Senataxin suppresses the antiviral transcriptional response and controls viral biogenesis.

The human helicase senataxin (SETX) has been linked to the neurodegenerative diseases amyotrophic lateral sclerosis (ALS4) and ataxia with oculomotor apraxia (AOA2). Here we identified a role for SETX in controlling the antiviral response. Cells that had undergone depletion of SETX and SETX-deficient cells derived from patients with AOA2 had higher expression of antiviral mediators in response to infection than did wild-type cells. Mechanistically, we propose a model whereby SETX attenuates the activity of RNA polymerase II (RNAPII) at genes stimulated after a virus is sensed and thus controls the magnitude of the host response to pathogens and the biogenesis of various RNA viruses (e.g., influenza A virus and West Nile virus). Our data indicate a potentially causal link among inborn errors in SETX, susceptibility to infection and the development of neurologic disorders.

Cell- and pathway-specific disruptions in the accumbens of Fragile X mouse.

Fragile X syndrome (FXS) is a genetic cause of intellectual disability and autism spectrum disorder (ASD), associated with social deficits. The mesocorticolimbic system, which includes the prefrontal cortex (PFC), basolateral amygdala (BLA), and nucleus accumbens core (NAcC), is essential for regulating socio-emotional behaviors. We employed optogenetics to compare the functional properties of the BLA→NAcC, PFC→NAcC, and reciprocal PFC↔BLA pathways in Fmr1-/y::Drd1a-tdTomato male mice. In FXS mice, the PFC↔BLA reciprocal pathway was unaffected, while significant synaptic modifications occurred in the BLA/PFC→NAcC pathways. We observed distinct changes in D1 striatal projection neurons (SPNs) and separate modifications in D2 SPNs. In FXS mice, the BLA/PFC→NAcC-D2 SPNs pathways demonstrated heightened synaptic strength. Focusing on the BLA→NAcC pathway, linked to autistic symptoms, we found increased AMPAR and NMDAR currents, and elevated spine density in D2 SPNs. Conversely, the amplified firing probability of BLA→NAcC-D1 SPNs was not accompanied by increased synaptic strength, AMPAR and NMDAR currents, or spine density. These pathway-specific alterations resulted in an overall enhancement of excitatory-to-spike coupling, a physiologically relevant index of how efficiently excitatory inputs drive neuronal firing, in both BLA→NAcC-D1 and BLA→NAcC-D2 pathways. Finally, the absence of FMRP led to impaired long-term depression specifically in BLA→D1 SPNs. These distinct alterations in synaptic transmission and plasticity within circuits targeting the NAcC highlight the potential role of postsynaptic mechanisms in selected SPNs in the observed circuit-level changes. This research underscores the heightened vulnerability of the NAcC in the context of FMRP deficiency, emphasizing its pivotal role in the pathophysiology of FXS.Significance Statement Fragile X Syndrome is a neurodevelopmental disorder characterized by significant emotional dysregulation and social challenges. The mesocorticolimbic system is a key socioemotional regulator. Nevertheless, its functioning in this condition is still poorly understood. Our study investigates connections between the basolateral amygdala (BLA), prefrontal cortex (PFC), and nucleus accumbens core (NAcC). We observed that while the PFC↔BLA reciprocal connections remained unaffected, their projections onto the NAcC showed target cell-specific changes. Specifically, D2 SPNs exhibited increased synaptic transmission and spine density, whereas D1 SPNs showed heightened firing probability and impaired long-term depression, alongside enhanced neuronal firing efficiency in both SPN types. These findings emphasize the NAcC's crucial role as a neurobiological substrate in the pathophysiology of Fragile X Syndrome.

Electromagnetic Forces and Torques: From Dielectrophoresis to Optical Tweezers.

