CDK4 Amplification Research Articles

Soft Tissue Sarcomas with Chromosomal Alterations in the 12q13-15 Region: Differential Diagnosis and Therapeutic Implications.

The chromosomal region 12q13-15 is rich in oncogenes and contains several genes involved in the pathogenesis of various mesenchymal neoplasms. Notable genes in this region include MDM2, CDK4, STAT6, DDIT3, and GLI1. Amplification of MDM2 and CDK4 genes can be detected in various mesenchymal and nonmesenchymal neoplasms. Therefore, gene amplification alone is not entirely specific for making a definitive diagnosis and requires the integration of clinical, radiological, morphological, and immunohistochemical findings. Neoplasms with GLI1 alterations may exhibit either GLI1 rearrangements or amplifications of this gene. Despite the diagnostic implications that the overlap of genetic alterations in neoplasms with changes in genes within the 12q13-15 region could create, the discovery of coamplifications of MDM2 with CDK4 and GLI1 offers new therapeutic targets in neoplasms with MDM2/CDK4 amplification. Lastly, it is worth noting that MDM2 or CDK4 amplification is not exclusive to mesenchymal neoplasms; this genetic alteration has also been observed in other epithelial neoplasms or melanomas. This suggests the potential use of MDM2 or CDK4 inhibitors in neoplasms where alterations in these genes do not aid the pathological diagnosis but may help identify potential therapeutic targets. In this review, we delve into the diagnosis and therapeutic implications of tumors with genetic alterations involving the chromosomal region 12q13-15, mainly MDM2, CDK4, and GLI1.

Clinicopathological and Molecular Characteristics of IDH-Wildtype Glioblastoma with FGFR3::TACC3 Fusion.

The World Health Organization Classification of Tumors of the Central Nervous System recently incorporated histological features, immunophenotypes, and molecular characteristics to improve the accuracy of glioblastoma (GBM) diagnosis. FGFR3::TACC3 (F3T3) fusion has been identified as an oncogenic driver in IDH-wildtype GBMs. Recent studies have demonstrated the potential of using FGFR inhibitors in clinical trials and TACC3-targeting agents in preclinical models for GBM treatment. However, there is limited information on the clinicopathological and genetic features of IDH-wildtype GBMs with F3T3 fusion. The aim of this study was to comprehensively investigate the clinical manifestations, histological features, and mutational profiles of F3T3-positive GBMs. Between September 2017 and February 2023, 25 consecutive cases (5.0%) of F3T3-positive GBM were extracted from 504 cases of IDH-wildtype GBM. Clinicopathological information and targeted sequencing results obtained from 25 primary and 4 recurrent F3T3-positive GBMs were evaluated and compared with those from F3T3-negative GBMs. The provisional grades determined by histology only were distributed as follows: 4 (26/29; 89.7%), 3 (2/29; 6.9%), and 2 (1/29; 3.4%). Grade 2-3 tumors were ultimately diagnosed as grade 4 GBMs based on the identification of the TERT promoter mutation and the combined gain of chromosome 7 and loss of chromosome 10 (7+/10-). F3T3-positive GBMs predominantly affected women (2.6 females per male). The mean age of patients with an F3T3-positive GBM at initial diagnosis was 62 years. F3T3-positive GBMs occurred more frequently in the cortical locations compared to F3T3-negative GBMs. Imaging studies revealed that more than one-third (12/29; 41.4%) of F3T3-positive GBMs displayed a circumscribed tumor border. Seven of the seventeen patients (41.2%) whose follow-up periods exceeded 20 months died of the disease. Histologically, F3T3-positive GBMs more frequently showed curvilinear capillary proliferation, palisading nuclei, and calcification compared to F3T3-negative GBMs. Molecularly, the most common alterations observed in F3T3-positive GBMs were TERT promoter mutations and 7+/10-, whereas amplifications of EGFR, PDGFRA, and KIT were not detected at all. Other genetic alterations included CDKN2A/B deletion, PTEN mutation, TP53 mutation, CDK4 amplification, and MDM2 amplification. Our observations suggest that F3T3-positive GBM is a distinct molecular subgroup of the IDH-wildtype GBM. Both clinicians and pathologists should consider this rare entity in the differential diagnosis of diffuse astrocytic glioma to make an accurate diagnosis and to ensure appropriate therapeutic management.

