53 for proteasome-mediated proteolysis [3] [4]. A close relative of p53, p73, has recently been identified [5] [6]. Here, we report that, like p53, p73alpha and the alternative transcription product p73beta also bind MDM2. Interaction between MDM2 and p53 represents a key step in the regulation of p53, as MDM2 promotes the degradation of p53. In striking contrast to p53, the half-life of p73 was found to be increased by binding to MDM2. Like MDM2, the MDM2-related protein MDMX also bound p73 and stabilized the level of p73. Moreover, the growth suppression functions of p73 and the induction of endogenous p21, a major mediator of the p53-dependent growth arrest pathway, were enhanced in the presence of MDM2. These differences between the regulation of p53 and p73 by MDM2/MDMX may highlight a physiological difference in their action.

p53 regulates Cdc2 independently of inhibitory phosphorylation to reinforce radiation-induced G2 arrest in human cells.

We have investigated the influence of p53 on radiation-induced G2 cell cycle arrest using human H1299 cells expressing temperature-sensitive p53. Gamma-irradiated cells lacking p53 arrested transiently in G2 with Cdc2 extensively phosphorylated at the inhibitory sites Thr14 and Tyr15, and with both Cdc2 and cyclin B1 restricted to the cytoplasm. Activation of p53 by temperature shift resulted in a more protracted G2 arrest that could not be overridden by checkpoint-abrogating drugs. Surprisingly, this enhancement of G2 arrest was associated with a marked lack of inhibitory phosphorylation of Cdc2 and with the nuclear localization of both Cdc2 and cyclin B1. While transient expression of an A14F15 mutant form of Cdc2 that is not subject to inhibitory phosphorylation induced mitotic catastrophe in cells lacking p53, the p53-expressing cells were relatively refractory to this effect. Enforced expression of p21WAF1/CJP1 was sufficient to confer nuclear localization on Cdc2 in the p53 null cells, though immunodepletion experiments demonstrated that only a small proportion of Cdc2 was stably associated with p21WAF1/CJP1 in the p53-expressing cells. We conclude that a p53-dependent pathway can operate after exposure of human cells to ionising radiation to promote G2 arrest accompanied by nuclear translocation rather than inhibitory phosphorylation of Cdc2.

p53 regulates the minimal promoter of the human topoisomerase IIalpha gene.

DNA topoisomerase IIalpha is an essential enzyme for chromosome segregation during mitosis. Consistent with a cell division-specific role, the expression of the topoisomerase IIalpha gene is strongly influenced by the proliferation status of cells. The p53 protein is one of the most important regulators of cell cycle progression in mammals, with an apparent dual role in the induction of cell cycle arrest following cytotoxic insults and in the regulation of the apoptotic cell death pathway. We have analysed whether p53 plays a role in regulating expression of the human topoisomerase IIalpha gene. We show that wild-type, but not mutant, p53 is able to decrease substantially the activity of the full length topoisomerase IIalpha gene promoter. Using a series of constructs comprising various deleted or mutated versions of the promoter lacking critical cis-acting elements, we show that this p53-specific regulation of the topoisomerase IIalpha promoter is independent of all characterised transcription factor binding sites and is directed at the minimal gene promoter. We conclude that expression of wild-type p53 induces downregulation of the human topoisomerase IIalpha promoter by acting on the basal transcription machinery. These findings implicate topoisomerase II as one of the downstream targets for p53-dependent regulation of cell cycle progression in human cells.

Inactivation of Cdc2 increases the level of apoptosis induced by DNA damage.

A number of lines of evidence have suggested a possible involvement of the mitosis-promoting protein kinase Cdc2 in the process of apoptotic cell death, and one recent study concluded that premature activation of Cdc2 is required for apoptosis. Here we have used a temperature-sensitive murine Cdc2 mutant cell line and Cdc2 inhibitor compounds to study the effect of inhibition of this protein kinase on apoptosis induced by DNA-damaging drugs. Inhibition of Cdc2 activity before or during exposure to DNA strand break-inducing drugs had the effect of increasing the level of subsequent apoptosis, as assessed by electron microscopy and flow cytometry. We conclude that, far from being required for cell death, a form of mammalian Cdc2 suppresses apoptosis induced by DNA damage. This form of Cdc2 appears to be active in G2-arrested cells and is therefore presumably distinct from the mitosis-promoting Cdc2-cyclin B heterodimer.