Electromagnetic forces and torques enable many key technologies, including optical tweezers or dielectrophoresis. Interestingly, both techniques rely on the same physical process: the interaction of an oscillating electric field with a particle of matter. This work provides a unified framework to understand this interaction both when considering fields oscillating at low frequencies─dielectrophoresis─and high frequencies─optical tweezers. We draw useful parallels between these two techniques, discuss the different and often unstated assumptions they are based upon, and illustrate key applications in the fields of physical and analytical chemistry, biosensing, and colloidal science.

Dissecting the difference in tree species richness between Africa and South America.

SignificanceOur full-scale comparison of Africa and South America's lowland tropical tree floras shows that both Africa and South America's moist and dry tree floras are organized similarly: plant families that are rich in tree species on one continent are also rich in tree species on the other continent, and these patterns hold across moist and dry environments. Moreover, we confirm that there is an important difference in tree species richness between the two continents, which is linked to a few families that are exceptionally diverse in South American moist forests, although dry formations also contribute to this difference. Plant families only present on one of the two continents do not contribute substantially to differences in tree species richness.

Expanding and improving nanobody repertoires using a yeast display method: Targeting SARS-CoV-2.

COVID-19, caused by the coronavirus SARS-CoV-2, represents a serious worldwide health issue, with continually emerging new variants challenging current therapeutics. One promising alternate therapeutic avenue is represented by nanobodies, small single-chain antibodies derived from camelids with numerous advantageous properties and the potential to neutralize the virus. For identification and characterization of a broad spectrum of anti-SARS-CoV-2 Spike nanobodies, we further optimized a yeast display method, leveraging a previously published mass spectrometry-based method, using B-cell complementary DNA from the same immunized animals as a source of VHH sequences. Yeast display captured many of the sequences identified by the previous approach, as well as many additional sequences that proved to encode a large new repertoire of nanobodies with high affinities and neutralization activities against different SARS-CoV-2 variants. We evaluated DNA shuffling applied to the three complementarity-determining regions of antiviral nanobodies. The results suggested a surprising degree of modularity to complementarity-determining region function. Importantly, the yeast display approach applied to nanobody libraries from immunized animals allows parallel interrogation of a vast number of nanobodies. For example, we employed a modified yeast display to carry out massively parallel epitope binning. The current yeast display approach proved comparable in efficiency and specificity to the mass spectrometry-based approach, while requiring none of the infrastructure and expertise required for that approach, making these highly complementary approaches that together appear to comprehensively explore the paratope space. The larger repertoires produced maximize the likelihood of discovering broadly specific reagents and those that powerfully synergize in mixtures.

Adjuvant treatment with anti-PD-1 in acral melanoma: A nationwide study.

Previous studies demonstrated limited efficacy of immune checkpoint inhibitors in unresectable acral melanoma (AM); it remains unclear how this translates to the adjuvant setting. This study investigates clinical outcomes of acral compared to cutaneous melanoma (CM) patients treated with adjuvant anti-PD-1 after complete resection. All stages III-IV AM and CM patients receiving adjuvant anti-PD-1 after complete resection between 2018 and 2022 were included from the prospective nationwide Dutch Melanoma Treatment Registry. We analyzed recurrence-free survival (RFS), distant metastasis-free survival (DMFS), and overall survival (OS). A multivariable Cox regression analysis of RFS was performed to adjust for potential confounders. We included 1958 (86 AM and 1872 CM) patients. At baseline, AM patients more frequently had KIT mutations, higher disease stages, and Eastern Cooperative Oncology Group Performance Status, and fewer BRAF and NRAS mutations. Median RFS was 14.8 months (95% confidence interval [CI]: 11.5-29.3) in AM and 37.4 months (95% CI: 34.6 to not reached) in CM (p = .002). After correcting for potential confounders, AM remained associated with a higher risk of recurrence (HRadj 1.53; 95% CI: 1.07-2.17; p = .019). Two-year DMFS tended to be worse for AM than for CM: 64.5% versus 79.7% (p = .050). Two-year OS was significantly lower in AM (71.5% vs. 84.3%; p = .027). The results of this study suggest a poorer outcome of adjuvant-treated AM compared to CM. Studies assessing the added value of adjuvant treatment in AM are needed. Future research should investigate alternative treatment strategies to improve outcomes of high-risk AM.