To Quiesce or Senesce, That Is the Question for Dedifferentiated Liposarcoma.

Dedifferentiated liposarcoma (DDLPS) has an appealing therapeutic target due to its CDK4 amplification on chromosome 12q. The understanding of geroconversion from quiescent cells to senescent cells defines a patient's response to CDK4 inhibitors. This new observation will inform not only the ongoing phase III clinical trial of abemaciclib, but all future clinical trials in DDLPS. See related article by Gleason et al., p. 703.

Abstract A035: Comparing the clinical and genomic landscapes of acral, mucosal, and cutaneous metastatic melanomas treated with immune checkpoint inhibitors

Abstract Background: Melanoma is a melanocytic malignancy that is classified into different subtypes, including cutaneous (CM), acral (AM), and mucosal melanoma (MM). Immune checkpoint inhibitors (ICI) targeting PD-1 and/or CTLA-4 have emerged as the standard of care for advanced metastatic melanoma. We investigated responses to ICIs, genomic profiles, and molecular differences among these different subtypes of melanoma. Methods: We performed a multi-center retrospective cohort study including patients with metastatic melanoma who received anti-PD1 +/- CTLA4 inhibitor ICI for metastatic disease. We employed the AACR GENIE (v13.1) cancer database, DepMap portal, and Enrichr web tool with MSigDB Hallmark 2020 database to analyze the incidence and distribution of significant alterations, differentially expressed genes, and pathway enrichment within distinct melanoma subtypes. Results: We identified 337 patients with advanced melanoma who received anti-PD1 +/- anti-CTLA4 for metastatic disease. CM was the most frequent melanoma subtype (81%), followed by MM (12%), and AM (8%). Patients with AM had the shortest OS and PFS CM (OS 3.4 years, PFS of 1.1 years), AM median (OS 1.4 years, PFS 3.8 months), and MM (OS 1.7 years, PFS 6 months; OS, P= 0.028; PFS P=0.029). Patients with CM or MM experienced longer OS with anti-PD1 +/- CTLA4 vs. anti-PD1 monotherapy, but no survival advantage was observed in patients with AM. In the GENIE dataset, we identified 2015/374/177 samples with CM/MM/AM, respectively. AM and MM patients were significantly more likely to be Female, Black or Asian than CM patients. BRAF V600 mutations were most frequent in CM (40%), followed by MM (7%) and AM (14%). However, MM and AM had an increase in alterations in cell cycle regulator genes. The incidence of CDK4 and CCND1 amplification, respectively was highest in AM (17%, 16%), followed by MM (6%, 6%) and CM (2%, 3%). We compared the pathways represented by differentially altered genes in AM vs. CM: G2M-Checkpoint (Q<0.05) and E2F targets (Q<0.05) pathways were enriched in AM. We also compared RNAseq data from n=4) AM and n=20 CM cell lines, and found the same pathways enriched amongst genes that were differentially expressed between AM and CM: G2M-Checkpoint (Q<0.01) and E2F targets (Q<0.01) Conclusions: Our findings highlight comparatively poor outcomes with ICIs in Acral Melanoma. AM has lower rates of actionable BRAF mutations; as such, very few patients with metastatic AM have access to effective 1st or 2nd line therapies. Thus, there is an urgent need to explore alternative therapeutic targets. Our data implicate cell cycle inhibitors as potentially important components of novel treatment strategies that could augment immunotherapy efficacy for patients with metastatic AM. Citation Format: Sadaf Solati, April A. N. Rose, Anna Spreafico, Adrian Sacher, Denis Yahiaoui, Wilson H. Miller. Comparing the clinical and genomic landscapes of acral, mucosal, and cutaneous metastatic melanomas treated with immune checkpoint inhibitors [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2023 Oct 1-4; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Immunol Res 2023;11(12 Suppl):Abstract nr A035.

