Abstract Background: Glioblastoma (GBM), a notably aggressive brain cancer, presents significant treatment challenges due to its propensity for deep, diffuse infiltration into healthy brain tissue. This trait enables GBM to resist and survive following surgery and conventional adjuvant therapies. Emerging evidence suggests that signaling between GBM stem cells (GSCs) may be instrumental in this invasive behavior. Our study aimed to unravel the intricacies of these intercellular communication networks within glioblastoma by employing comprehensive proteomic profiling across a diverse array of patient-derived GSC lines. Methods: We conducted our study using a sex-stratified cohort comprising 17 patient-derived GSC lines. Proteomic profiling encompassed analyses of both GSC lysates and conditioned media under various oxygen conditions (normoxia at 20% O2, hypoxia at 1% and 0.2% O2). Additionally, we incorporated the glioma cerebral organoid (GLICO) system, facilitating co-cultures of GSCs with human cerebral organoids. For in-depth proteomic analysis, we utilized advanced TMT-based liquid chromatography-tandem mass spectrometry (LC-MS/MS; Q-Exactive, Thermo Fisher). Results: Our findings corroborate that GSCs adhere to the tripartite model of proliferation, invasion, and hypoxic response. A salient discovery was the consistent enrichment of integrin signaling pathways in GSCs under hypoxic conditions. Specifically, we observed a significant association between upregulation in integrin signaling (BIOCARTA_INTEGRIN_PATHWAY) and elevated KRAS activity (SWEET_KRAS_TARGETS_UP: r=0.43, p<0.01), as well as hypoxic responses (WINTER_HYPOXIA_UP: r=0.76, p<0.0001) in GSCs. Notably, in the GLICO model, both integrin signaling and KRAS pathways were markedly upregulated in the presence of GSCs compared to cerebral organoids alone (p<0.05). Conclusions: This research highlights the pivotal role of integrin signaling in mediating the invasive attributes of glioblastoma. The data underscore the importance of further exploring key modulators within the integrin signaling pathway, presenting a promising avenue for the discovery of novel therapeutic targets in glioblastoma treatment. Citation Format: Lennart J. van Winden, Rifat Shahriar Sajid, Okty Abbasi Borhani, Evelyn Rose Kamski-Hennekam, Nakita Gopal, Alberto Leon, Phedias Diamandis. Global proteomics reveals integrin signaling as a driver of infiltration in glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1842.

The growing number of portable consumer-grade electroencephalography (EEG) wearables offers potential to track brain activity and neurological disease in real-world environments. However, accompanying open software tools to standardize custom recordings and help guide independent operation by users is lacking. To address this gap, we developed HEROIC, an open-source software that allows participants to remotely collect advanced EEG data without the aid of an expert technician. The aim of HEROIC is to provide an open software platform that can be coupled with consumer grade wearables to record EEG data during customized neurocognitive tasks outside of traditional research environments. This article contains a description of HEROIC’s implementation, how it can be used by researchers and a proof-of-concept demonstration highlighting the potential for HEROIC to be used as a scalable and low-cost EEG data collection tool. Specifically, we used HEROIC to guide healthy participants through standardized neurocognitive tasks and captured complex brain data including event-related potentials (ERPs) and powerband changes in participants’ homes. Our results demonstrate HEROIC’s capability to generate data precisely synchronized to presented stimuli, using a low-cost, remote protocol without reliance on an expert operator to administer sessions. Together, our software and its capabilities provide the first democratized and scalable platform for large-scale remote and longitudinal analysis of brain health and disease.

Collection of electroencephalographic (EEG) data provides an opportunity to non-invasively study human brain plasticity, learning and the evolution of various neuropsychiatric disorders. Traditionally, due to sophisticated hardware, EEG studies have been largely limited to research centers which restrict both testing contexts and repeated longitudinal measures. The emergence of low-cost "wearable" EEG devices now provides the prospect of frequent and remote monitoring of the human brain for a variety of physiological and pathological brain states. In this manuscript, we survey evidence that EEG wearables provide high-quality data and review various software used for remote data collection. We then discuss the growing body of evidence supporting the feasibility of remote and longitudinal EEG data collection using wearables including a discussion of potential biomedical applications of these protocols. Lastly, we discuss some additional challenges needed for EEG wearable research to gain further widespread adoption.

