Biological, diagnostic and therapeutic relevance of the MET receptor signaling in head and neck cancer.

Head and neck cancer constitutes the 6th most common malignancy worldwide and affects the crucial anatomical structures and physiological functions of the upper aerodigestive tract. Classical therapeutic strategies such as surgery and radiotherapy carry substantial toxicity and functional impairment. Moreover, the loco-regional control rates as well as overall survival still need to be improved in subgroups of patients. The scatter-factor/hepatocyte growth factor receptor tyrosine kinase MET is an established effector in the promotion, maintenance and progression of malignant transformation in a wide range of human malignancies, and has been gaining considerable interest in head and neck cancer over the last 15 years. Aberrant MET activation due to overexpression, mutations, tumor-stroma paracrine loops, and cooperative/redundant signaling has been shown to play prominent roles in epithelial-to-mesenchymal transition, angiogenesis, and responses to anti-cancer therapeutic modalities. Accumulating preclinical and translational evidence highly supports the increasing interest of MET as a biomarker for lymph node and distant metastases, as well as a potential marker of stratification for responses to ionizing radiation. The relevance of MET as a therapeutic molecular target in head and neck cancer described in preclinical studies remains largely under-evaluated in clinical trials, and therefore inconclusive. Also in the context of anti-cancer targeted therapy, a large body of preclinical data suggests a central role for MET in treatment resistance towards multiple therapeutic modalities in malignancies of the head and neck region. These findings, as well as the potential use of combination therapies including MET inhibitors in these tumors, need to be further explored.

Multimodal treatment for high-risk prostate cancer with high-dose intensity-modulated radiation therapy preceded or not by radical prostatectomy, concurrent intensified-dose docetaxel and long-term androgen deprivation therapy: results of a prospective phase II trial.

BACKGROUND The optimal management of high-risk prostate cancer remains uncertain. In this study we assessed the safety and efficacy of a novel multimodal treatment paradigm for high-risk prostate cancer. METHODS This was a prospective phase II trial including 35 patients with newly diagnosed high-risk localized or locally advanced prostate cancer treated with high-dose intensity-modulated radiation therapy preceded or not by radical prostatectomy, concurrent intensified-dose docetaxel-based chemotherapy and long-term androgen deprivation therapy. Primary endpoint was acute and late toxicity evaluated with the Common Terminology Criteria for Adverse Events version 3.0. Secondary endpoint was biochemical and clinical recurrence-free survival explored with the Kaplan-Meier method. RESULTS Acute gastro-intestinal and genito-urinary toxicity was grade 2 in 23% and 20% of patients, and grade 3 in 9% and 3% of patients, respectively. Acute blood/bone marrow toxicity was grade 2 in 20% of patients. No acute grade ≥ 4 toxicity was observed. Late gastro-intestinal and genito-urinary toxicity was grade 2 in 9% of patients each. No late grade ≥ 3 toxicity was observed. Median follow-up was 63 months (interquartile range 31-79). Actuarial 5-year biochemical and clinical recurrence-free survival rate was 55% (95% confidence interval, 35-75%) and 70% (95% confidence interval, 52-88%), respectively. CONCLUSIONS In our phase II trial testing a novel multimodal treatment paradigm for high-risk prostate cancer, toxicity was acceptably low and mid-term oncological outcome was good. This treatment paradigm, thus, may warrant further evaluation in phase III randomized trials.

Synthesis and cellular characterization of novel isoxazolo- and thiazolohydrazinylidene-chroman-2,4-diones on cancer and non-cancer cell growth and death

