High LC3 expression correlates with poor survival in patients with oral squamous cell carcinoma

doi:10.1016/j.humpath.2013.06.017

Human Pathology

Volume 44, Issue 11, November 2013, Pages 2558-2562

https://doi.org/10.1016/j.humpath.2013.06.017 Get rights and content

Summary

Oral squamous cell carcinoma (OSCC) is a destructive disease with very poor prognosis and no effective treatment. Autophagy is a dynamic cellular process involved in various physiological processes and diseases including cancer that degrades cytoplasmic proteins and organelles. The role of autophagy in the pathogenesis of OSCC is not yet understood. Microtubule-associated protein light chains 3 (LC3) is a reliable autophagosome markers for monitoring autophagy. In the present study, LC3 expression was determined in a cohort of 90 OSCC samples by immunohistochemistry. The results were correlated with clinical and pathological characteristics of patients. High LC3 expression (N = 57; 63.3%) correlated with stage (P < .0001), tumor size (P < .0001), and lymph node involvement (P = .0003) and with an increased risk of death (P < .0001; hazard ratio, 3.59) in a univariate analysis. In the multivariate analysis adjusted for grade, stage, and alcohol, betel, and tobacco consumption, high LC3 expression retained statistical significance with regard to survival (P = .0043; hazard ratio, 2.99). The Kaplan-Meier survival curve also showed that high LC3 expression was significantly associated with poor overall survival (P = .0001). Elevated LC3 expression, which corresponds to increased level of autophagy activity, is a frequent event and an indicator of poor prognosis in human OSCC.

Introduction

Oral squamous cell carcinoma (OSCC), a form of head and neck squamous cell cancer, is one of the most common cancers in the world [1]. In South and Southeast Asian countries where most cases are reported, OSCC is a major threat to the public health. The advances in treatment modalities (chemotherapy, radiation and surgery) failed to significantly improve the prognosis of patients with OSCC. The 5-year survival rate for OSCC has remained at approximately 50% in the last few decades [2], [3]. The failure of conventional therapies underscores the need to identify novel molecular targets that can improve the prognosis and therapeutic possibilities associated with OSCC.

Autophagy is a process of intracellular degradation in which cytoplasmic constituents are enveloped in double-membrane vesicles, termed autophagosomes, that deliver the contents to the lysosome for degradation [4]. Autophagy is generally activated during periods of metabolic stress to maintain homeostasis and viability, but it has also been associated with a number of physiological processes including differentiation, development, neurodegenerative disorders, myopathies, infection, and cancer [5].

Autophagy marker light chain 3 (LC3), originally identified as a subunit of microtubule-associated proteins 1A and 1B, is a mammalian homologue of the yeast Atg8 that becomes lapidated and tightly associated with the autophagosomal membranes [6], [7]. LC3 is essential for autophagosome formation and is indicative of autophagic activity [8]. In cancer, autophagy has been shown to act both as a tumor suppressor and as a mechanism to sustain survival [9], [10]. The role of autophagy in the context of OSCC is not yet clear. The present study investigated autophagy activity in OSCC by assessing the expression of LC3 in tumor samples and correlated the results with clinical and pathological characteristics of patients.

Section snippets

Patients and samples

Ninety patients with OSCC were identified who had undergone surgical resections and for whom both clinical data and tumor tissue were available. None of the patients received radiotherapy, chemotherapy or any other treatment prior to surgery. All tumors were staged according to American Joint Committee on Cancer 7th edition criteria [11]. Clinical data collected included patient demographics, survival, and tobacco, betel, and alcohol consumption. This study was approved by the Kaohsiung Medical

Results

Ninety patients' tumors were tested for LC3 immunoreactivity with representative staining illustrated in Fig. 1. A cutoff value of 6 was established by receiver operating characteristic curve analysis in which a total score of equal or less than 6 was regarded as low LC3 expression, whereas a score of greater than 6 was regarded as high LC3 expression. Table 1 summarizes the demographic, clinical and pathological findings. Thirty-three patients (36.7%) showed negative/low LC3 expression, while

Discussion

Autophagy, the type II programmed cell death that degrades cellular organelles and proteins from the cells, is an evolutionarily conserved and highly regulated process that contributes to an array of vital cellular functions [12], [13], [14]. Dysregulation of autophagy disrupts normal physiological processes and is associated with various diseases including cancer [9], [15], [16]. The role of autophagy in cancer has been shown to be context dependent. Many lines of evidence support a role for

References (27)

