nitrophenols and Carcinoma--Squamous-Cell

Head and neck squamous cell carcinomas (HNSCC) are common human malignancies with poor clinical outcomes. The 5-year survival rates for patients with advanced stage HNSCC have not changed appreciably in the past few decades, underscoring a dire need for improved therapeutic options. HNSCC is frequently characterized by overexpression of anti-apoptotic Bcl-2 family members. Increased levels of these anti-apoptotic proteins have been associated with radio- and chemoresistance and poor clinical outcome. The aim of this study was to evaluate combined effects of radiation and ABT-737, a BH3-mimetic molecule, in HNSCC. Although ABT-737, as a single agent, was largely ineffective at promoting HNSCC cell death, we found that combining ABT-737 and radiation induced strong synergistic apoptosis in HNSCC cell lines and delayed tumoral growth in vivo. Moreover, we demonstrated for the first time that ABT-737, alone or in combination with radiation, can efficiently eliminate cancer stem cells (CSCs). Altogether, our results indicate that therapy targeting anti-apoptotic Bcl-2 family members could be a highly effective potential adjuvant to radiotherapy capable of targeting CSCs in HNSCC and therefore overcoming cancer recurrence and metastasis.

Topics: Animals; Apoptosis; Biphenyl Compounds; Carcinoma, Squamous Cell; Cell Line, Tumor; Head and Neck Neoplasms; Humans; Mice; Mice, Inbred NOD; Mice, Nude; Mice, SCID; Neoplastic Stem Cells; Nitrophenols; Piperazines; Radiation-Sensitizing Agents; Squamous Cell Carcinoma of Head and Neck; Sulfonamides; Xenograft Model Antitumor Assays

Mammalian target of rapamycin (mTOR) is a therapeutic target for head and neck squamous cell carcinoma (HNSCC). Here, we evaluated the activity of AZD-2014, a potent mTOR complex 1/2 (mTORC1/2) dual inhibitor, against HNSCC cells. We showed that AZD-2014 blocked mTORC1/2 activation in established and primary human HNSCC cells, where it was anti-proliferative and pro-apoptotic. Yet, AZD-2014 was non-cytotoxic to the human oral epithelial cells with low basal mTORC1/2 activation. In an effect to identify possible AZD-2014 resistance factors, we showed that the anti-apoptosis protein Bcl-2 was upregulated in AZD-2014-resistant SQ20B HNSCC cells. Inhibition of Bcl-2 by ABT-737 (a known Bcl-2 inhibitor) or Bcl-2 shRNA dramatically potentiated AZD-2014 lethality against HNSCC cells. On the other hand, exogenous overexpression of Bcl-2 largely attenuated AZD-2014's activity against HNSCC cells. For the in vivo studies, we showed that oral gavage of AZD-2014 suppressed SQ20B xenograft growth in severe combined immunodeficient (SCID) mice. It also significantly improved mice survival. Importantly, AZD-2014's anti-HNSCC activity in vivo was potentiated with co-administration of ABT-737. The preclinical results of this study suggest that AZD-2014 could be further tested as a valuable anti-HNSCC agent, either alone or in combination with Bcl-2 inhibitors.

Topics: Aged; Benzamides; Biphenyl Compounds; Carcinoma, Squamous Cell; Head and Neck Neoplasms; Humans; Male; Middle Aged; Morpholines; Nitrophenols; Piperazines; Proto-Oncogene Proteins c-bcl-2; Pyrimidines; Squamous Cell Carcinoma of Head and Neck; Sulfonamides; TOR Serine-Threonine Kinases

We investigated the effects of recombinant human bone morphogenetic protein-7 (rhBMP-7) on the behaviour of oral keratinocytes and head and neck squamous cell carcinoma (SCC) cells in vitro. Expression of all three BMP receptors was high (p<0.01), and rhBMP-7 exhibited significant dose-related inhibitory effects on the doubling time and viability of cancer cells (p<0.01), but not on the proliferation or viability of oral keratinocytes. It elicited no significant effect on the invasion of Matrigel in SCC of the head and neck. Results indicate that in cell culture, rhBMP-7 exerts antineoplastic effects. This should be tested in an orthotopic animal model to more closely replicate in vivo effects.