Coumarins are extensively studied anticoagulants that exert additional effects such as anticancerogenic and even anti-inflammatory. In order to find new drugs with anticancer activities, we report here the synthesis and the structural analysis of new coumarin derivatives which combine the coumarin core and five member heterocycles in hydrazinylidene-chroman-2,4-diones. The derivatives were prepared by derivatization of the appropriate heterocyclic amines which were used as electrophiles to attack the coumarin ring. The structures were characterized by spectroscopic techniques including IR, NMR, 2D-NMR and MS. These derivatives were further characterized especially in terms of a potential cytotoxic and apoptogenic effect in several cancer cell lines including the breast and prostate cancer cell lines MCF-7, MDA-MB-231, PC-3, LNCaP, and the monocytic leukemia cell line U937. Cell viability was determined after 48 h and 72 h of treatment with the novel compounds by MTT assay and the 50% inhibitory concentrations (EC50 values) were determined. Out of the 8 novel compounds screened for reduced cell viability, 4c, 4d and 4e were found to be the most promising and effective ones having EC50 values that were several fold reduced when compared to the reference substance 4-hydroxycoumarin. However, the effects were cancer cell line dependent. The breast cancer MDA-MB-231 cells, the prostate cancer LNCaP cells, and U937 cells were most sensitive, MCF-7 cells were less sensitive, and PC-3 cells were more resistant. Reduced cell viability was accompanied by increased apoptosis as shown by PARP-1 cleavage and reduced activity of the survival protein kinase Akt. In summary, this study has identified three novel coumarin derivatives that in comparison to 4-hydroxycoumarin have a higher efficiency to reduce cancer cell viability and trigger apoptosis and therefore may represent interesting novel drug candidates

NOTCH ligands JAG1 and JAG2 as critical pro-survival factors in childhood medulloblastoma.

Medulloblastoma (MB), the most common pediatric malignant brain cancer, typically arises as pathological result of deregulated developmental pathways, including the NOTCH signaling cascade. Unlike the evidence supporting a role for NOTCH receptors in MB development, the pathological functions of NOTCH ligands remain largely unexplored. By examining the expression in large cohorts of MB primary tumors, and in established in vitro MB models, this research study demonstrates that MB cells bear abnormal levels of distinct NOTCH ligands. We explored the potential association between NOTCH ligands and the clinical outcome of MB patients, and investigated the rational of inhibiting NOTCH signaling by targeting specific ligands to ultimately provide therapeutic benefits in MB. The research revealed a significant over-expression of ligand JAG1 in the vast majority of MBs, and proved that JAG1 mediates pro-proliferative signals via activation of NOTCH2 receptor and induction of HES1 expression, thus representing an attractive therapeutic target. Furthermore, we could identify a clinically relevant association between ligand JAG2 and the oncogene MYC, specific for MYC-driven Group 3 MB cases. We describe for the first time a mechanistic link between the oncogene MYC and NOTCH pathway in MB, by identifying JAG2 as MYC target, and by showing that MB cells acquire induced expression of JAG2 through MYC-induced transcriptional activation. Finally, the positive correlation of MYC and JAG2 also with aggressive anaplastic tumors and highly metastatic MB stages suggested that high JAG2 expression may be useful as additional marker to identify aggressive MBs.

Targeting class IA PI3K isoforms selectively impairs cell growth, survival, and migration in glioblastoma.

The phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway is frequently activated in human cancer and plays a crucial role in glioblastoma biology. We were interested in gaining further insight into the potential of targeting PI3K isoforms as a novel anti-tumor approach in glioblastoma. Consistent expression of the PI3K catalytic isoform PI3K p110α was detected in a panel of glioblastoma patient samples. In contrast, PI3K p110β expression was only rarely detected in glioblastoma patient samples. The expression of a module comprising the epidermal growth factor receptor (EGFR)/PI3K p110α/phosphorylated ribosomal S6 protein (p-S6) was correlated with shorter patient survival. Inhibition of PI3K p110α activity impaired the anchorage-dependent growth of glioblastoma cells and induced tumor regression in vivo. Inhibition of PI3K p110α or PI3K p110β also led to impaired anchorage-independent growth, a decreased migratory capacity of glioblastoma cells, and reduced the activation of the Akt/mTOR pathway. These effects were selective, because targeting of PI3K p110δ did not result in a comparable impairment of glioblastoma tumorigenic properties. Together, our data reveal that drugs targeting PI3K p110α can reduce growth in a subset of glioblastoma tumors characterized by the expression of EGFR/PI3K p110α/p-S6.

