Alexander C. Gagnon

The Genetic Evolution of Melanoma from Precursor Lesions

BACKGROUND The pathogenic mutations in melanoma have been largely catalogued; however, the order of their occurrence is not known. METHODS We sequenced 293 cancer-relevant genes in 150 areas of 37 primary melanomas and their adjacent precursor lesions. The histopathological spectrum of these areas included unequivocally benign lesions, intermediate lesions, and intraepidermal or invasive melanomas. RESULTS Precursor lesions were initiated by mutations of genes that are known to activate the mitogen-activated protein kinase pathway. Unequivocally benign lesions harbored BRAF V600E mutations exclusively, whereas those categorized as intermediate were enriched for NRAS mutations and additional driver mutations. A total of 77% of areas of intermediate lesions and melanomas in situ harbored TERT promoter mutations, a finding that indicates that these mutations are selected at an unexpectedly early stage of the neoplastic progression. Biallelic inactivation of CDKN2A emerged exclusively in invasive melanomas. PTEN and TP53 mutations were found only in advanced primary melanomas. The point-mutation burden increased from benign through intermediate lesions to melanoma, with a strong signature of the effects of ultraviolet radiation detectable at all evolutionary stages. Copy-number alterations became prevalent only in invasive melanomas. Tumor heterogeneity became apparent in the form of genetically distinct subpopulations as melanomas progressed. CONCLUSIONS Our study defined the succession of genetic alterations during melanoma progression, showing distinct evolutionary trajectories for different melanoma subtypes. It identified an intermediate category of melanocytic neoplasia, characterized by the presence of more than one pathogenic genetic alteration and distinctive histopathological features. Finally, our study implicated ultraviolet radiation as a major factor in both the initiation and progression of melanoma. (Funded by the National Institutes of Health and others.).

Exome sequencing of desmoplastic melanoma identifies recurrent NFKBIE promoter mutations and diverse activating mutations in the MAPK pathway

Desmoplastic melanoma is an uncommon variant of melanoma with sarcomatous histology, distinct clinical behavior and unknown pathogenesis. We performed low-coverage genome and high-coverage exome sequencing of 20 desmoplastic melanomas, followed by targeted sequencing of 293 genes in a validation cohort of 42 cases. A high mutation burden (median of 62 mutations/Mb) ranked desmoplastic melanoma among the most highly mutated cancers. Mutation patterns strongly implicate ultraviolet radiation as the dominant mutagen, indicating a superficially located cell of origin. Newly identified alterations included recurrent promoter mutations of NFKBIE, encoding NF-κB inhibitor ɛ (IκBɛ), in 14.5% of samples. Common oncogenic mutations in melanomas, in particular in BRAF (encoding p.Val600Glu) and NRAS (encoding p.Gln61Lys or p.Gln61Arg), were absent. Instead, other genetic alterations known to activate the MAPK and PI3K signaling cascades were identified in 73% of samples, affecting NF1, CBL, ERBB2, MAP2K1, MAP3K1, BRAF, EGFR, PTPN11, MET, RAC1, SOS2, NRAS and PIK3CA, some of which are candidates for targeted therapies.

Primary U distribution in scleractinian corals and its implications for U series dating

In this study we use microsampling techniques to explore diagenetic processes in carbonates. These processes are important as they can affect the accuracy of U series chronometry. Fission track maps of deep-sea scleractinian corals show a threefold difference between the minimum and maximum [U] in modern corals, which is reduced to a factor of 2 in fossil corals. We use micromilling and MC-ICP-MS to make detailed analyses of the [U] and δ234Uinitial distributions in corals from 218 ka to modern. Within each fossil coral we observe a large range of δ234Uinitial values, with high δ234Uinitial values typically associated with low [U]. A simple model shows that this observation is best explained by preferential movement of alpha-decay produced 234U atoms (alpha-recoil diffusion). Open-system addition of 234U may occur when alpha-recoil diffusion is coupled with a high [U] surface layer, such as organic material. This process can result in large, whole-coral δ234Uinitial elevations with little effect on the final age. The diagenetic pathways that we model are relevant to both shallow-water and deep-sea scleractinian corals since both exhibit primary [U] heterogeneity and may be subject to U addition.