Background: The correlation between blood-based tumor mutation burden (bTMB) and tissue-based tumor mutation burden(tTMB) has not been broadly tested in a multicancer cohort. Here, we assess the correlation between bTMB with tTMB in phase I trial patients treated with immunotherapy. As an exploratory analysis, we evaluated circulating tumor DNA (ctDNA) dynamics in responders. Methods: Patients treated with immunotherapy at the Princess Margaret phase I trials unit were enrolled. Pretreatment plasma ctDNA and matched normal blood controls were collected. Available archival tissue formalin-fixed paraffin-embedded (FFPE) samples were analyzed. A 425-gene panel was used to sequence both ctDNA and FFPE samples. Samples with TMB within the highest tertile were considered as high TMB. Results: Thirty-eight patients were accrued from 25 different trials, 86.8% of which involved an anti-PD-1/PD-L1 agent. Thirty patients (78.9%) had detectable mutations in ctDNA, of which the median (range) bTMB was 5 (1-53) mutations per megabase (mut/Mb). Of the 22 patients with available FFPE samples, mutations were detected in 21 (95.4%); the median (range) tTMB was 6 (2-124) mut/Mb. Among the 16 patients with detectable mutations in both FFPE and ctDNA, a statistically significant correlation between bTMB and tTMB was observed (rho = 0.71; P = .002). High TMB was not associated with better survival. All 3 responders had a decrease in the variant allele frequency of mutations detected in ctDNA at a second timepoint relative to baseline, indicating a potential early marker of response. Conclusions: In this small series, bTMB correlated with tTMB. An on-treatment decrease in VAF of mutations detected in ctDNA at baseline was observed in responders. Larger studies to verify our findings are warranted.

Abstract Background In high grade serous ovarian cancer (HGSOC), there is a spectrum of sensitivity to first line platinum‐based chemotherapy. This study molecularly characterizes HGSOC patients from two distinct groups of chemotherapy responders (good vs. poor). Methods Following primary debulking surgery and intravenous carboplatin/paclitaxel, women with stage III–IV HGSOC were grouped by response. Patients in the good response (GR) and poor response (PR) groups respectively had a progression‐free intervals (PFI) of ≥12 and ≤6 months. Analysis of surgical specimens interrogated genomic and immunologic features using whole exome sequencing. RNA‐sequencing detected gene expression outliers and inference of immune infiltrate, with validation by targeted NanoString arrays. PD‐L1 expression was scored by immunohistochemistry (IHC). Results A total of 39 patient samples were analyzed (GR = 20; PR = 19). Median PFI for GR and PR patient cohorts was 32 and 3 months, respectively. GR tumors were enriched for loss‐of‐function BRCA2 mutations and had a significantly higher nonsynonymous mutation rate compared to PR tumors (p = 0.001). Samples from the PR cohort were characterized by mutations in MGA and RAD51B and trended towards a greater rate of amplification of PIK3CA, MECOM, and ATR in comparison to GR tumors. Gene expression analysis by NanoString correlated increased PARP4 with PR and increased PD‐L1 and EMSY with GR. There was greater tumor immune cell infiltration and higher immune cell PD‐L1 protein expression in the GR group. Conclusions Our research demonstrates that tumors from HGSOC patients responding poorly to first line chemotherapy have a distinct molecular profile characterized by actionable drug targets including PARP4.