TWIST1 and TWIST2 promoter methylation and protein expression in tumor stroma influence the epithelial-mesenchymal transition-like tumor budding phenotype in colorectal cancer.

Tumor budding in colorectal cancer is likened to an epithelial-mesenchymal transition (EMT) characterized predominantly by loss of E-cadherin and up-regulation of E-cadherin repressors like TWIST1 and TWIST2. Here we investigate a possible epigenetic link between TWIST proteins and the tumor budding phenotype. TWIST1 and TWIST2 promoter methylation and protein expression were investigated in six cell lines and further correlated with tumor budding in patient cohort 1 (n = 185). Patient cohort 2 (n = 112) was used to assess prognostic effects. Laser capture microdissection (LCM) of tumor epithelium and stroma from low- and high-grade budding cancers was performed. In colorectal cancers, TWIST1 and TWIST2 expression was essentially restricted to stromal cells. LCM results of a high-grade budding case show positive TWIST1 and TWIST2 stroma and no methylation, while the low-grade budding case was characterized by negative stroma and strong hypermethylation. TWIST1 stromal cell staining was associated with adverse features like more advanced pT (p = 0.0044), lymph node metastasis (p = 0.0301), lymphatic vessel invasion (p = 0.0373), perineural invasion (p = 0.0109) and worse overall survival time (p = 0.0226). Stromal cells may influence tumor budding in colorectal cancers through expression of TWIST1. Hypermethylation of the tumor stroma may represent an alternative mechanism for regulation of TWIST1.

Investigation of IL-23 (p19, p40) and IL-23R identifies nuclear expression of IL-23 p19 as a favorable prognostic factor in colorectal cancer: a retrospective multicenter study of 675 patients.

IL-23 is a heterodimeric cytokine involved in inflammatory diseases; its role in cancer progression is controversial. Here we analyse the expression of IL-23 subunits (p40 and p19) and IL-23R in colorectal cancer with regard to disease progression, clinical-pathological and molecular aspects. Immunohistochemistry for IL-23p19, IL-23p40, IL-23R and CD8 was performed on a multi-punch tissue microarray of 195 colorectal cancers (cohort 1), matched normal tissue, adenoma and lymph node metastases. Results were compared with clinical-pathological features and CD8+ T-cell counts, then validated on two patient cohorts (cohort 2: n=341, cohort 3: n=139). Cytoplasmic/membranous expression of IL-23 (p19 and p40 subunits) and IL-23R, respectively were over-expressed in carcinomas versus adenomas and normal tissues (p<0.0001) but were reduced in lymph node metastases (p<0.0001). Nuclear IL-23p19 expression was observed in 23.1% and was associated with early TNM stage (p=0.0186), absence of venous (p=0.0124) and lymphatic invasion (p=0.01493), favorable survival (p=0.014) and absence of distant metastasis (p=0.0146; specificity: 100%). This unexpected cellular localization was confirmed by cell fractionation. The beneficial effect of nuclear IL-23p19 was restricted to tumours with CD8+ high counts. Results were validated on Cohorts 2/3. This multicenter study underlines the possible CD8(+)--dependency and beneficial effect of nuclear IL-23p19 on overall patient survival.

Resistance prediction in AML: analysis of 4601 patients from MRC/NCRI, HOVON/SAKK, SWOG and MD Anderson Cancer Center

