Symptomatic brain metastases in melanoma.

Although clinical outcomes in metastatic melanoma have improved in recent years, the morbidity and mortality of symptomatic brain metastases remain challenging. Response rates and survival outcomes of patients with symptomatic melanoma brain metastases (MBM) are significantly inferior to patients with asymptomatic disease. This review focusses upon the specific challenges associated with the management of symptomatic MBM, discussing current treatment paradigms, obstacles to improving clinical outcomes and directions for future research.

Heterogeneity in cancer: cancer stem cells versus clonal evolution.

The identification and characterization of cancer stem cells might lead to more effective treatments for some cancers by focusing therapy on the most malignant cells. To achieve this goal it will be necessary to determine which cancers follow a cancer stem cell model and which do not, to address technical issues related to tumorigenesis assays, and to test the extent to which cancer cell heterogeneity arises from genetic versus epigenetic differences.

Whole-genome landscapes of major melanoma subtypes.

Melanoma of the skin is a common cancer only in Europeans, whereas it arises in internal body surfaces (mucosal sites) and on the hands and feet (acral sites) in people throughout the world. Here we report analysis of whole-genome sequences from cutaneous, acral and mucosal subtypes of melanoma. The heavily mutated landscape of coding and non-coding mutations in cutaneous melanoma resolved novel signatures of mutagenesis attributable to ultraviolet radiation. However, acral and mucosal melanomas were dominated by structural changes and mutation signatures of unknown aetiology, not previously identified in melanoma. The number of genes affected by recurrent mutations disrupting non-coding sequences was similar to that affected by recurrent mutations to coding sequences. Significantly mutated genes included BRAF, CDKN2A, NRAS and TP53 in cutaneous melanoma, BRAF, NRAS and NF1 in acral melanoma and SF3B1 in mucosal melanoma. Mutations affecting the TERT promoter were the most frequent of all; however, neither they nor ATRX mutations, which correlate with alternative telomere lengthening, were associated with greater telomere length. Most melanomas had potentially actionable mutations, most in components of the mitogen-activated protein kinase and phosphoinositol kinase pathways. The whole-genome mutation landscape of melanoma reveals diverse carcinogenic processes across its subtypes, some unrelated to sun exposure, and extends potential involvement of the non-coding genome in its pathogenesis.

Yield of baseline imaging for distant metastases in high-risk primary melanoma.

BACKGROUND: The yield of baseline imaging in patients presenting with higher risk primary tumours, at least American Joint Committee on Cancer 8th edition stage IIC or III melanoma, is unclear. METHODS: This retrospective study included patients referred to the Victorian Melanoma Service from January 2017 to April 2020, diagnosed with at least stage IIC or stage III melanoma. Patients with a T4b tumour and no sentinel lymph node biopsy were included as 'T4bNX'. RESULTS: One hundred and sixty-four patients (median age 65 years) with baseline imaging (T4bNX: 19, IIC: 30, IIIA: 21, IIIB: 43, IIIC: 50, IIID: 1) were included. The majority were male (73%), and those with T4bNX melanoma tended to be older (median age 79 years). Distant metastases were detected in 21% (4/19) of T4bNX, 3% (1/30) of stage IIC, 0% (0/21) of stage IIIA, and 6% (6/94) of stages IIIB-D melanoma patients. All stage III patients with distant metastases had palpable lymphadenopathy a presentation. Two patients had brain metastases, both of whom had T4bNX melanoma and synchronous extra-cranial metastases. CONCLUSIONS: Compared to stage IIC, baseline imaging detects higher rates of extra-cranial distant disease in stages IIIB-D and T4bNX melanoma. Intracranial imaging has greater yield in patients with distant extra-cranial disease.

Melanoma recurrence patterns and management after adjuvant targeted therapy: a multicentre analysis.

