ATRX Mutations Research Articles

Mutant ATRX: pathogenesis of ATRX syndrome and cancer

The transcriptional regulator ATRX, a genetic factor, is associated with a range of disabilities, including intellectual, hematopoietic, skeletal, facial, and urogenital disabilities. ATRX mutations substantially contribute to the pathogenesis of ATRX syndrome and are frequently detected in gliomas and many other cancers. These mutations disrupt the organization, subcellular localization, and transcriptional activity of ATRX, leading to chromosomal instability and affecting interactions with key regulatory proteins such as DAXX, EZH2, and TERRA. ATRX also functions as a transcriptional regulator involved in the pathogenesis of neuronal disorders and various diseases. In conclusion, ATRX is a central protein whose abnormalities lead to multiple diseases.

Radiogenomic profiling of global DNA methylation associated with molecular phenotypes and immune features in glioma

BackgroundThe radiogenomic analysis has provided valuable imaging biomarkers with biological insights for gliomas. The radiogenomic markers for molecular profile such as DNA methylation remain to be uncovered to assist the molecular diagnosis and tumor treatment.MethodsWe apply the machine learning approaches to identify the magnetic resonance imaging (MRI) features that are associated with molecular profiles in 146 patients with gliomas, and the fitting models for each molecular feature (MoRad) are developed and validated. To provide radiological annotations for the molecular profiles, we devise two novel approaches called radiomic oncology (RO) and radiomic set enrichment analysis (RSEA).ResultsThe generated MoRad models perform well for profiling each molecular feature with radiomic features, including mutational, methylation, transcriptional, and protein profiles. Among them, the MoRad models have a remarkable performance in quantitatively mapping global DNA methylation. With RO and RSEA approaches, we find that global DNA methylation could be reflected by the heterogeneity in volumetric and textural features of enhanced regions in T2-weighted MRI. Finally, we demonstrate the associations of global DNA methylation with clinicopathological, molecular, and immunological features, including histological grade, mutations of IDH and ATRX, MGMT methylation, multiple methylation-high subtypes, tumor-infiltrating lymphocytes, and long-term survival outcomes.ConclusionsGlobal DNA methylation is highly associated with radiological profiles in glioma. Radiogenomic global methylation is an imaging-based quantitative molecular biomarker that is associated with specific consensus molecular subtypes and immune features.

The 2022 WHO classification of tumors of the pituitary gland: An update on aggressive and metastatic pituitary neuroendocrine tumors.

The vast majority of pituitary neuroendocrine tumors (PitNETs) are benign and slow growing with a low relapse rate over many years after surgical resection. However, about 40% are locally invasive and may not be surgically cured, and about one percentage demonstrate an aggressive clinical behavior. Exceptionally, these aggressive tumors may metastasize outside the sellar region to the central nervous system and/or systemically. The 2017 (4th Edition) WHO Classification of Pituitary Tumors abandoned the terminology "atypical adenoma" for tumors previously considered to have potential for a more aggressive behavior since its prognostic value was not established. The 2022 (5th Edition) WHO Classification of the Pituitary Tumors emphasizes the concept that morphological features distinguish indolent tumors from locally aggressive ones, however, the proposed histological subtypes are not consistent with the real life clinical characteristics of patients with aggressive tumors/carcinomas. So far, no single clinical, radiological or histological parameter can determine the risk of growth or malignant progression. Novel promising molecular prognostic markers, such as mutations in ATRX, TP53, SF3B1, and epigenetic DNA modifications, will need to be verified in larger tumor cohorts. In this review, we provide a critical analysis of the WHO guidelines for prognostic stratification and diagnosis of aggressive and metastatic PitNETs. In addition, we discuss the new WHO recommendations for changing ICD-O and ICD-11 codes for PitNET tumor behavior from a neoplasm either "benign" or "unspecified, borderline, or uncertain behavior" to "malignant" neoplasm regardless of the clinical presentation, histopathological subtype, and tumor location. We encourage multidisciplinary initiatives for integrated clinical, histological and molecular classification, which would enable early recognition of these challenging tumors and initiation of more appropriate and aggressive treatments, ultimately improving the outcome.

Characteristics of H3K27M-mutant diffuse gliomas with a non-midline location.

Diffuse midline gliomas (DMG) with H3K27 alterations (H3K27M-DMG) are a highly aggressive form of brain cancer. In rare cases, H3K27 mutations have been observed in diffuse non-midline gliomas (DNMG). It is currently unclear how these tumors should be classified. Herein, we analyze the characteristics of DNMG with H3K27M mutations. We reviewed the clinical, radiological and histological characteristics of all patients with an H3K27M mutated diffuse glioma diagnosed in our institution, between 2016 and 2023, to identify cases with a non-midline location. We then performed a molecular characterization (DNA methylation profiling, whole genome and transcriptome sequencing or targeted sequencing) of patients with an H3K27M-mutant DNMG and reviewed previously reported cases. Among 51 patients (18 children and 33 adults) diagnosed with an H3K27M diffuse glioma, we identified two patients (4%) who had a non-midline location. Including our two patients, 39 patients were reported in the literature with an H3K27M-mutant DNMG. Tumors were most frequently located in the temporal lobe (48%), affected adolescents and adults, and were associated with a poor outcome (median overall survival was 10.3 months (0.1-84)). Median age at diagnosis was 19.1 years. Tumors frequently harbored TP53 mutations (74%), ATRX mutations (71%) and PDGFRA mutations or amplifications (44%). In DNA methylation analysis, H3K27M-mutant DNMG clustered within or close to the reference group of H3K27M-mutant DMG. Compared to their midline counterpart, non-midline gliomas with H3K27M mutations seemed more frequently associated with PDGFRA alterations. DNMG with H3K27M mutations share many similarities with their midline counterpart, suggesting that they correspond to a rare anatomical presentation of these tumors. This is of paramount importance, as they may benefit from new therapeutic approaches such as ONC201.