Therapeutic resistance remains the principal problem in acute myeloid leukemia (AML). We used area under receiver-operating characteristic curves (AUCs) to quantify our ability to predict therapeutic resistance in individual patients, where AUC=1.0 denotes perfect prediction and AUC=0.5 denotes a coin flip, using data from 4601 patients with newly diagnosed AML given induction therapy with 3+7 or more intense standard regimens in UK Medical Research Council/National Cancer Research Institute, Dutch–Belgian Cooperative Trial Group for Hematology/Oncology/Swiss Group for Clinical Cancer Research, US cooperative group SWOG and MD Anderson Cancer Center studies. Age, performance status, white blood cell count, secondary disease, cytogenetic risk and FLT3-ITD/NPM1 mutation status were each independently associated with failure to achieve complete remission despite no early death (‘primary refractoriness’). However, the AUC of a bootstrap-corrected multivariable model predicting this outcome was only 0.78, indicating only fair predictive ability. Removal of FLT3-ITD and NPM1 information only slightly decreased the AUC (0.76). Prediction of resistance, defined as primary refractoriness or short relapse-free survival, was even more difficult. Our limited ability to forecast resistance based on routinely available pretreatment covariates provides a rationale for continued randomization between standard and new therapies and supports further examination of genetic and posttreatment data to optimize resistance prediction in AML.

Vision impairment at diagnosis of cancer - choroidal metastasis

Choroidal metastasis represents the most common type of intraocular malignancy and preferably involves the posterior uveal tract. Breast and lung cancer - known or so far undiagnosed - are most frequently identified as the underlying tumor disease. The majority of patients diagnosed with uveal metastasis have additional metastatic manifestations elsewhere, so re-staging before treatment is recommended. The importance of a multidisciplinary approach is obvious. Early diagnosis and timely initiation of treatment are mandatory in case of vision-threatening situations. External beam radiation remains the therapy of choice. Overall survival of patients with uveal metastasis is limited, averaging six to twelve months. The other eye is frequently enough affected as well, justifying regular ophthalmologic follow-up during the further course of the disease.

Tyrosine kinase receptor B (TrkB) expression in colorectal cancers highlights anoikis resistance as a survival mechanism of tumour budding cells.

AIMS Tumour buds in colorectal cancer represent an aggressive subgroup of non-proliferating and non-apoptotic tumour cells. We hypothesize that the survival of tumour buds is dependent upon anoikis resistance. The role of tyrosine kinase receptor B (TrkB), a promoter of epithelial-mesenchymal transition and anoikis resistance, in facilitating budding was investigated. METHODS AND RESULTS Tyrosine kinase receptor B immunohistochemistry was performed on a multiple-punch tissue microarray of 211 colorectal cancer resections. Membranous/cytoplasmic and nuclear expression was evaluated in tumour and buds. Tumour budding was assessed on corresponding whole tissue slides. Relationship to Ki-67 and caspase-3 was investigated. Analysis of Kirsten Ras (KRAS), proto-oncogene B-RAF (BRAF) and cytosine-phosphate-guanosine island methylator phenotype (CIMP) was performed. Membranous/cytoplasmic and nuclear TrkB were strongly, inversely correlated (P < 0.0001; r = -0.41). Membranous/cytoplasmic TrkB was overexpressed in buds compared to the main tumour body (P < 0.0001), associated with larger tumours (P = 0.0236), high-grade budding (P = 0.0011) and KRAS mutation (P = 0.0008). Nuclear TrkB was absent in buds (P <0.0001) and in high-grade budding cancers (P =0.0073). Among patients with membranous/cytoplasmic TrkB-positive buds, high tumour membranous/cytoplasmic TrkB expression was a significant, independent adverse prognostic factor [P = 0.033; 1.79, 95% confidence interval (CI) 1.05-3.05]. Inverse correlations between membranous/cytoplasmic TrkB and Ki-67 (r = -0.41; P < 0.0001) and caspase-3 (r =-0.19; P < 0.05) were observed. CONCLUSIONS Membranous/cytoplasmic TrkB may promote an epithelial-mesenchymal transition (EMT)-like phenotype with high-grade budding and maintain viability of buds themselves.

The tumor border configuration of colorectal cancer as a histomorphological prognostic indicator.