Topics: Antineoplastic Agents; Bone Morphogenetic Protein 7; Bone Morphogenetic Protein Receptors, Type I; Bone Morphogenetic Protein Receptors, Type II; Carcinoma, Squamous Cell; Cell Culture Techniques; Cell Death; Cell Line; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cellular Senescence; Culture Media, Serum-Free; Humans; Indicators and Reagents; Keratinocytes; Mouth Neoplasms; Neoplasm Invasiveness; Nitrophenols; Organophosphorus Compounds

Oral and oropharyngeal cancers are the sixth most common cancers worldwide. Despite intensive investigation, oral squamous cell carcinomas (OSCC) represent a clinical challenge resulting in significant morbidity and mortality. Resistance to cell death is common in OSCC and is often mediated by the Bcl-2 family proteins. Among all anti-apoptotic Bcl-2 family members, Mcl-1 functions as a major survival factor, particularly in solid cancers. Despite the confirmed importance of Mcl-1 in several neoplasms, the role of Mcl-1 in OSCC survival has yet to be explored. In this study, we found that knocking down Mcl-1 sensitized OSCC cells to ABT-737, which binds to Bcl-2/Bcl-xL but not Mcl-1. We report for the first time that a BH3 mimetic, Sabutoclax, which functions as an inhibitor of all anti-apoptotic Bcl-2 proteins, induced cancer-specific cell death in an Mcl-1-dependent manner through both apoptosis and toxic mitophagy. In vivo studies demonstrated that Sabutoclax alone decreased tumor growth in a carcinogen-induced tongue OSCC mouse model. In a combination regimen, Sabutoclax and COX-2 inhibitor, Celecoxib, synergistically inhibited the growth of OSCC in vitro and also significantly reduced OSCC tumor growth in vivo. Overall, these results identify Mcl-1 as a therapeutic prospective target in OSCC.

Topics: 4-Nitroquinoline-1-oxide; Animals; Apoptosis; Apoptosis Regulatory Proteins; Autophagy-Related Protein 5; Autophagy-Related Protein 7; bcl-X Protein; Beclin-1; Biphenyl Compounds; Carcinoma, Squamous Cell; Celecoxib; Cell Line, Tumor; Cyclooxygenase 2 Inhibitors; Female; Gossypol; Humans; Membrane Proteins; Mice; Mice, Inbred BALB C; Mice, Nude; Microtubule-Associated Proteins; Mitophagy; Mouth Neoplasms; Myeloid Cell Leukemia Sequence 1 Protein; Nitrophenols; Piperazines; Proto-Oncogene Proteins; Quinolones; Random Allocation; RNA Interference; RNA, Small Interfering; Sulfonamides; Ubiquitin-Activating Enzymes; Xenograft Model Antitumor Assays

To date, many different chemotherapeutic agents have been widely used as common treatments for oral cancers. However, their therapeutic effects have been disappointing, and these agents may have unwanted side effects. Among the many regulatory factors, overexpression of pro-survival Bcl-2 family members may promote resistance to chemotherapeutic drugs in many tumors. The BH3 domain-only proteins effectively antagonize their apoptotic activities. Therefore, there is substantial interest in developing chemotherapeutic drugs that directly target pro-survival Bcl-2 proteins by mimicking the BH3 domain and unleashing pro-apoptotic molecules in tumor cells. Among the numerous available small molecule BH3 mimetics, ABT-737, a potent small molecule that binds to Bcl-2/Bcl-xL with high affinity, has anti-tumor activity in a wide variety of cancer cells. However, the effects of ABT-737 on human oral cancers and the underlying molecular mechanisms have not previously been elucidated. In the present study, we observed that inactivation of the ERK1/2 signaling pathway using ABT-737 dramatically increased the expression of pro-apoptotic protein Bim via transcriptional and/or posttranslational regulation, in a cell type-dependent manner, inducing mitochondria-mediated apoptosis of human oral cancer cells. To the best of our knowledge, this is the first demonstration of the antitumor effects of ABT-737 on human oral cancers.

