Brindha Shivalingam

Epigenome-wide DNA methylation landscape of melanoma progression to brain metastasis reveals aberrations on homeobox D cluster associated with prognosis

Melanoma brain metastasis (MBM) represents a frequent complication of cutaneous melanoma. Despite aggressive multi-modality therapy, patients with MBM often have a survival rate of <1 year. Alteration in DNA methylation is a major hallmark of tumor progression and metastasis; however, it remains largely unexplored in MBM. In this study, we generated a comprehensive DNA methylation landscape through the use of genome-wide copy number, DNA methylation and gene expression data integrative analysis of melanoma progression to MBM. A progressive genome-wide demethylation in low CpG density and an increase in methylation level of CpG islands according to melanoma progression were observed. MBM-specific partially methylated domains (PMDs) affecting key brain developmental processes were identified. Differentially methylated CpG sites between MBM and lymph node metastasis (LNM) from patients with good prognosis were identified. Among the most significantly affected genes were the HOX family members. DNA methylation of HOXD9 gene promoter affected transcript and protein expression and was significantly higher in MBM than that in early stages. A MBM-specific PMD was identified in this region. Low methylation level of this region was associated with active HOXD9 expression, open chromatin and histone modifications associated with active transcription. Demethylating agent induced HOXD9 expression in melanoma cell lines. The clinical relevance of this finding was verified in an independent large cohort of melanomas (n = 145). Patients with HOXD9 hypermethylation in LNM had poorer disease-free and overall survival. This epigenome-wide study identified novel methylated genes with functional and clinical implications for MBM patients.

Whole brain radiotherapy after local treatment of brain metastases in melanoma patients--a randomised phase III trial.

BackgroundCerebral metastases are a common cause of death in patients with melanoma. Systemic drug treatment of these metastases is rarely effective, and where possible surgical resection and/or stereotactic radiosurgery (SRS) are the preferred treatment options. Treatment with adjuvant whole brain radiotherapy (WBRT) following neurosurgery and/or SRS is controversial. Proponents of WBRT report prolongation of intracranial control with reduced neurological events and better palliation. Opponents state melanoma is radioresistant; that WBRT yields no survival benefit and may impair neurocognitive function. These opinions are based largely on studies in other tumour types in which assessment of neurocognitive function has been incomplete.Methods/DesignThis trial is an international, prospective multi-centre, open-label, phase III randomised controlled trial comparing WBRT to observation following local treatment of intracranial melanoma metastases with surgery and/or SRS. Patients aged 18 years or older with 1-3 brain metastases excised and/or stereotactically irradiated and an ECOG status of 0-2 are eligible. Patients with leptomeningeal disease, or who have had previous WBRT or localised treatment for brain metastases are ineligible. WBRT prescription is at least 30 Gy in 10 fractions commenced within 8 weeks of surgery and/or SRS. Randomisation is stratified by the number of cerebral metastases, presence or absence of extracranial disease, treatment centre, sex, radiotherapy dose and patient age. The primary endpoint is the proportion of patients with distant intracranial failure as determined by MRI assessment at 12 months. Secondary end points include: survival, quality of life, performance status and neurocognitive function.DiscussionAccrual to previous trials for patients with brain metastases has been difficult, mainly due to referral bias for or against WBRT. This trial should provide the evidence that is currently lacking in treatment decision-making for patients with melanoma brain metastases. The trial is conducted by the Australia and New Zealand Melanoma Trials Group (ANZMTG-study 01-07), and the Trans Tasman Radiation Oncology Group (TROG) but international participation is encouraged. Twelve sites are open to date with 43 patients randomised as of the 31st March 2011. The target accrual is 200 patients.Trial registrationAustralia and New Zealand Clinical Trials Register (ANZCTR): ACTRN12607000512426

Oligoastrocytomas: throwing the baby out with the bathwater?

biology, clinical trial design, and future treatment deci-sions. OA should not be ignored in the proposed new clas-sification guidelines [2].Sahm and colleagues studied 43 cases diagnosed as OA based on histology. But in 30 of these cases, immunostain-ing for IDH1 (R132H) mutation was restricted to oligo-dendroglial areas only. The astrocytic component of these tumours was re-interpreted to be reactive in nature [4], and they were reclassified as oligodendroglioma. We propose that this finding indicates that ‘true’ OAs (i.e. tumours com-posed of two distinct

Symptomatic Histologically Proven Necrosis of Brain following Stereotactic Radiation and Ipilimumab in Six Lesions in Four Melanoma Patients

Four cases previously treated with ipilimumab with a total of six histologically confirmed symptomatic lesions of RNB without any sign of active tumour following stereotactic irradiation of MBM are reported. These lesions were all originally thought to be disease recurrence. In two cases, ipilimumab was given prior to SRT; in the other two ipilimumab was given after SRT. The average time from first ipilimumab to RNB was 15 months. The average time from SRT to RNB was 11 months. The average time from first diagnosis of MBM to last follow-up was 20 months at which time three patients were still alive, one with no evidence of disease. These cases represent approximately three percent of the total cases of melanoma and ten percent of those cases treated with ipilimumab irradiated in our respective centres collectively. We report this to highlight this new problem so that others may have a high index of suspicion, allowing, if clinically warranted, aggressive surgical salvage, possibly resulting in increased survival. Further studies prospectively collecting data to understand the denominator of this problem are needed to determine whether this problem is just the result of longer survival or whether there is some synergy between these two modalities that are increasingly being used together.

