Prognostic Value of Different Allelic Polymorphism of Aldosterone Synthase Receptor in a Congestive Heart Failure European Continental Ancestry Population.

Aldosterone synthase (CYP11B2) is as an 9-exon gene on chromosome 8q22 and exists as a common single nucleotide polymorphism C-T transition for position -344. The aim of this study was to assess the -344T/C polymorphism of the aldosterone synthase promoter in a European continental ancestry congestive heart failure (CHF) population. METHODS: Patients discharged after an acute decompensation were enrolled and underwent echocardiography, determination of BNP, evaluation of non-invasive cardiac outputs and determination of -344 T/C SNP in the aldosterone synthase gene. RESULTS: 175 patients (137 male; age 69.9 ± 10.2 years) were enrolled. The genotype distribution of -344 T/C SNP demonstrated a TT genotype in 61 patients (34.9%), CT in 80 (45.7%) and finally CC in 34 (19.4%) CHF patients. According to presence of C allele, CHF patients were divided into C group (-CT/CC genotype, 114 subjects) and T Group (-TT genotype, 61 subjects). The two groups did not differ in term of age, non-invasive cardiac output at rest, creatinine level or end-systolic or diastolic left ventricle diameter, LVEF and BNP. In group C patients in comparison than in group T a higher degree of disability (Barthel Index p = 0.004), NYHA class (p = 0.02) and a lower cardiac index (p = 0.01) emerged. Moreover, the two groups showed a similar clinical outcome (death for any cause/hospital readmission for CHF) at 48 month follow-up (p = 0.16; log-rank 1.99). CONCLUSIONS: In European continental ancestry patients the C allele (CC or CT) at -344T/C SNP in the aldosterone synthase gene does not significantly influence clinical prognosis of CHF.

Elevated basal antibody-dependent cell-mediated cytotoxicity (ADCC) and high epidermal growth factor receptor (EGFR) expression predict favourable outcome in patients with locally advanced head and neck cancer treated with cetuximab and radiotherapy.

BACKGROUND: Antibody-dependent cell-mediated cytotoxicity (ADCC) may contribute to the antitumor activity of cetuximab. However, the extent of this contribution is unclear. In this study, we investigated the impact of baseline ADCC on the outcome of patients with locally advanced squamous cell carcinoma treated with cetuximab and radiotherapy. METHODS: We determined baseline ADCC in 28 patients treated with cetuximab and radiotherapy and in 15 patients treated with chemoradiation. We linked the values observed with complete response and with overall survival. We also considered the role of epidermal growth factor receptor (EGFR) expression and studied the combined effect of EGFR and ADCC. RESULTS: We observed a wide range of baseline values of ADCC. Complete response did not correlate with either ADCC or EGFR expression. However, when ADCC and EGFR were considered together using a mixed score, they significantly correlated with achieving a complete response (p = 0.04). High baseline ADCC significantly correlated with outcome compared to low (p = 0.03), but not in patients treated without cetuximab. Patients showing high baseline levels of both ADCC and EGFR3+ achieved the best outcome compared to the others (p = 0.02). CONCLUSIONS: In this study, patients treated with cetuximab and radiotherapy, showing high baseline of both ADCC and EGFR3+, have significant higher probability of achieving a complete response and a long overall survival compared to the others.

Telomeric repeat-binding factor 2: a marker for survival and anti-EGFR efficacy in oral carcinoma.