BACKGROUND: Adjuvant targeted therapy (TT) improves relapse free survival in patients with resected BRAF mutant stage III melanoma. The outcomes and optimal management of patients who relapse after adjuvant TT is unknown. METHODS: Patients from twenty-one centres with recurrent melanoma after adjuvant TT were included. Disease characteristics, adjuvant therapy, recurrence, treatment at relapse and outcomes were examined. RESULTS: Eighty-five patients developed recurrent melanoma; nineteen (22%) during adjuvant TT. Median time to first recurrence was 18 months and median follow-up from first recurrence was 31 months. Fifty-eight (68%) patients received immunotherapy (IT) or TT as 1st line systemic therapy at either first or subsequent recurrence and had disease that was assessable for response. Response to anti-PD-1 (±trial agent), combination ipilimumab-nivolumab, TT rechallenge and ipilimumab monotherapy was 63%, 62% 25% and 10% respectively. Twenty-eight (33%) patients had died at census, all from melanoma. Two-year OS was 84% for anti-PD-1 therapy (±trial agent), 92% for combination ipilimumab and nivolumab, 49% for TT and 45% for ipilimumab monotherapy (p = 0.028). CONCLUSIONS: Patients who relapse after adjuvant TT respond well to subsequent anti-PD-1 based therapy and have outcomes similar to those seen when first line anti-PD-1 therapy is used in stage IV melanoma.

Whole-genome characterization of chemoresistant ovarian cancer.

Patients with high-grade serous ovarian cancer (HGSC) have experienced little improvement in overall survival, and standard treatment has not advanced beyond platinum-based combination chemotherapy, during the past 30 years. To understand the drivers of clinical phenotypes better, here we use whole-genome sequencing of tumour and germline DNA samples from 92 patients with primary refractory, resistant, sensitive and matched acquired resistant disease. We show that gene breakage commonly inactivates the tumour suppressors RB1, NF1, RAD51B and PTEN in HGSC, and contributes to acquired chemotherapy resistance. CCNE1 amplification was common in primary resistant and refractory disease. We observed several molecular events associated with acquired resistance, including multiple independent reversions of germline BRCA1 or BRCA2 mutations in individual patients, loss of BRCA1 promoter methylation, an alteration in molecular subtype, and recurrent promoter fusion associated with overexpression of the drug efflux pump MDR1.

Parity reduces mammary repopulating activity but does not affect mammary stem cells defined as CD24 + CD29/CD49fhi in mice.

BACKGROUND: Breast cancer (BCa) mortality is decreasing with early detection and improvement in therapies. The incidence of BCa, however, continues to increase, particularly estrogen-receptor-positive (ER +) subtypes. One of the greatest modifiers of ER + BCa risk is childbearing (parity), with BCa risk halved in young multiparous mothers. Despite convincing epidemiological data, the biology that underpins this protection remains unclear. Parity-induced protection has been postulated to be due to a decrease in mammary stem cells (MaSCs); however, reports to date have provided conflicting data. METHODS: We have completed rigorous functional testing of repopulating activity in parous mice using unfractionated and MaSC (CD24midCD49fhi)-enriched populations. We also developed a novel serial transplant method to enable us to assess self-renewal of MaSC following pregnancy. Lastly, as each pregnancy confers additional BCa protection, we subjected mice to multiple rounds of pregnancy to assess whether additional pregnancies impact MaSC activity. RESULTS: Here, we report that while repopulating activity in the mammary gland is reduced by parity in the unfractionated gland, it is not due to a loss in the classically defined MaSC (CD24+CD49fhi) numbers or function. Self-renewal was unaffected by parity and additional rounds of pregnancy also did not lead to a decrease in MaSC activity. CONCLUSIONS: Our data show instead that parity impacts on the stem-like activity of cells outside the MaSC population.

A non-canonical role for desmoglein-2 in endothelial cells: implications for neoangiogenesis.

