Abstract Introduction: Cyclin-dependent kinase 4/6 inhibitors (CDK4/6i) in combination with endocrine therapy (ET) represent the standard of care for patients with oestrogen receptor-positive (ER+) metastatic breast cancer (MBC). However, a significant proportion of patients either present intrinsic resistance or acquired resistance over time, limiting the clinical benefit of these treatments. Understanding the molecular mechanisms driving resistance and identifying effective second-line therapies remain major clinical challenges. Objectives: The main objectives of this study are: 1) To generate and characterise patient-derived xenograft (PDX) models from tumor biopsies collected before and after treatment with CDK4/6i+ET. 2) To investigate mechanisms of intrinsic and acquired resistance to CDK4/6i+ET using genomic, transcriptomic, proteomic and epigenetic approaches. Methodology: A total of 17 PDX models were established from ER+ breast cancer patients treated with CDK4/6i+ET. These models were comprehensively characterised through targeted exome sequencing (IMPACT-MSKCC), PAM50-based transcriptomic classification, and immunohistochemistry. To evaluate drug response, PDXs were treated with CDK4/6i (Ribociclib or Abemaciclib) in combination with ET (Letrozole + ovariectomy or the oral SERD Elacestrant). Models that initially responded were further chronically exposed to treatment to induce acquired resistance, mimicking clinical progression. Results: Among the 17 PDX models, 8 showed intrinsic resistance while 9 were initially sensitive to CDK4/6i+ET. Acquired resistance was successfully generated in several initially sensitive models through continuous in vivo drug exposure. Molecular profiling identified both known and novel alterations associated with resistance, including mutations in NOTCH2, supporting its potential role as a resistance driver. These findings highlight the heterogeneity of resistance mechanisms and underscore the importance of individualised approaches. Conclusions: PDX models derived from ER+ MBC patients constitute a valuable preclinical platform to explore both intrinsic and acquired resistance to CDK4/6i+ET. These models not only recapitulate clinical phenotypes but also enable the identification of predictive biomarkers and the testing of targeted therapies. Our study provides mechanistic insights and supports the use of PDXs to develop personalised treatment strategies aimed at overcoming resistance in hormone receptor-positive metastatic breast cancer. Citation Format: C. Molina, L. Monserrat, A. Òdena, C. Viaplana, F. Brasó-Maristany, S. Chandarlapaty, A. Prat, M. Bellet, C. Saura, V. Serra, M. Malumbres. Preclinical Modeling of CDK4/6 Inhibitor and Endocrine Therapy Resistance in ER+ Metastatic Breast Cancer Using Patient-Derived Xenografts [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PD3-03.

Alport syndrome is an inherited kidney disease caused by pathogenic variants in COL4A3, COL4A4, or COL4A5, encoding type IV collagen chains of the glomerular basement membrane. Genetic confirmation is essential for its diagnosis, prognosis, and donor selection; however, conventional sequencing may fail to detect pathogenic variants, leaving a subset of clinically suspected patients without a genetic diagnosis. We report a 16-year-old girl with persistent hematuria and proteinuria, initially diagnosed with Alport syndrome at 2years of age based on kidney biopsy findings of a basket-weave glomerular basement membrane and mosaic α5(IV) staining. Despite a family history compatible with this diagnosis, targeted exome sequencing at 8years of age did not identify pathogenic variants. Whole-genome sequencing at 16years revealed a deep intronic variant in COL4A5 (NM_000495.5) (c.276 + 1306G > A). In silico analysis with SpliceAI predicted the creation of de novo acceptor and donor splice sites, leading to the inclusion of a 103-bp cryptic exon within intron 4. A minigene assay confirmed aberrant splicing and demonstrated that the inserted exon resulted in a frameshift and premature stop codon (p.Gly93Phefs*99). This case highlights the diagnostic utility of whole-genome sequencing in combination with splicing prediction and functional validation to resolve suspected cases of Alport syndrome in which the genetic cause has remained unclear. As sequencing technologies advance, such integrative approaches may become increasingly important in clinical practice, thereby improving diagnostic yield and supporting the development of RNA-based therapies.

