Glutamine is essential for overcoming the immunosuppressive microenvironment in malignant salivary gland tumors.

Rationale: Immunosuppression in the tumor microenvironment (TME) is key to the pathogenesis of solid tumors. Tumor cell-intrinsic autophagy is critical for sustaining both tumor cell metabolism and survival. However, the role of autophagy in the host immune system that allows cancer cells to escape immune destruction remains poorly understood. Here, we determined if attenuated host autophagy is sufficient to induce tumor rejection through reinforced adaptive immunity. Furthermore, we determined whether dietary glutamine supplementation, mimicking attenuated host autophagy, is capable of promoting antitumor immunity. Methods: A syngeneic orthotopic tumor model in Atg5+/+ and Atg5flox/flox mice was established to determine the impact of host autophagy on the antitumor effects against mouse malignant salivary gland tumors (MSTs). Multiple cohorts of immunocompetent mice were used for oncoimmunology studies, including inflammatory cytokine levels, macrophage, CD4+, and CD8+ cells tumor infiltration at 14 days and 28 days after MST inoculation. In vitro differentiation and in vivo dietary glutamine supplementation were used to assess the effects of glutamine on Treg differentiation and tumor expansion. Results: We showed that mice deficient in the essential autophagy gene, Atg5, rejected orthotopic allografts of isogenic MST cells. An enhanced antitumor immune response evidenced by reduction of both M1 and M2 macrophages, increased infiltration of CD8+ T cells, elevated IFN-γ production, as well as decreased inhibitory Tregs within TME and spleens of tumor-bearing Atg5flox/flox mice. Mechanistically, ATG5 deficiency increased glutamine level in tumors. We further demonstrated that dietary glutamine supplementation partially increased glutamine levels and restored potent antitumor responses in Atg5+/+ mice. Conclusions: Dietary glutamine supplementation exposes a previously undefined difference in plasticity between cancer cells, cytotoxic CD8+ T cells and Tregs.

Efficacy and toxicity of sorafenib in patients with adenoid cystic carcinoma of the head and neck: a case series of five patients.

INTRODUCTION: Adenoid cystic carcinoma (ACC) of the head and neck is a rare malignancy of the salivary glands that accounts for approximately 10% of salivary gland carcinoma. Despite aggressive local therapy, local recurrence and distant metastases occur frequently. Response rates (RR) to potential curative and palliative chemotherapy are limited, so new strategies are needed. CASE REPORTS: We describe five case reports of patients with unresectable locally advanced or metastatic ACC of the head and neck who have been treated with sorafenib, a multi-tyrosine kinase inhibitor (mTKI). RESULTS: In this case series, we found that three out of five patients treated with sorafenib survived, respectively, 16, 35 and 35 months. Two patients showed a partial response (PR) and one patient had a prolonged stable disease (SD) for almost three years. Grade 3 adverse events (AE) occur under sorafenib so adequate toxicity management is essential. This retrospective case series hints towards the possibility of clinical benefit for treating ACC patients with sorafenib. Efficacy of sorafenib should be studied in a prospective-randomized clinical trial which is a challenging task due to the rarity of the disease.

A multiplex preclinical model for adenoid cystic carcinoma of the salivary gland identifies regorafenib as a potential therapeutic drug.

Adenoid cystic carcinomas (ACC) are rare salivary gland cancers with a high incidence of metastases. In order to study this tumor type, a reliable model system exhibiting the molecular features of this tumor is critical, but none exists, thereby inhibiting in-vitro studies and the analysis of metastatic behavior. To address this deficiency, we have coupled an efficient method to establish tumor cell cultures, conditional reprogramming (CR), with a rapid, reproducible and robust in-vivo zebrafish model. We have established cell cultures from two individual ACC PDX tumors that maintain the characteristic MYB translocation. Additional mutations found in one ACC culture also seen in the PDX tumor. Finally, the CR/zebrafish model mirrors the PDX mouse model and identifies regorafenib as a potential therapeutic drug to treat this cancer type that mimic the drug sensitivity profile in PDX model, further confirming the unique advantages of multiplex system.

A phase II study of sorafenib in recurrent and/or metastatic salivary gland carcinomas: Translational analyses and clinical impact.