Topics: Antineoplastic Agents; Apoptosis; Apoptosis Regulatory Proteins; Bcl-2-Like Protein 11; BH3 Interacting Domain Death Agonist Protein; Biomimetics; Biphenyl Compounds; Carcinoma, Mucoepidermoid; Carcinoma, Squamous Cell; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; Humans; MAP Kinase Signaling System; Membrane Proteins; Molecular Targeted Therapy; Mouth Neoplasms; Nitrophenols; Piperazines; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Sulfonamides

Tumour progression and therapy resistance in squamous cell carcinoma of the skin (SCC) is strongly associated with resistance to intrinsic mitochondrial apoptosis. We thus investigated the role of various anti-apoptotic Bcl-2 proteins for apoptosis protection in SCC using the BH3 agonist ABT737 that can overcome multidomain Bcl-2 protein protection. Sensitive SCC cells underwent rapid loss of mitochondrial membrane potential (MMP), subsequent apoptosis concomitant with caspase-3 activation and an early release of mitochondria-derived cytochrome c and smac/DIABLO. In contrast, ABT737 resistance in subsets of SCC cells was not explained by XIAP, important for protection from DR-induced apoptosis in SCC. Of note, ABT737 did not prime SCC cells to DR-induced apoptosis. Interestingly, the ratio of Mcl-1 and Noxa determined sensitivity to ABT737: loss of Mcl-1 rendered resistant cells sensitive to ABT737, whereas loss of Noxa promoted resistance in sensitive cells. In line, suppression of Mcl-1 by the pan-Bcl-2 inhibitor Obatoclax or overexpression of Noxa rendered resistant SCC cells sensitive to BH3 mimetics. Our data indicate that targeting of the Mcl-1/Noxa axis is important to overcome resistance to mitochondrial apoptosis in SCC. Therefore, combination treatment of ABT737 or derivatives with Mcl-1 inhibitors, or inducers of Noxa, may represent a novel option of targeted therapy in metastatic SCC of the skin.

Topics: Antineoplastic Agents; Apoptosis; Biphenyl Compounds; Carcinoma, Squamous Cell; Cell Line, Tumor; Cells, Cultured; Drug Resistance, Neoplasm; Humans; Keratinocytes; Myeloid Cell Leukemia Sequence 1 Protein; Nitrophenols; Piperazines; Proto-Oncogene Proteins c-bcl-2; Skin Neoplasms; Sulfonamides

In this issue of Cancer Discovery, He and colleagues determined that Mcl-1 levels are a key factor in the response to histone deacetylase (HDAC) inhibitors, and FBXW7 mutation is a biomarker for sensitivity to HDAC inhibition. They also present evidence for synergy between treatment with HDAC inhibitors and Bcl-2-targeted therapeutics. These data provide an exciting new biomarker and combination therapy that should be evaluated clinically.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Biphenyl Compounds; Carcinoma, Squamous Cell; Cell Cycle Proteins; F-Box Proteins; F-Box-WD Repeat-Containing Protein 7; Histone Deacetylase Inhibitors; Histone Deacetylases; Humans; Myeloid Cell Leukemia Sequence 1 Protein; Nitrophenols; Piperazines; Proto-Oncogene Proteins c-bcl-2; Sulfonamides; Ubiquitin-Protein Ligases