Survival and prognostic factors for patients with melanoma brain metastases in the era of modern systemic therapy

Historically, the prognosis of patients with melanoma brain metastases is poor, with median overall survival (OS) of 4‐6 months. Little is known of OS in the era of modern systemic therapies and local therapy with stereotactic radiosurgery (SRS) or surgery. Patients diagnosed with melanoma brain metastases at Melanoma Institute Australia from January 2011 to December 2014 were included. OS and prognostic factors were analysed using Cox regression and Kaplan‐Meier survival analyses.355 patients were included. The median OS was 7.1 months (95% confidence interval [CI] 6.0‐8.1). Median OS differed by treatment modality: systemic therapy and SRS and/or surgery 14.9 months (95% CI 10.7‐19.0), SRS and/or surgery with or without whole brain radiotherapy (WBRT) 6.4 months (95% CI 5.4‐7.5), systemic therapy 5.4 months (95% CI 3.1‐7.7), systemic therapy and WBRT 5.2 months (95% CI 4.1‐6.4), WBRT 4.4 months (95% CI 2.4‐6.3), and best supportive care 1.8 months (95% CI 1.2‐2.3). OS for patients with melanoma brain metastases appears improved in the modern era, particularly for patients who are candidates for systemic therapy with SRS and/or surgery.

Deep sequencing of circulating exosomal microRNA allows non-invasive glioblastoma diagnosis

Exosomes are nano-sized extracellular vesicles released by many cells that contain molecules characteristic of their cell-of-origin, including microRNA. Exosomes released by glioblastoma cross the blood-brain-barrier into the peripheral circulation, and carry molecular cargo distinct to that of ‘free-circulating’ miRNA. In this pilot study, serum exosomal-microRNAs were isolated from glioblastoma (n=12) patients and analyzed using unbiased deep sequencing. Results were compared to sera from age- and gender-matched healthy controls, and to grades II-III (n=10) glioma patients. Significant differentially expressed microRNAs were identified, and the predictive power of individual and subsets of microRNAs were tested using univariate and multivariate analyses. Additional sera from glioblastoma patients (n=4) and independent sets of healthy (n=9) and non-glioma (n=10) controls were used to further test the specificity and predictive power of this unique exosomal-microRNA signature. Twenty-six microRNAs were differentially expressed in serum exosomes from glioblastoma patients’ relative to healthy controls. Random forest modeling and data partitioning selected seven miRNAs (miR-182-5p, miR-328-3p, miR-339-5p, miR-340-5p, miR-485-3p, miR-486-5p and miR-543) as the most stable for classifying glioblastoma. Strikingly, within this model, six iterations of these miRNA classifiers could distinguish glioblastoma patients from controls with perfect accuracy. The seven-miRNA panel was able to correctly classify all specimens in validation cohorts (n=23). Also identified were 23 dysregulated miRNAs in IDHMUT gliomas, a partially overlapping yet distinct signature of lower grade glioma. Serum exosomal-miRNA signatures can accurately diagnose glioblastoma preoperatively. miRNA signatures identified are distinct from previously reported ‘free-circulating’ miRNA studies in GBM patients, and appear to be superior.

Chapter 5 – Brain Metastases from Melanoma

Brain metastases are common in patients with metastatic melanoma and the majority of patients who die of the disease do so as a direct result of them. Not only does the diagnosis of brain metastasis signify a very poor prognosis, but it portends a severe impact on the patient’s quality of life. The major clinical problem of melanoma brain metastases has in the past been addressed by treatment with local therapies—surgery and radiation therapy—but the results have been unimpressive, with median survival times of just a few months reported in most series, little better than the survival times of patients offered supportive care only. However, in recent times more effective and less morbid techniques for both the surgical management of brain metastases and the treatment of them using new radiation therapy methodologies have been introduced and, at last, systemic therapies that can treat melanoma brain metastases effectively have been discovered. In this chapter the range of treatments now available to treat melanoma brain metastases will be discussed, outcomes achieved by their use will be presented, and the importance of a multidisciplinary approach to management of the problem will be emphasized.