Multifocality in sporadic medullary thyroid carcinoma is associated with low tumor grade and no difference in somatic driver mutations or overall survival when compared with unifocal tumors.

Background: Familial hypercholesterolemia (FH) is a genetic disorder characterized by elevated low-density lipoprotein cholesterol (LDL-C) levels, leading to premature cardiovascular disease (CVD). This study aimed to identify genetic variants associated with FH in patients from Telangana State, India.Methods: Probands with suspected FH were identified using the Dutch Lipid Clinic Network (DLCN) score, followed by cascade screening of their first-degree relatives. Targeted exome sequencing and pedigree analysis were performed to identify FH-associated genetic variants.Results: We identified both novel and known high-impact mutations in genes implicated in FH pathogenesis, including stop-gain mutations in LPL (6/30; 20%) and LDLR (4/30; 13.3%), as well as splice donor site mutations in SLCO1B1 (1/30; 3.3%) and CETP (3/30; 10%). Notably, a novel frameshift mutation in LDLR was identified in two siblings (2/30; 6.7%), one of whom (50%) exhibited a homozygous variant and met the "Definite FH" classification based on the DLCN criteria. Additionally, moderate-impact variants rs2075291 (APOA5) and rs193922571 (LDLR) showed strong correlations with the DLCN score, suggesting increased susceptibility to FH. In contrast, rs6756629 (ABCG5) and rs11887534 (ABCG8) were strongly negatively correlated with LDL-C levels and the DLCN score, indicating potential protective effects against FH.Conclusions: These findings highlight the genetic heterogeneity of FH and emphasize the importance of identifying novel pathogenic variants. Moreover, the study underscores the role of moderate-impact variants in FH susceptibility. Overall, this research enhances our understanding of the genetic landscape of FH in the Indian population, with implications for improved diagnosis, risk assessment, and personalized management.

Clinical, cytogenetic and molecular predictors of early mortality in acute promyelocytic leukemia (APL): Real-world analysis

Clinicopathologic, Immunohistochemical, and Molecular Analysis of Primary Ovarian Carcinoid Tumors With Correlation of Ki67 Proliferation Index With Patient Outcomes.

Genomic and Epigenomic Signatures Can Distinguish Aggressive Chromophobe Renal Cell Carcinoma from Indolent Renal Oncocytic Tumors in Clinical-grade Samples.

A teenage girl born to non-related parents presented with progressive weakness in her lower limbs followed by her upper limbs over the past 5 years. Proximal muscle weakness was more significant than distal muscle weakness. Examination revealed weakness in both proximal and distal muscles, with an asymmetric distribution, more pronounced on the right side and in the lower limbs compared with the upper limbs. She also had mild scoliosis. Tests revealed that serum creatine phosphokinase was normal. Muscle biopsy revealed the presence of reducing bodies, as demonstrated by menadione nitro blue tetrazolium (MNBT) staining. The targeted exome sequencing identified a pathogenic missense mutation c.311G>A (p.C104Y) in the FHL1 gene. Although her clinical signs indicated muscle disease, her serum creatine phosphokinase levels were normal. Nonetheless, a muscle biopsy indicated myopathy with reducing bodies, leading to a diagnosis of FHL1-related reducing body myopathy confirmed by genetic testing.