Invasive Pancreatic Adenocarcinoma with Squamous Differentiation Arising in an IOPN with Novel MDM2 Amplification

Abstract Introduction/Objective Intraductal oncocytic papillary neoplasms (IOPNs) are rare pancreatic tumors that are genetically distinct from intraductal papillary mucinous neoplasms (IPMNs). Morphologic and molecular characterization of invasive adenocarcinomas associated with IOPNs remains limited due to the rarity of this entity. Methods/Case Report We present a 68-year-old female who initially presented with abdominal pain and jaundice. MRI revealed a 2.6 cm mass in the pancreatic head with abutment of the superior mesenteric vein and dilation of the biliary and pancreatic duct. Biopsy revealed adenocarcinoma with mucinous features. The patient was started on gemcitabine and abraxane. Her treatment course was complicated by biliary and duodenal obstruction requiring common bile duct and duodenal stenting. Surgical management with pancreaticoduodenectomy was performed six months following initial diagnosis. Gross examination revealed a 2.5 cm ill-defined tumor surrounding the main pancreatic duct adjacent to the common bile duct. Microscopy revealed an IOPN arising within the pancreatic duct exhibiting a complex papillary growth pattern. There was an associated moderately to poorly differentiated invasive carcinoma with mucinous and squamous differentiation with lymph node metastasis. Molecular testing (FoundationOne®CDx) revealed ARID1A mutation and amplification of MDM2, CDK4, and ERBB3. Patient is alive and without radiologic evidence of disease recurrence at 6 months follow-up. Results (if a Case Study enter NA) NA Conclusion To our knowledge, our case is the first report of an invasive pancreatic adenocarcinoma with squamous differentiation arising in association with an IOPN. In addition, our case showed MDM2 amplification which has not been previously identified in pancreatic adenocarcinoma arising in IOPNs. Although recent literature demonstrated that less than 30% of IOPN showed invasive disease and even fewer cases with lymph node metastasis, our case showed extensive invasion with metastases to 2 lymph nodes. This may suggest that MDM2 amplification may play a role in more aggressive disease since it is implicated with poorer outcome in conventional pancreatic adenocarcinoma. Further molecular studies of IOPN may help to identify prognostic features of this entity.

Genetic alterations that deregulate RB and PDGFRA signaling pathways drive tumor progression in IDH2-mutant astrocytoma

In IDH-mutant astrocytoma, IDH2 mutation is quite rare and biological mechanisms underlying tumor progression in IDH2-mutant astrocytoma remain elusive. Here, we report a unique case of IDH2 mutant astrocytoma, CNS WHO grade 3 that developed tumor progression. We performed a comprehensive genomic and epigenomic analysis for primary and recurrent tumors and found that both tumors harbored recurrent IDH2R172K and TP53R248W mutation with CDKN2A/B hemizygous deletion. We also found amplifications of CDK4 and MDM2 with PDGFRA gain in the recurrent tumor and upregulated protein expressions of these genes. We further developed, for the first time, a xenograft mouse model of IDH2R172K and TP53R248W mutant astrocytoma from the recurrent tumor, but not from the primary tumor. Consistent with parent recurrent tumor cells, amplifications of CDK4 and MDM2 and PDGFRA gain were found, while CDKN2A/B was identified as homozygous deletion in the xenografts, qualifying for integrated diagnosis of astrocytoma, IDH2-mutant, CNS WHO grade 4. Cell viability assay found that CDK4/6 inhibitor and PDGFR inhibitor potently decreased cell viability in recurrent tumor cells, as compared to primary tumor cells. These findings suggest that gene alterations that activate retinoblastoma (RB) signaling pathways and PDGFR may drive tumor progression and xenograft formation in IDH2-mutant astrocytoma, which is equivalent to progressive IDH1-mutant astrocytoma. Also, our findings suggest that these genomic alterations may represent therapeutic targets in IDH2-mutant astrocytoma.