Peptide Receptor Radionuclide Therapy versus Capecitabine/Temozolomide for the Treatment of Metastatic Pancreatic Neuroendocrine Tumors.

Background: Peptide Receptor Radionuclide Therapy (PRRT), a form of Radioligand Therapy (RLT), and Capecitabine/Temozolomide (CAPTEM) are cornerstones of systemic therapy for metastatic pancreatic neuroendocrine tumors (PNETs). Data regarding comparative efficacy are lacking. Herein, we compare the efficacy of PRRT vs. CAPTEM as second-line/beyond regimens and treatment sequencing. Methods: Clinicopathologic, radiographic, and genomic data were captured for metastatic PNETs seen in our multi-disciplinary NET clinic between 2013 and 2023. The primary outcome was progression-free survival (PFS) after progression on a previous line of systemic therapy. The secondary outcomes were objective response rate (ORR), time to response (TTR), and overall survival (OS). Results: Fifty-nine cases were included. PFS was similar in the PRRT (n = 29) and CAPTEM (n = 30) groups (PRRT = 21.90 months vs. CAPTEM = 20.03 months; HR 0.99; p = 0.97). On subgroup analysis, PRRT had longer PFS in cases without extrahepatic metastases (26.47 months vs. 17.67 months; p = 0.03) and cases with a mutation in the MEN1, DAXX, and/or ATRX genes (28.43 months vs. 18.67 months; p = 0.03). PRRT had reduced PFS in patients with grade 3 disease (7.83 months vs. 16.33 months; p = 0.02). ORR did not vary significantly (34.78% vs. 40.91%; p = 0.67). CAPTEM responders showed shorter TTR (6.03 months vs. 11.15 months; p = 0.03). In patients who received both, OS did not vary based on the sequence (HR 1.20; p = 0.75). Conclusions: PFS, ORR, and OS are similar when using PRRT vs. CAPTEM as second-line-and-beyond therapy for patients with metastatic PNETs. However, patients with MEN1, DAXX, and/or ATRX mutations or without extrahepatic metastases might better benefit from PRRT and patients with grade 3 disease from CAPTEM. Candidates for surgical debulking or with tumor-induced symptoms may benefit from initial treatment with CAPTEM due to shorter TTR.

ERBB2/ ERBB3-mutated S100/ SOX10-positive unclassified high-grade uterine sarcoma: first detailed description of a novel entity.

With the increasing use of innovative next generation sequencing (NGS) platforms in routine diagnostic and research settings, the genetic landscape of uterine sarcomas has been dynamically evolving during the last two decades. Notably, the majority of recently recognized genotypes in uterine sarcomas represent gene fusions, while recurrent oncogene mutations of diagnostic and/ or therapeutic value have been rare. Recently, a distinctive aggressive uterine sarcoma expressing S100 and SOX10, but otherwise lacking diagnostic morphological, immunophenotypic and molecular features of other uterine malignancies has been presented in a scientific abstract form (USCAP, 2023), but detailed description and delineation of the entity is still missing. We herein describe two high-grade unclassified uterine sarcomas characterized by spindle to round cell morphology and diffuse expression of S100 and SOX10, originating in the uterine body and cervix of 53- and 45-year-old women and carrying an ERBB3 (p.Glu928Gly) and an ERBB2 (p.Val777Leu) mutation, respectively. Both tumors harbored in addition genomic HER2 amplification, ATRX mutation and CDKN2A deletion. Methylation studies revealed a methylome most similar to MPNST-like tumors, but distinct from melanoma, MPNST, clear cell sarcoma, and endometrial stromal sarcoma. Case 1 died of progressive peritoneal metastases after multiple trials of chemotherapy 47months after diagnosis. Case 2 is a recent case who presented with a cervical mass, which was biopsied. This study defines a novel heretofore unrecognized aggressive uterine sarcoma with unique phenotypic and genotypic features. Given the potential value of targeting HER2, recognizing this tumor type is mandatory for appropriate therapeutic strategies and for better future delineation of the entity.

Pediatric Hemispheric High-Grade Gliomas and H3.3-G34 Mutation: A Review of the Literature on Biological Features and New Therapeutic Strategies.

