Concurrent paclitaxel and radiation in the treatment of locally advanced head and neck cancer.

PURPOSE: To determine the feasibility of an organ preservation regimen consisting of infusional paclitaxel administered concurrently with radiotherapy to patients with locally advanced head and neck squamous cell carcinoma (HNSCC). PATIENTS AND METHODS: Thirty-three previously untreated patients with stage III or IV tumors were enrolled onto the study. Paclitaxel was administered as a 120-hour continuous infusion every 3 weeks during the course of radiation therapy. Sixteen patients received a paclitaxel dose of 105 mg/m(2), and 17 patients received 120 mg/m(2). Radiation was delivered in a standard format at 1.8 Gy/d to a total dose of 70.2 to 72 Gy. RESULTS: Three months after therapy, a 76% complete response (CR) at the primary site and a 70% overall CR was achieved. At 36 months, locoregional control was 55.7%, overall survival was 57.8%, and disease-free survival was 51.1%. The median survival duration for all 33 patients was greater than 50 months at the time of this report. Local toxicities including mucositis, dysphagia, and skin reactions were severe but tolerable. All patients retained functional speech, and all but four patients were swallowing food 3 months after treatment. Steady-state plasma concentrations for paclitaxel were not achieved during a 120-hour infusion, suggesting a nonlinear process. Tumor volume quantified by pretreatment computerized tomography imaging was associated with likelihood of response and survival. CONCLUSION: Paclitaxel administered as a 120-hour continuous infusion in combination with radiotherapy is a feasible and promising treatment for patients with advanced HNSCC.

Cisplatin and radiation sensitivity in human head and neck squamous carcinomas are independently modulated by glutathione and transcription factor NF-kappaB.

BACKGROUND: Response to neoadjuvant cisplatin-based chemotherapy has been used to predict overall response to chemoradiation therapy and to select patients with head and neck squamous cell carcinoma (HNSCC) for organ preservation therapy in NCI and VA cooperative group trials. However, different molecular determinants have been reported to contribute to sensitivity of cells to cisplatin and radiation, including glutathione (GSH), and activation of nuclear factor-kappaB (NF-kappaB), a transcription factor that regulates cytoprotective genes. We have reported that NF-kappaB is constitutively activated in HNSCC, but the relationship of NF-kappaB to GSH and to cisplatin and radiation sensitivity in HNSCC is unknown. METHODS: We examined human HNSCC lines to define the relationship of cisplatin and radiation sensitivity to intracellular GSH and NF-kappaB and determined whether HNSCC could be sensitized to these modalities by lowering the concentration of glutathione with L-buthionine sulfoximine or inhibiting activation of NF-kappaB by expression of a degradation-resistant mutant inhibitor-kappaBalpha. RESULTS: Cisplatin resistance did not predict radiation resistance in three HNSCC cell lines, UM-SCC-9, 11B, and, 38. Resistance to cisplatin correlated with intracellular GSH, and depletion of GSH by treatment with L-BSO sensitized UM-SCC-9 cells to cisplatin but not radiation. Conversely, radiation resistance was correlated with activation of NF-kappaB. Expression of a mutant Inhibitor-kappaB after gene transfer inhibited NF-kappaB and sensitized UM-SCC-9 cells to radiation but not cisplatin. CONCLUSIONS: GSH and transcription factor NF-alphaB can contribute independently to cisplatin and radiation sensitivity of human HNSCC. These results highlight the need to define molecular determinants of chemotherapy and radiation sensitivity for use in the selection of patients and as novel targets for therapy in future chemoradiation therapy trials for organ preservation in patients with HNSCC.

Inhibition of transcription factor nuclear factor-kappaB by a mutant inhibitor-kappaBalpha attenuates resistance of human head and neck squamous cell carcinoma to TNF-alpha caspase-mediated cell death.