PATH-34. CLINICOPATHOLOGIC CHARACTERIZATION OF DIFFUSE PEDIATRIC-TYPE HIGH-GRADE GLIOMA, H3-WILDTYPE AND IDH-WILDTYPE: AN AGGRESSIVE AND GENOMICALLY COMPLEX GROUP OF TUMORS AFFECTING CHILDREN AND ADULTS

Abstract Diffuse pediatric-type high-grade glioma, H3-wildtype and IDH-wildtype (pHGG) was newly introduced in the 2021 WHO CNS5. pHGG is further divided into distinct methylation subtypes (MYCN, A/B, RTK1, RTK2). Compared to adult-type diffuse gliomas, relatively little is known about pHGG, especially in the adult population and we therefore aimed to more fully understand the clinicopathological spectrum of pHGG. Our cohort of methylation-profiling-confirmed pHGGs (n=299) included pediatric (n=211) and adult (n=88) populations (age range=0-79 years). The mean age was highest in the A/B subtypes (27.7 years) and lowest in the MYCN subtype (13.5 years). Median survival (18 months for the entire cohort), MGMT promoter methylation status, and history of prior radiation did not differ between the pediatric and adult populations. MGMT promoter methylation was highest in RTK1C (51%) and lowest in subtype A (0%). All those with history of prior radiation (n=37) were classified as RTK1, with subclass distribution differing between patients with history of prior radiation (RTK1A=7.3%, RTK1B=41.8%, RTK1C=50.9%) versus those without (RTK1A=41.8%, RTK1B=21.5%, RTK1C=36.7%). Interestingly, the genomic somatic copy number alteration (SCNA) load did not differ for RTK1 based on prior radiation history status. However, adult pHGG did harbor a higher SCNA load than pediatric pHGG (p< 0.01). The higher SCNA load was driven by genomic copy number losses rather than amplifications, including adult pHGG harboring more CDKN2A/B and RB1 deletions. Amplifications of MET, CDK4, PDGFRA, MYC, and KIT were present across all subtypes, but were enriched in RTK1. MYCN amplification was enriched in the MYCN subtype. EGFR, MDM4, and CDK6 amplifications did not have subtype predilection. Overall, while the pediatric and adult pHGG have similar outcomes, there are differences in their genomic structures, most notably in adult pHGG harboring a higher SCNA load. Overall, pHGG represent a biologically diverse set of clinically aggressive tumors that warrant further intensive study.

Osteosarcomas With Few Chromosomal Alterations or Adult Onset Are Genetically Heterogeneous

Osteosarcoma is the most common primary bone malignancy, often detected in children and adolescents and commonly associated with TP53 alterations along with a high number of chromosomal rearrangements. However, osteosarcoma can affect patients of any age, and some tumors display less genetic complexity. Besides TP53 variants, data on key driving mutations are lacking for many osteosarcomas, particularly those affecting adults. To detect osteosarcoma-specific alterations, we screened transcriptomic and genomic sequencing and copy number data from 150 bone tumors originally diagnosed as osteosarcomas. To increase the precision in gene fusion detection, we developed a bioinformatic tool denoted as NAFuse, which extracts gene fusions that are verified at both the genomic and transcriptomic levels. Apart from the already reported genetic subgroups of osteosarcoma with TP53 structural variants, or MDM2 and/or CDK4 amplification, we did not identify any recurrent genetic driver that signifies the remaining cases. Among the plethora of mutations identified, we found genetic alterations characteristic of, or similar to, those of other bone and soft tissue tumors in 8 cases. These mutations were found in tumors with relatively few other genetic alterations or in adults. Due to the lack of clinical context and available tissue, we can question the diagnosis only on a genetic basis. However, our findings support the notion that osteosarcomas with few chromosomal alterations or adult onset seem genetically distinct from conventional osteosarcomas of children and adolescents.