Desmogleins (DSG) are a family of cadherin adhesion proteins that were first identified in desmosomes and provide cardiomyocytes and epithelial cells with the junctional stability to tolerate mechanical stress. However, one member of this family, DSG2, is emerging as a protein with additional biological functions on a broader range of cells. Here we reveal that DSG2 is expressed by non-desmosome-forming human endothelial progenitor cells as well as their mature counterparts [endothelial cells (ECs)] in human tissue from healthy individuals and cancer patients. Analysis of normal blood and bone marrow showed that DSG2 is also expressed by CD34(+)CD45(dim) hematopoietic progenitor cells. An inability to detect other desmosomal components, i.e., DSG1, DSG3 and desmocollin (DSC)2/3, on these cells supports a solitary role for DSG2 outside of desmosomes. Functionally, we show that CD34(+)CD45(dim)DSG2(+) progenitor cells are multi-potent and pro-angiogenic in vitro. Using a 'knockout-first' approach, we generated a Dsg2 loss-of-function strain of mice (Dsg2 (lo/lo)) and observed that, in response to reduced levels of Dsg2: (i) CD31(+) ECs in the pancreas are hypertrophic and exhibit altered morphology, (ii) bone marrow-derived endothelial colony formation is impaired, (iii) ex vivo vascular sprouting from aortic rings is reduced, and (iv) vessel formation in vitro and in vivo is attenuated. Finally, knockdown of DSG2 in a human bone marrow EC line reveals a reduction in an in vitro angiogenesis assay as well as relocalisation of actin and VE-cadherin away from the cell junctions, reduced cell-cell adhesion and increased invasive properties by these cells. In summary, we have identified DSG2 expression in distinct progenitor cell subpopulations and show that, independent from its classical function as a component of desmosomes, this cadherin also plays a critical role in the vasculature.

Socrates: identification of genomic rearrangements in tumour genomes by re-aligning soft clipped reads.

MOTIVATION: Methods for detecting somatic genome rearrangements in tumours using next-generation sequencing are vital in cancer genomics. Available algorithms use one or more sources of evidence, such as read depth, paired-end reads or split reads to predict structural variants. However, the problem remains challenging due to the significant computational burden and high false-positive or false-negative rates. RESULTS: In this article, we present Socrates (SOft Clip re-alignment To idEntify Structural variants), a highly efficient and effective method for detecting genomic rearrangements in tumours that uses only split-read data. Socrates has single-nucleotide resolution, identifies micro-homologies and untemplated sequence at break points, has high sensitivity and high specificity and takes advantage of parallelism for efficient use of resources. We demonstrate using simulated and real data that Socrates performs well compared with a number of existing structural variant detection tools. AVAILABILITY AND IMPLEMENTATION: Socrates is released as open source and available from http://bioinf.wehi.edu.au/socrates CONTACT: papenfuss@wehi.edu.au Supplementary information: Supplementary data are available at Bioinformatics online.

Post-operative survival following metastasectomy for patients receiving BRAF inhibitor therapy is associated with duration of pre-operative treatment and elective indication.

INTRODUCTION: Metastasectomy can provide durable disease control for selected patients with metastatic melanoma. Vemurafenib is a BRAF kinase inhibitor which has demonstrated significant improvement in disease-specific survival in patients with metastatic melanoma with a BRAF gene mutation. This study examined the efficacy and safety of metastasectomy during treatment with vemurafenib. METHODS: A retrospective review was performed of all patients receiving vemurafenib at Peter MacCallum Cancer Centre. Patient records were reviewed to identify patients undergoing surgery within 30 days of vemurafenib therapy. Descriptive statistics and survival analysis were performed. RESULTS: Nineteen patients underwent 21 metastasectomies including craniotomy (57%), spinal decompression (14%), small bowel resection (14%), lung resection (9.5%) and neck dissection (4.5%). Indications for surgery were: an isolated residual focus of disease (n = 2); isolated progressive disease in the setting of stability elsewhere (n = 9); and symptomatic disease (n = 8). Grade 2 or higher surgical complications occurred in 19% of cases and there was one peri-operative death. Median post-operative survival was seven months. There was a trend toward improved post-operative survival for patients with longer duration of vemurafenib therapy (P = 0.04) and for those undergoing elective surgery (P = 0.07). CONCLUSION: Resection of oligometastatic disease during BRAF-targeted therapy is safe. Selected patients have durable post-operative disease control.

Gata-3 is an essential regulator of mammary-gland morphogenesis and luminal-cell differentiation.