A prediction model for response to immune checkpoint inhibition in advanced melanoma.

Predicting who will benefit from treatment with immune checkpoint inhibition (ICI) in patients with advanced melanoma is challenging. We developed a multivariable prediction model for response to ICI, using routinely available clinical data including primary melanoma characteristics. We used a population-based cohort of 3525 patients with advanced cutaneous melanoma treated with anti-PD-1-based therapy. Our prediction model for predicting response within 6 months after ICI initiation was internally validated with bootstrap resampling. Performance evaluation included calibration, discrimination and internal-external cross-validation. Included patients received anti-PD-1 monotherapy (n = 2366) or ipilimumab plus nivolumab (n = 1159) in any treatment line. The model included serum lactate dehydrogenase, World Health Organization performance score, type and line of ICI, disease stage and time to first distant recurrence-all at start of ICI-, and location and type of primary melanoma, the presence of satellites and/or in-transit metastases at primary diagnosis and sex. The over-optimism adjusted area under the receiver operating characteristic was 0.66 (95% CI: 0.64-0.66). The range of predicted response probabilities was 7%-81%. Based on these probabilities, patients were categorized into quartiles. Compared to the lowest response quartile, patients in the highest quartile had a significantly longer median progression-free survival (20.0 vs 2.8 months; P < .001) and median overall survival (62.0 vs 8.0 months; P < .001). Our prediction model, based on routinely available clinical variables and primary melanoma characteristics, predicts response to ICI in patients with advanced melanoma and discriminates well between treated patients with a very good and very poor prognosis.

BRAF/MEK inhibitor rechallenge in advanced melanoma patients.

BACKGROUND: Effectivity of BRAF(/MEK) inhibitor rechallenge has been described in prior studies. However, structured data are largely lacking. METHODS: Data from all advanced melanoma patients treated with BRAFi(/MEKi) rechallenge were retrieved from the Dutch Melanoma Treatment Registry. The authors analyzed objective response rate (ORR), progression-free survival (PFS), and overall survival (OS) for both first treatment and rechallenge. They performed a multivariable logistic regression and a multivariable Cox proportional hazards model to assess factors associated with response and survival. RESULTS: The authors included 468 patients in the largest cohort to date who underwent at least two treatment episodes of BRAFi(/MEKi). Following rechallenge, ORR was 43%, median PFS was 4.6 months (95% confidence interval [CI], 4.1-5.2), and median OS was 8.2 months (95% CI, 7.2-9.4). Median PFS after rechallenge for patients who discontinued first BRAFi(/MEKi) treatment due to progression was 3.1 months (95% CI, 2.7-4.0) versus 5.2 months (95% CI, 4.5-5.9) for patients who discontinued treatment for other reasons. Discontinuing first treatment due to progression and lactate dehydrogenase (LDH) levels greater than two times the upper limit of normal were associated with lower odds of response and worse PFS and OS. Symptomatic brain metastases were associated with worse survival, whereas a longer treatment interval between first treatment and rechallenge was associated with better survival. Responding to the first BRAFi(/MEKi) treatment was not associated with response or survival. CONCLUSIONS: This study confirms that patients benefit from rechallenge. Elevated LDH levels, symptomatic brain metastases, and discontinuing first BRAFi(/MEKi) treatment due to progression are associated with less benefit from rechallenge. A prolonged treatment interval is associated with more benefit from rechallenge.

Planar 16-band metasurface-enhanced spectral filter for integrated image sensing.