Phase II trial of CDK4/6 inhibitor palbociclib in advanced sarcoma based on mRNA expression of CDK4/ CDKN2A

Cyclin-dependent kinases 4 and 6 (CDK4/6) inhibitors demonstrated activity in terms of progression-free survival (PFS) in advanced dedifferentiated liposarcoma (DD-LPS), a sarcoma with CDK4 amplification. CDK4 overexpression is by far more common than amplification in sarcomas and it might be a rational target for CDK inhibitors. Preclinical investigators of this study found that CDK4 overexpression, while not of CDKN2A, was the most consistent predictive factor for palbociclib efficacy in sarcomas. Advanced adult-type soft-tissue sarcoma, excluding DD-LPS, or bone sarcoma patients, progressing after at least one systemic line, whose tumors overexpressed CDK4, but not CDKN2A at baseline biopsy, were accrued in this single-arm phase II trial (EudraCT number: 2016-004039-19). With the main endpoint of a 6-month PFS rate, 40% was considered promising in this population. Palbociclib was administered orally at 125 mg/day for 21 days in 28-day cycles. A total of 214 patients with 236 CDK4/CDKN2A determinations were assessed for prescreening, archival material (141), and screening, baseline biopsy (95). There were 28 (29%) with favorable mRNA profiles from 95 screened patients at baseline. From 23 enrolled patients, 21 evaluable, the 6-month PFS rate was 29% (95% CI 9–48), and there were 6 patients out of 21 with a PFS longer than 6 months. The median PFS and overall survival were 4.2 (95% CI 3.6–4.8) and 12 (95% CI 8.7–15.4) months, respectively. Translational research showed a significant correlation between CDK4 mRNA and protein expression. Palbociclib was active in a variety of sarcoma subtypes, selected by CDK4/CDKN2A, and deserves further investigation in the sarcoma context.

Liposarcoma Involving Serous Fluid Cavities-A Case Series Illustrating Clinical Implications and the Diagnostic Role of Exfoliative Cytology.

Introduction: Cytological diagnosis of sarcomas requires detailed cytomorphological assessment and integration of immunocytochemistry and/or molecular testing. The role of exfoliative cytology, as compared to aspiration cytology, is less understood. This case series describes well-differentiated/dedifferentiated liposarcomas in effusions, with cytomorphological features, ancillary test results and clinical outcomes detailed. Methods: A computerized search of the department pathology archives was performed for sarcomatous effusions with histological diagnosis or clinical history of well-differentiated/dedifferentiated liposarcoma. Clinical progress, cytology slides, immunocytochemistry and molecular test results were reviewed. Results: Six patients were identified. In 5 patients with clinical follow up, 4 (80%) were deceased within 5 months of malignant effusion. One patient was alive with 12 years disease-free survival after radical resection with adjuvant radiotherapy. Three patients showed dedifferentiation on histology, and high-grade (dedifferentiated) tumor cells were present in effusion cytology of 2 patients. Two showed well-differentiated components only on biopsy, but high-grade (dedifferentiated) tumor cells were identified in cytology. The high-grade tumor cells displayed marked nuclear irregularity, enlargement, size variation, with macronucleoli and multinucleation. Well-differentiated lipomatous components were demonstrated in 4 patients (66.7%), comprising of multivacuolated lipoblasts and atypical lipocytes. CDK4 and MDM2 immunoreactivity in all 3 cases with cell blocks, and CDK4 and MDM2 amplification in one were successfully demonstrated. Conclusion: Lipomatous and dedifferentiated components can be sampled and cytomorphologically identified on effusion fluids of liposarcomas, with sufficient cellularity for immunocytochemistry and molecular testing. Although generally associated with poor prognosis, long disease-free survival with sarcomatous effusion is possible with radical surgery and adjuvant treatment.