Histomorphological features of colorectal cancers (CRC) represent valuable prognostic indicators for clinical decision making. The invasive margin is a central feature for prognostication shaped by the complex processes governing tumor-host interaction. Assessment of the tumor border can be performed on standard paraffin sections and shows promise for integration into the diagnostic routine of gastrointestinal pathology. In aggressive CRC, an extensive dissection of host tissue is seen with loss of a clear tumor-host interface. This pattern, termed "infiltrative tumor border configuration" has been consistently associated with poor survival outcome and early disease recurrence of CRC-patients. In addition, infiltrative tumor growth is frequently associated with presence of adverse clinicopathological features and molecular alterations related to aggressive tumor behavior including BRAFV600 mutation. In contrast, a well-demarcated "pushing" tumor border is seen frequently in CRC-cases with low risk for nodal and distant metastasis. A pushing border is a feature frequently associated with mismatch-repair deficiency and can be used to identify patients for molecular testing. Consequently, assessment of the tumor border configuration as an additional prognostic factor is recommended by the AJCC/UICC to aid the TNM-classification. To promote the assessment of the tumor border configuration in standard practice, consensus criteria on the defining features and method of assessment need to be developed further and tested for inter-observer reproducibility. The development of a standardized quantitative scoring system may lay the basis for verification of the prognostic associations of the tumor growth pattern in multivariate analyses and clinical trials. This article provides a comprehensive review of the diagnostic features, clinicopathological associations, and molecular alterations associated with the tumor border configuration in early stage and advanced CRC.

Bcl-2 predicts response to neoadjuvant chemotherapy and is overexpressed in lymph node metastases of urothelial cancer of the bladder

PURPOSE To assess whether Bcl-2, an inhibitor of the apoptotic cascade, can predict response to neoadjuvant chemotherapy in patients with urothelial cancer of the bladder (UCB). METHODS Bcl-2 expression was analyzed in 2 different tissue microarrays (TMAs). One TMA was constructed of primary tumors and their corresponding lymph node (LN) metastases from 152 patients with chemotherapy-naive UCB treated by cystectomy and pelvic lymphadenectomy (chemotherapy-naive TMA cohort). The other TMA was constructed of tumor samples obtained from 55 patients with UCB before neoadjuvant chemotherapy (transurethral resection of the bladder cancer) and after cystectomy with pelvic lymphadenectomy (residual primary tumor [ypT+], n = 38); residual LN metastases [ypN+], n = 24) (prechemotherapy/postchemotherapy TMA cohort). Bcl-2 overexpression was defined as 10% or more cancer cells showing cytoplasmic immunoreactivity. RESULTS In both TMA cohorts, Bcl-2 overexpression was significantly (P<0.05) more frequent in LN metastases than in primary tumors (chemotherapy-naive TMA group: 18/148 [12%] in primary tumors vs. 39/143 [27%] in metastases; postchemotherapy TMA: ypT+7/35 [20%] vs. ypN+11/19 [58%]). In the neoadjuvant setting, patients with Bcl-2 overexpression in transurethral resection of the bladder cancer specimens showed significantly (P = 0.04) higher ypT stages and less regression in their cystectomy specimens than did the control group, and only one-eighth (13%) had complete tumor regression (ypT0 ypN0). In survival analyses, only histopathological parameters added significant prognostic information. CONCLUSIONS Bcl-2 overexpression in chemotherapy-naive primary bladder cancer is related to poor chemotherapy response and might help to select likely nonresponders.

The molecular signature of the stroma response in prostate cancer-induced osteoblastic bone metastasis highlights expansion of hematopoietic and prostate epithelial stem cell niches