Pediatric high-grade glioma (pHGG) encompasses a wide range of gliomas with different genomic, epigenomic, and transcriptomic features. Almost 50% of pHGGs present a mutation in genes coding for histone 3, including the subtype harboring the H3.3-G34 mutation. In this context, histone mutations are frequently associated with mutations in TP53 and ATRX, along with PDGFRA and NOTCH2NL amplifications. Moreover, the H3.3-G34 histone mutation induces epigenetic changes in immune-related genes and exerts modulatory functions on the microenvironment. Also, the functionality of the blood-brain barrier (BBB) has an impact on treatment response. The prognosis remains poor with conventional treatments, thus eliciting the investigation of additional and alternative therapies. Promising molecular targets include PDGFRA amplification, BRAF mutation, EGFR amplification, NF1 loss, and IDH mutation. Considering that pHGGs harboring the H3.3-G34R mutation appear to be more susceptible to immunotherapies (ITs), different options have been recently explored, including immune checkpoint inhibitors, antibody mediated IT, and Car-T cells. This review aims to summarize the knowledge concerning cancer biology and cancer-immune cell interaction in this set of pediatric gliomas, with a focus on possible therapeutic options.

MEN1/DAXX/ATRX mutations enhance progression-free survival in gastroenteropancreatic neuroendocrine tumors treated with peptide receptor radionuclide therapy.

Pre-clinical data suggest that mutations in the MEN1, DAXX, and/or ATRX genes may potentially increase radiation efficacy in cancer cells. Herein, we explore the association between response to peptide receptor radionuclide therapy (PRRT) and those mutations in patients with gastroenteropancreatic neuroendocrine tumors (GEP-NETs). We analyzed tissue-based next generation sequencing (NGS) assay results and clinicopathologic data from 28 patients with GEP-NETs treated with PRRT. Findings were correlated with progression-free survival (PFS) and objective response rate (ORR). Patients with mutations in MEN1, DAXX, and/or ATRX (n = 13) had a longer median PFS (26.47 vs 12.13 months; P = 0.014) than wild-type (n = 15) patients when adjusted for surgery prior to PRRT, tumor grade, and presence of TP53 mutation. Alterations in MEN1 along with a concurrent mutation in either DAXX or ATRX (n = 6) trended toward longer PFS compared to patients without concurrent mutations (31.53 vs 17.97 months; P = 0.09). ORR was higher in patients with a mutation in MEN1, DAXX, or ATRX (41.67% vs 15.38%). In pancreatic NET patients, these target mutations also showed a longer PFS (28.43 vs 9.83 months; P = 0.04). TP53 alterations showed a shorter PFS than wild-type cases (11.17 vs 20.47 months; P = 0.009). Mutations in MEN1/DAXX/ATRX are associated with improved PFS in patients with GEP-NETs receiving PRRT and might be used as a biomarker for treatment response.

Molecular alterations and prognosis of breast cancer with cutaneous metastasis

PurposeCutaneous metastasis (CM) accounts for 5–30% of patients with breast cancer (BC) and presents unfavorable response to treatment and poor prognosis. A better understanding of the molecular alterations involved in metastasis is essential, which would help identify diagnostic and efficacy biomarkers for CM.Materials: We retrospectively reviewed a total of 13 patients with histological or cytological diagnosis of breast cancer and CM. Clinical information was extracted from the medical records. The mutational landscape of matched primary tumors with their lymph nodes or CM tissues were analyzed using next-generation sequencing (NGS) of 425 cancer-relevant genes. All tissues were also analyzed by immunohistochemistry (IHC). The association of prognosis with various clinical and molecular factors was also evaluated.ResultsMore than half of the patients were Ki67 low (< 50%, 53.7%). Most patients (12, 92.3%) had other metastasis sites other than skin. The median time from diagnosis to the presentation of CM (T1) was 15 months (range: 0–94 months) and the median time from CM to death (T2) was 13 months (range 1–78). The most frequently altered genes across the three types of tissues were TP53 (69.6%, 16/23), PIK3CA (34.8%, 8/23), and MYC (26.1%). The number of alterations in CM tends to be higher than in primary tumors (median 8 vs. 6, P = 0.077). Copy number loss in STK11, copy number gain in FGFR4, TERT, AR, FLT4 and VEGFA and mutations in ATRX, SRC, AMER1 and RAD51C were significantly enriched in CM (all P < 0.05). Ki67 high group (> 50%) showed significantly shorter T1 than the Ki67 low group (≤ 50%) (median 12.5 vs. 50.0 months, P = 0.036). TP53, PIK3CA mutations, and TERT amplification group were associated with inferior T2 (median 11 vs. 36 months, P = 0.065; 8 vs. 36 months, P = 0.013, 7 vs. 36 months, P = 0.003, respectively). All p values were not adjusted.ConclusionWe compared the genomic features of primary breast cancer tissues with their corresponding CM tissues and discussed potential genes and pathways that may contribute to the skin metastasis of advanced breast cancers patients. TP53, PIK3CA mutant, and TERT amplification may serve as biomarkers for poor prognosis for CM patients.

