Biology subjects

Scolyer, R. A.

Publications and source records attributed to Scolyer, R. A..

7 recordsLinked to original sources

bioRxiv2020

Cross-Platform Omics Prediction procedure enables precision medicine in patients with stage-III melanoma

There is no consensus methodology that can account for the variation in omics signatures when they are acquired across different platforms and times. This poses a significant barrier to the implementation of valuable biomarkers into clinical practice. We present a novel procedure (Cross-Platform Omics Prediction) that accounts for these variations and demonstrate its utility in three risk models for different diseases that is suitable for prospective and multi-centre clinical implementation.

bioinformatics↗

bioRxiv2020

Anatomic position determines oncogenic specificity in melanoma

Oncogenic alterations to DNA are not transforming in all cellular contexts1,2. This may be due to pre-existing transcriptional programs in the cell of origin. Here, we define anatomic position as a major determinant of why cells respond to specific oncogenes. Cutaneous melanoma arises throughout the body, whereas the acral subtype arises on the palms of the hands, soles of the feet, or under the nails3. We sequenced the DNA of cutaneous and acral melanomas from a large cohort of human patients and found a specific enrichment for BRAF mutations in cutaneous melanoma but CRKL amplifications in acral melanoma. We modeled these changes in transgenic zebrafish models and found that CRKL-driven tumors predominantly formed in the fins of the fish. The fins are the evolutionary precursors to tetrapod limbs, indicating that melanocytes in these acral locations may be uniquely susceptible to CRKL. RNA profiling of these fin/limb melanocytes, compared to body melanocytes, revealed a positional identity gene program typified by posterior HOX13 genes. This positional gene program synergized with CRKL to drive tumors at acral sites. Abrogation of this CRKL-driven program eliminated the anatomic specificity of acral melanoma. These data suggest that the anatomic position of the cell of origin endows it with a unique transcriptional state that makes it susceptible to only certain oncogenic insults.

cancer biology↗

bioRxiv2020

The deacylase SIRT5 supports melanoma viability by regulating chromatin dynamics

Cutaneous melanoma remains the most lethal skin cancer, and ranks third among all malignancies in terms of years of life lost. Despite the advent of immune checkpoint and targeted therapies, only roughly half of patients with advanced melanoma achieves a durable remission. SIRT5 is a member of the sirtuin family of protein deacylases that regulate metabolism and other biological processes. Germline Sirt5 deficiency is associated with mild phenotypes in mice. Here we show that SIRT5 is required for proliferation and survival across all cutaneous melanoma genotypes tested, as well as uveal melanoma, a genetically distinct melanoma subtype that arises in the eye and is incurable once metastatic. Likewise, SIRT5 is required for efficient tumor formation by melanoma xenografts and in an autochthonous mouse Braf;Pten-driven melanoma model. Via metabolite and transcriptomic analyses, we find that SIRT5 is required to maintain histone acetylation and methylation levels in melanoma cells, thereby promoting proper gene expression. SIRT5-dependent genes notably include MITF, a key lineage-specific survival oncogene in melanoma, and the c-MYC proto-oncogene. SIRT5 may represent a novel, druggable genotype-independent addiction in melanoma.

cancer biology↗

bioRxiv2020

Cryopreservation of human cancers conserves tumour heterogeneity for single-cell multi-omics analysis

BackgroundHigh throughput single-cell RNA sequencing (scRNA-Seq) has emerged as a powerful tool for exploring cellular heterogeneity amongst complex human cancers. scRNA-Seq studies using fresh human surgical tissue is logistically difficult, precludes histopathological triage of samples and limits the ability to perform batch processing. This hinderance can often introduce technical biases when integrating patient datasets and increase experimental costs. Although tissue preservation methods have been previously explored to address such issues, it is yet to be examined on complex human tissues, such as solid cancers, and on high throughput scRNA-Seq platforms. ResultsWe show that the viable cryopreservation of human cancers provides high quality single-cell transcriptomes using the Chromium 10X platform. We sequenced a total of [~]120,000 cells from fresh and cryopreserved replicates across three breast cancers, two prostate cancers and a cutaneous melanoma. Importantly, tumour heterogeneity identified from fresh tissues was largely conserved in cryopreserved replicates. We show that sequencing of single cells prepared from cryopreserved tissue fragments or from cryopreserved cell suspensions is comparable to sequenced cells prepared from fresh tissue, with cryopreserved cell suspensions displaying higher correlations with fresh tissue in gene expression. We then show that cryopreservation had minimal impacts on results of downstream analyses such as biological pathway enrichment. Further, we demonstrate the advantage of cryopreserving whole-cells for immunophenotyping methods such as CITE-Seq, which is impossible using other preservation methods such as single nuclei-sequencing. ConclusionsOur study guides new experimental designs for tissue biobanking for future clinical single-cell RNA sequencing studies.

cancer biology↗

bioRxiv2020

Hotspot KRAS mutations in brain metastases at the first metastatic recurrence of cutaneous melanoma