BACKGROUND:Both genetic and methylation analysis have been shown to provide insight into the diagnosis and prognosis of many brain tumors. However, the implication of methylation profiling and its interaction with genetic alterations in pediatric low-grade gliomas (PLGGs) are unclear. METHODS:We performed a comprehensive analysis of PLGG with long-term clinical follow-up. In total 152 PLGGs were analyzed from a range of pathological subtypes, including 40 gangliogliomas. Complete molecular analysis was compared with genome-wide methylation data and outcome in all patients. For further analysis of specific PLGG groups, including BRAF p.V600E mutant gliomas, we compiled an additional cohort of clinically and genetically defined tumors from 3 large centers. RESULTS:Unsupervised hierarchical clustering revealed 5 novel subgroups of PLGG. These were dominated by nonneoplastic factors such as tumor location and lymphocytic infiltration. Midline PLGG clustered together while deep hemispheric lesions differed from lesions in the periphery. Mutations were distributed throughout these location-driven clusters of PLGG. A novel methylation cluster suggesting high lymphocyte infiltration was confirmed pathologically and exhibited worse progression-free survival compared with PLGG harboring similar molecular alterations (P = 0.008; multivariate analysis: P = 0.035). Although the current methylation classifier revealed low confidence in 44% of cases and failed to add information in most PLGG, it was helpful in reclassifying rare cases. The addition of histopathological and molecular information to specific methylation subgroups such as pleomorphic xanthoastrocytoma-like tumors could stratify these tumors into low and high risk (P = 0.0014). CONCLUSION:The PLGG methylome is affected by multiple nonneoplastic factors. Combined molecular and pathological analysis is key to provide additional information when methylation classification is used for PLGG in the clinical setting.

Whole genome and transcriptome sequencing (WGTS) enables comprehensive detection of cancer genome variation from a single cancer specimen, however, clinical reporting of these assays is challenged by the need for significant computational infrastructure and breadth of genomic algorithms needed for analysis. Here, we report on the deployment of a clinical bioinformatics process for comprehensive genomic profiling of a patient's tumour.

5561 Background: Patients with advanced high grade serous ovarian cancer (SOC) who undergo a suboptimal debulking primary surgery typically have adverse clinical outcomes. However, a spectrum of sensitivity to first line platinum-based chemotherapy is observed but poorly understood. In this study, we perform molecular characterization of two groups of responders (extreme versus poor) to first line carboplatin/taxol chemotherapy in suboptimally debulked SOC patients. Methods: Suboptimally debulked SOC patients with advanced disease (stage III-IV) were grouped by response to first-line chemotherapy and clinicopathologic data collected. Extreme platinum-sensitive (PS) responders had a PFI (progression-free interval) > 12 months (mo) and platinum-resistant (PR) responders had a PFI < 6mo. Tissue specimens were used to interrogate the molecular features of both PS and PR cohorts using whole exome and transcriptome sequencing. Sequence alignment and variant calling were performed using GATK and annotation was performed using Variant Effect Predictor for assessment of non-synonymous tumor mutation burden (TMB) and discovery of novel mutational signatures to predict platinum response. Results: There were 39 patient samples analyzed from primary surgery (PS group = 20; PR group = 19). Median PFI for PS and PR patient cohorts was 30 mo and 3 mo (p < 0.001), respectively. In all tumors, in addition to BRCA and TP53 mutations, additional oncogenic mutations were noted in genes associated with PI3K/AKT/mTOR signaling and in epigenetic regulation. The PS samples were characterized by mutations in BRCA1/2 and the PR samples by mutations in MGA. Compared to tumors in the PR cohort, PS tumors had a significantly higher non-synonymous mutation rate using TMP analysis (p < 0.05) with a trend towards increased immune response. Additional bioinformatics analysis is ongoing and will include copy number variation analysis, immune inference using ESTIMATE and Gene Set Enrichment Analysis. Conclusions: Contracting a mutational signature is feasible from patient tumors at primary surgery and helps to elucidate extreme responses to platinum-based chemotherapy.