HGG-48. THE BLOOD-BASED TELOMERIC DNA C-CIRCLE ASSAY CAN DETECT ALTERNATIVE LENGTHENING OF TELOMERES (ALT) IN PEDIATRIC HIGH-GRADE GLIOMAS

Abstract BACKGROUND High-grade gliomas (HGG) comprise ~10% of pediatric brain tumors; 40% of pediatric HGG use the alternative lengthening of telomeres (ALT) mechanism to maintain replicative immortality. ALT cancers share a unique biology providing potential therapeutic targets. Extrachromosomal telomere DNA repeats, termed C-circles, can be detected by a unique PCR assay, providing a sensitive and specific biomarker for ALT cancers. C-circles have been detected in ALT tumors and serum of ALT patients. We have adapted the C-circle assay (CCA) to provide a sensitive and specific assay with cfDNA in patient plasma. Here we determined if cfDNA in plasma can be used to detect patients with ALT-positive HGG. METHODS Frozen plasma samples were collected in Streck tubes at time of surgery prior to resection from patients with HGG. DNA was extracted and quantified. Isothermal rolling C-circle amplification and real-time telomere PCR was performed followed by analysis. The C-circle positive neuroblastoma cell line, CHLA-90, and C-circle negative neuroblastoma cell line, CHLA-20, served as the positive and negative controls respectively. CCA values in plasma of 2 and above indicate positivity for C-circles. RESULTS Plasma samples were analyzed by CCA from three pediatric patients whose brain tumors were previously biopsied and identified as ATRX mutation positive. ATRX mutation indicates ALT however, not all ALT cancers have ATRX mutations. All plasma samples were above the 2.0 cutoff at 3.37, 3.13, and 4.59 and thus classified as C-circle positive. CONCLUSIONS This preliminary cohort shows that the CCA has potential for identifying ALT pediatric brain tumors using cfDNA from plasma. As novel therapies effective against ALT HGG are developed, the plasma CCA may allow accelerated initiation of neoadjuvant therapy to shrink tumor prior to surgical resection and lessen or prevent the need for tumor biopsy. Expansion of the initial test cohort is underway. Supported by NCI UO1 CA63988.

OTHR-25. ANAPLASTIC TRANSFORMATION OF DIFFUSE LEPTOMENINGEAL GLIONEURONAL TUMORS: TWO PEDIATRIC CASES

Abstract BACKGROUND Diffuse Leptomeningeal Glioneuronal Tumors (DLGT) typically have a clinical course with slow progression; however, there have been cases of anaplastic transformation. We describe the clinical presentation, radiographic features, and molecular characteristics in 2 pediatric DLGT cases that underwent aggressive transformation. METHODS Patient 1: A 20 y/o male with DLGT with BRAF-KIAA1549 fusion and loss of 1p who was first diagnosed at 2.5 y/o. Over the course of 17 years, he was treated with numerous therapies due to multiple progressions, including temozolomide, carboplatin/vincristine, selumetinib, and vinblastine. At age 17, he developed significant growth of a posterior fossa mass with solid and cystic characteristics. Resection revealed new anaplastic pathologic features. Patient 2: He was diagnosed with DLGT at age 2 y/o. Imaging demonstrated intramedullary spinal nodules at T4 and T5 with abnormal thickening and enhancement over the cerebral leptomeningeal space. A biopsy revealed DLGT with a BRAF-KIAA1549 fusion, 1p loss and Ki67 5%. He received multiple therapies including carboplatin/vincristine, vinblastine, and selumetinib. RESULTS Patient 1: Molecular characterization revealed new ATRX mutation, and CDKN2A/B homozygous deletion. Over the following 2 years, he had multiple treatments, progressions, and surgeries. Final pathology revealed significant anaplasia with Ki67 90%, ATRX loss, and mutation of PPM1D and ARID1A. He rapidly declined. Patient 2: Approximately 2.5 years after diagnosis, he developed rapid progression in the 4th and lateral ventricles. Pathology confirmed recurrent DLGT, but with high-grade transformation, H3K27M (-), H3k27m3 loss, ATRX (+), Ki67 40%. He was treated with focal irradiation followed by temozolomide/CCNU. CONCLUSION In rare cases, DLGT can undergo anaplastic transformation. Once this occurs, it behaves clinically like a high-grade glioma. Understanding the molecular underpinnings and prognostic features that lead to this transformation are important for future study.

HGG-01. DURABLE RESPONSE TO IMMUNOTHERAPY IN AN ADOLESCENT PATEINT WITH CONSTITUTIONAL MISMATCH REPAIR DEFICIENCY CMMRD AND SYNCHRONOUS THREE PRIMARY MALIGNANCIES