The transcription factor Gata-3 is a defining marker of the 'luminal' subtypes of breast cancer. To gain insight into the role of Gata-3 in breast epithelial development and oncogenesis, we have explored its normal function within the mammary gland by conditionally deleting Gata-3 at different stages of development. We report that Gata-3 has essential roles in the morphogenesis of the mammary gland in both the embryo and adult. Through the discovery of a novel marker (beta3-integrin) of luminal progenitor cells and their purification, we demonstrate that Gata-3 deficiency leads to an expansion of luminal progenitors and a concomitant block in differentiation. Remarkably, introduction of Gata-3 into a stem cell-enriched population induced maturation along the alveolar luminal lineage. These studies provide evidence for the existence of an epithelial hierarchy within the mammary gland and establish Gata-3 as a critical regulator of luminal differentiation.

Efficient tumour formation by single human melanoma cells.

A fundamental question in cancer biology is whether cells with tumorigenic potential are common or rare within human cancers. Studies on diverse cancers, including melanoma, have indicated that only rare human cancer cells (0.1-0.0001%) form tumours when transplanted into non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice. However, the extent to which NOD/SCID mice underestimate the frequency of tumorigenic human cancer cells has been uncertain. Here we show that modified xenotransplantation assay conditions, including the use of more highly immunocompromised NOD/SCID interleukin-2 receptor gamma chain null (Il2rg(-/-)) mice, can increase the detection of tumorigenic melanoma cells by several orders of magnitude. In limiting dilution assays, approximately 25% of unselected melanoma cells from 12 different patients, including cells from primary and metastatic melanomas obtained directly from patients, formed tumours under these more permissive conditions. In single-cell transplants, an average of 27% of unselected melanoma cells from four different patients formed tumours. Modifications to xenotransplantation assays can therefore dramatically increase the detectable frequency of tumorigenic cells, demonstrating that they are common in some human cancers.

Immune Regulation and Immune Therapy in Melanoma: Review with Emphasis on CD155 Signalling.

Melanoma is commonly diagnosed in a younger population than most other solid malignancies and, in Australia and most of the world, is the leading cause of skin-cancer-related death. Melanoma is a cancer type with high immunogenicity; thus, immunotherapies are used as first-line treatment for advanced melanoma patients. Although immunotherapies are working well, not all the patients are benefitting from them. A lack of a comprehensive understanding of immune regulation in the melanoma tumour microenvironment is a major challenge of patient stratification. Overexpression of CD155 has been reported as a key factor in melanoma immune regulation for the development of therapy resistance. A more thorough understanding of the actions of current immunotherapy strategies, their effects on immune cell subsets, and the roles that CD155 plays are essential for a rational design of novel targets of anti-cancer immunotherapies. In this review, we comprehensively discuss current anti-melanoma immunotherapy strategies and the immune response contribution of different cell lineages, including tumour endothelial cells, myeloid-derived suppressor cells, cytotoxic T cells, cancer-associated fibroblast, and nature killer cells. Finally, we explore the impact of CD155 and its receptors DNAM-1, TIGIT, and CD96 on immune cells, especially in the context of the melanoma tumour microenvironment and anti-cancer immunotherapies.

Globe Salvage and Vision Preservation by Neoadjuvant Darovasertib and Crizotinib in Uveal Melanoma.

PURPOSE: To report the effective use of neoadjuvant darovasertib and crizotinib in a patient with a large uveal melanoma (UM) in his only functional eye. DESIGN: Case report. SUBJECTS: One patient with T4b UM. INTERVENTION: Neoadjuvant darovasertib and crizotinib, followed by plaque brachytherapy. MAIN OUTCOME MEASURES: Objective tumor response and conversion from planned enucleation to placement of fovea- and optic nerve-sparing plaque brachytherapy. RESULTS: A patient with a history of left eye blindness from retinal artery occlusion presented with rapidly declining right eye vision due to a primary UM measuring 18 mm in maximal diameter and 16.5 mm in maximal thickness. To salvage vision, neoadjuvant treatment was initiated using darovasertib and crizotinib. After 6 months of neoadjuvant treatment, which included intraocular lens replacement for tumor-associated cataract, the tumor regressed to 14.1 mm in maximal diameter and 2.6 mm in maximal thickness, enabling treatment with plaque brachytherapy rather than enucleation. CONCLUSIONS: The combination of darovasertib and crizotinib for UM is an effective neoadjuvant strategy that warrants further investigation as an approach to improve visual outcomes from the treatment of primary UM. FINANCIAL DISCLOSURE: The other authors have no proprietary or commercial interest in any materials discussed in this article.