Disclosure: M.F. Stecchini: None. A.B. Arruda: None. J. Marrero-Gutiérrez: None. M.R. Campos: None. I.A. Cardinalli: None. C.A. Scrideli: None. C.A. molina: None. S. Tucci: None. F.B. Coeli-Lacchini: None. A.C. Moreira: None. L.N. Ramalho: None. S.R. Brandalise: None. A.J. Yunes: None. R.Z. Vencio: None. F. Chahud: None. M. De Castro: None. S.R. Rauber Antonini: None.Background: ATRX protein, a chromatin remodeler and transcriptional regulator, plays a crucial role in essential cellular pathways by cooperating with DAXX and promoting histone modifications in the telomere area. Abnormal functioning ATRX and DAXX, as well as in telomeres, have been recognized as important elements for tumorigenesis and prognosis in different types of cancer. However, their roles in pediatric adrenocortical tumors (pACT) require broader investigation. Objective: To evaluate the presence and clinical significance of ATRX, DAXX, and TERT genotype, gene expression, protein expression, telomere length, and the alternative lengthening of telomeres (ALT) mechanism in primary pACT samples. Methods: Tumor ATRX, DAXX, and TERT genotypes were assessed by targeted exome sequencing and their relative gene expression by qPCR. IHC assessed ATRX and DAXX protein expression. Telomere length was assessed by qPCR and ALT by FISH. Normal adrenal cortices were used as controls. Patients' progression-free (PFS) and overall survival (OS) were calculated using Kaplan-Meier curves. A significance level of α=0.05 was used. Results: Most of the analyzed 106 patients with pACT were female (71.7%) and harbored the TP53 p.R337H germline variant (84.9%). The median age at the diagnosis was 22.1 months. Age at diagnosis below four years was associated with a better prognosis, while advanced disease (IPACTR stages 3 and 4) and Wieneke score higher than three were associated with a worse prognosis. ATRX and TERT somatic variants were identified in 46% and 36% of the pACT, all from TP53 p.R337H germline variant carriers, and harbored the corresponding somatic TP53 variant. No DAXX variants were identified. ATRX variants were not associated with ATRX gene expression. None of the samples expressed TERT. Compared to control adrenals, ATRX gene expression was decreased (p<0.01), while DAXX gene expression was increased (p<0.01) in pACT. ATRX protein expression was lost in most samples (95.6%) but was positive in two wild-type samples. DAXX protein expression was lost in a minority (22.2%). There was a weak negative correlation between ATRX (ρ=-0.35; p=0.008) and DAXX (ρ=-0,3; p=0.023) gene expression and telomere length. ALT was present in 27.3% of the ACT. ATRX or TERT variants were not associated with telomere length nor with ALT. Patients whose tumors had both ATRX and TERT variants (n=5) had reduced PFS (p=0.03) and OS (p=0.01). There was no association between gene/protein expression, telomere length, or ALT, and PFS or OS. Conclusions: Abnormal ATRX and DAXX gene and protein expression are frequent in pACT and appear to influence telomere length but not the patient's clinical outcome. However, the co-occurrence of somatic variants in ATRX and TERT is associated with an unfavorable prognosis for these patients.Presentation: Saturday, July 12, 2025