Abstract BACKGROUND Familial Cancer Syndrome FCS is of particular concern in Saudi Arabia due to the high rates of consanguinity. Constitutional mismatch repair deficiency Syndrome (CMMRD) is an autosomal recessive FCS with a wide spectrum of malignancies Caused by Biallelic germline mutations in one of 4 mismatch repair genes (MLH1, PMS2, MSH2, and MSH6). METHODS we report 13 yr. Saudi boy, previously healthy with multiple café-au-lait (CAL) macules, and Consanguineous parents with a positive family history of Malignancy. presented with progressive headache, vomiting, abdominal pain, weight loss, and microcytic anemia. MRI Brain showed a Large left hemispheric necrotic mass, MRI abdomen: Circumferential gastric body wall thickening Distal transverse colon intraluminal polypoid mass, and liver hypodense lesion. He had a Resection of the brain tumor and Upper GI endoscopy + colonoscopy with polypectomy: which showed an Ugly gastric ulcer, and multiple colonic polyps. RESULTS Brain tumor molecular pathology: Gliosarcoma WHO grade 4 with negative IHC expression in PMS2 protein, hypermutant tumor with somatic PMS2 ATRX, TP53, NFI, and PIK3R1 genes mutation. stomach biopsy confirmed invasive gastric adenocarcinoma and Colonic polyp biopsy: Tubulovillous adenoma. Genetic testing Detect pathogenic Biallelic germline PMS2 mutation. He received adjuvant focal Brain radiation therapy 59.4Gy/33 F concurrent with immunotherapy (PD-1) inhibitor Nivolumab. Follow-up Abdominal imaging showed excellent response to immunotherapy with no significant residual lesion. MRI of the brain showed surgical bed multi-cystic lesions with thick nodular enhancement suggestive either of radiation necrosis vs. progressions/pseudoprogression. repeated MRI confirmed further progression of the enhancing surgical cavity lesion which was unresectable. CTLA-4 Inhibitor ipilimumab was added to Nivolumab to optimize therapy. UpToDate our patient is clinically stable on Maintenance Nivolumab with no side effects. CONCLUSION Our case confirms immunotherapy’s efficacy for treating CMMRD-related cancers, which can improve survival while reducing toxicity from less effective conventional therapies.

Convolutional neural network and glioma genotyping: A systematic review.

e14038 Background: Gliomas are the most common primary intracranial tumors. Overall, there are 23 different glioma types, and the grading and prognosis of each type are determined by their specific mutations. Several identified mutations are IDH gene status, CDKN2A/B homozygote delegation, PTEN mutation, p53 mutation, TERT promotor mutation, H3K27M mutation, MGMT promotor methylation, trisomy 7/monozomy10, EGFR amplification, ATRX mutation, 1p/19q codeletion, and BRAF mutation. These specific mutations and genomic features can have their distinctive radiologic features seen in different magnetic resonance imaging (MRI) sequences. By using a Convolutional Neural Network (CNN) for feature extraction, the genotyping power of MRI for glioma could be increased. To address this, we systematically reviewed the use of different CNN-based models to detect the genetic context of glioma based on the MRI images. Methods: We conducted a systematic search in PubMed, Embase, Scopus, Web of Science, and Google Scholar. We included original peer-reviewed human studies (retrospective or prospective), focusing on the use of CNN in individuals with glioma. We excluded animal studies, studies with no CNN-based models, reviews, book chapters, conference abstracts, and case reports. JBI checklists were utilized for quality assessment. Results: The initial search resulted in 973 articles. after removing duplicates, 563 studies entered the screening process by title and abstract. 59 studies met our inclusion criteria (56 retrospective and 3 prospective cohort studies) with a total of 26434 glioma patients included (studies ranged from 21 – 2648 individuals). 31 studies used CNN methods only in different stages of image processing (tumor segmentation, feature extraction, image encoder) and 28 studies used CNN as a classifier and mutation predictor. ResNet was the most commonly used CNN-based model (n = 14). U-net (n = 7) and DenseNet models (n = 4) were other commonly used CNN-based models. Included studies targeted IDH status (n=44), 1p/19q codeletion (n=18), MGMT promotor methylation (n=11), ATRX mutation (n=5), TERT promotor mutation (n=5), CDKN2A/B homozygote deletion (n=3), PTEN mutation (n=3), p53 mutation (n=3), H3K27M mutation (n=2), trisomy 7/monozomy10 (n=2), EGFR amplification (n=2), and BRAF mutation (n=1). Highly severe methodological heterogeneity among these studies excluded the possibility of meta-analysis. Conclusions: Current studies have shown that, as a non-invasive method, using CNN-based models on multiple MRI sequences increases the power of early detection and prediction of glioma genetic types before pathological confirmation and has promising results. However, DL performance varies among mutations. Combining multiple DL models with clinical and radiomic features in different and larger databases can further increase the final results.