S.S. Mann et al.
Molecular characterization of light chain 3. A microtubule binding subunit of MAP1A and MAP1B
J Biol Chem
(1994)
B. Levine et al.
Development by self-digestion: molecular mechanisms and biological functions of autophagy
Dev Cell
(2004)
Z. Yang et al.
Mammalian autophagy: core molecular machinery and signaling regulation
Curr Opin Cell Biol
(2010)
K. Degenhardt et al.
Autophagy promotes tumor cell survival and restricts necrosis, inflammation, and tumorigenesis
Cancer Cell
(2006)
N.M. Mazure et al.
Hypoxia-induced autophagy: cell death or cell survival?
Curr Opin Cell Biol
(2010)
J.J. Lum et al.
Growth factor regulation of autophagy and cell survival in the absence of apoptosis
Cell
(2005)
V.M. Aita et al.
Cloning and genomic organization of beclin 1, a candidate tumor suppressor gene on chromosome 17q21
Genomics
(1999)
H. Nomura et al.
Overexpression and altered subcellular localization of autophagy-related 16-like 1 in human oral squamous-cell carcinoma: correlation with lymphovascular invasion and lymph-node metastasis
Hum Pathol
(2009)
V. Kapoor et al.
Deregulation of Beclin 1 in patients with tobacco-related oral squamous cell carcinoma
Biochem Biophys Res Commun
(2012)
A. Jemal et al.
Global cancer statistics
CA Cancer J Clin
(2011)

D.M. Parkin et al.

Global cancer statistics, 2002

CA Cancer J Clin

(2005)

S.H. Landis et al.

Cancer statistics, 1999

CA Cancer J Clin

(1999)

N. Mizushima

Autophagy: process and function

Genes Dev

(2007)