Sunflower (Helianthus annuus L.) seed oil with high palmitic acid content has enhanced thermo-oxidative stability, which makes it well suited to high-temperature uses. CAS-5 is a sunflower mutant line that accumulates over 25 % palmitic acid in its seed oil, compared to 5–8 % in conventional cultivars. The objective of this study was to investigate the molecular basis of the high-palmitic acid trait in CAS-5 through both candidate gene and QTL mapping approaches. An F2 population derived from the cross between CAS-5 and the conventional line HA-89 was developed. A 3-ketoacyl-ACP synthase II (KASII) locus on a telomeric region of linkage group (LG) 9 of the sunflower genetic map was found to co-segregate with palmitic acid content in this population. The KASII locus explained the vast majority of the phenotypic variation (98 %) of the trait. Two minor QTL affecting palmitic acid content were also found on the lower half of LG 9 and on LG 17. Additionally, QTL associated with other major fatty acids (stearic, oleic, and linoleic acid) were identified on LG 1, 6, and 10. This result may reflect untapped genetic variation that could exist among sunflower cultivars for genes determining fatty acid composition. In addition to demonstrating the major role of a KASII locus in the accumulation of high levels of palmitic acid in CAS-5 seeds, this study stressed the importance of characterizing genes with minor effects on fatty acid profile in order to establish optimal breeding strategies for modifying fatty acid composition in sunflower seed oil.

Genetic resistance to broomrape (Orobanche cumana Wallr.) in sunflower is mainly monogenic and dominant. Massive use of vertical resistance has led to the progressive extension of increasingly virulent races of the parasite. Additional introduction of horizontal resistance genes is crucial to develop more durable resistance. The objective of this research was to study the inheritance of resistance to broomrape race F in sunflower line K-96 and to identify QTL of potential value for marker assisted pyramiding of resistance genes. The inheritance of broomrape resistance was studied in crosses with the susceptible line P-21 and the line P-96, with oligogenic recessive resistance. Crosses with P-96 revealed broad transgressive segregation for susceptibility in the F2, indicating that both lines possess different resistance alleles. Crosses with P-21 suggested that the trait is mainly controlled by a dominant-recessive epistasis at two loci. Additionally, segregation for plant height was identified and measured in a non-inoculated F2 population. Five QTL on LG 2, 3, 4, 5, and 6 were associated with broomrape resistance traits. Two of them at LG 4 and 5 were also associated with plant height, suggesting an alleged pleiotropic effect of plant height on broomrape resistance. The latter two QTL had been previously identified in the cross P-21 × P-96, whereas the other QTL seem to be involved in K-96 but not in P-96 resistance. This study concluded that K-96 and P-96 have complementary QTL with minor effect on broomrape resistance. They are, therefore, good donor sources for marker-assisted pyramiding programs.

Considering the vast knowledge gained in crop physiology, breeding and biotechnology, a framework integrating these complementary disciplines could be important to achieve steeper gains in plant breeding. In this context, this chapter focuses on the contribution of crop physiology to plant breeding and biotechnology. Our aim is to highlight the value of crop physiology to the understanding and interpretation of complex genetic and physiological mechanisms with strong interaction with the environment. We discuss the contribution of the discipline in (1) analyzing past achievements of plant breeding, (2) identifying relevant traits for yield potential and yield stability across environments, and (3) disentangling complex interactions between traits and the environment, and between relevant traits. A major challenge and a limiting factor for the success of the interdisciplinary approach is the identification of the main traits that confer yield potential and yield stability in different environments. In this context, crop physiology is crucial to understanding and extrapolating processes and mechanisms occurring at different levels of organization and to explain and predict complex interactions. We also discuss the potential contributions of biotechnology to facilitate crop physiology studies as well as the use of physiological traits in plant breeding. Evidence is emerging on the contribution of this type of integrated multidisciplinary framework to higher and more stable yields.

Mutagenesis has gained popularity in plant genetics research in the last century as a powerful tool for inducing genetic variation. Induced mutations have been applied to produce mutant cultivars in sunflower, significantly increasing crop yield and quality, and many characteristics of modern genotypes have been obtained through this approach. Induced mutations were widely used in sunflower breeding programs due to its low cost and technology requirement, and its adaptation to a wide spectrum of breeding objectives. Mutagenesis does not substitute for conventional breeding methods, but rather complements them. Mutations can be classified as: (1) point mutations, (2) structural mutations, or (3) mutations changing the chromosome number. The most usual kind of point mutation is the base pair substitution, which can result in a changed codon or a changed amino acid. Mutagenesis has allowed for the development of several features of modern sunflower genotypes, but it has been particularly important for the progress of the sunflower oil market and in the creation of herbicide-resistant genotypes.