Association of mutational status with metastatic pattern, race, ethnicity, and overall survival (OS) in leiomyosarcoma (LMS).

e23553 Background: LMS, one of the most common soft tissue sarcomas, can arise from the uterus, retroperitoneum, vessels, or extremities. It is unclear if the metastatic pattern and potentially the mutational status differs among the different types of LMS. We investigated the association between sites of metastases, mutations, race, ethnicity, and OS in different LMS types. Methods: This single-center retrospective study evaluated patients with biopsy-proven LMS that underwent genomic testing between January 2009 and May 2023. Association between genomic profile, sites of metastases, types of LMS (extremity, retroperitoneal, uterine, vascular), self-reported race and ethnicity was performed using Fisher’s exact test. Survival curves for self-reported race and ethnicity, mutations, sites of metastases, and types of LMS were estimated using the Kaplan-Meier method and compared with a log-rank test. Cox proportional hazard regression models were used to examine the association between OS with types of LMS, self-reported race and ethnicity, mutational status, and sites of metastases. Results: The study included 110 subjects (F/M:81/29; median age 57 years, 99/110 with metastatic disease). 27/110 (25%) subjects self-reported as Hispanic, and 27/110 (25%) as Black. The most common type of LMS was uterine (50/110, 45%), followed by retroperitoneal (27/110, 25%), extremities (23/110, 21%) and vascular (10/110, 9%). The most common mutation was TP53 (74/110, 67%), followed by RB1 (24/110, 22%). ER/PR expression was present in 25/110 subjects (23%). The lungs were the most common site of metastasis (79/99, 80%), followed by the liver (37/99, 37%), and pelvis (29/99, 29%). Peritoneal and pelvic metastases were more common in uterine LMS (p = 0.0001 and 0.002, respectively). Liver metastases were more common in retroperitoneal LMS (p = 0.03). TP53 mutation was more common in vascular LMS (p = 0.03), whereas intermediate/high tumor burden was more common in LMS arising from the extremities (p = 0.004). BRCA1/2 mutations were more common in subjects with liver (p = 0.02) and intramuscular metastases (p = 0.04). Pleural metastases (11/99, 11%) were associated with lower OS (Risk Ratio: 2.7; 95% CI: 1.13, 6.32; p = 0.02). ATRX mutation (17/110, 15%) was more common in Hispanic subjects (p = 0.005), in which it was associated with lower OS (p = 0.0007). Conclusions: The mutational status and metastatic pattern of LMS varies based on the LMS types, and potentially provides prognostic data. In our sample, OS was reduced in Hispanic subjects with ATRX mutation, and the presence of pleural metastases was also independently associated with shorter OS.

Longitudinal tumor-wide sampling of glioblastoma reveals diverse genomic drivers of the earliest clonal expansion at diagnosis and recurrence.

2009 Background: Despite decades of clinical trials, glioblastoma (GBM) remains a rapidly fatal disease. Much of therapy resistance in GBM is attributed to intratumoral heterogeneity in which subclones rapidly evolve and treatment-resistant cell populations emerge. Understanding heterogeneity at diagnosis and its relationship to the origins of recurrence are critical to selecting therapies that are efficacious across the entire tumor. However, few genomic studies go beyond analyzing single tumor samples per patient. Methods: A spatially oriented, whole tumor sampling approach was used to obtain 43 biopsies from 3 GBMs at diagnosis and recurrence. Pyclone and ClonEvol were applied to whole exome sequencing to reconstruct clonal evolution, FACETs identified copy number variations and HATCHet estimated tumor purity. Candidate driver mutations were evaluated in relation to the Catalogue of Somatic Mutations In Cancer. Results: A single founding clone and multiple subclones were identified for each diagnosis-recurrence pair. Tumor-wide clonal alterations representing initial clonal expansions of these GBMs included both canonical changes (Chr 7 gain, Chr 10 loss, CDKN2A deletion, EGFR amplification) and a diverse set of large-scale copy number variations (Chr 19, 20 gain), driver mutations (PTEN, KDR, CDH11, CNTNAP2), and fusions (LIMCH1::UCHL1, KANK::DOCK8). A second subset of alterations (Chr 8 gain, ATRX mutation), appeared to be tumor wide at diagnosis but were not identified at recurrence. Cancer drivers were also present subclonally, including CDKN2A deletion, MDM2 amplification, and mutations in NF1, and GRM3. Evolutionary trees consisted of 5 generations of clones in Patient 323 (P323), 3 in P454, and 4 in P534. Divergence of the recurrent tumors from their matched primary occurred in the second generation in P454 and P534 and in the third in P323. As a result, an average of 37% of potential drivers of oncogenesis and clonal expansion across the cohort appear after divergence. Furthermore, each recurrent tumor contained at least one tumor wide driver alteration that was subclonal or undetected at diagnosis. Conclusions: Whole-tumor sampling of three GBM patients at both diagnosis and recurrence identified a diversity of genomic drivers and deeper and more complex genetic roots of individual GBM than previously seen in single-biopsy studies. Tumor recurrences consistently arose from a single subclone that diverged early in evolution of the primary tumor and contained clonal drivers either not detected or subclonal in the primary—suggesting a role for these drivers in persistence and expansion. In the age of personalized medicine, our study highlights the clinical potential of tumor wide sampling in identifying therapeutic targets that could avoid heterogeneity-related therapeutic failures.

NGS of brush cytology samples improves the detection of high-grade dysplasia and cholangiocarcinoma in patients with primary sclerosing cholangitis: A retrospective and prospective study.