We study theoretically and demonstrate experimentally a 16-band narrow band wavelength selective filter in the near-infrared range. The combination of a pair of distributed Bragg reflectors with a sub-wavelength grating metasurface embedded in the intra-cavity provides a narrow response which can be tuned by adjusting the geometry of the sub-wavelength grating metasurface. The key advantage of this approach is its ease of fabrication, where the spectral response is tuned by merely changing the grating period, resulting in a perfectly planar geometry that can be easily integrated with a broad variety of photodetectors, thus enabling attractive applications such as bio-imaging, time-of-flight sensors and LiDAR. The experimental results are supported by numerical simulations and effective medium theory that unveil the mechanisms that lead to the optical response of the device. It is also shown how the polarization dependence of the structure can be used to determine very accurately the polarization of incoming light.

Phylogenomics of Brachystegia: Insights into the origin of African miombo woodlands.

PREMISE: Phylogenetic approaches can provide valuable insights on how and when a biome emerged and developed using its structuring species. In this context, Brachystegia Benth, a dominant genus of trees in miombo woodlands, appears as a key witness of the history of the largest woodland and savanna biome of Africa. METHODS: We reconstructed the evolutionary history of the genus using targeted-enrichment sequencing on 60 Brachystegia specimens for a nearly complete species sampling. Phylogenomic inferences used supermatrix (RAxML-NG) and summary-method (ASTRAL-III) approaches. Conflicts between species and gene trees were assessed, and the phylogeny was time-calibrated in BEAST. Introgression between species was explored using Phylonet. RESULTS: The phylogenies were globally congruent regardless of the method used. Most of the species were recovered as monophyletic, unlike previous plastid phylogenetic reconstructions where lineages were shared among geographically close individuals independently of species identity. Still, most of the individual gene trees had low levels of phylogenetic information and, when informative, were mostly in conflict with the reconstructed species trees. These results suggest incomplete lineage sorting and/or reticulate evolution, which was supported by network analyses. The BEAST analysis supported a Pliocene origin for current Brachystegia lineages, with most of the diversification events dated to the Pliocene-Pleistocene. CONCLUSIONS: These results suggest a recent origin of species of the miombo, congruently with their spatial expansion documented from plastid data. Brachystegia species appear to behave potentially as a syngameon, a group of interfertile but still relatively well-delineated species, an aspect that deserves further investigations.

Sales revenues for new therapeutic agents approved by the US Food and Drug Administration from 1995 to 2014: a retrospective study.

OBJECTIVES: To analyze worldwide sales of new therapeutic agents and to estimate the time it takes for product sales to exceed industry-wide average drug development costs. METHODS: Data obtained from company reports were analyzed to track worldwide sales of new medicines approved by the US Food and Drug Administration from 1995 to 2014. All sales figures were reported in 2019 US dollars. Kaplan-Meier curves were used to evaluate the time it took for discounted product sales to exceed the average costs associated with developing one new drug (accounting for the costs of failed trials), using published estimates of these costs. RESULTS: Based on data for 361 of 558 new therapeutic agents approved over the study period (median follow-up: 13.2 years), mean sales revenue per product was $15.2 billion through the end of 2019; the median was $6.7 billion. These products jointly generated global sales of $5.5 trillion since approval. Revenues were highly skewed, with the 25 best selling products (7%, 25/361) accounting for 38% of this amount ($2.1 trillion/$5.5 trillion). About 47% of products had discounted sales that exceeded the estimated industry-wide average costs of development within 5 years of approval, and 75% within 10 years. After attributing potential production, marketing, and other costs, these numbers dropped to 21% of products within 5 years of approval, and 46% within 10 years. CONCLUSIONS: Sales of new medicines approved from 1995 to 2014 were highly skewed, but a majority of products had net discounted sales that exceeded the industry-wide average costs of development within 10 years of approval. An understanding of how sales revenues accrue in the years after initial approval, alongside data on business costs, can inform discussions about how to incentivize private investment in innovation while ensuring affordable prices for patients and the health-care system.