BACKGROUND: Pre-clinical and clinical evidence suggests a rationale for the use of anti-angiogenic agents, including sorafenib, in recurrent and/or metastatic salivary gland carcinomas (RMSGCs). This study evaluates the activity of sorafenib in patients with RMSGCs and also investigates whether the activity of sorafenib could be related to its main tailored targets (i.e. BRAF, vascular endothelial growth factor receptor 2 [VEGFR2], platelet-derived growth factor receptor α [PDGFRα] and ß, RET, KIT). PATIENTS AND METHODS: Patients received sorafenib at 400 mg BID. The primary end-point was response rate (RR) including complete response or partial response (PR); secondary end-points included RR according to Choi criteria, disease control rate (DCR), overall survival (OS), and progression-free survival (PFS). RESULTS: Thirty-seven patients (19 adenoid cystic cancers, ACC) were enrolled. Six PRs were recorded. RR was 16% (95% confidence interval [CI]: 6-32; 11% in ACC and 22% in non-ACC). Choi criteria could be applied in 30 out of 37 cases with a RR of 50% (95% CI: 31-69%); DCR was 76% (95% CI: 59-88%). Incidence of ≥G3 adverse events was 29.7%. Median PFS and OS for the entire population were 5.9 months and 23.4 months, respectively. Median PFS and OS were 8.9 and 26.4 months for ACC versus 4.2 and 12.3 months for non-ACC patients. All the cases showed expression of PDGFRß in the stroma and VEGFR2 in endothelial cells; PDGFRα positivity was found in the stroma of four (27%) cases. All except for two cases showed no PDGFRß, VEGFR2 and PDGFRα expression in the tumour cells. KIT expression was restricted to ACC and a weak RET expression was limited to one adenocarcinoma, not otherwise specified (NOS). No BRAF mutation was found. No correlation was observed between the sorafenib activity and the expression of its markers although all six responders (two ACC, one adenocarcinoma, NOS, one salivary duct cancer [SDC], one high-grade mucoepidermoid [HG-MEC] and one poorly-differentiated cancer) are enriched in the stromal component showing a PDGFRß immunodecoration. In ACCs, immunohistochemistry revealed MYB protein expression in 15/16 cases (94%) and the MYB-NFIB fusion oncogene was observed in 9/14 (64%). CONCLUSIONS: Sorafenib is the first anti-angiogenic agent to demonstrate activity in RMSGC patients, particularly in some histotypes such as HG-MEC, SDC and adenocarcinoma, NOS. The PDGFRß-positive rich stromal component characterising these histotypes and the lack of correlation between the activity of sorafenib and its targets suggests anti-angiogenic effect as the prevalent mechanism of action of sorafenib in SGCs.

Convallaria keiskei as a novel therapeutic alternative for salivary gland cancer treatment by targeting myeloid cell leukemia-1.

BACKGROUND: Various chemotherapeutic agents have been used largely for the treatment of salivary gland cancer. However, results are disappointing, and these agents can cause some serious side effects. Therefore, recent studies have focused on the possible roles of natural products to overcome these limitations. METHODS: Salivary gland cancer cells treated with or without Convallaria keiskei (MECK) for 24 hours. Apoptotic changes were evaluated by live/dead assay, immunoblotting, and expression levels of caspase-3 and B-cell lymphoma-2 family member. RESULTS: MECK significantly inhibited salivary gland cancer growth. At the molecular level, MECK dramatically reduced myeloid cell leukemia-1 (Mcl-1) in a translation-dependent manner and thereby induced apoptosis through Bax/Bid. Furthermore, we found that Mcl-1 could be a potential therapeutic target of MECK-induced apoptosis and its stability is regulated by extracellular signal-regulated kinases 1/2 (ERK1/2) signaling CONCLUSION: MECK can be used as a safe and efficient therapeutic alternative for the treatment of salivary gland cancer. © 2015 Wiley Periodicals, Inc. Head Neck 38: E761-E770, 2016.

Phase II trial of sorafenib in advanced salivary adenoid cystic carcinoma of the head and neck.

BACKGROUND: There is a need to improve the systemic treatment of advanced adenoid cystic carcinoma (ACC). Response rates to chemotherapy are poor and preliminary investigations of molecularly targeted agents have been disappointing. In this study, we evaluate sorafenib, an oral multikinase inhibitor, which has an attractive targeting profile for this disease. METHODS: In a single-arm phase II trial, patients with unresectable locally recurrent and/or metastatic ACC were treated with sorafenib 400 mg bid. RESULTS: Twenty-three patients, median age 51 years, were recruited from 2009 to 2011. Median progression-free survival (PFS) and overall survival (OS) were 11.3 and 19.6 months, respectively. PFS at 6 and 12 months were 69.3% and 46.2%, respectively. Sorafenib was only reasonably well tolerated, and 13 patients (57%) experienced grade 3 toxicity. CONCLUSION: Sorafenib showed modest activity in ACC with a 12-month PFS of 46.2%. Sorafenib 400 mg bid was associated with significant toxicity and, taken together with limited effectiveness, cannot be enthusiastically recommended for further evaluation.