Oral Squamous Cell Carcinoma (OSCC) is the most common oral cancer worldwide. Treatments including surgery, radio- and chemo-therapies mostly result in debilitating side effects. Thus, a more accurate evaluation of patients at risk of recurrence after radio/chemo treatment is important for preserving their quality of life. We assessed whether the Telomeric Repeat-binding Factor 2 (TERF2) influences tumor aggressiveness and treatment response. TERF2 is over-expressed in many cancers but its correlation to patient outcome remains controversial in OSCC. Our retrospective study on sixty-two patients showed that TERF2 overexpression has a negative impact on survival time. TERF2-dependent survival time was independent of tumor size in a multivariate analysis. In vitro, TERF2 knockdown by RNA interference had no effect on cell proliferation, migration, senescence and apoptosis. Instead, TERF2 knockdown increased the expression of cytokines implicated in inflammation and angiogenesis, except for vascular endothelial growth factor. TERF2 knockdown resulted in a decrease vascularization and growth of xenograft tumors. Finally, response to erlotinib/Tarceva and cetuximab/Erbitux treatment was increased in TRF2 knocked-down cells. Hence, TERF2 may represent an independent marker of survival for OSCC and a predictive marker for cetuximab/Erbitux and erlotinib/Tarceva efficacy.

E1 detection as prognosticator in human papillomavirus-positive head and neck cancers.

PURPOSE: HPV-related locally advanced head and neck cancers (LA-HNCs) show a good prognosis. This study aimed to investigate the HPV prevalence in LA-HNCs and compare the prognostic value of E1, E6 and L1 genomic viral fragments and p16, individually and in combination, in order to find the best prognosticator in terms of overall survival (OS) and progression-free survival (PFS). PATIENTS AND METHODS: HPV16 was searched in 255 LA-HNC formalin-fixed paraffin-embedded tumor tissues, 89 oropharyngeal cancers (OPCs), and 166 non-OPCs by DNA-PCR with 3 primer pairs. p16 was analyzed by immunohistochemistry in 235 patients. RESULTS: The prevalence of positive samples decreased constantly from E6 to L1 and E1 in both OPCs and non-OPCs. Each LA-HNC patient highlighted variable positivity for each fragment. OPCs showed a higher prevalence of positive samples compared to non-OPCs.Positive coexistence of all the fragments was more common in OPCs (31.5%) than non-OPCs (4.2%), and E1 detection was always associated with E6 and L1. E1-positive OPCs showed improved OS (p = 0.012) and PFS (p = 0.036), while L1- or E6-positive ones did not. p16-positive patients were more prevalent in the OPC (29.8%) than the non-OPC group (7.3%) (p<0.0001) and its prognostic value was not superior to that of E1. However, the multivariate Cox analysis which included E1, L1, E6 status and p16 expression did not show a significant p value. CONCLUSIONS: Though HPV16 positivity measured by DNA-PCR was higher for L1 and E6, they performed weakly as prognosticators; E1 might become a strong prognostic marker for OS and PFS in OPCs.

Reoxygenation Reverses Hypoxia-related Radioresistance in Head and Neck Cancer Cell Lines.

BACKGROUND/AIM: Head and neck cancer (HNC) is characterized by epidermal growth factor receptor (EGFR) overexpression and radiotherapy (RT) resistance. Cancer cells are able to survive and proliferate in hypoxic conditions. Hypoxia can be transiently interrupted by phases of reoxygenation. This work aimed to analyze the reoxygenation effect on proliferation in response to radiation in HNC cells. MATERIALS AND METHODS: HNC cell lines CAL33 and CAL166 were subjected to an 8-Gy radiation dose in hypoxia and/or after reoxygenation. Cell proliferation and molecular factors involved in response to treatments were studied. RESULTS: Cytotoxicity test confirmed radioresistance in hypoxia and highlighted that reoxygenation before RT restores sensitivity in both cell lines. Our results showed a similar proliferation inhibition effect and EGFR modulation but a different cell death mechanism in the two cell lines after treatment. CONCLUSION: Reoxygenation before RT rescued radiosensitivity in HNC cells.

Genetic and epigenetic factors affect RET gene expression in breast cancer cell lines and influence survival in patients.