IMPORTANCEBrain metastases occur in 60% of patients with advanced melanoma and are a major cause of melanoma-related mortality and morbidity. Although our understanding of the molecular alterations associated with melanoma progression is improving, there are currently no validated biomarkers which might help identify those patients at highest risk of developing brain metastases. OBJECTIVETo examine the somatic mutational and copy-number landscape of brain metastases that develop as the isolated first visceral site of recurrence - "early brain-metastasis" compared to extracranial melanoma metastases. DESIGN, SETTING AND PARTICIPANTSWhole-exome sequencing of 50 tumors from patients undergoing surgical resection of one or more brain metastasis occurring as the first site of visceral relapse were identified from prospectively maintained databases in Sydney, Wellington, New York and Cambridge. Whole exome sequencing analyses allowed mutational profiles to be compared to cutaneous melanomas in The Cancer Genome Atlas (SKCM-TCGA; n=358) and the Memorial Sloan Kettering (SKCM-MSK-IMPACT; n=186) datasets. An external dataset comprising a further 18 patients with surgically resected early brain metastasis from two additional academic centers served as an independent validation cohort. MAIN OUTCOMES AND MEASURESTo assess the frequency of driver mutations in early brain metastasis and their influence on survival. RESULTSIn concordance with the landmark melanoma sequencing studies, we identified mutations in BRAF (21/50, 42%), NRAS (14/50, 28%) and NF1 (11/50, 22%) as the most frequently mutated melanoma driver genes. When compared to the mutational landscape of cutaneous melanomas in TCGA (SKCM-TCGA), KRAS was the most significantly enriched driver gene, with 5/50 (10%) of brain metastases harboring non-synonymous mutations, of which 4/5 (80%) were in the hotspot positions of codons 12 and 61. This was significantly higher than the corresponding frequency of KRAS-mutations within the entire SKCM-TCGA (2% (7/358), p=0.009, Fishers Exact Test) as well as the SKCM-MSK-IMPACT cohort (1.6% (3/186), p=0.016). Variants in KRAS were mutually exclusive from BRAFV600, NRAS and HRAS mutations and were associated with a significantly reduced overall survival from resection of brain metastasis (relative to KRAS-wild type brain metastases) in multivariate Cox proportional hazard models (HR 1.80, 95% CI 1.46-24.89, p=0.013). Mutations in KRAS were also clonal and concordant with extracranial disease, which suggests these mutations are present within the primary tumor CONCLUSIONS AND RELEVANCEOur analysis, the largest to date, suggests that early metastases to the brain (presenting as the first site of visceral relapse) are characterized by significant enrichment of hotspot KRAS mutations, potentially implicating constitutive RAS-driven cellular programs in neurotropic metastatic behavior in these cases. Based on these data, we suggest that screening for KRAS mutations might help identify those patients with primary melanoma at higher risk of brain metastases or poor survival, and could help inform future surveillance strategies. Key PointsO_ST_ABSQuestionC_ST_ABSWhat is the frequency of driver mutations in early melanoma brain metastases? FindingsIn this study of 50 patients with melanoma metastasizing first to the brain, KRAS mutations were the most significantly enriched driver gene (n=5, 10% of patients) when compared to landmark cutaneous melanoma studies. The high KRAS mutation frequency was also observed in an external validation cohort of 18 patients with early brain metastases. Mutations in KRAS were mutually exclusive from mutations in the key RAS signaling genes and conferred a worse overall survival from resection of brain metastasis. MeaningHotspot KRAS mutations could help identify those patients with primary melanoma at higher risk of brain metastases that may benefit from more intensive, protracted surveillance as well as earlier use of adjuvant therapy.

genomics↗

bioRxiv2019

Temporal and spatial modulation of the immune response of the murine Gl261 glioma tumor microenvironment

Glioblastoma, the most aggressive form of glioma, has a 5-year survival rate of <5%. While radiation and immunotherapies are routinely studied in the murine Gl261 glioma model, little is known about its inherent immune response. This study quantifies the temporal and spatial localization of immune cell populations and mediators during glioma development. Eight-week old male C57Bl/6 mice were orthotopically inoculated with 1x106 Gl261 cells and tumor morphology, local and systemic immune cell populations, and plasma cytokines/chemokines assessed at Day-0, 1, 3, 7, 14, and 21 post-inoculation by magnetic resonance imaging, chromogenic immunohistochemistry, multiplex immunofluorescent immunohistochemistry, flow cytometry and multiplex immunoassay respectively. From Day-3 tumors were distinguishable with >30% Ki67 and increased tissue vascularization (p<0.05). Increasing tumor proliferation/malignancy and vascularization were associated with significant temporal changes in immune cell populations within the tumor (p<0.05) and systemic compartments (p=0.02 to p<0.0001). Of note, at Day-14 16/24 plasma cytokine/chemokines levels decreased coinciding with an increase in tumor cytotoxic T cells, natural killer and natural killer/T cells. Data derived provide baseline characterization of the local and systemic immune response during glioma development. They reveal that type II macrophages and myeloid-derived suppressor cells are more prevalent in tumors than regulatory T cells, highlighting these cell types for further therapeutic exploration.

cancer biology↗

bioRxiv2018

Epigenetic Profiling for the Molecular Classification of Metastatic Brain Tumors

Optimal treatment of brain metastases is often hindered by limitations in diagnostic capabilities. To meet these challenges, we generated genome-scale DNA methylomes of the three most frequent types of brain metastases: melanoma, breast, and lung cancers (n=96). Using supervised machine learning and integration of multiple DNA methylomes from normal, primary, and metastatic tumor specimens (n=1,860), we unraveled epigenetic signatures specific to each type of metastatic brain tumor and constructed a three-step DNA methylation-based classifier (BrainMETH) that categorizes brain metastases according to the tissue of origin and therapeutically-relevant subtypes. BrainMETH predictions were supported by routine histopathologic evaluation. We further characterized and validated the most predictive genomic regions in a large cohort of brain tumors (n=165) using quantitative methylation-specific PCR. Our study highlights the importance of brain tumor-defining epigenetic alterations, which can be utilized to further develop DNA methylation profiling as a critical tool in the histomolecular stratification of patients with brain metastases.

cancer biology↗