Development and characterization of nuclear microsatellite markers for the African walnut Coula edulis Baill (Coulaceae).

PREMISE OF THE STUDY: Coula edulis Baill (Coulaceae) is a common tree species in the Guineo-Congolian forests producing an edible fruit known as African walnut, which is an important food and income resource for rural populations. However, the species suffers from a deficit of natural regeneration. We developed here nuclear microsatellite markers for C. edulis to be able to study the genetic structure of its natural populations and gene flow. METHODS AND RESULTS: A genomic library was obtained using the Illumina platform, and 21 polymorphic microsatellite loci were developed. The polymorphic microsatellites displayed eight to 22 alleles per locus (average: 14.2), with a mean expected heterozygosity ranging from 0.33 to 0.72 in five populations from Central and West Africa. CONCLUSIONS: The high polymorphism of the nuclear microsatellite markers developed makes them useful to investigate gene flow and the organization of genetic diversity in C. edulis, and to assess whether particular genetic resources require conservation efforts.

Contrasting genetic signal of recolonization after rainforest fragmentation in African trees with different dispersal abilities.

Although today the forest cover is continuous in Central Africa, this may have not always been the case, as the scarce fossil record in this region suggests that arid conditions might have significantly reduced tree density during the ice ages. Our aim was to investigate whether the dry ice age periods left a genetic signature on tree species that can be used to infer the date of the past fragmentation of the rainforest. We sequenced reduced representation libraries of 182 samples representing five widespread legume trees and seven outgroups. Phylogenetic analyses identified an early divergent lineage for all species in West Africa (Upper Guinea) and two clades in Central Africa: Lower Guinea-North and Lower Guinea-South. As the structure separating the Northern and Southern clades-congruent across species-cannot be explained by geographic barriers, we tested other hypotheses with demographic model testing using δαδι. The best estimates indicate that the two clades split between the Upper Pliocene and the Pleistocene, a date compatible with forest fragmentation driven by ice age climatic oscillations. Furthermore, we found remarkably older split dates for the shade-tolerant tree species with nonassisted seed dispersal than for light-demanding species with long-distance wind-dispersed seeds. Different recolonization abilities after recurrent cycles of forest fragmentation seem to explain why species with long-distance dispersal show more recent genetic admixture between the two clades than species with limited seed dispersal. Despite their old history, our results depict the African rainforests as a dynamic biome where tree species have expanded relatively recently after the last glaciation.

Resistance of African tropical forests to an extreme climate anomaly.

The responses of tropical forests to environmental change are critical uncertainties in predicting the future impacts of climate change. The positive phase of the 2015-2016 El Niño Southern Oscillation resulted in unprecedented heat and low precipitation in the tropics with substantial impacts on the global carbon cycle. The role of African tropical forests is uncertain as their responses to short-term drought and temperature anomalies have yet to be determined using on-the-ground measurements. African tropical forests may be particularly sensitive because they exist in relatively dry conditions compared with Amazonian or Asian forests, or they may be more resistant because of an abundance of drought-adapted species. Here, we report responses of structurally intact old-growth lowland tropical forests inventoried within the African Tropical Rainforest Observatory Network (AfriTRON). We use 100 long-term inventory plots from six countries each measured at least twice prior to and once following the 2015-2016 El Niño event. These plots experienced the highest temperatures and driest conditions on record. The record temperature did not significantly reduce carbon gains from tree growth or significantly increase carbon losses from tree mortality, but the record drought did significantly decrease net carbon uptake. Overall, the long-term biomass increase of these forests was reduced due to the El Niño event, but these plots remained a live biomass carbon sink (0.51 ± 0.40 Mg C ha-1 y-1) despite extreme environmental conditions. Our analyses, while limited to African tropical forests, suggest they may be more resistant to climatic extremes than Amazonian and Asian forests.

How does V1 population activity inform perceptual certainty?