Germline and somatic mutations play a crucial role in breast cancer (BC), driving the initiation, progression, response to therapy and outcome of the disease. Hormonal therapy is limited to patients with tumors expressing steroid hormone receptors, such as estrogen receptor (ER), nevertheless resistance often limits its success.The RET gene is known to be involved in neurocristopathies such as Hirschsprung disease and Multiple Endocrine Neoplasia type 2, in the presence of loss-of-function and gain-of-function mutations, respectively. More recently, RET over-expression has emerged as a new player in ER-positive (ER+) BC, and as a potential target to enhance sensitivity and avoid resistance to tamoxifen therapy.Therefore, targeting the RET pathway may lead to new therapies in ER+ BC. To this end, we have investigated the molecular mechanisms which underlie RET overexpression and its possible modulation in two BC cell lines, MCF7 and T47D, showing different RET expression levels. Moreover, we have carried out a pilot association study in 93 ER+ BC patients. Consistent with the adverse role of RET over-expression in BC, increased overall survival was observed in carriers of the variant allele of SNP rs2435357, a RET polymorphism already known to be associated with reduced RET expression.

Schedule-dependent interaction between temsirolimus and cetuximab in head and neck cancer: a preclinical study.

Aberrant epidermal growth factor receptor (EGFR) signaling is associated with tumor growth in head and neck squamous cell carcinoma (HNSCC) and is a major focus of targeted therapy. The phosphatidylinositol-3-kinase/AKT/mammalian target of the rapamycin (PI3K/AKT/mTOR) signaling pathway is frequently mutated in HNSCC and is involved in disease progression and resistance to EGFR inhibitors. The aim of this study was to assess the antiproliferative effects of mTOR inhibition (temsirolimus) combined with the anti-EGFR monoclonal antibody cetuximab, administered according to different combination schedules. Antiproliferative effects of the combination of temsirolimus and cetuximab were determined on the representative HNSCC CAL33 cell line (PI3KCA H1047R mutated and K-RAS wild-type). In addition, key proteins related to the EGFR pathway (pEGFR/EGFR, pAKT/AKT) and the mTOR pathway (p-p70S6K1, p4E-BP1) were determined to explain the cytotoxic effects. Temsirolimus and cetuximab showed a synergistic effect when administered in combination. Supra-additive effect was lost when the two drugs were administered sequentially, irrespective of which drug was administered first. Synergistic effect of the combination was corroborated by a marked downregulation of pEGFR, significant downregulation of pAKT expression, and a marked diminution of p70S6K1 and p4E-BP1 expression. Our study demonstrated a synergistic effect of temsirolimus and cetuximab administered in combination, well illustrated by a simultaneous blockade of intracellular signaling pathways regulating cell proliferation and survival. These results establish the notion of a schedule dependency for the combined treatment, which can be of importance at the clinical level.

Correlation of TP53 and MDM2 genotypes and clinical outcome in platinum-treated head and neck cancer patients with more than 10 years' follow-up.

PURPOSE: Adequate biomarkers are still required to optimize therapy in patients with locally advanced head and neck squamous carcinomas (HNSCC) treated with chemoradiotherapy (CRT). METHODS: We updated the follow-up of 66 HNSCC patients treated with CRT we described more than 10 years ago, focusing on SNP Arg/Pro (R/P) at codon 72 and somatic mutations in TP53 and on SNP309 in the MDM2 gene. RESULTS: In wild-type TP53 cases, overall survival (OS) was longer in 72RR and less favorable in 72PP (p = 0.005); when TP53 was mutated, OS was longest in 72PP and less favorable in 72RR and 72RP (p = 0.058). Median OS was significantly shorter in patients with MDM2 SNP309 GG or GT genotypes compared with the TT genotype (p = 0.002). CONCLUSIONS: TP53 SNP72 may be useful in selecting patients for CRT, but has to be related to somatic TP53 mutations. The MDM2 SNP309, easily determined in peripheral blood, might be more convenient as a predictive biomarker.

Evaluation of antibody-dependent cell-mediated cytotoxicity activity and cetuximab response in KRAS wild-type metastatic colorectal cancer patients.

