PDGFRα up-regulation mediated by sonic hedgehog pathway activation leads to BRAF inhibitor resistance in melanoma cells with BRAF mutation

Control of BRAF(V600E) metastatic melanoma by BRAF inhibitor (BRAF-I) is limited by intrinsic and acquired resistance. Growth factor receptor up-regulation is among the mechanisms underlying BRAF-I resistance of melanoma cells. Here we demonstrate for the first time that PDGFRα up-regulation causes BRAF-I resistance. PDGFRα inhibition by PDGFRα-specific short hairpin (sh)RNA and by PDGFRα inhibitors restores and increases melanoma cells'' sensitivity to BRAF-I in vitro and in vivo. This effect reflects the inhibition of ERK and AKT activation which is associated with BRAF-I resistance of melanoma cells. PDGFRα up-regulation is mediated by Sonic Hedgehog Homolog (Shh) pathway activation which is induced by BRAF-I treatment. Similarly to PDGFRα inhibition, Shh inhibition by LDE225 restores and increases melanoma cells'' sensitivity to BRAF-I. These effects are mediated by PDGFRα down-regulation and by ERK and AKT inhibition. The clinical relevance of these data is indicated by the association of PDGFRα up-regulation in melanoma matched biopsies of BRAF-I +/- MEK inhibitor treated patients with shorter time to disease progression and less tumor regression. These findings suggest that monitoring patients for early PDGFRα up-regulation will facilitate the identification of those who may benefit from the treatment with BRAF-I in combination with clinically approved PDGFRα or Shh inhibitors.     

MAPK pathway in melanoma part II—secondary and adaptive resistance mechanisms to BRAF inhibition

BRAF mutation can be identified in about 45% of the patients with metastatic melanoma. In these patients, BRAF and MEK inhibitors are able to induce rapid responses and to prolong survival. However, a significant percentage of patients will develop resistance to targeted therapy and will have progressive disease.MAPK pathway is the most important pathway involved in BRAF/MEK inhibition resistance, particularly MAPK pathway reactivation.Resistance mechanisms can be classified as 1) primary or intrinsic characterised by no response to therapy, 2) secondary or acquired with MAPK pathway reactivation after a time of tumour regression and 3) as adaptive with initial response and early resistance.BRAF inhibition also alters the immune response. Several publications have described immune effects of BRAF inhibition in melanoma tumours, showing that combining targeted and immunotherapy can improve response, despite a possible cross-resistance.Here, we continue the review on resistance mechanisms to BRAF/MEK inhibition and focus on the secondary and adaptive mechanisms.     

Prospective evaluation of follow-up in melanoma patients in Germany – Results of a multicentre and longitudinal study

BackgroundPatient numbers requiring long-term melanoma surveillance are constantly rising. Surveillance is costly and guideline recommendations vary substantially.MethodsIn this German nationwide study, information on surveillance and treatment of patients diagnosed with melanoma and melanoma in situ (MMis) between April and June 2008 was prospectively collected over four years. Additionally, patient self-report questionnaires were evaluated to assess anxiety, depression, health-related quality of life, socio-demographic information and use of disease specific health information sources at year 4 after primary diagnosis.ResultsComplete data was available for 668 patients from 67 centres, of whom 96.0% were in regular melanoma surveillance. In year 3–4 of surveillance, only 55.6% of locoregionary metastases were detected during surveillance visits. Only 33.3% were self-detected by the patient even though 69.4% were documented as being clinically visible or palpable. Costs of 4 year surveillance of 550 patients without tumour recurrence (stage I–IIC and MMis) accumulated to 228,155.75 €. Guideline-adherence for follow-up frequency, lymph node ultrasound, S100 serum level tests and diagnostic imaging recommendations was approximately 60% in year 3–4 of surveillance. Multivariate regression analysis showed that certain patient/tumour characteristics and regional differences were significantly associated with guideline deviations. The percentage of patients who exceeded published cut-off scores indicating clinically relevant symptoms of anxiety and depression were significantly increased. Patients frequently reported lack of psychosocial support and education but ascribed great importance to these.ConclusionsWe recommend further reduction of melanoma follow-up in low-risk melanoma patients and improvement of psycho-social support and patient education for all melanoma patients.     

Evaluation of patients’ engagement in radiation therapy safety

PurposeTreatment safety has become a priority in health policies after several incidents occurred around the world in radiation oncology departments. The aim of this study was to analyse the patients’ contribution in that field and to understand which actions empower the patient in that regard.MethodsSeveral methods were used in a general hospital and in a comprehensive cancer centre to analyse the activities of the radiation therapists and the patients and the interactions between them: treatment session observations, semidirective interviews with radiation therapists and patients, self and alloconfrontation with radiation therapists and explanatory interviews with patients.ResultsCooperation of the patients in treatment safety acts as an additional step that contributes to safer treatments. Radiation therapy sessions are a creative opportunity for the patient to observe, learn and analyse what is happening. Changes between treatment sessions are a source of anxiety for the patients. This study highlights the factors that favour the patients’ participation. A trusting relationship and support from the health professionals can be leveraged in that manner.ConclusionThere is a common will shared between the patients and the health professionals towards better treatment safety. The cooperation is still not well-known and underused. This empowerment of the patient cannot be mandatory but should be promoted and developed.     

