Response-adapted volume de-escalation (RAVD) in locally advanced head and neck cancer.

BACKGROUND: Efforts to reduce the late toxicity associated with chemoradiation (CRT) for locally advanced head and neck squamous cell cancer (LA-HNSCC) have focused on radiotherapy (RT) dose de-escalation. In this phase I/II protocol investigating the addition of everolimus to induction chemotherapy (IC), we incorporated a novel response-adapted volume de-escalation (RAVD) approach using IC response to guide the extent of RT volume reduction. PATIENTS AND METHODS: Patients with measurable LA-HNSCC received two cycles of IC (cisplatin, paclitaxel, cetuximab ± everolimus). Patients with ≥50% reduction in the sum of tumor diameters [good response (GR)] received TFHX (paclitaxel, fluorouracil, hydroxyurea, and 1.5 Gy twice daily RT every other week) to a dose of 75 Gy with the single planning target volume (PTV1) encompassing exclusively gross disease. Patients with <50% response [non-response (NR)] were treated with TFHX encompassing PTV1 and the next nodal station at risk (PTV2) to a dose of 45 Gy followed by a sequential boost to PTV1 to a dose of 75 Gy. RESULTS: Ninety-four patients were enrolled. Randomization to everolimus was discontinued on interim analysis after 50 patients due to futility. IC response was evaluable in 89 patients. Thirty-seven patients (41.6%) had GR and 52 (58.4%) had NR. There was a trend for improved progression-free (P = 0.086) but not overall survival (P = 0.94) for GR versus NR. The 2-year PFS and OS were 86.0% and 83.5% for GR and 68.7% and 85.4% for NR, respectively. NR were significantly more likely to undergo G-tube placement during treatment (50.0% GR versus 73.5% NR, P = 0.040) and be G-tube dependent at 6-month follow-up (5.7% GR versus 32.6% NR, P = 0.005). CONCLUSIONS: The addition of everolimus to IC was not beneficial. The elimination of elective nodal coverage in patients with GR to IC did not appear to compromise outcomes and resulted in significantly decreased late toxicity. Further investigation of RAVD is warranted. CLINICALTRIALSGOV: NCT01133678.

STAT3 mediates IL-6-induced growth inhibition in the human prostate cancer cell line LNCaP.

BACKGROUND: In prostate cancer, we and others have observed distinct phenotypic responses to interleukin-6 (IL-6), which acts either as a paracrine growth inhibitor in the LNCaP cell line or as an autocrine growth stimulator in PC-3, DU145, and TSU cell lines. To understand the underlying mechanism responsible for this phenotypic difference, we investigated differences in the IL-6-induced Janus kinase-signal transducers and activators of transcription (JAK-STAT) signal transduction pathway between these two phenotypes. METHODS: Prostate cancer cell lines were assayed for STAT3 activity by immunoblotting, electrophoretic gel shift assays (EMSA), and a luciferase reporter assay to test for STAT3 protein expression, phosphorylation, DNA binding, and transcriptional activity. To address the physiological role of STAT3, we introduced a dominant-negative mutant of STAT3 into LNCaP cells and assayed the effects of IL-6 on cell growth of this stable transfectant by cell counting, clonogenic assays, and c-myc expression. RESULTS: IL-6 induced transcriptional activity of STAT3 only in LNCaP. STAT3 was transcriptionally inactive in PC-3, TSU, and DU145 at the level of protein expression, tyrosine phosphorylation, and DNA binding/transcriptional activity, respectively. An isolated LNCaP subclone containing a dominant-negative mutant of STAT3, LNCaP-SF, did not show STAT3-DNA binding or transcriptional activity. LNCaP-SF exhibited a proliferative response to IL-6 as compared to the control LNCaP-neo clone, which underwent growth arrest. Unlike LNCaP-neo, LNCaP-SF was able form colonies and to maintain c-myc expression in the presence of IL-6. CONCLUSIONS: STAT3 transcriptional activation correlates with the growth-inhibitory signal of IL-6 in LNCaP, suggesting that STAT3 transcriptional activity is an important determinant in the different phenotypic responses to IL-6 in prostate cancer.

STAT3 mediates IL-6-induced neuroendocrine differentiation in prostate cancer cells .