AIM: To investigate the prognostic role of invariant natural killer T (iNKT) cells and antibody-dependent cell-mediated cytotoxicity (ADCC) in wild type KRAS metastatic colorectal cancer (mCRC) patients treated with cetuximab. METHODS: Forty-one KRAS wt mCRC patients, treated with cetuximab and irinotecan-based chemotherapy in II and III lines were analyzed. Genotyping of single nucleotide polymorphism (SNP)s in the FCGR2A, FCGR3A and in the 3' untranslated regions of KRAS and mutational analysis for KRAS, BRAF and NRAS genes was determined either by sequencing or allelic discrimination assays. Enriched NK cells were obtained from lymphoprep-peripheral blood mononuclear cell and iNKT cells were defined by co-expression of CD3, TCRVα24, TCRVß11. ADCC was evaluated as ex vivo NK-dependent activity, measuring lactate dehydrogenase release. RESULTS: At basal, mCRC patients performing ADCC activity above the median level (71%) showed an improved overall survival (OS) compared to patients with ADCC below (median 16 vs 8 mo; P = 0.026). We did not find any significant correlation of iNKT cells with OS (P = 0.19), albeit we observed a trend to a longer survival after 10 mo in patients with iNKT above median basal level (0.382 cells/microliter). Correlation of OS and progression-free survival (PFS) with interesting SNPs involved in ADCC ability revealed not to be significant. Patients carrying alleles both with A in FCGR2A and TT in FCGR3A presented a trend of longer PFS (median 9 vs 5 mo; P = 0.064). Chemotherapy impacted both iNKT cells and ADCC activity. Their prognostic values get lost when we analysed them after 2 and 4 mo of treatment. CONCLUSION: Our results suggest a link between iNKT cells, basal ADCC activity, genotypes in FCGR2A and FCGR3A, and efficacy of cetuximab in KRAS wt mCRC patients.

p16 cutoff in head and neck squamous cell carcinoma: correlation between tumor and patient characteristics and outcome.

BACKGROUND: p16 has been indicated as a suitable surrogate biomarker of HPV infection. The prognosis of p16-positive oropharynx tumors (OTs) of squamous cell carcinoma (SCC) histology is better than that of p16-negative tumors. METHODS: We analyzed 209 samples of head and neck SCC to establish a predictive cutoff for p16 and determine the role of p16 positivity in OTs versus non-OTs. We compared the outcomes of tumors harboring any percentage of p16-positive cells (≥1%) with those of p16-negative OTs. We then considered 3 cutoffs (10%, 50% and 70% positive cells) to evaluate the outcome of OTs/non-OTs with similar p16 expression and p16-positive versus p16-negative tumors stratified by patient age. RESULTS: p16-negative tumors among OTs and non-OTs were 29% and 49%, respectively (p = 0.0054). The cumulative distribution showed that the positive values were located around 2 focus points: 2% and 96%. Subgroup analysis showed that only OTs occurred in young patients (aged <65 years) and that there was a ≥70% gain in survival in cases with p16-positive cells. CONCLUSIONS: p16 positivity influences outcome only in young patients and OTs (p = 0.048).

Combination of mTOR and EGFR targeting in an orthotopic xenograft model of head and neck cancer.