Neural population activity in sensory cortex informs our perceptual interpretation of the environment. Oftentimes, this population activity will support multiple alternative interpretations. The larger the spread of probability over different alternatives, the more uncertain the selected perceptual interpretation. We test the hypothesis that the reliability of perceptual interpretations can be revealed through simple transformations of sensory population activity. We recorded V1 population activity in fixating macaques while presenting oriented stimuli under different levels of nuisance variability and signal strength. We developed a decoding procedure to infer from V1 activity the most likely stimulus orientation as well as the certainty of this estimate. Our analysis shows that response magnitude, response dispersion, and variability in response gain all offer useful proxies for orientation certainty. Of these three metrics, the last one has the strongest association with the decoder's uncertainty estimates. These results clarify that the nature of neural population activity in sensory cortex provides downstream circuits with multiple options to assess the reliability of perceptual interpretations.

Demonstration of a Plasmonic Nonlinear Pseudodiode.

We demonstrate a nonlinear plasmonic metasurface that exhibits strongly asymmetric second-harmonic generation: nonlinear scattering is efficient upon excitation in one direction, and it is substantially suppressed when the excitation direction is reversed, thus enabling a diode-like functionality. A significant (approximately 10 dB) extinction ratio of SHG upon opposite excitations is measured experimentally, and those findings are substantiated with full-wave simulations. This effect is achieved by employing a combination of two commonly used metals─aluminum and silver─producing a material composition asymmetry that results in a bianisotropic response of the system, as confirmed by performing homogenization analysis and extracting an effective susceptibility tensor. Finally, we discuss the implications of our results from the more fundamental perspectives of reciprocity and time-reversal asymmetry.

Adjuvant treatment of in-transit melanoma: Narrowing the knowledge gap left by clinical trials.

Few clinical trials address efficacy of adjuvant systemic treatment in patients with in-transit melanoma (ITM). This study describes adjuvant systemic therapy of ITM patients beyond clinical trials. In this study, we included stage III adjuvant-treated melanoma patients registered in the nationwide Dutch Melanoma Treatment Registry between July 2018 and December 2020. Patients were divided into three groups: nodal disease only, ITM only and ITM and nodal disease. Recurrence patterns, recurrence-free survival (RFS) and overall survival (OS) at 12-months were analyzed. In our study population of 1037 patients, 66.8% had nodal disease only, 16.7% had ITM only and 16.2% had ITM with nodal disease. RFS at 12-months was comparable in the nodal only and ITM only group (72.2% vs70.1%, P = .97) but lower in ITM and nodal disease patients (57.8%; P = .01, P < .01). Locoregional metastases occurred as first recurrence in 38.9% nodal disease only, 71.9% of ITM-only and 44.0% of ITM and nodal disease patients. Distant recurrences occurred in 42.3%, 18.8% and 36.0%, respectively (P = .02). 12-months OS was not significantly different for nodal disease only patients compared with ITM-only (94.4% vs 97.6%, P = .06) but was significantly higher for ITM-only compared with ITM and nodal disease patients (97.6% vs 91.0%, P < .01). In conclusion, we showed that in the adjuvant setting, RFS rates in ITM-only patients are similar to non-ITM, though better than in ITM and nodal disease patients. Adjuvant-treated ITM-only patients less often experience distant recurrences and have a superior OS compared with ITM and nodal disease patients.

Time interval from primary melanoma to first distant recurrence in relation to patient outcomes in advanced melanoma.