Effective targeted therapeutics for squamous cell carcinoma (SCC) are lacking. Here, we uncover Mcl-1 as a dominant and tissue-specific survival factor in SCC, providing a roadmap for a new therapeutic approach. Treatment with the histone deacetylase (HDAC) inhibitor vorinostat regulates Bcl-2 family member expression to disable the Mcl-1 axis and thereby induce apoptosis in SCC cells. Although Mcl-1 dominance renders SCC cells resistant to the BH3-mimetic ABT-737, vorinostat primes them for sensitivity to ABT-737 by shuttling Bim from Mcl-1 to Bcl-2/Bcl-xl, resulting in dramatic synergy for this combination and sustained tumor regression in vivo. Moreover, somatic FBW7 mutation in SCC is associated with stabilized Mcl-1 and high Bim levels, resulting in a poor response to standard chemotherapy but a robust response to HDAC inhibitors and enhanced synergy with the combination vorinostat/ABT-737. Collectively, our findings provide a biochemical rationale and predictive markers for the application of this therapeutic combination in SCC.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Biphenyl Compounds; Carcinoma, Squamous Cell; Cell Cycle Proteins; Cell Line, Tumor; Drug Synergism; F-Box Proteins; F-Box-WD Repeat-Containing Protein 7; Histone Deacetylase Inhibitors; Histone Deacetylases; Humans; Hydroxamic Acids; Mice; Mice, Nude; Myeloid Cell Leukemia Sequence 1 Protein; Nitrophenols; Piperazines; Proto-Oncogene Proteins c-bcl-2; Sulfonamides; Transfection; Ubiquitin-Protein Ligases; Vorinostat; Xenograft Model Antitumor Assays

Overexpression of Bcl-X(L), an antiapoptotic Bcl-2 family member, occurs in a majority of head and neck squamous cell carcinomas (HNSCCs) and correlates with chemotherapy resistance in this disease. Overexpression of Bcl-2 is also observed in HNSCC, albeit less frequently. We have previously shown that peptides derived from the BH3 domains of proapoptotic proteins can be used to target Bcl-X(L) and Bcl-2 in HNSCC cells, promoting apoptosis. In this report, we examined the impact of ABT-737 (for structure, see Nature 435: 677-681, 2005 ), a potent small-molecule inhibitor of Bcl-X(L) and Bcl-2, on HNSCC cells. As a single agent, ABT-737 was largely ineffective at promoting HNSCC cell death. By contrast, ABT-737 strongly synergized with the chemotherapy drugs cisplatin and etoposide to promote HNSCC cell death and loss of clonogenic survival. Synergism between ABT-737 and chemotherapy was associated with synergistic activation of caspase-3 and cleavage of poly(ADP-ribose) polymerase. Treatment with ABT-737 plus chemotherapy resulted in dramatic up-regulation of proapoptotic Noxa protein, and small interfering RNA (siRNA)-mediated inhibition of Noxa up-regulation partially attenuated cell death by the synergistic combination. Treatment with cisplatin or etoposide, alone or in combination with ABT-737, resulted in substantial down-regulation of Mcl-1L, a known inhibitor of ABT-737 action. Further down-regulation of Mcl-1L using siRNA failed to enhance killing by the cisplatin/ABT-737 synergistic combination, indicating that chemotherapy treatment of HNSCC cells is sufficient to remove this impediment to ABT-737. Together, our results demonstrate potent synergy between ABT-737 and chemotherapy drugs in the killing of HNSCC cells and reveal an important role for Noxa in mediating synergism by these agents.

Topics: Antineoplastic Agents; Apoptosis; bcl-X Protein; Biphenyl Compounds; Boronic Acids; Bortezomib; Carcinoma, Squamous Cell; Cell Line, Tumor; Cisplatin; Drug Synergism; Head and Neck Neoplasms; Humans; Myeloid Cell Leukemia Sequence 1 Protein; Nitrophenols; Piperazines; Proto-Oncogene Proteins c-bcl-2; Pyrazines; Signal Transduction; Sulfonamides