 Verticillium dahliae (Kleb) is a soil-borne pathogen widely distributed which causes the premature death and stem broken of sunflower. In Argentina this disease is considered a major problem that produces up to 30 percent of yield reduction in susceptible commercial hybrids. In 2004, two main physiological Argentinean races (Varg1 and Varg2) were described. The objective of this work was to increase V. dahliae resistance by pyramiding of resistance QTLs to the main pathogenic races of Argentina.  Sources of resistance were detected for each race and the quantitative trait loci (QTL) were located in different chromosomes by mapping populations. Near isogenic lines (NILs) with the resistance QTL for each race were obtained from backcross program with molecular markers and phenotypic selection. NILs (L1: Varg1 resistance, L2: Varg2 resistance), families F3 and F4 (from a cross between L1 and L2) and the susceptible line were evaluated under natural infection conditions and artificial inoculation in growth chamber, with each separately race.  Typical disease symptoms were observed at flowering stage in the field. The susceptible line had 1753 % of incidence, L1 showed 0-6 % and L2 had 5-27 % of diseased plants. Fifty percent of F3 families with both QTLs were totally resistant and the rest exhibited 6-19 % susceptible plants. When the isolines were artificially inoculated, they had the expected behavior according to the race used in each trial. On the other hand, F4 selected families with both QTLs were resistant to both races. No significant differences were observed between selected families and the original line with respect to oil % and yield, in healthy plants.  These results show the presence of different races of this fungus in the field and reveal that the combined QTLs increase the level of resistance to the disease. Nevertheless the search of resistance sources to minor races is being investigated.  This research shows the importance of pyramiding QTLs, with the assistance of molecular tools and phenotypic selection, in breeding programs to obtain a more durable resistance in sunflower materials without yield drag.

Background: Sunflower (Helianthus annuus L.) is an important oilseed crop grown widely in various areas of the world. Classical genetic studies have been extensively undertaken for the improvement of this particular oilseed crop. Pertaining to this endeavor, we developed a "chemically induced mutated genetic resource for detecting SNP by TILLING" in sunflower to create new traits.Results: To optimize the EMS mutagenesis, we first conducted a "kill curve" analysis with a range of EMS dose from 0.5% to 3%. Based on the observed germination rate, a 50% survival rate i.e. LD50, treatment with 0.6% EMS for 8 hours was chosen to generate 5,000 M2 populations, out of which, 4,763 M3 plants with fertile seed set. Phenotypic characterization of the 5,000 M2 mutagenised lines were undertaken to assess the mutagenesis quality and to identify traits of interest. In the M2 population, about 1.1% of the plants showed phenotypic variations. The sunflower TILLING platform was setup using Endo-1-nuclease as mismatch detection system coupled with an eight fold DNA pooling strategy. As proof-of-concept, we screened the M2 population for induced mutations in two genes related to fatty acid biosynthesis, FatA an acyl-ACP thioesterase and SAD the stearoyl-ACP desaturase and identified a total of 26 mutations.Conclusion: Based on the TILLING of FatA and SAD genes, we calculated the overall mutation rate to one mutation every 480 kb, similar to other report for this crop so far. As sunflower is a plant model for seed oil biosynthesis, we anticipate that the developed genetic resource will be a useful tool to identify novel traits for sunflower crop improvement.