Computational repurposing of oncology drugs through off-target drug binding interactions from pharmacological databases.

PURPOSE: Systematic repurposing of approved medicines for another indication may accelerate drug development in oncology. We present a strategy combining biomarker testing with drug repurposing to identify new treatments for patients with advanced cancer. METHODS: Tumours were sequenced with the Illumina TruSight Oncology 500 (TSO-500) platform or the FoundationOne CDx panel. Mutations were screened by two medical oncologists and pathogenic mutations were categorised referencing literature. Variants of unknown significance were classified as potentially pathogenic using plausible mechanisms and computational prediction of pathogenicity. Gain of function (GOF) mutations were evaluated through repurposing databases Probe Miner (PM), Broad Institute Drug Repurposing Hub (Broad Institute DRH) and TOPOGRAPH. GOF mutations were repurposing events if identified in PM, not indexed in TOPOGRAPH and excluding mutations with a known Food and Drug Administration (FDA)-approved biomarker. The computational repurposing approach was validated by evaluating its ability to identify FDA-approved biomarkers. The total repurposable genome was identified by evaluating all possible gene-FDA drug-approved combinations in the PM dataset. RESULTS: The computational repurposing approach was accurate at identifying FDA therapies with known biomarkers (94%). Using next-generation sequencing molecular reports (n = 94), a meaningful percentage of patients (14%) could have an off-label therapeutic identified. The frequency of theoretical drug repurposing events in The Cancer Genome Atlas pan-cancer dataset was 73% of the samples in the cohort. CONCLUSION: A computational drug repurposing approach may assist in identifying novel repurposing events in cancer patients with no access to standard therapies. Further validation is needed to confirm a precision oncology approach using drug repurposing.

Predictive Impact of Tumor Mutational Burden on Real-World Outcomes of First-Line Immune Checkpoint Inhibition in Metastatic Melanoma.

PURPOSE: The choice of threshold and reliability of high tumor mutational burden (TMB) to predict outcomes and guide treatment choice for patients with metastatic melanoma receiving first-line immune checkpoint inhibitor (ICI) therapy in the real world is not well known. METHODS: Using a deidentified nationwide (US-based) melanoma clinicogenomic database, we identified a real-world cohort of patients with metastatic melanoma (N = 497) who received first-line monotherapy anti-PD-1 (n = 240) or dual anti-PD-1 and anti-CTLA-4 ICI (n = 257) and had a tissue-based comprehensive genomic profiling test TMB score. RESULTS: TMB-high (TMB-H; ≥10 mutations per megabase [muts/Mb], n = 352, 71%) was independently predictive of superior real-world progression-free survival and overall survival versus TMB-low (<10 mut/Mb, n = 145, 29%) in both mono ICI (hazard ratio [HR], 0.45 [95% CI, 0.32 to 0.63]; P < .001; HR, 0.61 [95% CI, 0.41 to 0.90]; P = .01, respectively) and dual ICI (HR, 0.67 [95% CI, 0.49 to 0.90]; P = .009; HR, 0.61 [95% CI, 0.42 to 0.88]; P = .007, respectively) patients. Dual ICI offered no significant advantage in BRAFwt patients and unexpectedly demonstrated greatest benefit in the TMB 10-19 mut/Mb group, identifying a TMB-very high (≥20 mut/Mb, n = 247, 50%) BRAFmut patient subgroup for whom mono ICI may be preferable. CONCLUSION: TMB-H predicts superior outcomes on ICI while coassessment of BRAF status and TMB may inform first-line regimen choice.