Abstract INTRODUCTION: Young-onset pancreatic cancer is increasing in incidence. Age is the strongest risk factor for pancreatic cancer; yet, tobacco smoking and inherited genetic factors are known to influence disease risk, particularly in younger individuals. METHODS: To investigate how aging, tobacco exposure, and inherited susceptibility contribute to disease onset, we analyzed germline and somatic whole-genome, whole-exome, and targeted exome sequencing data from &amp;gt;1000 patients with pancreatic cancer across multiple cohorts from local and publicly available datasets. Mutational signature analyses quantified endogenous (e.g., aging-related) and exogenous (e.g., tobacco-related) mutational processes, which were assessed by age at diagnosis and smoking status. In a subset of tumors with DNA methylation data, we applied an epigenetic mitotic clock to estimate stem cell division rates based on age-related methylation at CpG-rich promoters. We also identified pathogenic or likely pathogenic (P/LP) germline variants in 21 pancreatic cancer risk genes using ClinVar annotations, evaluated somatic second hits in matched tumors, and calculated a weighted polygenic risk score (PRS) from 22 known low-penetrance susceptibility variants, defining a higher PRS as above the median in healthy individuals. RESULTS: In non-hypermutated tumors, total somatic mutation burden significantly increased with age, primarily driven by clock-like signatures (e.g., SBS1, SBS5, ID1, ID2, ID5). These signatures accounted for an estimated 78% of mutations in older-onset cases (&amp;gt;60 years), increasingly accounting for mutations in main driver genes (KRAS, CDKN2A, SMAD4, TP53). In contrast, smoking-related mutational changes were exclusive to young-onset cases (≤60 years). Young-onset smokers had elevated mutation burdens despite lacking the canonical tobacco signature (SBS4), instead showing increased SBS1 and ID2 signatures, linked to stem cell divisions and DNA replication, as well as higher stem cell division rates inferred from epigenomic analyses. Young smokers also had higher frequencies of TP53 mutations compared to non-smokers (83% vs. 64%; P=.001), particularly missense mutations at CpG sites. Additionally, P/LP germline variants were more frequent in young-onset than older-onset cases (13% vs. 7%; P=.002), with enrichment most evident for BRCA2, and contributed to younger onset only when accompanied by a somatic second hit. A higher PRS was modestly associated with reduced median age of cancer onset (by 4 years), with greater reductions when combined with smoking (7 years earlier) or P/LP variants (14 years earlier), compared to patients with neither of these factors. CONCLUSIONS: Our findings support a multifactorial model for pancreatic cancer development: aging-related mutational processes drive older-onset disease, while young-onset disease is more strongly influenced by environmental and inherited factors. These insights have important implications for risk stratification and interception strategies across age groups. Citation Format: Chen Yuan, Harshabad Singh, Kevin S. Kapner, Andressa Dias Costa, Anuradha B. Chittenden, Xinran Qi, Vidya Madineedi, Leigh Culnane, Lauren K. Brais, Douglas A. Rubinson, James M. Cleary, Brandon M. Huffman, Joseph D. Mancias, Thomas E. Clancy, Simona Cristea, Kimmie Ng, Matthew B. Yurgelun, David C. Linehan, Richard F. Dunne, Albert C. Koong, Daniel T. Chang, Aram F. Hezel, Andrew J. Aguirre, Jonathan A. Nowak, Brian M. Wolpin. Young- and older-onset pancreatic cancers arise through distinct mechanisms: Insights from genomic and germline analyses [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research—Emerging Science Driving Transformative Solutions; Boston, MA; 2025 Sep 28-Oct 1; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2025;85(18_Suppl_3):Abstract nr B075.

Plain Language SummaryCongenital hypogonadotropic hypogonadism (CHH) is a condition that affects the production and action of a hormone called gonadotropin-releasing hormone (GnRH), leading to problems with puberty and other health issues. Many genes are involved in CHH, making it a genetically diverse condition. This study investigated the genetic causes of CHH in 35 patients at a specialized endocrinology clinic. We used next-generation sequencing (NGS) to analyze 52 genes related to CHH. We found a genetic cause in 20 of the patients, which is about 57%. Among these, some had variations in multiple genes, indicating that the condition can be inherited from more than one gene. We discovered new harmful gene variations that could contribute to CHH. This research highlights how NGS can help identify complex genetic conditions like CHH and suggests that multiple gene involvement may explain the wide range of symptoms seen in patients. By expanding the gene panel for testing, we may find more connections to other disorders in the brain, which could improve early diagnosis, genetic counseling, and treatment options for patients.

BackgroundAlport syndrome (AS) is an inherited disorder affecting basement membrane collagen IV. AS is characterized by hematuria and progressive renal failure, accompanied by high-frequency sensorineural deafness and ocular changes. AS is caused by collagen type IV α3 chain (COL4A3), α4 chain (COL4A4), and α5 chain (COL4A5) variants. We aimed to identify the genetic variants in a cohort of 20 children with clinically suspected AS in Southwest China.ResultsWe detected 21 COL4A3, COL4A4, and COL4A5 variants in 20 probands. Of these variants, 16 (16/21, 76.2%) were classified as pathogenic/likely pathogenic and 5 (5/21, 23.8%) of uncertain significance according to the American College of Medical Genetics and Genomics criteria. A total of 11 (11/21, 52.4%) and 10 (10/21, 47.6%) variants were known and novel, respectively.ConclusionWe performed molecular diagnosis on 15 patients using targeted exome sequencing. Our findings indicate additional COL4A3, COL4A4, and COL4A5 variants as involved in AS, having implications for genetic diagnosis, therapy, and counseling of affected families.