OBJECTIVES/HYPOTHESIS: Recent preclinical and clinical studies on head and neck squamous cell carcinoma (HNSCC) revealed synergistic effects when combining anti-EGFR agents with conventional chemotherapeutic drugs. Activation of the PI3-kinase/AKT/mTOR signaling pathway has been identified as an important mechanism implicated in tumor progression and resistance to EGFR inhibitors. The aim of this study was to investigate the effects of combining the mTOR inhibitor temsirolimus (Tem) with the anti-EGFR agent cetuximab (Cet) and conventional chemotherapeutic drugs (cisplatin and fluorouracil (C/F)) on an orthotopic model of HNSCC. STUDY DESIGN: Preclinical in vivo study. METHODS: We evaluated the anti-tumor efficacy (measured tumor volume) of Tem, Cet, and C/F, administered alone or in combination. Investigations were performed using a human HNSCC cell line, CAL33, injected into the mouth floor of nude mice. RESULTS: As compared with the control, the combination of Tem and Cet led to the highest tumor inhibition and induced almost complete tumor growth arrest (P = 0.001). Tem significantly enhanced the impact of the Cet-C/F combination on tumor growth (P < 0.001). The highest inhibitory effects of treatments on cell proliferation (Ki67 labeling), MAPK (pP42/44 labeling), and PI3K/AKT/mTOR (pS6R labeling) signaling pathways were found with the Tem-Cet association. CONCLUSION: In this orthotopic HNSCC model, the combination of Tem with Cet produced synergistic effects on tumor growth. These results were corroborated by a strong inhibition of both MAPK and PI3K-mTOR signaling pathways. LEVEL OF EVIDENCE: N/A.

The relevance of ADCC for EGFR targeting: A review of the literature and a clinically-applicable method of assessment in patients.

BACKGROUND: Advances in the understanding of tumor biology have led to the development of targeted therapies as monoclonal antibodies (MoAbs) in clinical oncology. Among their suggested mechanisms of action monoclonal antibodies (IgG1) selectively directed against tumor membrane receptors mediate of antibody-dependent cellular cytotoxicity (ADCC) by triggering Fc-γRIII on natural killer (NK) cells. METHODS: This study reviews the clinical context of ADCC measurement with a particular focus on EGFR targeting and describes an ex vivo ADCC method applied to MoAbs (cetuximab and panitumumab), against epidermal growth factor receptor (EGFR). The test performance was evaluated on different target cells lines (CAL166, A431, HNO91, CAL27), with different effector cells (peripheral blood mononuclear cells or natural killers -NK-) and in various experimental conditions, in order to establish a truly clinically applicable method. RESULTS: Using the experience available in the published literature, we optimized all variables involved in the experimental design: target cells type, numbers and ratio target cells and NK cells (effector cells) per well, time of exposure and repeatability. CONCLUSION: ADCC measurement may be of clinical relevance in the context of treatment with MoAbs. This study describes a non-radioactive method which has proven satisfactory in terms of sensitivity, reproducibility, feasibility and cost effectiveness for the measurement of ADCC activity mediated by NK with an orientation towards the EGFR target.

MGMT promoter methylation and glioblastoma: a comparison of analytical methods and of tumor specimens.

It is already well known that hypermethylation of the O6-methylguanine DNA methyltransferase (MGMT) gene promoter is a predictive biomarker of response to temozolomide treatment and of favorable outcomes in terms of overall survival (OS) and progression-free survival (PFS) in glioblastoma (GBM) patients. Nevertheless, MGMT methylation status has not currently been introduced into routine clinical practice, as the choice of the ideal technique and tissue sample specimen is still controversial. The aim of this study was to compare 2 analytical methods, methylation-specific polymerase chain reaction (MSP) and pyrosequencing (PSQ), and their use on 2 different tissue type samples, snap-frozen and formalin-fixed paraffin-embedded (FFPE), obtained from a single-center and uniformly treated cohort of 46 GBM patients. We obtained methylation data from all frozen tissues, while no results were obtained for 5 FFPE samples. The highest concordance for methylation was found on frozen tissues (88.5%, 23/26 samples), using PSQ (76.7%, 23/30 samples). Moreover, we confirmed that OS and PFS for patients carrying methylation of the MGMT promoter were longer than for patients with an unmethylated promoter. In conclusion, we considered MSP a limited technique for FFPE tissues due to the high risk of false-positive results; in contrast, our data indicated PSQ as the most powerful method to stratify methylated/unmethylated patients as it allows reaching quantitative results with high sensitivity and specificity. Furthermore, frozen tumor tissues were shown to be the best specimens for MGMT methylation analysis, due to the low DNA degradation and homogeneity in methylation throughout the tumor.