Ninety-seven inbred lines of sunflower were screened in the field by treatment with a combination of imazamox and malathion, an inhibitor of cytochrome P450 monooxygenases (P450s), to identify sunflower lines with natural tolerance to the herbicide reversed by malathion. One tolerant line, named TolP450-1, was selected and characterized in the field and in the greenhouse to evaluate its response to the herbicides imazamox, prosulfuron, and atrazine at different plant development stages (germination, emergence, and third difoliate) with and without malathion. For all herbicides and all development stages analyzed, TolP450-1 showed significantly higher tolerance compared with the susceptible line RHA266. In all cases, the tolerance was reversed by malathion. This sunflower line, tolerant to multiple herbicides, may be useful in helping to manage herbicide-resistant weeds by allowing additional herbicides to be used in this oilseed crop.

Nucleotide binding site-leucine rich repeat (NBS-LRR) proteins are encoded by a ubiquitous gene family in sunflower and frequently harbor disease resistance genes. We investigated NBS-LRR-encoding resistance gene candidates (RGCs) flanking the downy mildew resistance genes Pl (8) and Pl (14) and the rust resistance gene R (Adv) , which map on the NBS-LRR clusters of linkage groups 1 and 13 in sunflower genome. We shotgun sequenced bacterial artificial chromosome (BAC) clones proximal to Pl (8) , Pl (14) , and R (Adv) and identified seven novel non-Toll/interleukin-1 receptor (TIR)-like NBS-LRR RGCs, which clustered with previously identified RGCs of linkage group 13 but were phylogenetically distant from the TIR- and non-TIR-NBS-LRR-encoding superfamilies of sunflower. Six of the seven predicted RGCs have intact open reading frames and reside in genomic segments with abundant transposable elements. The genomic localization and sequence similarity of the novel non-TIR-like predicted RGCs suggests that they originated from tandem duplications. RGCs in the proximity of Pl (8) and R (Adv) were likely introgressed from silverleaf sunflower genome, where the RGC cluster of linkage group 13 is duplicated in two independent chromosomes that have different architecture and level of recombination from the respective common sunflower chromosomes.

The discovery of unbranched, monocephalic natural variants was pivotal for the domestication of sunflower (Helianthus annuus L.). The branching locus (B), one of several loci apparently targeted by aboriginal selection for monocephaly, pleiotropically affects plant, seed and capitula morphology and, when segregating, confounds the discovery of favorable alleles for seed yield and other traits. The present study was undertaken to gain deeper insights into the genetics of branching and seed traits affected by branching. We produced an unbranched hybrid testcross recombinant inbred line (TC-RIL) population by crossing branched (bb) and unbranched (BB) RILs to an unbranched (BB) tester. The elimination of branching concomitantly eliminated a cluster of B-linked seed trait quantitative trait loci (QTL) identified by RIL per se testing. We identified a seed oil content QTL linked in repulsion and a 100-seed weight QTL linked in coupling to the B locus and additional unlinked QTL, previously masked by B-locus pleiotropy. Genomic segments flanking the B locus harbor multiple loci for domestication and post-domestication traits, the effects of which are masked by B-locus pleiotropy in populations segregating for branching and can only be disentangled by genetic analyses in unbranched populations. QTL analyses of NILs carrying wild B alleles substantiated the pleiotropic effects of the B locus. The effect of the B locus on branching was masked by the effects of wild alleles at independent branching loci in hybrids between monocephalic domesticated lines and polycephalic wild ecotypes; hence, the B locus appears to be necessary, but not sufficient, for monocephaly in domesticated sunflower.

Sunflower (Helianthus annuus L.) crops are affected by Sunflower chlorotic mottle virus (SuCMoV) which reduces yield parameters in commercial hybrids infected at early ontogenic stages. Sunflower isohybrids (between two near isogenic males differing in the Rcm-1 resistance gene and three different females) were mechanically inoculated with SuCMoV under field conditions in two locations (Balcarce and Venado Tuerto) and evaluated for symptom expression and agronomic characteristics. Symptoms were scarce chlorotic pinpoint in all resistant hybrid combinations and severe chlorotic mottling in all susceptible hybrids, independently of the female parents. Nevertheless, higher symptoms intensity was detected in Venado Tuerto. Morphological parameters were more affected in Venado Tuerto than in Balcarce and differed among hybrid combinations.