Activating MET kinase rearrangements in melanoma and Spitz tumours

Oncogenic gene fusions have been identified in many cancers and many serve as biomarkers or targets for therapy. Here we identify six different melanocytic tumors with genomic rearrangements of MET fusing the kinase domain of MET in-frame to six different N-terminal partners. These tumors lack activating mutations in other established melanoma oncogenes. We functionally characterize two of the identified fusion proteins (TRIM4-MET and ZKSCAN1-MET) and find that they constitutively activate the mitogen-activated protein kinase (MAPK), phosphoinositol-3 kinase (PI3K), and phospholipase C gamma 1 (PLCγ1) pathways. The MET inhibitors cabozantinib (FDA-approved for progressive medullary thyroid cancer) and PF-04217903 block their activity at nanomolar concentrations. MET fusion kinases thus provide a potential therapeutic target for a rare subset of melanoma for which currently no targeted therapeutic options currently exist.

Strong depth-related zonation of megabenthos on a rocky continental margin (∼700-4000 m) off southern Tasmania, Australia.

Assemblages of megabenthos are structured in seven depth-related zones between ∼700 and 4000 m on the rocky and topographically complex continental margin south of Tasmania, southeastern Australia. These patterns emerge from analysis of imagery and specimen collections taken from a suite of surveys using photographic and in situ sampling by epibenthic sleds, towed video cameras, an autonomous underwater vehicle and a remotely operated vehicle (ROV). Seamount peaks in shallow zones had relatively low biomass and low diversity assemblages, which may be in part natural and in part due to effects of bottom trawl fishing. Species richness was highest at intermediate depths (1000–1300 m) as a result of an extensive coral reef community based on the bioherm-forming scleractinian Solenosmilia variabilis. However, megabenthos abundance peaked in a deeper, low diversity assemblage at 2000–2500 m. The S. variabilis reef and the deep biomass zone were separated by an extensive dead, sub-fossil S. variabilis reef and a relatively low biomass stratum on volcanic rock roughly coincident with the oxygen minimum layer. Below 2400 m, megabenthos was increasingly sparse, though punctuated by occasional small pockets of relatively high diversity and biomass. Nonetheless, megabenthic organisms were observed in the vast majority of photographs on all seabed habitats and to the maximum depths observed - a sandy plain below 3950 m. Taxonomic studies in progress suggest that the observed depth zonation is based in part on changing species mixes with depth, but also an underlying commonality to much of the seamount and rocky substrate biota across all depths. Although the mechanisms supporting the extraordinarily high biomass in 2000–2500 m depths remains obscure, plausible explanations include equatorwards lateral transport of polar production and/or a response to depth-stratified oxygen availability.

NTRK3 kinase fusions in Spitz tumours

Oncogenic fusions in TRK family receptor tyrosine kinases have been identified in several cancers and can serve as therapeutic targets. We identified ETV6–NTRK3, MYO5A–NTRK3 and MYH9–NTRK3 fusions in Spitz tumours, and demonstrated that NTRK3 fusions constitutively activate the mitogen‐activated protein kinase, phosphoinositide 3‐kinase and phospholipase Cγ1 pathways in melanocytes. This signalling was inhibited by DS‐6051a, a small‐molecule inhibitor of NTRK1/2/3 and ROS1. NTRK3 fusions expand the range of oncogenic kinase fusions in melanocytic neoplasms and offer targets for a small subset of melanomas for which no targeted options currently exist. Copyright

Link between light-triggered Mg-banding and chamber formation in the planktic foraminifera Neogloboquadrina dutertrei

The relationship between seawater temperature and the average Mg/Ca ratios in planktic foraminifera is well established, providing an essential tool for reconstructing past ocean temperatures. However, many species display alternating high and low Mg-bands within their shell walls that cannot be explained by temperature alone. Recent experiments demonstrate that intrashell Mg variability in Orbulina universa, which forms a spherical terminal shell, is paced by the diurnal light/dark cycle. Whether Mg-heterogeneity is also diurnally paced in species with more complex shell morphologies is unknown. Here we show that high Mg/Ca-calcite forms at night in cultured specimens of the multi-chambered species Neogloboquadrina dutertrei. Our results demonstrate that N. dutertrei adds a significant amount of calcite, and nearly all Mg-bands, after the final chamber forms. These results have implications for interpreting patterns of calcification in N. dutertrei and suggest that diurnal Mg-banding is an intrinsic component of biomineralization in planktic foraminifera.