A phase I�CII study of the histone deacetylase inhibitor valproic acid plus chemoimmunotherapy in patients with advanced melanoma

We explored in a phase I/II clinical trial the combination of valproic acid (VPA), a clinically available histone deacetylase inhibitor, with standard chemoimmunotherapy in patients with advanced melanoma, to evaluate its clinical activity, to correlate the clinical response with the biological activity of VPA and to assess toxicity. Patients were treated initially with VPA alone for 6 weeks. The inhibition of the target in non-tumour peripheral blood cells (taken as a potential surrogate marker) was measured periodically, and valproate dosing adjusted with the attempt to reach a measurable inhibition. After the treatment with valproate alone, dacarbazine plus interferon-α was started in combination with valproate. Twenty-nine eligible patients started taking valproate and 18 received chemoimmunotherapy and are assessable for response. We observed one complete response, two partial remissions and three disease stabilisations lasting longer than 24 weeks. With the higher valproate dosages needed to reach a measurable inhibition of the target, we observed an increase of side effects in those patients who received chemoimmunotherapy. The combination of VPA and chemoimmunotherapy did not produce results overtly superior to standard therapy in patients with advanced melanoma and toxicity was not negligible, casting some doubts on the clinical use of VPA in this setting (at least in the administration schedule adopted).     

Non‐invasive sensitive detection of KRAS and BRAF mutation in circulating tumor cells of colorectal cancer patients

Characterization of genetic alterations in tumor biopsies serves as useful biomarkers in prognosis and treatment management. Circulating tumor cells (CTCs) obtained non‐invasively from peripheral blood could serve as a tumor proxy. Using a label‐free CTC enrichment strategy that we have established, we aimed to develop sensitive assays for qualitative assessment of tumor genotype in patients. Blood consecutively obtained from 44 patients with local and advanced colorectal cancer and 18 healthy donors were enriched for CTCs using a size‐based microsieve technology. To screen for CTC mutations, we established high‐resolution melt (HRM) and allele‐specific PCR (ASPCR) KRAS‐codon 12/13‐ and BRAF‐codon 600‐ specific assays, and compared the performance with pyrosequencing and Sanger sequencing. For each patient, the resulting CTC genotypes were compared with matched tumor and normal tissues. Both HRM and ASPCR could detect as low as 1.25% KRAS‐ or BRAF‐mutant alleles. HRM detected 14/44 (31.8%) patients with KRAS mutation in CTCs and 5/44 (11.3%) patients having BRAF mutation in CTCs. ASPCR detected KRAS and BRAF mutations in CTCs of 10/44 (22.7%) and 1/44 (2.3%) patients respectively. There was an increased detection of mutation in blood using these two methods. Comparing tumor tissues and CTCs mutation status using HRM, we observed 84.1% concordance in KRAS genotype (p = 0.000129, Fishers'' exact test; OR = 38.7, 95% CI = 4.05–369) and 90.9% (p = 0.174) concordance in BRAF genotype. Our results demonstrate that CTC enrichment, coupled with sensitive mutation detection methods, may allow rapid, sensitive and non‐invasive assessment of tumor genotype.     

What is the impact of antithrombotic therapy and risk factors on the frequency of thrombovascular events in patients with metastatic breast cancer receiving epoetin beta?

Purpose, patients and methodsThis retrospective analysis of the BRAVE study evaluated the impact of baseline risk factors and antithrombotic therapy on the risk of thrombovascular events (TVEs) in patients receiving epoetin compared to patients not receiving epoetin.ResultsBaseline risk factors have a significant impact on TVE risk under epoetin therapy. More than 2 risk factors increased the risk of TVEs in patients receiving epoetin (hazard ratio [HR] 2.89, confidence interval [CI] 1.04–8.02, p value [p] = 0.04). In patients on epoetin without antithrombotic therapy, the risk for TVEs was higher (HR 4.11, CI 1.37–12.4, p = 0.01) compared to those who received antithrombotics (HR 1.37, CI 0.59–3.18, p = 0.45).ConclusionsOur analysis has identified several risk factors which may impact the risk of TVEs under epoetin therapy. These data suggest that antithrombotic therapy may have the potential to reduce the risk of TVEs under epoetin therapy. These findings are hypothesis-generating and need to be confirmed in a prospective, randomised study.     