ObjectiveThis study aimed to identify tumor-derived DNA in Papanicolaou (Pap) smears and plasma specimens from patients with endometrial cancer or endometrial intraepithelial neoplasia (EC/EIN).MethodsTumor tissue, peripheral blood, and Pap smear samples were collected from 84 patients with EC and EIN. Somatic mutations in tumor specimens were analyzed using targeted exome sequencing with a 363-cancer-related gene panel. Circulating single-molecule amplification and resequencing technology (cSMART) was used to evaluate somatic mutations in Pap smear and plasma circulating cell-free DNA (cfDNA).ResultsHigher pathological grades and lymph node metastases in EC were associated with elevated plasma cfDNA concentrations (p < 0.05 for both). Mutations corresponding to tissue samples were identified in 42.9% of plasma cfDNA and 77.4% of Pap smear cfDNA, with Pap smears demonstrating a higher detection rate (p < 0.05). In patients with EC, the detection rate of consistent mutations in cfDNA from peripheral blood was significantly elevated in those with higher pathological grades, lymphovascular space involvement, and lymph node metastasis (p < 0.05). The detection rate of consistent mutations in cfDNA from Pap smears was significantly higher in the EC group (p < 0.001).ConclusionMulti-gene panels can detect tumor-derived DNA in cfDNA from both blood and Pap smears of patients with EC, with Pap smear cfDNA potentially offering higher efficacy for liquid biopsies in EC. This approach could complement tissue biopsies for early diagnosis and risk stratification, warranting further investigation into the clinical utility of liquid biopsies for EC management.

KRAS is mutationally activated in 45–50% of colorectal cancer (CRC) cases, and while KRAS-targeted therapies have shown some clinical promise, upfront and acquired resistance limit their efficacy. To explore the acute response and mechanisms underlying KRAS inhibitor resistance, we used targeted exome sequencing and single-cell spatial transcriptomics to analyze patient-matched pre-treatment, on-treatment, and progression biopsies from patients treated with combined KRASG12C and EGFR inhibition. Acquired genetic events were identified in most patients at progression but were often subclonal and coexisted with transcriptional adaptive states. Mesenchymal, YAP, and fetal-like transcriptional signatures predominated in resistant tumors, while tumor cell-intrinsic inflammatory programs were induced in the early treatment phase. Single-cell spatial analysis revealed significant intratumoral heterogeneity, with diverse adaptive states predominating in different zones of individual tumors. Using human and murine organoid models, we show that these drug-induced inflammatory programs are cancer-cell autonomous and precede the emergence of regenerative fetal-like programs associated with drug resistance. We uncover TBK1 as a promising target to abrogate the early inflammatory adaptive phase and enhance responses to KRAS inhibition.

Background/Objectives: Lung cancer is a highly diverse disease, and reliable preclinical models that accurately reflect tumor characteristics are essential for studying lung cancer biology and testing new therapies. This study aimed to establish patient-derived tumor organoids (PDTOs) using small biopsy samples and surgical specimens to create a model system that preserves the genetic and histological features of the original tumors. Methods: PDTOs were generated from 163 lung cancer specimens, including 109 samples obtained using endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) or bronchoscopy, 52 surgical specimens, and 2 pleural fluid samples. The organoid establishment rate beyond passage three was assessed, and histological subtypes and genetic profiles were analyzed using immunohistochemical staining and targeted exome sequencing. Results: The overall PDTO establishment rate was 34.4% (56/163), and 44.6% (25/56) of these organoids retained the histological and genetic features of the parental tumors. Genetic analysis identified key mutations, including KRAS G12C, EGFR L858R, MET exon 14 skipping mutation, and ROS1 fusion. PDTOs successfully formed tumors in mice while maintaining the genetic characteristics of the original tumors. Co-culture of PDTOs with cancer-associated fibroblasts (CAFs) resulted in increased resistance to paclitaxel. In the co-culture model of PDTOs with immune cells, dose-dependent growth inhibition of PDTOs was observed in response to immune checkpoint inhibitors. Conclusions: PDTOs established from small biopsy and surgical specimens serve as a valuable model for studying lung cancer biology, tumor microenvironment interactions, and drug response. This model has the potential to improve personalized treatment strategies.