Nanometer-Scale Chemistry of a Calcite Biomineralization Template: Implications for Skeletal Composition and Nucleation

Significance Many marine organisms build complex CaCO3 shells, which record an archive of past climate in their trace chemistry. Organic–mineral interactions are a crucial, poorly understood aspect of shell formation, which may alter shell composition and bias climate records. We map the chemistry of an organic template preserved within a type of shell that is widely used in studies of past climate. We find that the organic template embedded within this shell is enriched in Na and Mg, and can locally influence shell composition. Atom-scale maps of template chemistry offer a detailed view of the chemical interactions at an organic mineral template, and suggest that elements other than Ca2+ may be important in defining the energetics of CaCO3 nucleation during biomineralization. Plankton, corals, and other organisms produce calcium carbonate skeletons that are integral to their survival, form a key component of the global carbon cycle, and record an archive of past oceanographic conditions in their geochemistry. A key aspect of the formation of these biominerals is the interaction between organic templating structures and mineral precipitation processes. Laboratory-based studies have shown that these atomic-scale processes can profoundly influence the architecture and composition of minerals, but their importance in calcifying organisms is poorly understood because it is difficult to measure the chemistry of in vivo biomineral interfaces at spatially relevant scales. Understanding the role of templates in biomineral nucleation, and their importance in skeletal geochemistry requires an integrated, multiscale approach, which can place atom-scale observations of organic-mineral interfaces within a broader structural and geochemical context. Here we map the chemistry of an embedded organic template structure within a carbonate skeleton of the foraminifera Orbulina universa using both atom probe tomography (APT), a 3D chemical imaging technique with Ångström-level spatial resolution, and time-of-flight secondary ionization mass spectrometry (ToF-SIMS), a 2D chemical imaging technique with submicron resolution. We quantitatively link these observations, revealing that the organic template in O. universa is uniquely enriched in both Na and Mg, and contributes to intraskeletal chemical heterogeneity. Our APT analyses reveal the cation composition of the organic surface, offering evidence to suggest that cations other than Ca2+, previously considered passive spectator ions in biomineral templating, may be important in defining the energetics of carbonate nucleation on organic templates.

Coral calcification feels the acid

As the world’s largest reservoir of exchangeable carbon on millennial timescales, the oceans play a dominant role in global change. Indeed, the oceans currently act as a major sink for anthropogenic carbon (1), partially moderating increases in atmospheric CO2 at the expense of a more acidic ocean. One of the great challenges for the next century is understanding how this shift in seawater chemistry will affect marine systems. Serving as a vivid microcosm for the ocean as whole, coral reefs display the beauty, diversity, and complexity of the ocean, while also exhibiting the ocean’s sensitivity to environmental perturbations. Built from a framework of CaCO3 skeletons, coral reefs are particularly sensitive to ocean acidification because acidified seawater tends to slow skeletal growth (2). Despite the threat posed by ocean acidification to reef health, the detailed mechanisms responsible for this sensitivity are still poorly understood. In PNAS, Venn et al. (3) address a key component of this problem, providing a detailed view of the chemical-scale changes that link coral skeletal growth and ocean acidification.

Growth kinetics and distribution of trace elements in precious corals

The concentration and spatial distribution of major (Ca, Mg) and trace elements (Na, Sr, S, Li, Ba, Pb, and U) in different Corallium skeletons (C. rubrum, C. japonicum, C. elatius, C. konojoi) have been studied by electron microprobe (EMP) and laser ablation inductively coupled plasma mass spectrometry (LA-ICPMS). EMP data show positive Na-Mg and negative Na-S and Mg-S correlations in all skeletons. LA-ICPMS data display additional Sr-Mg, Li-Mg, and U-Mg positive correlations. Medullar zones in the skeletons, corresponding to fast growing zones, are systematically richer in Mg, Na, Sr, Li, and U and poorer in S than the surrounding slow growing zones. These spatial distributions are mostly interpreted in terms of growth kinetics combined with steric effects influencing the incorporation of impurities in biogenic calcites. This interpretation is in agreement with available experimental data on kinetic effects on the incorporation of elements in calcite. At a different scale, annual growth rings in annular slow growing zones show oscillations in Mg, Na, Sr, and S. These chemical oscillations probably result from growth rate variations: fast growth would produce rings enriched in Mg, Sr, and Na, while slow growth would produce rings enriched in Ca, S and organic matter. From previous studies in C. rubrum, the Mg-rich rings would develop during the spring to fall period while the S-rich rings would form immediately after (late fall and winter). Analytical traverses performed in annular zones of different Corallium skeletons indicate that Mg, Na, Sr, Li, and U decrease from core to rim. This observation indicates that radial growth rate decreases as the colony gets older. Contrary to Mg, Na, Sr, Li, S, and U, barium and lead concentrations are identical in medullar and annular zones and appear independent of growth kinetics. Thus, concentrations in Corallium skeletons could provide indications on Ba and Pb contents in the oceans. Barium and lead concentrations are higher in Mediterranean than in Pacific precious corals, these two elements can be used to discriminate C. rubrum from C. japonicum, and contribute enforcing regulations on the trade of precious corals.