Inhibition of autophagy augments chemotherapy in human salivary adenoid cystic carcinoma.

Although cisplatin (DDP)-based adjuvant chemotherapy is widely used in the treatment of salivary adenoid cystic carcinoma (SACC), SACCs have developed resistance to cisplatin, resulting in chemotherapy failure. Autophagy serves as a critical adaptive response, which was increased in tumor cells in chemotherapy. However, the function of autophagy is not clear in SACC. In this study, apoptosis induced by DDP in SACC high metastatic cell line (ACC-M) was revealed using MTT assay, flow cytometry, and caspase-3 immunoblotting. The autophagy activation induced by DDP treatment was measured by transmission electron microscopy, green fluorescent protein-light chain 3 plasmid transfection LC3 immunoblotting and p62 immunoblotting. 3-methyladenine (3-MA) or small interference RNA targeting beclin 1 (beclin 1 siRNA) inhibited autophagy and significantly enhanced DDP-induced apoptosis. ACC-M xenografts in nude mice further verified the synergistic effect of DDP and 3-MA. In conclusion, autophagy activation was caused to protect cancer cells from DDP-induced apoptosis and autophagy inhibition could be a promising strategy for adjuvant chemotherapy in SACC.

Mechanisms of apple polyphenols-induced proliferation inhibiting and apoptosis in a metastatic oral adenoid cystic carcinoma cell line.

Adenoid cystic carcinoma (ACC) is characterized by intensive local invasion and high incidence of distant metastases. Conventional chemotherapy for ACC produces a poor result. We aimed to evaluate the effect of apple polyphenols (APs), a novel nutraceutical agent, on the proliferation and apoptosis levels in a metastatic oral ACC cell line. A metastatic ACC (ACC-M) cell line and control cells (MRC-5 cells derived from normal lung tissue) were treated with APs at different concentrations. MTT assay was used to determine the in vitro cytotoxicity. The cell cycle distribution and apoptosis levels were measured by flow cytometry. To evaluate the mechanism of APs, vascular endothelial growth factor receptor-2 (VEGFR-2) and caspase-3 messenger ribonucleic acid (mRNA) and protein levels were evaluated by reverse transcription-polymerase chain reaction and Western blots, respectively. After cells were cultured for 24 hours or 48 hours, the critical concentration of cytotoxicity of APs in MRC-5 cells was found to be 250 µg/mL. In contrast, in the concentration range of 100-250 µg/mL, the cytotoxicity of APs in ACC-M cells was time- and dose-dependent: ACC-M cell proliferation declined at 100 µg/mL when cultured for 48 hours, whereas growth was not inhibited at the concentrations of APs below 200 µg/mL when cultured for 24 hours. In selected time and dose patterns (ACC-M cells cultured at the concentrations of 150 and 250 µg/mL for 48 hours), the flow cytometry performance showed that apoptosis and necrosis occurred in APs-treated ACC-M cells. Also, in these patterns, VEGFR-2 mRNA and protein levels decreased whereas the levels of caspase-3 increased. In summary, APs could inhibit proliferation and induce apoptosis in ACC-M cells in vitro. These effects may be related to the downregulation of VEGFR-2 expression and the activation of caspase-3 expression.

Adjuvant chemoradiotherapy for locoregionally advanced and high-risk salivary gland malignancies.