Rationale:Chronic lymphoproliferative disorder of natural killer cells is a rare heterogeneous indolent hematological disease, characterized by persistent clonal increase of mature NK cells with a typical large granular lymphocyte pattern. Chronic lymphoproliferative disorder of natural killer cells was revised to NK-large granular lymphocytic leukemia (NK-LGLL) in 2022 WHO classification. Renal involvement in NK-LGLL is extremely rare. Here, we report a woman diagnosed with NK-LGLL and nephrotic syndrome.Patient concerns:A 54-year-old woman had no obvious symptoms except for persistent peripheral lymphocytosis and neutropenia before kidney involvement. Then she presented with nephrotic syndrome, acute kidney injury and Epstein-Barr virus infection.Diagnoses:Bone marrow displayed clonal increase of mature NK cells with a typical large granular lymphocyte pattern. Renal biopsy showed pauci-immune crescentic glomerulonephritis and renal infiltration by NK-LGLL after exclusion of other diseases. Pathogenic N642H mutation of STAT5B was detected by targeted exome sequencing. A319T mutation in RELN and R500W mutation in INTS1 were also identified. Hence, she was diagnosed with crescentic glomerulonephritis associated with NK-LGLL.Interventions and outcome:We planned to combine methylprednisolone and cyclophosphamide in the treatment of this case. Unfortunately, our patient died of severe cerebral hemorrhage shortly after the diagnosis of NK-LGLL. We had no opportunity to use immunosuppressive drugs for therapy.Lessons:In short, we report a unique case diagnosed with crescentic glomerulonephritis associated with NK-LGLL, with pathogenic N642H mutation in STAT5B, Epstein-Barr virus infection and poor prognosis, different from typical inert type. Close monitoring of renal function is suggested for similar NK-LGLL patients.

A real-world comparison between the diagnostic yield of trio-whole exome sequencing and proband-only targeted exome sequencing in complex childhood epilepsy.

Single-nucleotide variants (SNVs) of ABCB1 gene encoding glycoprotein P might be connected with increased or decreased levels of direct oral anticoagulants (DOAC), and therefore with DOAC-related adverse bleeding or embolism. The aim of this study was to perform a targeted exome sequencing (TES) of ABCB1 gene in DOAC-treated patients with atrial fibrillation, who experienced a DOAC-related adverse event (AE) during the therapy. Targeted next generation sequencing was employed to examine SNVs in ABCB1 gene, which served as the basis for haplotype analysis. The study enrolled 33 patients with an AE (13 patients with bleeding and 20 patients with embolic stroke) and 33 patients tolerating long-term DOAC therapy without any AE (controls). The PLINK software was used to compare the differences between the groups. Fisher's test was employed for a standard case/control allelic association, and the chi-squared test was applied to test haplotype associations in contingency tables. In patients with DOAC-related bleeding compared with controls, no significant differences were found in all the examined SNVs; however, there were significant differences in the presence of AAAGAGCT (11.5%vs.1.6%,p<0.05) and AGAG (11.1vs.1.7%,p<0.05) haplotypes. Compared to controls, patients with stroke had a minor allele observed more frequently in rs2235033 (62.5%vs.39.4%,p<0.05), and significant differences were also found in the presence of AAAGAACT (19.3vs.39.0%,p<0.05), GGCGGGAT (19.5vs.6.4%,p<0.05), AGAA (30.8vs.51.4%,p<0.05), CGGG (23.1vs.7.6%,p<0.05) haplotypes. This study found significant differences in the selected rs2235033 SNV and in several ABCB1 gene common haplotypes in patients with DOAC-related AE.