Tumour necrosis factor-alpha (TNF-alpha) is a cytokine that can induce cell death of different cancers via a cellular cascade of proteases, the caspases. However, TNF-alpha has been detected in tumour and serum of patients with head and neck squamous cell carcinoma (HNSCC), and tumour cell lines derived from this environment often exhibit resistance to TNF-alpha-induced cell death. Cell death mediated by TNF-alpha and caspases may be inhibited by cytoprotective genes regulated by transcription factor nuclear factor-kappaB (NF-kappaB). We recently showed that NF-kappaB is constitutively activated in HNSCC, and that inhibition of NF-kappaB by expression of a nondegradable mutant inhibitor of NF-kappaB, IkappaBalphaM, markedly decreased survival and growth of HNSCC cells in vivo. In the present study, we examined the TNF-alpha sensitivity and response of HNSCC with constitutively active NF-kappaB, and of HNSCC cells in which NF-kappaB is inhibited by stable expression of a dominant negative mutant inhibitor, IkappaBalphaM. Human lines UM-SCC-9, 11B and 38, previously shown to exhibit constitutive activation of NF-kappaB, were found to be highly resistant to growth inhibition by up to 10(4)U/ml of TNF-alpha in 5 day MTT assay. These TNF-alpha resistant HNSCC lines expressed TNF receptor I, and exhibited constitutive and TNF-alpha-inducible activation of NF-kappaB as demonstrated by nuclear localization of NF-kappaB p65 by immunohistochemistry. UM-SCC-9 I11 cells which stably expressed an inhibitor of NF-kappaB, IkappaBalpham, were susceptible to TNF-alpha-induced growth inhibition. DNA cell cycle analysis revealed that TNF-alpha induced growth inhibition was associated with accumulation of cells with sub-G0/G1 DNA content. Cell death was demonstrated by trypan blue staining, and was blocked by caspase inhibitor. We conclude that HNSCC that exhibit constitutive and TNF-alpha-inducible activation of transcription factor NF-kappaB are resistant to TNF-alpha, and that inhibition of NF-kappaB sensitizes HNSCC to TNF-alpha caspase-mediated cytotoxicity. The demonstration of the role of activation of NF-kappaB in resistance of HNSCC to TNF-alpha may be helpful in the identification of potential targets for pharmacological, molecular and immune therapy of HNSCC.

Expression of proinflammatory and proangiogenic cytokines in patients with head and neck cancer.

Altered immune, inflammatory, and angiogenesis responses are observed in patients with head and neck squamous cell carcinoma (HNSCC), and many of these responses have been linked with aggressive malignant behavior and a decrease in prognosis. In this study, we examined the hypothesis that HNSCC cells produce cytokines that regulate immune, inflammatory, and angiogenesis responses. We identified important regulatory cytokines in supernatants of well-defined and freshly cultured HNSCC cell lines by ELISA and determined whether these cytokines are detected in tumor cell lines and tissue specimens by immunohistochemistry. The serum concentration of the cytokines and cytokine-dependent acute phase inflammatory responses (i.e., fibrinogen, C-reactive protein, and erythrocyte sedimentation rate) from patients with HNSCC was determined, and the potential relationship of serum cytokine levels to tumor volume was analyzed. Cytokines interleukin (IL)-1alpha, IL-6, IL-8, granulocyte-macrophage colony-stimulating factor (GM-CSF), vascular endothelial growth factor (VEGF), and basic fibroblast growth factor were detected in similar concentration ranges in the supernatants of a panel of established University of Michigan squamous cell carcinoma (UM-SCC) cell lines and supernatants of freshly isolated primary HNSCC cultures. Evidence for the expression of IL-1alpha, IL-6, IL-8, granulocyte-macrophage colony-stimulating factor, and VEGF in HNSCC cells within tumor specimens in situ was obtained by immunohistochemistry. In a prospective comparison of the cytokine level and cytokine-inducible acute-phase proteins in serum, we report that cytokines IL-6, IL-8, and VEGF were detected at higher concentrations in the serum of patients with HNSCC compared with patients with laryngeal papilloma or age-matched control subjects (at P < 0.05). The serum concentrations of IL-8 and VEGF were found to be weakly correlated with large primary tumor volume (R2 = 0.2 and 0.4, respectively). Elevated IL-1- and IL-6-inducible acute-phase responses were also detected in cancer patients but not in patients with papilloma or control subjects (at P < 0.05). We therefore conclude that cytokines important in proinflammatory and proangiogenic responses are detectable in cell lines, tissue specimens, and serum from patients with HNSCC. These cytokines may increase the pathogenicity of HNSCC and prove useful as biomarkers or targets for therapy.

Expression of a dominant-negative mutant inhibitor-kappaBalpha of nuclear factor-kappaB in human head and neck squamous cell carcinoma inhibits survival, proinflammatory cytokine expression, and tumor growth in vivo.