The Fight Tumour Blindness Registry: Efficient capture of high-quality real-world data in uveal melanoma.

OBJECTIVE: To describe the development of a web-based data collection tool to track the management and outcomes of uveal melanoma patients. DESIGN: Description of a clinical registry. PARTICIPANTS: Patients with uveal melanoma. METHODS: A panel of expert ocular oncologists, with input from other relevant specialties and individuals with expertise in registry development, collaborated to formulate a minimum data set to be collected to track patient centred, real-world outcomes in uveal melanoma. This data set was used to create the Fight Tumour Blindness! (FTB!) registry within Save Sight Registries. RESULTS: The data set to be collected includes patient demographics and medical history, baseline visit, follow-up visit including tumour treatment, metastatic staging and surveillance, pathology, and patient-reported questionnaires. The inbuilt mechanisms to ensure efficient and complete data collection are described. CONCLUSIONS: The FTB! registry can be used to monitor outcomes for patients with uveal melanoma. It allows benchmarking of outcomes and comparisons between different clinics and countries.

Evaluating two infiltration gallery designs for managed aquifer recharge using secondary treated wastewater.

As managed aquifer recharge (MAR) becomes increasingly considered for augmenting water-sensitive urban areas, fundamental knowledge of the achievable scale, longevity and maintenance requirements of different options will become paramount. This paper reports on a 39 month pilot scale MAR scheme that infiltrated secondary treated wastewater through unsaturated sand into a limestone and sand aquifer. Two types of infiltration gallery were constructed to compare their hydraulic performance, one using crushed, graded gravel, the other using an engineered leach drain system (Atlantis Leach System(®)). Both galleries received 25 kL of nutrient-rich, secondary treated wastewater per day. The Atlantis gallery successfully infiltrated 17 ML of treated wastewater over three years. The slotted distribution pipe in the gravel gallery became clogged with plant roots after operating for one year. The infiltration capacity of the gravel gallery could not be restored despite high pressure cleaning, thus it was replaced with an Atlantis system. Reduction in the infiltration capacity of the Atlantis system was only observed when inflow was increased by about 3 fold for two months. The performance of the Atlantis system suggests it is superior to the gravel gallery, requiring less maintenance within at least the time frame of this study. The results from a bromide tracer test revealed a minimum transport time of 3.7 days for the recharged water to reach the water table below 9 m of sand and limestone. This set a limit on the time available for attenuation by natural treatment within the unsaturated zone before it recharged groundwater.

Immune-related adverse events are associated with therapeutic efficacy of immunotherapy in patients with melanoma brain metastases.

Immunotherapy with T-cell checkpoint inhibitors have changed the treatment landscape for patients with melanoma brain metastases (MBMs), offering increased survival compared with historical outcomes. We sought to identify clinical features associated with intracranial tumour responses or progression-free survival (PFS) in patients with MBMs treated with immunotherapy. Patients with MBMs treated with immunotherapy from August 2013 to March 2020 were identified through local databases. Melanoma disease burdens and immune-related adverse events (irAEs) were assessed retrospectively by review of patient medical records. Efficacy was evaluated by determining objective response rates (ORRs) in brain metastases using immune-Response Evaluation Criteria in Solid Tumours criteria, MBM-specific survival and overall PFS. Twenty-six patients were identified as eligible for this study. The presence and volume of extracranial metastases (ECM) were associated with a non-significant trend of reduced intracranial ORRs and PFS. Patients with irAEs, on the other hand, had significantly increased intracranial ORRs and PFS compared to those without irAEs. Severe, grade ≥3 irAEs and co-occurrence of ≥2 irAEs were also significantly associated with longer PFS. The presence and volume of ECM correlated inversely with development and severity of irAEs. We report a strong association between the development of irAEs and favourable melanoma-specific outcomes in patients with MBMs receiving immunotherapy. Contrary to previous studies, we found that co-occurrence of ECM in these patients was associated with fewer irAEs and reduced treatment efficacy.