207P Palbociclib (P) in patients (pts) with solid tumors with CDK4 or CDK6 amplification (amp): Results from the Targeted Agent and Profiling Utilization Registry (TAPUR) study

207P Palbociclib (P) in patients (pts) with solid tumors with CDK4 or CDK6 amplification (amp): Results from the Targeted Agent and Profiling Utilization Registry (TAPUR) study

OS12.5.A HIGH-GRADE GLIOMAS OCCURRING IN TEENAGERS AND YOUNG ADULT PATIENTS COMPRISE NOVEL MOLECULAR SUBGROUPS

Abstract BACKGROUND High-grade gliomas (HGG) arise in any central nervous system location, in adults and children, with a poor prognosis. HGGs in teenagers and young adults (TYA) are understudied; this project aimed to characterise these tumours and identify potential therapeutic targets. MATERIAL AND METHODS HGG samples (n=195, FFPE/FF, 13-30 years) were collected from national/international collaborators, excluding well-characterised entities (histone/IDH-mutant). DNA methylation profiling (Illumina EPIC BeadArrays, brain tumour classifier (MNP v12.5 R package), n=195) classified cases against a reference cohort. Calibrated scores guided workflows to characterise mutational landscapes including RNA-based ArcherDx fusion panel (n=112), whole exome sequencing (n=84), and histological reviews in parallel. Well-characterised entities (n=50, including IDH-mutant tumours (n=18), PXA (n=12)) were excluded. RESULTS Of the cases scoring >0.6, 64% classified as paediatric-type subgroups (pedHGG_RTK1A/B/C (30%), pedHGG_MYCN (10%), pedHGG_A/B (7%), pedHGG_RTK2A (6%)). 29% classified as subgroups more frequently seen in adults (GBM_MES (12%), GBM_RTK1/2 (3%)). 54% classified with a score <0.6. 38% cases were assigned to novel, recently identified, poorly-characterised subgroups with distinct methylation profiles and molecular features (HGG_B, HGG_E, ANTCON, GBM_CBM). Frequency comparisons to publicly available methylation data showed GBM_MES_ATYP tumours are TYA age-specific, in contrast to adult age-specific GBM_MES_TYP. Copy number profiling identified frequent changes across the cohort, including chromosomal gains (chr1q (54%), chr2 (22%), chr7 (41%)) and losses (chr10 (40%), chr13 (64%)). Focal amplifications included PDGFRA (12%), CDK4 (7%), MYCN/ID2 (3%), MYC (2%) and EGFR (0.7%) and the most common focal deletion was CDKN2A/p16 (12%). The most frequent glioma-associated somatic variants included TP53, PDGFRA, EGFR, and NF1. BCOR alterations were enriched compared to adult and paediatric reference cohorts. 10% cases showed a hypermutator phenotype, enriched in HGG_E. CDK4 amplifications were recurrent in GBM_RTK1 and GBM_CBM. Histology showed variable cytological and architectural features within novel subgroups. CONCLUSION TYA HGG comprise novel subgroups with distinct methylation profiles and molecular characteristics, representing opportunities to refine future treatment.

P02.23.A MDM2/X ALTERATIONS IN A MULTICOHORT RETROSPECTIVE ANALYSIS: A POTENTIAL TARGET OF TREATMENT IN PATIENTS WITH PRIMARY BRAIN TUMORS