The reciprocal interaction between cancer cells and the tissue-specific stroma is critical for primary and metastatic tumor growth progression. Prostate cancer cells colonize preferentially bone (osteotropism), where they alter the physiological balance between osteoblast-mediated bone formation and osteoclast-mediated bone resorption, and elicit prevalently an osteoblastic response (osteoinduction). The molecular cues provided by osteoblasts for the survival and growth of bone metastatic prostate cancer cells are largely unknown. We exploited the sufficient divergence between human and mouse RNA sequences together with redefinition of highly species-specific gene arrays by computer-aided and experimental exclusion of cross-hybridizing oligonucleotide probes. This strategy allowed the dissection of the stroma (mouse) from the cancer cell (human) transcriptome in bone metastasis xenograft models of human osteoinductive prostate cancer cells (VCaP and C4-2B). As a result, we generated the osteoblastic bone metastasis-associated stroma transcriptome (OB-BMST). Subtraction of genes shared by inflammation, wound healing and desmoplastic responses, and by the tissue type-independent stroma responses to a variety of non-osteotropic and osteotropic primary cancers generated a curated gene signature ("Core" OB-BMST) putatively representing the bone marrow/bone-specific stroma response to prostate cancer-induced, osteoblastic bone metastasis. The expression pattern of three representative Core OB-BMST genes (PTN, EPHA3 and FSCN1) seems to confirm the bone specificity of this response. A robust induction of genes involved in osteogenesis and angiogenesis dominates both the OB-BMST and Core OB-BMST. This translates in an amplification of hematopoietic and, remarkably, prostate epithelial stem cell niche components that may function as a self-reinforcing bone metastatic niche providing a growth support specific for osteoinductive prostate cancer cells. The induction of this combinatorial stem cell niche is a novel mechanism that may also explain cancer cell osteotropism and local interference with hematopoiesis (myelophthisis). Accordingly, these stem cell niche components may represent innovative therapeutic targets and/or serum biomarkers in osteoblastic bone metastasis.

Molecular markers for urothelial bladder cancer prognosis: toward implementation in clinical practice

OBJECTIVES To summarize the current status of clinicopathological and molecular markers for the prediction of recurrence or progression or both in non-muscle-invasive and survival in muscle-invasive urothelial bladder cancer, to address the reproducibility of pathology and molecular markers, and to provide directions toward implementation of molecular markers in future clinical decision making. METHODS AND MATERIALS Immunohistochemistry, gene signatures, and FGFR3-based molecular grading were used as molecular examples focussing on prognostics and issues related to robustness of pathological and molecular assays. RESULTS The role of molecular markers to predict recurrence is limited, as clinical variables are currently more important. The prediction of progression and survival using molecular markers holds considerable promise. Despite a plethora of prognostic (clinical and molecular) marker studies, reproducibility of pathology and molecular assays has been understudied, and lack of reproducibility is probably the main reason that individual prediction of disease outcome is currently not reliable. CONCLUSIONS Molecular markers are promising to predict progression and survival, but not recurrence. However, none of these are used in the daily clinical routine because of reproducibility issues. Future studies should focus on reproducibility of marker assessment and consistency of study results by incorporating scoring systems to reduce heterogeneity of reporting. This may ultimately lead to incorporation of molecular markers in clinical practice.

Treatment and outcomes of urethral recurrence of urinary bladder cancer in women after radical cystectomy and orthotopic neobladder: a series of 12 cases

INTRODUCTION The incidence, treatment, and outcome of urethral recurrence (UR) after radical cystectomy (RC) for muscle-invasive bladder cancer with orthotopic neobladder in women have rarely been addressed in the literature. PATIENTS AND METHODS A total of 12 patients (median age at recurrence: 60 years) who experienced UR after RC with an orthotopic neobladder were selected for this study from a cohort of 456 women from participating institutions. The primary clinical and pathological characteristics at RC, including the manifestation of the UR and its treatment and outcome, were reviewed. RESULTS The primary bladder tumors in the 12 patients were urothelial carcinoma in 8 patients, squamous cell carcinoma and adenocarcinoma in 1 patient each, and mixed histology in 2 patients. Three patients (25%) had lymph node-positive disease at RC. The median time from RC to the detection of UR was 8 months (range 4-55). Eight recurrences manifested with clinical symptoms and 4 were detected during follow-up or during a diagnostic work-up for clinical symptoms caused by distant metastases. Treatment modalities were surgery, chemotherapy, radiotherapy, and bacillus Calmette-Guérin urethral instillations. Nine patients died of cancer. The median survival after the diagnosis of UR was 6 months. CONCLUSIONS UR after RC with an orthotopic neobladder in females is rare. Solitary, noninvasive recurrences have a favorable prognosis when detected early. Invasive recurrences are often associated with local and distant metastases and have a poor prognosis. © 2014 S. Karger AG, Basel.