Treatment effect of buparlisib, cetuximab and irradiation in wild-type or PI3KCA-mutated head and neck cancer cell lines.

Introduction In complement to anti-EGFR therapy, the targeting of PI3K/AKT/mTOR signaling pathway is of particular interest in the management of Head and Neck Squamous Cell Carcinoma (HNSCC). Here, we assess the effects of PI3K inhibition combined with anti-EGFR monoclonal antibody cetuximab and/or irradiation (RT). Material and methods Anti-proliferative effects of the combination of buparlisib (a specific PI3K inhibitor), cetuximab and RT was determined in two HNSCC cell lines (CAL33, PI3KCA H1047R-mutated and CAL27, PI3KCA wild-type). We examined biochemical factors related to proliferation, apoptosis (caspases), DNA repair (ERCC1, XRCC1) and the PI3K pathway (pEGFR/EGFR, pAKT/AKT, p-p70, p4EBP1). Results The best synergistic combined treatment in inhibiting cell proliferation was sequence 2 (cetuximab followed by buparlisib) in both cell lines. Addition of RT increased sequence 2 anti-proliferative effect only in CAL27. Data on protein expression indicated a possible activation of mTORC2 complex and caspases proteins in CAL27 not seen in CAL33. In CAL33, the synergistic anti-proliferative effect of the two drugs may derive from the higher sensitivity of mutated cells to PI3K targeting. Conclusions Our study demonstrates a synergistic effect of cetuximab followed by buparlisib in both PI3KCA wild-type and mutated cells, even with different intracellular signaling cross-talk depending on mutational status.

Role of the renin-angiotensin-aldosterone system and the glutathione S-transferase Mu, Pi and Theta gene polymorphisms in cardiotoxicity after anthracycline chemotherapy for breast carcinoma.

BACKGROUND: Anthracyclines are among the most active drugs against breast cancer, but can exert cardiotoxic effects eventually resulting in congestive heart failure (CHF). Identifying breast cancer patients at high risk of developing cardiotoxicity after anthracycline therapy would be of value in guiding the use of these agents. AIMS: We determined whether polymorphisms in the renin-angiotensin-aldosterone system (RAAS) and in the glutathione S-transferase (GST) family of phase II detoxification enzymes might be useful predictors of left ventricular ejection fraction (LVEF) kinetics and risk of developing CHF. We sought correlations between the development of cardiotoxicity and gene polymorphisms in 48 patients with early breast cancer treated with adjuvant anthracycline chemotherapy. METHODS: We analyzed the following polymorphisms: p.Met235Thr and p.Thr174Met in angiotensinogen (AGT), Ins/Del in angiotensin-converting enzyme (ACE), A1166C in angiotensin II type-1 receptor (AGTR1A), c.-344T>C in aldosterone synthase (CYP11B2), p.Ile105Val in GSTP1. Additionally, we analyzed the presence or absence of the GSTT1 and GSTP1 genes. A LVEF <50% was detected at least once during the 3 years of follow-up period in 13 out of 48 patients (27.1%). CONCLUSION: RAAS gene polymorphisms were not significantly associated with the development of cardiotoxicity. GSTM1may be useful as a biomarker of higher risk of cardiotoxicity, as demonstrated in our cohort of patients (p=0.147).

Differential molecular mechanism of docetaxel-octreotide combined treatment according to the docetaxel-resistance status in PC3 prostate cancer cells.