BACKGROUND: To report the outcomes of patients with locoregionally advanced and high- risk salivary gland malignancies treated with surgery followed by adjuvant chemoradiotherapy. METHODS: From 09/1991 - 06/2007, 24 high-risk salivary gland cancer patients were treated with surgery, followed by adjuvant chemoradiotherapy for high-risk pathologic features including, perineural involvement, nodal involvement, positive margins, or T3/T4 tumors. Chemoradiotherapy was delivered for 4-6 alternating week cycles: the most common regimen, TFHX, consisted of 5 days paclitaxel (100 mg/m² on d1), infusional 5-fluorouracil (600 mg/m²/d × 5d), hydroxyurea (500 mg PO BID), and 1.5 Gy twice daily irradiation followed by a 9-day break without treatment. RESULTS: Median follow-up was 42 months. The parotid gland was more frequently involved (n = 17) than minor (n = 4) or submandibular (n = 3) glands. The median radiation dose was 65 Gy (range 55-68 Gy). Acute treatment related toxicity included 46% grade 3 mucositis and 33% grade 3 hematologic toxicity. Six patients required feeding tubes during treatment. One patient progressed locally, 8 patients progressed distantly, and none progressed regionally. Five-year locoregional progression free survival was 96%. The 3 and 5 year overall survival was 79% and 59%, respectively. Long-term complications included persistent xerostomia (n = 5), esophageal stricture requiring dilatation (n = 1), and tempromandibular joint syndrome (n = 1). CONCLUSIONS: Surgical resection followed by adjuvant chemoradiotherapy results in promising locoregional control for high-risk salivary malignancy patients.

Sustained response of carcinoma ex pleomorphic adenoma treated with trastuzumab and capecitabine.

BACKGROUND: Carcinoma ex pleomorphic adenoma is a rare histologic subtype of salivary gland cancer with an overall poor prognosis. Limited histopathologic analyses have shown that some such tumors exhibit significant HER2/neu immunoreactivity, suggesting a potential role for HER2-based therapy. We report here a case of a 58-year old man with metastatic carcinoma ex pleomorphic adenoma who achieved a sustained long term response to combination therapy with trastuzumab and capecitabine. CASE PRESENTATION: A 58 year old man presented with T1N2bM0 carcinoma ex pleomorphic adenoma and underwent surgery followed by adjuvant radiation therapy. Multiple metastases to bone were documented one year later. Since the original tumor was strongly HER2/neu positive by immunohistochemistry, the patient was treated with trastuzumab, capecitabine, and zoledronic acid. He experienced total resolution of symptoms and repeat FDG-PET scan after three cycles revealed interval disease resolution. Continued treatment has resulted in maintenance of disease control for over 2 years. CONCLUSION: This case illustrates the successful long term treatment of carcinoma ex pleomorphic adenoma with targeted therapy with trastuzumab in combination with chemotherapy. In the absence of definitive clinical trials which are unlikely to be performed due to the rarity of this tumor, case reports such as this one suggest potential utility for trastuzumab in combination with chemotherapy in the treatment of HER2/neu-overexpressing carcinoma ex pleomorphic adenoma.

Mammalian target of rapamycin pathway promotes tumor-induced angiogenesis in adenoid cystic carcinoma: its suppression by isoliquiritigenin through dual activation of c-Jun NH2-terminal kinase and inhibition of extracellular signal-regulated kinase.

Tumor-induced angiogenesis is essential for invasive growth and hematogenous metastasis of adenoid cystic carcinoma (ACC), a highly aggressive neoplasm mostly occurring in salivary glands. Previous studies have indicated that strategies directed against angiogenesis will help develop new therapeutic agents for ACC. The Chinese folk medicine licorice has been used for years as a natural remedy for angiogenesis-related diseases. In this study, we examined the effects of isoliquiritigenin (ISL), a flavonoid isolated from licorice, on the growth and viability of ACC cells and observed a concentration-dependent (0-20 microM) inhibition of cell growth without cell death at 24 h. In a further mimic coculture study, ISL effectively suppressed the ability of ACC cells to induce in vitro proliferation, migration, and tube formation of human endothelial hybridoma (EAhy926) cells as well as ex vivo and in vivo angiogenesis, whereas it exerted no effect on EAhy926 cells when added directly or in the presence of vascular endothelial growth factor (VEGF). The data also showed that the specific suppression of tumor angiogenesis by ISL was caused by down-regulation of mammalian target of rapamycin (mTOR) pathway-dependent VEGF production by ACC cells, correlating with concurrent activation of c-Jun NH(2)-terminal kinase (JNK) and inhibition of extracellular signal-regulated kinase (ERK). Most importantly, ISL also significantly decreased microvessel density within xenograft tumors, associating with the reduction of VEGF production and suppression of the mTOR pathway coregulated by JNK and ERK, as revealed by immunohistochemical studies and clustering analysis. Taken together, our results highlight the fact that ISL is a novel inhibitor of tumor angiogenesis and possesses great therapeutic potential for ACC.