We demonstrated recently that constitutive expression of proinflammatory cytokines interleukin (IL)-1alpha, IL-6, IL-8, and granulocyte-macrophage colony-stimulating factor in head and neck squamous cell carcinoma is correlated with activation of transcription factor nuclear factor (NF)-kappaB/Rel A (p50/p65), which binds the promoter region within each of the genes encoding this repertoire of cytokines. NF-kappaB can be activated after signal-dependent phosphorylation and degradation of inhibitor-kappaBalpha and has been reported to promote cell survival and growth. In the present study, we expressed a phosphorylation site mutant of inhibitor-kappaBalpha (IkappaBalphaM) in head and neck squamous cell carcinoma lines UM-SCC-9, -11B, and -38 to determine the effect of inhibition of NF-kappaB on cytokine expression, cell survival in vitro, and growth in vivo. After transfection with IKBalphaM, only a few UM-SCC-9 clones were obtained that stably expressed the mutant IkappaB, suggesting that expression of a mutant IkappaBalpha may affect survival of the transfected UM-SCC cell lines. After cotransfection of IkappaBalphaM with a Lac-Z reporter, we found that the number of surviving beta-galactosidase-positive cells in the three cell lines was reduced by 70-90% when compared with controls transfected with vector lacking the insert. In UM-SCC-9 cells that stably expressed IkappaBalphaM, inhibition of constitutive and tumor necrosis factor-a induced NF-kappaB activation, and production of all four cytokines was observed. Although UM-SCC-9 IkappaBalphaM-transfected cells proliferated at the same rate as vector-transfected cells in vitro, a significant reduction in growth of tumor xenografts was observed in SCID mice in vivo. The decreased growth of UM-SCC-9 IkappaBalphaM-transfected tumor cells accompanied decreased immunohistochemical detection of the activated form of NF-kappaB in situ. These results provide evidence that NF-KB and IkappaBalpha play an important role in survival, constitutive and inducible expression of proinflammatory cytokines, and growth of squamous cell carcinoma. NF-kappaB could serve as a potential target for therapeutic intervention against cytokine and other immediate-early gene responses that contribute to the survival, growth, and pathogenesis of these cancers.

Apoptotic vs. nonapoptotic cytotoxicity induced by hydrogen peroxide.

The regulation of cellular cytotoxicity induced by hydrogen peroxide (H2O2) over a wide concentration range was assessed. Three distinct patterns were detected: the highest concentrations (> 10 mM) rapidly induced a necrotic form of death characterized by smeared patterns of DNA digestion and morphological evidence of primary cytoplasm and plasma membrane damage; In contrast, 10 and 5 mM H2O2 induced endonucleosomal DNA digestion concurrently with cytotoxicity and target cell death was associated with morphologic evidence of apoptosis. Apoptosis was inhibited by cycloheximide, emetine, aminobenzamide (ABA), aurintricarboxylic acid, and calcium depletion. The lowest concentrations of H2O2 (0.5 and 0.1 mM)-induced delayed cytotoxicity (at 24 or 48 hr), which was not associated with DNA ladder formation or morphologic evidence of apoptosis, but was inhibited by ABA. Enforced expression of BCL-2 induced resistance to 0.5 and 0.1 mM H2O2 but had no effect on cytotoxicity induced by 5 and 10 mM. Exposure of isolated nuclei to H2O2 in the absence of calcium or magnesium failed to induce endonucleosomal fragmentation. These data indicate that distinct pathways of H2O2-induced cytotoxicity can be distinguished by their different concentration dependences, and that BCL-2 can protect against some forms of H2O2-induced cytotoxicity.

Regulation of transforming growth factor-beta 1-mediated apoptosis in head and neck squamous cell carcinoma.

The apoptotic cell death in Cal-27 cells induced by exposure to transforming growth factor-beta 1 was inhibited by the endonuclease inhibitor aurintricarboxylic acid (ATA) in a concentration-dependent fashion. In vitro studies of cytotoxicity, DNA fragmentation, and protein synthesis by Cal-27 cell lines were performed. Inhibition of cytotoxicity as well as endonucleolytic DNA cleavage was detected. ATA did not inhibit cytotoxicity either via transforming growth factor cell-surface-receptor alteration or by inhibition of macromolecular synthesis. ATA-sensitive events occurred late during treatment. These data suggest that endonucleolytic DNA cleavage is a mandatory event leading to cell death in this system.

Oral lichen planus and its association with squamous cell carcinoma: an update on pathogenesis and treatment implications.

Lichen planus of the oral mucosa (OLP) is characterized by lymphocytic mucositis, basal cell lysis, and lymphocyte transmigration into the epithelial compartment. Some reports have suggested a high incidence of oral squamous cell carcinoma (OSCCA) in OLP patients and have implicated OLP as a premalignant lesion. We describe five cases of OSCCA arising in patients with preexisting OLP. At our institution, the incidence of OSCCA in patients with OLP approximates that reported in other series. The immunopathologic basis for OLP, its potential association with malignancy, and the variable clinical picture of OSCCA in patients with OLP are reviewed. Specific recommendations are given for treatment and follow-up of lesions, including the role of future testing with viral and oncogene markers.

Clear cell odontogenic carcinoma. A clinicopathologic analysis.

Clear cell odontogenic tumor is a rare neoplasm of the jaws that histologically may be confused with metastatic renal cell carcinoma. A review of 17 cases revealed that these tumors are aggressive, with a tendency to recur locally after surgery; they also may metastasize both regionally and distantly. There is a marked female predilection, and most cases are discovered during the fifth to seventh decades of life. Seventy percent occur in the mandible, appearing as poorly marginated radiolucencies. We conclude that this tumor is malignant and should be referred to as clear cell odontogenic carcinoma; it requires an aggressive surgical approach.