To examine the effect and the molecular mechanisms of the combined treatment of the somatostatin (SST) analogue octreotide with docetaxel: analysis of proliferation, apoptosis and migration in the human prostate cancer cell line PC3, either sensitive (PC3wt) or made resistant to docetaxel (PC3R). We examined the effect of the two drugs individually or in combination on cell proliferation and migration by analysis of apoptosis and cell cycle proteins. The role of octreotide in modulating P-glycoprotein function was examined together with the modulation of SST receptors type 2 and 5 (SSTR2 and SSTR5). We observed an enhanced effect of docetaxel and octreotide given in combination or in sequence compared with either agent alone; this result was particularly evident when docetaxel was given before octreotide in PC3wt and when the two drugs were given together in PC3R cells. In contrast to lanreotide, our data indicate that octreotide does not act as a P-glycoprotein inhibitor in PC3R cells. A role of docetaxel and combined treatment in regulating SSTR2, SSTR5, proliferation and apoptosis gene expression is suggested as the possible mechanism for the enhanced effect observed. In addition, an evaluation of the effect of the combined treatment on cellular migration was examined, showing a moderate loss of invasive properties in PC3R cells. The present results confirm that SST analogues may be combined with docetaxel to increase the antitumour effect in patients with advanced prostate carcinoma.

Contrasted effects of the multitarget TKi vandetanib on docetaxel-sensitive and docetaxel-resistant prostate cancer cell lines.

OBJECTIVES: Overexpression of epidermal growth factor receptor (EGFR) and angiogenic factors is associated with the progression of androgen-independent prostate cancer (AIPC). We examined the effects of vandetanib, an inhibitor of vascular endothelial growth factor (VEGFR), EGFR, and rearranged during transfection (RET) tyrosine-kinase activities, alone or combined with docetaxel, on PC3 docetaxel-sensitive (PC3wt) or docetaxel-resistant (PC3R) AIPC cell growth in vivo and in vitro. METHODS: Mice bearing PC3wt or PC3R tumors were treated for 3 weeks with vandetanib (25 or 50 mg/kg/d p.o., 5 d/wk), docetaxel (10 or 30 mg/kg i.p., 1 d/wk), or their combination (low or high doses). Xenograft tumors were analyzed for expression of Ki-67, EGFR, VEGFR2, and production of VEGFA. RESULTS: On PC3wt, vandetanib at both doses stimulated tumor growth, whereas docetaxel at both doses exerted strong growth-inhibiting effects. The low-dose vandetanib-docetaxel combination resulted in tumor growth similar to that of control, whereas the high-dose combination induced a significant antiproliferative effect. In contrast, on PC3R, the low-dose of vandetanib had no effect on tumor growth, whereas the high-dose of vandetanib significantly inhibited tumor growth. Docetaxel at both doses exerted moderate and transient antitumor effects. The combination of high-dose vandetanib with high-dose docetaxel resulted in antiproliferative effects, which were lower than expected from the sum of individual drug effects. Importantly, tumor analyses revealed overexpression of the EGFR/VEGFR pathways in PC3R relative to PC3wt. CONCLUSION: Present results suggest that vandetanib should not be associated with docetaxel in treatment-naive or docetaxel-resistant prostate cancer (CaP). The use of high-dose vandetanib alone may warrant further investigation in patients with docetaxel-resistant AIPC overexpressing VEGFR/EGFR pathways.

Combination of docetaxel and vandetanib in docetaxel-sensitive or resistant PC3 cell line.