&lt;p&gt;Background: Sensorineural hearing loss is among the most common sensory defects worldwide. Nonsyndromic hearing loss (NSHL) accounts for 70% of inherited hearing loss. The genetic causes of NSHL are considered heterogeneous. The high rate of consanguineous marriages in Saudi Arabia increases the population's prevalence of autosomal recessive inheritance patterns.&lt;/p&gt; &lt;p&gt;Objective: To discover a novel variant for NSHL patients.&lt;/p&gt; &lt;p&gt;Methods: A family with two hearing-impaired children was recruited. Targeted exome sequencing, the Twist Exome 2.0 kit (Twist Bioscience) using the Novaseq X plus platform, was used to identify the variant. Sanger sequencing was carried out to confirm the finding and perform segregation analysis. MutationTaster tool was used to determine the pathogenicity effect on the protein structure.&lt;/p&gt; &lt;p&gt;Results: A homozygous two-bp duplication variant on the (c.8813_8814dup) MYO15A gene was identified in a Saudi family of two hearing-impaired children. Sanger sequencing confirmed the variant in the affected children and their parents. The prediction tool indicated the frameshift effect on the protein level, which leads to protein function disruption. Based on the American College of Medical Genetics and Genomics guidelines, it is classified as a pathogenic variant.&lt;/p&gt; &lt;p&gt;Conclusion: A novel biallelic frameshift variant in MYO15A causes NSHL in a Saudi family. This variant is considered rare and isolated to the Saudi population. Expanded genotype-phenotype correlations for hearing loss patients are likely to confirm the findings and reveal novel variants.&lt;/p&gt;

Abstract Breast cancer (BC) is one of the primary causes of cancer related deaths among women worldwide. According to Cancer Statistics, over 310,720 new breast cancer diagnoses are expected in the United States in 2024 with an estimate of over 42,250 expected BC related deaths. Invasive Lobular Carcinoma (ILC) accounts for the 2nd most frequently diagnosed histological subtype of BC, constituting 10%-15% of all reported cases. ILC is hormonally driven (estrogen and progesterone receptors) with a hallmark characteristic of ILC being lack of E-cadherin (CDH1) and with mutations in the CDH1 gene reported in over 90% of cases. Mutations in the CDH1 gene have also been associated with 39%–52% increased lifetime risk of developing breast cancer in women. Patient-derived organoid (PDO) models have been shown to offer significant advantages in terms of better recapitulating the tumor/microenvironment niche, however such models have not been available to study ILC. With a vision to develop 3D models to better understand ILC and develop specific therapeutic targets, here, we describe the establishment and comprehensive characterization of a diverse biobank of ILC PDOs. We have performed in-depth genomic, transcriptomic and histologic profiling of the established ILC PDOs and validated the models to be true representatives of ILC tumors. Comprehensive genomic profiling of ILC PDOs using a custom 143 cancer driver gene panel (targeted exome sequencing) identified ILC specific mutations in CDH1, PIK3CA, TBX3, RUNX, etc. in PDOs. Short Multiply Aggregated Sequence Homologies (SMASH) identified widely reported copy number gains and losses (gains on Chr1q and losses on Chr16q and Chr17p among others) in established ILC PDOs. Transcriptomic profiling performed on matched tumor-normal pairs revealed the upregulation of ESR1 signalling-related pathways known to drive ILC carcinogenesis. Multiplexed single-cell RNA sequencing revealed that the established PDOs retained their cellular complexity and heterogeneity even after multiple passages and contained hormone responsive, hormone secretory, basal, fibroblasts, and proliferating populations of cells representing major cell types in ILC tumors. Lastly, we also show that the established ILC PDOs when xenografted (either in mammary glands or intra-ductally) in NOD-SCID gamma mice retain their tumorigenicity in vivo and the single file growth pattern, a characteristic of invasiveness in ILC tumors. In summary, we have established a living biobank of ILC PDOs often with matched normals (adjacent and/or distal) that are representative of different subtypes of ILC (classic, pleomorphic, tubule-lobular, mixed, etc.). Overall, this study has optimized a pipeline for establishment, culturing and validation of ILC PDOs as robust models available for studying ILC biology with a focus on basic, translational, and personalized medicine. Citation Format: Minakshi Gandhi, Melissa Kramer, Jill Habel, Zihua Wang, Raditya Utama, Suzanne Russo, Payal Naik, Karen Kostroff, David L Spector. Characterization of a Living Biobank of Patient-Derived Invasive Lobular Carcinoma Organoids that Retain Tumor Heterogeneity in 3D Culture and Tumorigenicity In Vivo [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P3-06-24.