Outcomes of postoperative concurrent chemoradiotherapy for locally advanced major salivary gland carcinoma.

OBJECTIVE: To investigate the potential value of postoperative concurrent chemoradiation among patients with high-risk salivary gland carcinomas. DESIGN: Case control study based on retrospective medical record review. SETTING: A tertiary care comprehensive cancer center. PATIENTS: A total of 24 patients, 12 with major salivary gland carcinoma who were treated with postoperative concurrent chemoradiotherapy from 1998 to 2007 (chemoradiation group), and a control group of 12 patients treated with postoperative radiation alone. MAIN OUTCOME MEASURES: Overall survival, progression-free survival, toxic effects. RESULTS: All but 1 patient had stage III or IV disease; close or positive surgical margins were identified in 20 patients (83%). The median radiation dose was 63 Gy. In the chemoradiation group, platinum-based regimens were used in all. Treatment was well tolerated, but toxic effects, predominantly hematologic, were increased in the chemoradiation group. To date, 8 patients have died; the median overall survival was 53 months. The overall survival in the chemoradiation group was significantly better than in the radiation-alone group: 3-year survival rates were 83% and 44%, respectively (P = .05). CONCLUSIONS: Locally advanced or high-grade salivary gland carcinomas follow an aggressive clinical course. Based on our limited experience, postoperative chemoradiation with a platinum-based regimen seems to be effective in selected patients and warrants further investigation.

Epirubicin, cisplatin and protracted venous infusion 5-Fluorouracil chemotherapy for advanced salivary adenoid cystic carcinoma.

AIMS: Adenoid cystic carcinoma (ACC) is a rare tumour that usually arises in the salivary glands. Initial management is surgery often combined with adjuvant radiotherapy. Chemotherapy is reserved for treatment of symptomatic recurrence. We evaluated the combination of epirubicin, cisplatin and protracted venous infusion 5-fluorouracil (ECF) in the management of ACC. MATERIALS AND METHODS: Patients referred for treatment of advanced, symptomatic ACC were considered. The drugs given were epirubicin 50 mg/m(2) 3-weekly, cisplatin 60 mg/m(2) 3-weekly and protracted venous infusion 5-fluorouracil 200 mg/m(2)/day. RESULTS: Eight patients (median age 46 years) received a median of five cycles of chemotherapy. All patients had had previous surgery, seven had had previous radiotherapy and one had had previous chemotherapy. One patient showed a partial response (duration 34 months) and five showed stable disease (median duration 13.6 months [6.8-15.9+ months]). Median survival was 27 months (3.5-62.3 months). CONCLUSIONS: The activity of ECF in ACC of the head and neck seems to be similar to the combination of cisplatin and 5-fluorouracil and single-agent epirubicin.

Synergistic inhibitory effect of sulforaphane and 5-fluorouracil in high and low metastasis cell lines of salivary gland adenoid cystic carcinoma.

The present study aimed to evaluate the growth-inhibitory effect of sulforaphane (SFN) and a traditional chemotherapy agent, 5-fluorouracil (5-Fu), against the proliferation of salivary gland adenoid cystic carcinoma high metastatic cell line (ACC-M) and low metastasis cell line (ACC-2). Furthermore, the expression of nuclear factor kappa B (NF-kappaB) which induces resistance to anticancer chemotherapeutic agents was also detected. The combination effect of SFN and 5-Fu was quantitatively determined using the method of median effect principle and the combination index. The nuclear NF-kappaB p65 expression after treatment with the SFN-5-Fu combination was also evaluated by western blot analysis. The ACC-M and ACC-2 cells exhibited relative resistant to 5-Fu. Treatment ACCs cells with SFN and 5-Fu in combination, led to synergistic inhibition on cell growth and a decreased expression in nuclear NF-kappaB p65 protein. This synergistic inhibitory effect was more significant in ACC-M cells, which is associated with the greatly decreased expression of NF-kappaB p65 (almost 5-fold) after the combination treatment. Our results demonstrate synergism between SFN and 5-Fu at higher doses against the ACC-M and ACC-2 cells, which was associated with the decreased expression of nuclear NF-kappaB p65 protein.

Effective treatment of cutaneous and subcutaneous malignant tumours by electrochemotherapy.