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Immunoblotting, and confocal microscopy were used to examine the effect of tumor microenvironmental stressors on the autophagy activity. Cellular proliferation and migration assays were performed to assess the anti-cancer activity of standard chemotherapy and autophagy initiation inhibitors, either alone or in combination. High resolution mass-spectrometry based proteomic analysis was utilized to understand the mechanisms behind chemoresistance in OSCC models. Finally, immunohistochemistry was performed to determine associations between autophagy markers and clinicopathological characteristics.
Tumor microenvironmental stressors were shown to induce autophagy activity in OSCC cell lines. Novel combinations of chemotherapy and autophagy inhibitors as well as different classes of autophagy inhibitors were identified. Combination of MRT68921 and SAR405 demonstrated marked synergy in their anti-proliferative activity and also showed synergy with chemotherapy in chemoresistant OSCC cell models. Autophagy was identified as one of the key pathways involved in mediating chemoresistance in OSCC. Furthermore, TGM2 was identified as a key upstream regulator of chemoresistance in OSCC models. Finally, positive staining for autophagosome marker LC3 was shown to be associated with low histological grade OSCC.
In conclusion, this study identified a combination of novel autophagy inhibitors which can potently inhibit proliferation of both chemosensitive as well as chemoresistant OSCC cells and could be developed as a novel therapy against advanced OSCC tumors.
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Autophagy plays an important role in maintaining cellular homeostasis. Dysregulation of autophagy has been linked to a number of diseases, including cancer. We retrospectively evaluated immunohistochemical expression of the autophagy markers LC3B and p62 and the autophagy regulator mTOR as an indicator of autophagy in 100 surgically resected primary oral squamous cell carcinoma (OSCC) samples and sought associations with various clinicopathological factors. The expression of all three proteins was significantly higher in malignant squamous cells than in benign squamous cells in the free mucosal margin adjacent to the OSCC. Male sex, higher tumour (T) stage, node (N) stage and tumour, node, metastasis (TNM) stage were significantly associated with high marker expression; age and histological grade showed no significant association. LC3B, p62 and mTOR expression were positively correlated with one another in OSCCs, and the correlation was significant for LC3B and mTOR as well as for LC3B and p62. Disease-free survival showed an inverse correlation with high mTOR expression. Our data suggest that autophagy inhibitors and mTOR inhibitors may have a therapeutic role in the treatment of OSCCs.
Secretory clusterin promotes oral cancer cell survival via inhibiting apoptosis by activation of autophagy in AMPK/mTOR/ULK1 dependent pathway
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Secretory clusterin (sCLU) plays an important role in tumor development and cancer progression. However, the molecular mechanisms and physiological functions of sCLU in oral cancer is unclear. We examined the impact of sCLU-mediated autophagy in cell survival and apoptosis inhibition in oral cancer.
Immunohistochemical analysis was performed to analyze protein expression in patient samples. Autophagy and mitophagy was studied by immunofluorescence microscopy and Western blot. The gain and loss of function was studied by overexpression of plasmid and siRNA approaches respectively. Cellular protection against nutrient starvation and therapeutic stress by sCLU was studied by cell viability, caspase assay and meta-analysis.
The data from oral cancer patients showed that the expression levels of sCLU, ATG14, ULK1, and PARKIN increased in grade-wise manners. Interestingly, sCLU overexpression promoted autophagy through AMPK/Akt/mTOR signaling pathway leading to cell survival and protection from long exposure serum starvation induced-apoptosis. Additionally, sCLU was demonstrated to interact with ULK1 and inhibition of ULK1 activity by SBI206965 was found to abolish sCLU-induced autophagy indicating critical role of ULK1 in induction of autophagy. Furthermore, sCLU was observed to promote expression of mitophagy-associated proteins in serum starvation conditions to protect cells from nutrient deprivation. The meta-analysis elucidated that high CLU expression is associated with therapy resistance in cancer and we demonstrated that sCLU-mediated mitophagy was revealed to inhibit cell death by cisplatin.
The present investigation has highlighted the probable implications of the clusterin-induced autophagy in cell survival and inhibition of apoptosis in oral cancer.
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Previously, by comparing the cellular processes in which RKIP is implied and by analyzing the function of numerous molecular partners, we have suggested that RKIP may participate in regulating actin reorganization and membrane remodeling. Here, based on the role of RKIP in cancer, we describe the cellular function of five important partners of RKIP (GRK2, IQGAP, MT1-MMP, LC3, and Rab8). By their own activities, they may explain, at least in part, the multiple functions of RKIP as they are key actors in diverse cellular processes. All these proteins play a role in the formation or maturation of cellular protrusions such primary cilium, lamellipodia, filopodia, or invadopodia. Considering the close relationship between the actin cytoskeleton and the membrane during cell migration, we suggest that a major role of RKIP in cells is likely to associate with the main molecular assemblies, in order to drive the cortical actin/membrane complex in response to signaling pathways. In cancer cells, because of the lack of RKIP, this important regulation is lost.
Influence of interval between biopsy and surgery on prognosis of patients with early-stage oral squamous cell carcinoma: A preliminary study
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Citation Excerpt :
Oral squamous cell carcinoma (OSCC) accounts for 95% of all oral and oropharyngeal malignancies, and around 300,000 new cases are diagnosed every year, making this the sixth most common cancer in the world [1–3]. Despite advances in treatment strategy, the overall survival rate of OSCC remains poor, at approximately 50–60% [4,5]. This unfavorable prognosis generally stems from loco-regional recurrence and metastasis to cervical lymph nodes or distant organs.
Recent studies have suggested that preoperative interventions such as tooth extraction, incision and curettage may be associated with poor prognosis in patients with oral malignancies. Although biopsy is another preoperative intervention, its influence on the prognosis of patients with oral squamous cell carcinoma (OSCC) has remained unclear. We investigated the prognosis of stage I and II OSCC patients, with special reference to the influence of the interval between biopsy and surgery.
The cases of 47 patients diagnosed with primary early-stage (T1 or T2N0M0) OSCC were reviewed retrospectively.
Primary surgery was performed ≤21 days after biopsy for 17 patients, and >21 days after biopsy for 30 patients. Incidence of local recurrence or delayed cervical lymph node metastasis was significantly higher among the long-interval group (p = 0.03). Patients with local or neck failure displayed a significantly longer interval (33.6 days) than patients who were stable (21.9 days; p < 0.01). Five-year survival rates in groups with and without recurrence or metastasis were 66.7% and 90.6%, respectively (p < 0.05).
Treatment delay can worsen the prognosis of patients with early-stage OSCC by increasing the incidence of local and neck failure. Prompt treatment after biopsy is thus recommended to optimize the outcomes.
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Citation Excerpt :
Increasing incidence rate of oral cancer malignancies and late stage presentation have become global healthcare issues [1,2]. Further, the survival rate is reported to be < 60% without any major improvement over the last five decades [3]. South-Asian counties such as India, Sri Lanka, Bangladesh, and Pakistan are reported to have highest number of patients with the risk of oral cancer in diverse stages [4].
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^☆: This study was supported by a grant from the Department of Health, Taiwan (DOH102-TD-C-111-002).

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Original contributionHigh LC3 expression correlates with poor survival in patients with oral squamous cell carcinoma☆

Summary

Introduction

Section snippets

Patients and samples

Results

Discussion

J Biol Chem

Dev Cell

Curr Opin Cell Biol

Cancer Cell

Curr Opin Cell Biol

Cell

Genomics

Hum Pathol

Biochem Biophys Res Commun

Global cancer statistics

CA Cancer J Clin

Global cancer statistics, 2002

CA Cancer J Clin

Cancer statistics, 1999

CA Cancer J Clin

Autophagy: process and function

Genes Dev

Original contribution
High LC3 expression correlates with poor survival in patients with oral squamous cell carcinoma☆