Abstract BACKGROUND Primary brain tumors (pBTs) are a heterogeneous group of neoplasms with limited therapeutic alternatives. Disruption of the TP53 tumor suppressor pathway occurs in most high-grade gliomas. While TP53 loss-of-function mutations are poorly targetable, in 14% of patients this disruption is due to a Mouse double minute 2 homolog (MDM2) amplification. We aim to describe the characteristics of patients with MDM2 amplification and analyze its clinical and molecular factors using a cohort of patients with NGS data. MATERIAL AND METHODS Patients currently enrolled in the institutionally approved, retrospective multicenter study ESCAT-pBTs, which includes patients with pBT profiled used NGS methods (Foundation Medicine®, local NGS, Caris®, Oncomine®, copy number alterations (CNA), and fusion panels) were analyzed to determine the presence of MDM2 alterations (MDM2alt) and clinical factors including WHO 2021 diagnosis, sex, age, ECOG and other molecular findings. RESULTS A total of 493 pts with NGS and CNA were collected between Feb 2018 and January 2023 at 5 hospitals in Spain. The median age was 50.3 (range, 3.3-83.8), 14.2% had ECOG ≥2, 39.1% were women, 26.9% were 40 or younger, 21.5% had IDH1mut and 24.3% were methylated. The diagnosis of GBM (WHO 2021) was present in 68.8% of the patients and 33.8% had mutations or CNV in TP53. The overall prevalence of MDM2alt was 8.5%(42 pts), 92.9% gene amplifications, and 7,1% mutations. No differences were found in gender proportion between MDM2alt and no MDM2alt (Female: 37.1% vs 39.4% p:0.64) and no major variance in the percentage of ECOG ≥2 (16.6% vs 17.6% p:0.77). Prevalence in patients with GBM was: 11.8% vs 1.3% in patients with other pBTs. Concurrent mutations in MDM2alt patients were: CDK-4 copy number gain in 76,2%; a similar proportion of mutations in PI3KCA/PTEN was observed compared with the non-MDM2alt population in the pts with GBM (38,3% vs 50%, p:0.15). No significant variation was found in EGFR amplifications (42,9% vs 32% p:0.10) and EGFR mutations GBM MDM2 and no MDM2alt respectively (19% vs 18,7% p: 0.84). The median TMB was 5.22 (range from 2.24 to 13 mutations/megabase) in pts with MDM2alt and 4.76 (range from 1.26 to 53.7 mutations/megabase). 97,6% of the MDM2alt population was TP53 wild type. CONCLUSION MDM2alt is a common genomic event in patients with molecularly defined GBM and has a low prevalence in other pBT. The presence of MDM2alt is associated in our series with other genomic events such as PTEN loss, low TMB, and a high degree of CDK4 amplification. No statistically significant differences were found between the GBM MDM2alt and no MDM2alt subgroup in terms of molecular alterations and clinical aspects studied. These findings should inform the further clinical development of treatment strategies for MDM2alt pts.

The Landscape of Alterations from 1407 Ultra-Rare Sarcomas from the AACR GENIE Database: Clinical Implications.

Ultra-rare sarcomas (URS) comprise a group of orphan diseases with an incidence of ≤1/1,000,000 people per year. We aimed to assess clinically actionable genomic alterations in URS. Data were extracted from the GENIE database using cBioPortal. OncoKB was used to assess for clinical actionability of mutations. Tumor mutational burden (TMB) was inferred from clinical sequencing data. Soft tissue (ST) URS made up 23.5% of ST sarcoma cases, and bone URS made up 16.5% of bone sarcoma cases. The most commonly mutated gene in all four groups was TP53. The most common fusions involved EWSR1. The most common copy-number variations included deletions of CDKN2A and CDKN2B and amplifications of MDM2 and CDK4. TMB was generally low across all four categories of sarcoma, though there was considerable heterogeneity, with 3.8% of ST URS and 0.55% of bone URS having high TMB. We find Level 1 alterations (FDA-recognized biomarker predictive of response to an FDA-approved drug) in 10.0% of ST URS compared with 7.1% of ST non-URS, 1.1% of bone URS, and 4.5% of bone non-URS. Level 1-3 alterations (also include alterations for which there are standard-of-care drugs or clinical evidence supporting a drug) were seen in 27.8% of ST URS, 25.2% of ST non-URS, 20.9% of bone URS, and 17.4% of bone non-URS. Clinically actionable genomic alterations are seen in a substantial fraction of URS. Clinical sequencing in advanced URS has the potential to guide the treatment of a significant portion of patients with URS.