OBJECTIVE: To examine the anti-proliferative effect of the combination of docetaxel, the cornerstone of modern chemotherapy for prostate cancer, and vandetanib, a potent inhibitor of VEGFR-2 tyrosine kinase, applied to the representative hormone-refractory human prostate cancer cell line PC3. The aim is to analyze if a supra-additive/synergic effect of the combined treatment on cell viability exists and to understand the molecular key-factors involved. We first hypothesized an effect of vandetanib in modulation the function of MDR1, leading to a longer retention of docetaxel inside the cell. It may also be possible that vandetanib could modulate the docetaxel-induced changes in expression of prosurvival and proapoptotic proteins, leading to a positive balance forward cell death. MATERIALS AND METHODS: We used PC3 cells either wild type (PC3wt) or with acquired resistance to docetaxel (PC3R), characterized by a higher expression of MDR1. We studied both mRNA and protein, the expression of EGF and VEGF receptors at a basal level and after each treatment, as well as the expression of cell cycle and apoptosis related genes. RESULTS: Cell proliferation data suggested a supra-additive cytotoxic effect of the combination of docetaxel plus vandetanib, when given together or with the sequence vandetanib followed by docetaxel. We did not observe any effect of vandetanib on MDR1, in the PC3R cell lines, characterized by a higher pump expression than PC3wt. On the other side, we defined a number of key factors involved in the pro- and anti-survival balance, which regulation, by single drugs and/or by combined treatment, could explain the effect on cell cytotoxicity; also where there are apparently contradictory results. CONCLUSIONS: Our data suggest that combined treatment with vandetanib and docetaxel alters the balance of proapoptotic and prosurvival proteins, ultimately leading to potentiation of docetaxel-induced apoptosis in human prostate cancer cells in vitro, irrespective of cells being sensitive or resistant to docetaxel.

Impact of age on acute toxicity induced by bio- or chemo-radiotherapy in patients with head and neck cancer.

INTRODUCTION: The purpose of this study is to retrospectively analyze acute toxicity encountered in young and old patients treated with chemo-radiation or bio-radiation at the S. Croce General Hospital between 1997 and 2008, in daily clinical practice. MATERIAL AND METHODS: Three hundred and seventeen patients were allocated into two groups according to age (cut-off 65 years). The two groups were compared in terms of treatment related toxicities, treatment activity and efficacy. Epidermal Growth factor receptor (EGFr), Human papillomavirus (HPV) and p53 status were also considered. RESULTS: As expected, overall survival was significantly worse in elderly patients (p=0.005), but response rate, including complete response rate, was similar between the two age groups, as were most of the side effects analyzed. However, infections (p=0.011) and in particular pneumonias (p=0.002) were significantly more represented in elderly patients. CONCLUSION: Elderly patients treated with chemo-radiation or bio-radiation in our centre had a higher risk of infection and in particular, pneumonia. These data suggest a more careful follow-up, but age alone does not justify their exclusion from treatment.

MDM2 309 polymorphism predicts outcome in platinum-treated locally advanced head and neck cancer.

Chemo-radiotherapy (CRT) with cisplatin-based regimens is curative in a subset of patients with locally advanced (stage III and IV) squamous carcinomas of the head and neck (LAHNSCC), but causes considerable toxicity. To seek predictive biomarkers, we analysed single nucleotide polymorphisms (SNPs) in the p53 and MDM2 genes in LAHNSCC patients treated with cisplatin-based CRT. We analysed germ-line p53 72 Arg/Pro (R/P) and MDM2 309 SNPs and somatic p53 mutational status in 140 LAHNSCC and determined their utility as predictive biomarkers. In cases with wild-type p53, overall survival (OS) was longest in 72RR (median OS=60.8 months) and less favourable in 72PP (median OS=6.7 months, p<0.0001). OS in individuals with 72RP was intermediate between 72RR and 72PP, while in patients with missense p53 mutations, median OS did not reach statistical significance. Median OS was significantly shorter in patients with MDM2 309 SNP genotypes GG or GT, compared to TT (15 vs. 86 months; p<0.0001). The predictive effect of the G allele was maintained independent of age, gender, stage, primary site, nodal status, performance status, EGFR grade, HPV status, p53 mutation and p53 SNP (HR for death 3.241; 95% CI 1.90-5.52, p<0.001). The predictive utility of the MDM2 germ-line 309 SNP, which can be easily determined from peripheral blood, implies that it may be of value in the objective selection of patients for radical CRT. In contrast, the predictive utility of the 72 Arg/Pro SNP in p53 requires mutational analysis of p53, limiting its routine clinical use.