Biliary dysplasia, a precursor of cholangiocarcinoma (CCA), is a common complication of primary sclerosing cholangitis. Patients with high-grade dysplasia (HGD) or early CCA who have received oncological treatment are candidates for liver transplantation. The preoperative diagnosis of CCA or HGD is challenging, and the sensitivity of biliary brush cytology (BC) is limited. By using next-generation sequencing (NGS), we retrospectively analyzed archived tissue samples (n=62) obtained from explanted liver tissue and CCA samples to identify oncogenic mutations that occur during primary sclerosing cholangitis carcinogenesis. BC samples were prospectively collected from patients with primary sclerosing cholangitis (n=97) referred for endoscopic retrograde cholangiography to measure the diagnostic utility of NGS combined with BC compared with traditional cytology alone. Mutations in KRAS, GNAS, FLT3, RNF43, TP53, ATRX, and SMAD4 were detected in archived CCA or HGD samples. KRAS, GNAS, TP53, CDKN2A, FBXW7, BRAF, and ATM mutations were detected in prospectively collected brush samples from patients with histologically verified CCA or HGD. One patient with low-grade dysplasia in the explanted liver had KRAS and GNAS mutations in brush sample. No mutations were observed in brush samples or archived tissues in liver transplantation cases without biliary neoplasia. While KRAS mutations are common in biliary neoplasms, they were also observed in patients without biliary neoplasia during surveillance. In summary, NGS of BC samples increased the sensitivity of detecting biliary neoplasia compared with traditional cytology. Performing NGS on BC samples may help diagnose HGD or early CCA, benefiting the timing of liver transplantation.

Abstract 7614: Mutations in MEN1/DAXX/ATRX are associated with improved progression-free survival in patients with gastroenteropancreatic neuroendocrine tumors receiving peptide receptor radionuclide therapy

Abstract Background: Peptide Receptor Radionuclide Therapy (PRRT) using 177Lu-DOTATATE is an effective treatment for metastatic Gastroenteropancreatic Neuroendocrine Tumors (GEP-NETs). However, it remains unknown why some patients (pts) respond more favorably than others. Pre-clinical data suggests that mutations in the MEN1, DAXX, and/or ATRX genes influence DNA repair capabilities and increase radiation effect in cancer cells. We therefore thought to evaluate the association between response to PRRT and the presence of MEN1, DAXX, and/or ATRX mutations in patients with GEP-NETs. Methods: We retrospectively analyzed CLIA certified tissue-based Next Generation Sequencing (NGS) assay results and clinicopathological data from 28 pts with GEP-NETs treated with PRRT at our center from 2017 to 2023. Genetic findings were correlated with progression-free survival (PFS) using Kaplan-Meier estimators and Cox proportional hazards regression modeling. Objective response was assessed per RECIST v1.1 by investigator review. Results: Pts with mutations in MEN1, DAXX, or ATRX (n = 13, 46.42%) had a longer PFS [mPFS, 26.47 months (95% CI, 17.97 - NA) vs. 12.13 months (95% CI, 9.17 - NA); HR, 0.19; 95% CI, 0.06 - 0.68; p = 0.01] than wildtype (n = 15, 53.58%) pts, when adjusted for surgery prior to PRRT and tumor grade. Pts with mutations in MEN1 along with a concurrent mutation in either DAXX or ATRX (n = 6) trended towards better PFS compared to patients without concurrent mutations [only MEN1mt or DAXXmt/ATRXmt, but not both; mPFS, 31.53 months (95% CI, 28.43 - NA) vs. 17.97 months (95% CI, 11.17 - NA); HR, 0.25; 95% CI, 0.05 - 1.29; p = 0.09]. Among evaluable pts, objective response rate was higher in pts with a mt in MEN1, DAXX, or ATRX compared to wild-type pts (41.67% vs. 15.38%; p = 0.09). In the pancreatic NET (PNET) subset, mutations in either MEN1, DAXX, or ATRX were found in 11/16 pts (68.75%) and were also associated with a better PFS compared to wild type patients [mPFS, 28.43 months (95% CI, 11.50 - NA) vs. 9.83 months (95% CI, 7.83 - NA); HR, 0.21; 95% CI, 0.05 - 0.95; p = 0.04). Conclusions: Mutations in MEN1/DAXX/ATRX correlate with improved PFS in patients with GEP-NETs receiving PRRT. Concurrent mutations in MEN1 and DAXX/ATRX may have additive effects on response to PRRT. If validated in larger studies, these findings may represent a novel biomarker to predict PRRT response. Limitations: Limited sample size precludes generalization of results and might overestimate effect sizes. Citation Format: Rushabh Gujarathi, Sara Abou Azar, Joseph Tobias, Xavier Keutgen, Chih-Yi Liao. Mutations in MEN1/DAXX/ATRX are associated with improved progression-free survival in patients with gastroenteropancreatic neuroendocrine tumors receiving peptide receptor radionuclide therapy [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 7614.

Prognostic Value of ATRX and p53 Status in High-Grade Glioma Patients in Morocco.