Electrochemotherapy (ECT) enhances the effectiveness of chemotherapeutic agents by administering the drug in combination with short intense electric pulses. ECT is effective because electric pulses permeabilize tumour cell membranes and allow non-permeant drugs, such as bleomycin, to enter the cells. The aim of this study was to demonstrate the anti-tumour effectiveness of ECT with bleomycin on cutaneous and subcutaneous tumours. This article summarizes results obtained in independent clinical trials performed by five cancer centres. A total of 291 cutaneous or subcutaneous tumours of basal cell carcinoma (32), malignant melanoma (142), adenocarcinoma (30) and head and neck squamous cell carcinoma (87) were treated in 50 patients. Short and intense electric pulses were applied to tumours percutaneously after intravenous or intratumour administration of bleomycin. The tumours were measured and the response to the treatment evaluated 30 days after the treatment. Objective responses were obtained in 233 (85.3%) of the 273 evaluable tumours that were treated with ECT. Clinical complete responses were achieved in 154 (56.4%) tumours, and partial responses were observed in 79 (28.9%) tumours. The application of electric pulses to the patients was safe and well tolerated. An instantaneous contraction of the underlying muscles was noticed. Minimal adverse side-effects were observed. ECT was shown to be an effective local treatment. ECT was effective regardless of the histological type of the tumour. Therefore, ECT offers an approach to the treatment of cutaneous and subcutaneous tumours in patients with minimal adverse side-effects and with a high response rate.

Glucocorticoid-induced growth inhibition with enhanced expression of ductal epithelium of human salivary gland adenocarcinoma cells transplanted into athymic nude mice.

The effect of glucocorticoid on growth and the histologic features of adenocarcinoma induced in nude mouse by the inoculation of neoplastic epithelial cells of salivary duct origin (HSG) were studied. Subcutaneous injection of dexamethasone (low-dose group: 0.05 micrograms/g, high-dose group: 0.25 micrograms/g) four times a week for 8 weeks significantly (P less than 0.05) inhibited tumor growth, and in one mouse the tumor had almost completely disappeared by the middle of the sixth week of treatment. In addition, dexamethasone induced an apparent luminal structure in the tissue section of the tumor and enhanced the immunoperoxidase reaction to epithelial membrane antigen. The results indicate that dexamethasone inhibited the proliferation of HSG cells in the nude mouse transplantation system probably by inducing the cellular differentiation of the HSG cells toward the more differentiated ductal epithelia via glucocorticoid receptors.

Histopathological studies on experimental murine salivary gland tumours treated with adriamycin: evidence for an anti-myoepithelial cell action.

Mice bearing pleomorphic salivary tumours induced by polyoma virus underwent chemotherapy with Adriamycin. Animals were killed at 2 day intervals and the tumours examined histologically. Two days after a single dose of Adriamycin, gross vacuolation degeneration was seen around ductal cells at sites corresponding to myoepithelial cells. At subsequent time periods there was progressive degeneration of other tumour elements. It is proposed that myoepithelial cell death in salivary gland tumours induced by Adriamycin may be an important factor in the early response of this tumour to chemotherapy.

Combination chemotherapy of head and neck cancer.

A total of 77 patients with cancer of the head and neck area were treated with five different drug combination regimens. Five of the 77 patients had lymphoepithelioma; four had adenocystic carcinoma, and 68 had squamous-cell carcinoma of the head and neck (16 from the skin). Of these 77 patients, 16 had no previous treatment, five had surgery, 11 had radiotherapy, and 45 had surgery and radiotherapy. The first regimen consisted of a four-day Bleomycin infusion followed after a 24-hour rest, by cyclophosphamide (Cytoxan), Vincristine (Oncovin), methotrexate and 5-Fluorouracil (5-FU) (B-COMF). The next three regimens consisted of a four-day Bleomycin course, followed by either Cytoxan and methotrexate (B-CM), Cytoxan and 5-FU (B-CF) or Methotrexate and 5-FU (B-MF). The fifth regimen consisted of Bleomycin concomitant with Cytoxan, Methotrexate, and 5-FU (B-CMF). Of the 49 patients receiving B-COMF and B-CMF, 12 showed a complete response and 12 a partial response. Among the 28 patients receiving Bleomycin, followed by any one of the two drug regimens, only six showed a partial response. The severity of the thrombocytopenia, number of drugs, lymphoepithelioma histology and performance status of the patient influenced the rate of response. Drug toxicity consisted mostly in myelosuppression. The B-CMF combination is highly effective and can be used as an adjuvant to surgery and/or radiotherapy.