Na+,K+-ATPase (EC 3.6.1.37) is assumed to be involved in the transport of cisplatin [cis-diamminedichloroplatinum(II)] into cells and to act as a modulator of 5-fluorouracil (5-FU) in combination therapy of cisplatin and 5-FU. Whereas inhibition of Na+,K+-ATPase activity by cisplatin is expected to have effects on both anti-cancer therapy and nephrotoxicity, the inhibition mechanism remains to be elucidated. We studied the inhibition of Na+,K+-ATPase activity by cisplatin using an enzyme partially purified from Ca9-22 cells derived from a human squamous cell carcinoma of the gingiva. Cisplatin inhibited the Na+,K+-dependent ATP hydrolysis activity, and this inhibition depended on both the concentration of cisplatin and the preincubation time with cisplatin. The time-dependent inhibition was thought to be caused by a slow change of cisplatin from the inactive to the active form. We further tested the effect of cisplatin on the partial reactions of the enzyme, Na+-dependent ATP hydrolysis and K+-dependent pnitrophenylphosphate hydrolysis activities to determine which step in the reaction sequence of Na+,K+-ATPase was inhibited. Cisplatin inhibited both activities depending on its concentration and the preincubation time, whereas the Na+-dependent ATP hydrolysis activity was inhibited even at lower concentrations. Formation of a phosphointermediate of Na+,K+-ATPase was also inhibited by cisplatin depending on the concentration and preincubation time. Cisplatin (500 microM) and 8-fold higher concentration of 2-mercaptoethanol (2-ME; 4 mM) prevented inactivation of the enzyme by cisplatin, and the Na+,K+-ATPase activity inhibited by pretreatment with cisplatin was also recovered almost completely by 2-ME. These results suggest that the active form of cisplatin inhibits the Na+,K+-ATPase activity by inhibiting the formation of a phosphointermediate of the enzyme and that the inhibition by cisplatin is arrested by an addition of thiol group.

Topics: Antineoplastic Agents; Carcinoma, Squamous Cell; Cisplatin; Dose-Response Relationship, Drug; Enzyme Inhibitors; Gingival Neoplasms; Humans; Mercaptoethanol; Nitrophenols; Organophosphorus Compounds; Sodium-Potassium-Exchanging ATPase; Time Factors; Tumor Cells, Cultured

One hundred eighty-three patients with advanced solid neoplasms were tested for their ability to react to four common skin test antigens (tuberculin PPD, streptokinase-streptodornase, mumps, and Monilia) and their ability to develop delayed cutaneous hypersensitivity (DCH) to 2, 4 dinitrochlorobenzene (DNCB). All patients were followed for at least 6 months or until death. Histologic tumor types studied were: melanoma (65), sarcoma (28), squamous cell carcinoma (23), and adenocarcinoma (67). The rate of progression of disease within 6 months of testing was lower in patients who had a positive response to a challenging dose of 50 mug of DNCB. Reactivity to recall antigens had no prognostic value except in patients with adenocarcinomas. Among patients with adenocarcinoma, those who reacted strongly to DNCB and one or more skin test antigens had the best prognosis, while those who were nonreactive to all had the worst prognosis (progression rate: 18% vs. 78%). Peripheral lymphocyte counts were related to the results of DCH to DNCB and skin tests. The preseence or absence of lymphocytopenia (count less than 1000/mm3) had prognostic value in patients who had positive skin test(s). In such patients, the disease progression rate was much higher in patients who were anergic to DNCB and who were lymphocytopenic (90% vs. 40%). These data suggest that DCH to DNCB, recall antigens, and peripheral lymphocyte counts are useful immunologic measurements in patients with advanced cancer. Although the prognostic value of each individual test is relatively limited, the predictive worth can be increased when multiple tests are employed. Pertinent findings reported in the literature are reviewed.

Topics: Adenocarcinoma; Candida; Carcinoma, Squamous Cell; Humans; Hypersensitivity, Delayed; Immunity, Cellular; Leukocyte Count; Lymphocytes; Melanoma; Mumps virus; Neoplasms; Nitrophenols; Prognosis; Sarcoma; Skin Tests; Streptodornase and Streptokinase; Time Factors; Tuberculin Test

Topics: Animals; Antibodies; Arsphenamine; Carcinoembryonic Antigen; Carcinoma, Squamous Cell; Cell Line; Colonic Neoplasms; Cytotoxicity Tests, Immunologic; Glucose Oxidase; Goats; Haptens; HeLa Cells; Humans; Immunoglobulin G; Laryngeal Neoplasms; Nitrophenols; Peroxidases