Since the introduction of BRAF(/MEK) inhibition and immune checkpoint inhibition (ICI), the prognosis of advanced melanoma has greatly improved. Melanoma is known for its remarkably long time to first distant recurrence (TFDR), which can be decades in some patients and is partly attributed to immune-surveillance. We investigated the relationship between TFDR and patient outcomes after systemic treatment for advanced melanoma. We selected patients undergoing first-line systemic therapy for advanced melanoma from the nationwide Dutch Melanoma Treatment Registry. The association between TFDR and progression-free survival (PFS) and overall survival (OS) was assessed by Cox proportional hazard regression models. The TFDR was modeled categorically, linearly, and flexibly using restricted cubic splines. Patients received anti-PD-1-based treatment (n = 1844) or BRAF(/MEK) inhibition (n = 1618). For ICI-treated patients with a TFDR <2 years, median OS was 25.0 months, compared to 37.3 months for a TFDR >5 years (P = .014). Patients treated with BRAF(/MEK) inhibition with a longer TFDR also had a significantly longer median OS (8.6 months for TFDR <2 years compared to 11.1 months for >5 years, P = .004). The hazard of dying rapidly decreased with increasing TFDR until approximately 5 years (HR 0.87), after which the hazard of dying further decreased with increasing TFDR, but less strongly (HR 0.82 for a TFDR of 10 years and HR 0.79 for a TFDR of 15 years). Results were similar when stratifying for type of treatment. Advanced melanoma patients with longer TFDR have a prolonged PFS and OS, irrespective of being treated with first-line ICI or targeted therapy.

Protein Dielectrophoresis with Gradient Array of Conductive Electrodes Sheds New Light on Empirical Theory.

Dielectrophoresis (DEP) is a versatile tool for the precise microscale manipulation of a broad range of substances. To unleash the full potential of DEP for the manipulation of complex molecular-sized particulates such as proteins requires the development of appropriate theoretical models and their comprehensive experimental verification. Here, we construct an original DEP platform and test the Hölzel-Pethig empirical model for protein DEP. Three different proteins are studied: lysozyme, BSA, and lactoferrin. Their molecular Clausius-Mossotti function is obtained by detecting their trapping event via the measurement of the fluorescence intensity to identify the minimum electric field gradient required to overcome dispersive forces. We observe a significant discrepancy with published theoretical data and, after a very careful analysis to rule out experimental errors, conclude that more sophisticated theoretical models are required for the response of molecular entities in DEP fields. The developed experimental platform, which includes arrays of sawtooth metal electrode pairs with varying gaps and produces variations of the electric field gradient, provides a versatile tool that can broaden the utilization of DEP for molecular entities.

Is a History of Optimal Staging by Sentinel Lymph Node Biopsy in the Era Prior to Adjuvant Therapy Associated with Improved Outcome Once Melanoma Patients have Progressed to Advanced Disease?

INTRODUCTION: Sentinel lymph node biopsy (SLNB) is important for staging in patients with primary cutaneous melanoma. Did having previously undergone SLNB also affect outcomes in patients once they have progressed to metastatic melanoma in the era prior to adjuvant therapy? METHODS: Data were retrieved from the Dutch Melanoma Treatment Registry, a prospectively collected, nationwide database of patients with unresectable stage IIIC or IV (advanced) melanoma between 2012 and 2018. Melanoma-specific survival (MSS) was compared between patients with advanced cutaneous melanoma, previously treated with a wide local excision (WLE) or WLE combined with SLNB as initial treatment of their primary tumor. Cox regression analyses were used to analyze the influence of different variables on MSS. RESULTS: In total, 2581 patients were included, of whom 1412 were treated with a WLE of the primary tumor alone and 1169 in whom this was combined with SLNB. At a median follow-up of 44 months from diagnosis of advanced melanoma, MSS was significantly longer in patients who had previously undergone SLNB {median 23 months (95% confidence interval [CI] 19-29) vs. 18 months (95% CI 15-20) for patients treated with WLE alone; p = 0.002}. However, multivariate Cox regression did not identify SLNB as an independent favorable prognostic factor for MSS after diagnosis of advanced melanoma. CONCLUSION: Prior to the availability of adjuvant systemic therapy, once patients have unresectable stage IIIC or IV (advanced) melanoma, there was no difference in disease outcome for patients who were or were not previously staged with SLNB.