BEAC or BEAM for high-dose therapy in patients with non-Hodgkin's lymphoma? A single centre analysis on toxicity and efficacy

High-dose chemotherapy supported by autologous stem cell transplantation is widely used in patients with non-Hodgkin's lymphoma (NHL). Limited data is available on the comparative toxicity and efficacy of various high-dose regimens applied in NHL. We therefore analysed regimen-related toxicity and outcome in 71 consecutive NHL patients who received either BEAC (N = 36) or BEAM (N = 35) supported by peripheral blood progenitor cell infusion plus granulocyte colony-stimulating factor. The patients who received BEAM had significantly more often WHO grade > 2 mucositis (63 vs. 28%, P = 0.009) and diarrhoea grade >2 (29 vs. 8%, P = 0.062). Septicaemia also tended to be more frequent and the peak CRP value was higher in the BEAM group (140 vs. 113 mg/l, P = 0.034). Transplant-related mortality (< 100 d) was 3 and 9% in the BEAC and BEAM groups, respectively. No significant differences were observed in overall survival or progression free survival between these two groups. While BEAC and BEAM appears to have equal antitumour efficacy in patients with NHL, BEAM seems to be more toxic to the gastrointestinal tract. However, randomised studies are needed for more definitive conclusions on the relative merits of various high-dose regimens in patients with NHL.     

Is the toxicity of adjuvant aromatase inhibitor therapy underestimated? Complementary information from patient-reported outcomes (PROs)

This retrospective study investigates the correlation of intra-individual HER2 status between primary breast cancers and corresponding recurrences in a population derived cohort. The REMARK criteria were used as reference. In 151 breast cancer patients, primary tumors were analyzed for HER2 status on histopathology sections using immunohistochemistry (IHC) confirmed by fluorescence in situ hybridization (FISH) for IHC 2+ and 3+. Recurrences (loco regional and distant) were investigated by aspiration cytology, using HER2 immunocytochemistry (ICC) or FISH (ICC in 84 patients and FISH in 102 patients). In the 151 patients, sites of recurrence were bone/bone marrow 30%, liver 16%, local recurrence 18%, lung/pleura 10%, axillary lymph nodes 9%, skin (non-local) 7%, supra clavicular lymph nodes 5%, and other sites 7%. In 15 patients (10%) HER2 status changed, 7 of 108 patients (6%) from HER2 negative to HER2 positive and 8 of 43 (19%) from HER2 positive to HER2 negative. Intra-patient agreement in HER2 status was 76% (95% CI 64–87%), and the disagreement was 10% (95% CI 5–15%). The multivariable Cox analysis showed a significantly increased risk of dying in the patient group with changed HER2 status compared to patients with concordant positive HER2 status. Overall survival HR is 5.47 (95% CI 2.01–14.91) and survival from relapse HR is 3.22 (95% CI 1.18–8.77). The unstable status for HER2 in breast cancer is clinically significant and should motivate more frequent testing of recurrences.     

Bladder cancer therapy without toxicity—A dose-escalation study of alpha1-oleate

Potent chemotherapeutic agents are required to counteract the aggressive behavior of cancer cells and patients often experience severe side effects, due to tissue toxicity. Our study addresses if a better balance between efficacy and toxicity can be attained using the tumoricidal complex alpha1-oleate, formed by a synthetic, alpha-helical peptide comprising the N-terminal 39 amino acids of alpha-lactalbumin and the fatty acid oleic acid. Bladder cancer was established, by intravesical instillation of MB49 cells on day 0 and the treatment group received five instillations of alpha1-oleate (1.7-17 mM) on days 3 to 11. A dose-dependent reduction in tumor size, bladder size and bladder weight was recorded in the alpha1-oleate treated group, compared to sham-treated mice. Tumor markers Ki-67, Cyclin D1 and VEGF were inhibited in a dose-dependent manner, as was the expression of cancer-related genes. Remarkably, toxicity for healthy tissue was not detected in alpha1-oleate-treated, tumor-bearing mice or healthy mice or rabbits, challenged with increasing doses of the active complex. The results define a dose-dependent therapeutic effect of alpha1-oleate in a murine bladder cancer model.     

Low- and high-risk malignant melanoma—I. Evaluation of clinical and histological prognosticators in 585 cases

In 585 cases with primary cutaneous stage I malignant melanoma (294 disease-free for at least 5 yr, 291 with later metastases) prognostic parameters were examined. The most effective proved to be tumor thickness and mitotic activity, particularly when combined as a prognostic index. Furthermore, vascular invasion, ulceration in thick tumors (thickness ⩾ 3.0 mm), severe cellular atypia, the small, lymphocytic-like cell type and the absence of an inflammatory reaction were closely associated with a high rate of metastatic cases. Less relevant prognostic factors were the level of invasion, sex, site, tumor breadth, clinical diameters and infiltrative growth. Tumor type, age, duration and an adjacent nevocellular nevus were not significantly associated with the occurrence of later metastases. Furthermore, the growth-type (exo- or endophytic) did not have a bearing on the prognosis.