Glioblastoma and astrocytoma, grade 4, are the most common and aggressive brain tumors. Several biomarkers, such as the isocitrate dehydrogenase mutation (IDH-1), alpha-thalassemia/mental retardation, and the X-linked mutation (ATRX), enable more accurate glioma classification and facilitate patient management. This study aimed to determine the prognostic value of clinical and molecular factors (IDH, TP53, and ATRX mutations). We also studied the relationship between these molecular markers and the overall survival (OS) of 126 patients with grade 4 glioblastoma/astrocytoma. The immunohistochemical study was conducted using antibodies namely, IDH1, R132H, p53, and ATRX. Statistical tests were used to investigate factors that might influence overall survival using IBM SPSS Statistics, version 25.0 (IBM Corp., Armonk, NY). The median age at diagnosis was 51.5 years. Patients with a Karnofsky performance score (KPS) <70 presented less favorable survival outcomes compared to those with a KPS ≥70. The median OSfor patients was found to be 11.17 months. Expression ofIDH1 R132H was found in 13.5% of patients, p53 overexpression was identified in 55.6% of cases, and loss of ATRX expression was detected in 11.9%. The group of patients with IDH mutant/ATRX mutant/p53 wild-typehad the best prognosis (OS= 27.393 months; p = 0.015). Our results were in line with previous studies. The clinical value of IDH and ATRX mutations in prognostic assessment was confirmed (p ≤0.05). The overexpression of p53 had no significant impact on OS (p = 0.726). Therefore, p53 alone cannot predict survival in glioblastoma patients. Based on the results, these biomarkers may be a potential therapeutic target to prolong patient survival, hence the need for further investigations.

Developing Novel Genomic Risk Stratification Models in Soft Tissue and Uterine Leiomyosarcoma.

Leiomyosarcomas (LMS) are clinically and molecularly heterogeneous tumors. Despite recent large-scale genomic studies, current LMS risk stratification is not informed by molecular alterations. We propose a clinically applicable genomic risk stratification model. We performed comprehensive genomic profiling in a cohort of 195 soft tissue LMS (STLMS), 151 primary at presentation, and a control group of 238 uterine LMS (ULMS), 177 primary at presentation, with at least 1-year follow-up. In STLMS, French Federation of Cancer Centers (FNCLCC) grade but not tumor size predicted progression-free survival (PFS) or disease-specific survival (DSS). In contrast, in ULMS, tumor size, mitotic rate, and necrosis were associated with inferior PFS and DSS. In STLMS, a 3-tier genomic risk stratification performed well for DSS: high risk: co-occurrence of RB1 mutation and chr12q deletion (del12q)/ATRX mutation; intermediate risk: presence of RB1 mutation, ATRX mutation, or del12q; low risk: lack of any of these three alterations. The ability of RB1 and ATRX alterations to stratify STLMS was validated in an external AACR GENIE cohort. In ULMS, a 3-tier genomic risk stratification was significant for both PFS and DSS: high risk: concurrent TP53 mutation and chr20q amplification/ATRX mutations; intermediate risk: presence of TP53 mutation, ATRX mutation, or amp20q; low risk: lack of any of these three alterations. Longitudinal sequencing showed that most molecular alterations were early clonal events that persisted during disease progression. Compared with traditional clinicopathologic models, genomic risk stratification demonstrates superior prediction of clinical outcome in STLMS and is comparable in ULMS.

First-in-Human Study of the Ataxia Telangiectasia and Rad3-Related (ATR) Inhibitor Tuvusertib (M1774) as Monotherapy in Patients with Solid Tumors.

Tuvusertib (M1774) is a potent, selective, orally administered ataxia telangiectasia and Rad3-related (ATR) protein kinase inhibitor. This first-in-human study (NCT04170153) evaluated safety, tolerability, maximum tolerated dose (MTD), recommended dose for expansion (RDE), pharmacokinetics (PK), pharmacodynamics (PD), and preliminary efficacy of tuvusertib monotherapy. Ascending tuvusertib doses were evaluated in 55 patients with metastatic or locally advanced unresectable solid tumors. A safety monitoring committee determined dose escalation based on PK, PD, and safety data guided by a Bayesian 2-parameter logistic regression model. Molecular responses (MR) were assessed in circulating tumor DNA samples. Most common grade ≥3 treatment-emergent adverse events were anemia (36%), neutropenia, and lymphopenia (both 7%). Eleven patients experienced dose-limiting toxicities, most commonly grade 2 (n = 2) or 3 (n = 8) anemia. No persistent effects on blood immune cell populations were observed. The RDE was 180 mg tuvusertib QD (once daily), 2 weeks on/1 week off treatment, which was better tolerated than the MTD (180 mg QD continuously). Tuvusertib median time to peak plasma concentration ranged from 0.5 to 3.5 hours and mean elimination half-life from 1.2 to 5.6 hours. Exposure-related PD analysis suggested maximum target engagement at ≥130 mg tuvusertib QD. Tuvusertib induced frequent MRs in the predicted efficacious dose range; MRs were enriched in patients with radiological disease stabilization, and complete MRs were detected for mutations in ARID1A, ATRX, and DAXX. One patient with platinum- and PARP inhibitor-resistant BRCA wild-type ovarian cancer achieved an unconfirmed RECIST v1.1 partial response. Tuvusertib demonstrated manageable safety and exposure-related target engagement. Further clinical evaluation of tuvusertib is ongoing.