BACKGROUND: In the human prostate cancer cell line LNCaP, interleukin (IL)-6 has been shown to regulate both growth and neuroendocrine (NE) differentiation. We recently observed that IL-6 mediated growth arrest in LNCaP by activating STAT 3. Since differentiation and growth arrest are often associated processes, we investigated whether STAT3 also mediated NE differentiation in this prostate cancer cell line. METHODS: We treated previously characterized clones LNCaP-neo (neomycin-resistant LNCaP) and LNCaP-SF (LNCaP-STAT3 dominant negative mutant) with IL-6 and screened for NE differentiation by observing morphological changes and immunoblotting for two NE markers, neuron-specific enolase (NSE) and chromogranin A (ChA). To characterize further the role of STAT3 in growth arrest and differentiation, we transfected a wild-type STAT3 vector into PC-3 cells and generated a subclone PC-3-S3. In this clone, we assessed differentiation by observing morphological changes and determined growth responses by cell counting and clonogenic assays. RESULTS: We observed that IL-6 induced formation of neurite extensions, morphologic features associated with NE differentiation, and enhanced expression of neuronal markers ChA and NSE in LNCaP-neo cells. In contrast, LNCaP-SF, possessing a dominant negative mutant form of STAT3, exhibited no characteristics of IL-6 induced NE differentiation. Furthermore, expression of a constitutively phosphorylated wild-type STAT3 in PC-3 cells inhibited growth and induced the formation of neurite extensions and NSE expression. CONCLUSIONS: These results indicate that STAT3 is a mediator of both NE differentiation and growth inhibition in LNCaP and PC-3, suggesting a connection between growth inhibition and NE differentiation in prostate cancer.

Interleukin-6 activates phosphatidylinositol-3 kinase, which inhibits apoptosis in human prostate cancer cell lines.

BACKGROUND: A number of recent studies have identified interleukin (IL)-6 as an important regulator of prostate cancer growth. Here, we investigate the potential interaction of IL-6 with phosphatidylinositol (PI)-3 kinase, a key growth regulatory enzyme, in prostate cancer cell lines. METHODS: Tyrosine phosphorylation of p85, the regulatory subunit of PI-3 kinase, in the human prostate cancer cell lines LNCaP and PC-3 was assessed by sequential immunoprecipitation with anti-p85 antibody and immunoblotting with anti-phosphotyrosine. The effects of wortmannin, an inhibitor of PI-3 kinase, and/or IL-6 on cell growth were assessed by MTT assays. DNA laddering experiments were performed to assay for programmed cell death. RESULTS: Tyrosine phosphorylation of p85 is upregulated by IL-6 in both LNCaP and PC-3. IL-6 promotes coprecipitation of p85 with gp130, the signal-transducing component of the IL-6 receptor. Inhibition of PI-3 kinase with wortmannin induces programmed cell death in PC-3 cells. In contrast, wortmannin has no effect on LNCaP growth when used alone; however, combined with IL-6, wortmannin promotes apoptosis in these cells. CONCLUSIONS: PI-3 kinase is involved in IL-6 signal transduction and delivers an antiapoptotic signal in human prostate cancer cell lines.

Characterization of the role of IL-6 in the progression of prostate cancer.

BACKGROUND: We recently identified IL-6, a pleiotropic cytokine implicated in the neoplastic process of a variety of neoplasms, as a mediator of prostate cancer morbidity. In the present study, we investigated the expression of members of the IL-6 supergene family and related cytokines and the potential role of IL-6 in prostate cancer growth regulation. METHODS: Five established human prostate cancer cell lines were screened by ELISA for production of granulocyte colony-stimulating factor (G-CSF), leukemia inhibitory factor (LIF), ciliary neurotrophic factor (CNTF), oncostatin M (OSM), tumor necrosis factor (TNF), interleukin-1 (IL-1), and granulocyte macrophage colony-stimulating factor (GM-CSF). Expression of the ligand-binding component of the IL-6 receptor, IL-6Rp80, was evaluated by ELISA and RT-PCR. Sequential immunoprecipitation and immunoblotting were performed to assay for expression of the signal-transducing component of the IL-6 receptor, gp130. The effects of IL-6 on cell growth were assessed by MTT assays. RESULTS: The three hormone-refractory cell lines, DU-145, TSU, and PC-3, secreted distinct combinations of cytokines (DU-145: IL-6, GM-CSF; TSU: IL-6, LIF; PC-3: IL-6, G-CSF, LIF, IL-1, GM-CSF), each uniformly expressing IL-6. In contrast, neither of the two hormone-dependent cell lines, LNCaP-ATCC and LNCaP-GW, secreted significant quantities of any of the cytokines analyzed. None of the cell lines secreted detectable quantities of OSM, CNTF, or TNF. All cell lines, irrespective of hormone status, expressed both Il-6Rp80 and gp130. Addition of IL-6 in vitro inhibited growth of hormone-dependent cells, but had no effect on hormone-refractory lines. Anti-IL-6 neutralizing antibody inhibited growth of hormone-refractory cells. CONCLUSIONS: IL-6 appears to undergo a functional transition from paracrine growth inhibitor to autocrine growth stimulator during progression of prostate cancer to the hormone-refractory phenotype.