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. The mesocorticolimbic system, which includes the prefrontal cortex (PFC), basolateral amygdala (BLA), and nucleus accumbens core (NAcC), is essential for regulating socioemotional 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 SPN 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 fragile X messenger ribonucleoprotein 1 (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.

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.

Visual and Quantitative Analysis of the Trapping Volume in Dielectrophoresis of Nanoparticles.

Nanoparticle manipulation requires careful analysis of the forces at play. Unfortunately, traditional force measurement techniques based on the particle velocity do not provide sufficient resolution, while balancing approaches involving counteracting forces are often cumbersome. Here, we demonstrate that a nanoparticle dielectrophoretic response can be quantitatively studied by a straightforward visual delineation of the dielectrophoretic trapping volume. We reveal this volume by detecting the width of the region depleted of gold nanoparticles by the dielectrophoretic force. Comparison of the measured widths for various nanoparticle sizes with numerical simulations obtained by solving the particle-conservation equation shows excellent agreement, thus providing access to the particle physical properties, such as polarizability and size. These findings can be further extended to investigate various types of nano-objects, including bio- and molecular aggregates, and offer a robust characterization tool that can enhance the control of matter at the nanoscale.

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.

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.

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.

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.

Inbreeding depression affects the growth of seedlings of an African timber species with a mixed mating reproductive system, Pericopsis elata (Harms) Meeuwen.

Selfing or mating between related individuals can lead to inbreeding depression (ID), which can influence the survival, growth and evolution of populations of tree species. As selective logging involves a decrease in the density of congeneric partners, it could lead to increasing biparental inbreeding or self-fertilization, exposing the population to higher ID. We assessed the influence of inbreeding on the growth of a commercial timber species, Pericopsis elata (Fabaceae), which produced about 54% of self-fertilized seedlings in a natural population of the Congo basin. We followed the survival and growth of 540 plants raised in a plantation along a gradient of plant density (0.07-15.9 plants per m2). Parentage analysis allowed us distinguishing selfed and outcrossed seedlings. The annual growth was higher for outcrossed than selfed plants, on average by 10.8% for diameter and 12.9% for height growth. Based on the difference in above ground biomass between selfed and outcrossed seedlings after 41 months, we estimated the level of ID at δ = 0.33, while a lifetime estimate of ID based on the proportions of selfed plants at seedling and adult stages led to δ = 0.7. The level of ID on growth rate did not change significantly with age but tended to vanish under high competition. Pericopsis elata is a particularly interesting model because inbreeding depression is partial, with about 26% of reproducing adults resulting from selfing, contrary to most tropical tree species where selfed individuals usually die before reaching adulthood. Hence, the risks of ID must be considered in the management and conservation of the species.

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 United States Food and Drug Administration From 1995 to 2014.

OBJECTIVES: This study aimed 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 1 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 of 361) accounting for 38% of this amount ($2.1 trillion of $5.5 trillion). Approximately 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 many 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 healthcare system.

Integrating Price Benchmarks and Comparative Clinical Effectiveness to Inform the Medicare Drug Price Negotiation Program.

OBJECTIVES: By September 2024, the Centers for Medicare and Medicaid Services (CMS) will publicly report the negotiated prices (Maximum Fair Prices) for the first 10 drugs selected for price negotiation. We estimate initial price offers based on net prices, statutorily defined ceilings, and comparative effectiveness data for the 10 drugs and their therapeutic alternatives. METHODS: We utilized net prices and other price benchmarks for the 10 drugs and their therapeutic alternatives. We searched for data on comparative clinical effectiveness for the primary indications. We outlined a range of plausible initial price offers based on CMS guidance and our interpretation of regulatory intent. RESULTS: For ibrutinib and ustekinumab, statutorily defined ceiling prices will likely determine the initial price offers. The integration of net pricing and clinical evidence from comparator branded products will inform the initial price offers for apixaban, empagliflozin, etanercept, and insulin aspart. Rivaroxaban and sacubitril/valsartan have therapeutic alternatives that are generics; therefore, CMS may apply a discount to current net prices. To achieve savings in the negotiation of dapagliflozin and sitagliptin, CMS will have to leverage additional negotiation factors because statutory defined ceilings and net prices of therapeutic alternatives are similar or higher. CONCLUSIONS: This analysis sheds light on important price benchmarks and clinical evidence factors for the determination of the initial price offers. Although we were not able to simulate the offer and counter-offer process, our findings provide a transparent and systematic way to produce initial offers that are consistent with CMS guidance.

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.