Molecular-based targeted therapies in patients with hepatocellular carcinoma and hepato-cholangiocarcinoma refractory to atezolizumab/bevacizumab

The "French Medicine Genomic program 2025" has been designed to give patients with cancers that are refractory to systemic treatments access to off-label therapies adapted to their specific genomic profile. Herein, we reported the results of this program in patients with advanced hepatocellular carcinoma (HCC) and hepato-cholangiocarcinoma (H-CCK). In one center, all patients with HCC or H-CCK who progressed under atezolizumab/bevacizumab with available tumor frozen samples benefited from whole-genome/-exome and RNA sequencing. Targeted therapies were matched to genomic alterations following the recommendations of a molecular tumor board and radiological response and overall survival were assessed. Among 135 patients with HCC and H-CCK treated by atezolizumab/bevacizumab, 20 patients benefited from genomic analysis after progression (16 HCC; 4H-CCK). Nineteen patients had analyzable data, 70% were male, median age was 57 years, 65% had metastatic disease and 45% had vascular invasion. Among these 19 patients, 14 patients (76%) harbored at least one actionable genomic alteration and 9/14 received an adapted targeted therapy (45%). One patient with H-CCK showing CDK4 amplification was treated with palbociclib and achieved a partial radiological response for 16 months. Another patient with H-CCK, high HER2 overexpression and a high homologous recombination score was treated with trastuzumab/olaparib and had stable disease. One patient with an HCC and bi-allelic inactivation of TSC2 achieved a complete radiological response under everolimus. The remaining six treated patients (all HCC) had progressive disease, including three patients treated with trametinib, two with everolimus and one with olaparib. Molecular-based guided therapy is feasible in patients with HCC/H-CCK progressing under atezolizumab/bevacizumab and may be useful in a small subset of patients. The use of whole-genome/-exome and RNA sequencing in clinical practice has not been reported in patients with hepatocellular carcinoma and hepato-cholangiocarcinoma. Herein, we performed a pilot study which suggested that whole-genome/-exome and RNA sequencing is feasible on tumor biopsies from patients refractory to atezolizumab/bevacizumab, with a small subset of patients exhibiting at least one actionable genomic alteration and receiving an adapted targeted therapy. This proof-of-concept study suggests that this clinical strategy could benefit a small subset of patients. Finally, validation of this approach will be required in a larger cohort of patients.

A threshold for mitotic activity and post-surgical residual volume defines distinct prognostic groups for astrocytoma IDH-mutant.

The distinction between CNS WHO grade 2 and grade 3 is instrumental in choosing between observational follow-up and adjuvant treatment for resected astrocytomas IDH-mutant. However, the criteria of CNS WHO grade 2 versus 3 have not been updated since the pre-IDH era. Maximal mitotic activity in consecutive high-power fields corresponding to 3 mm2 was examined for 118 lower-grade astrocytomas IDH-mutant. The prognostic value for time-to-treatment (TTT) and overall survival (OS) of mitotic activity and other putative prognostic factors (including age, performance status, pre-surgical tumour volume, multilobar involvement, post-surgical residual tumour volume, midline involvement) was assessed for tumours with ATRX loss and the absence of CDKN2A homozygous deletion or CDK4 amplification, contrast enhancement, histological necrosis, and microvascular proliferation. Seventy-one per cent of the samples had < 6 mitoses per 3 mm2 . Mitotic activity, residual volume, and multilobar involvement were independent prognostic factors of TTT. The threshold of ≥ 6 mitoses per 3 mm2 identified patients with a shorter TTT (median 18.5 months). A residual volume ≥ 1 cm3 also identified patients with a shorter TTT (median 24.5 months). The group defined by < 6 mitoses per 3 mm2 and a residual volume < 1 cm3 had the longest TTT (median 73 months) and OS (100% survival at 7 years). These findings were confirmed in a validation cohort of 52 tumours. Mitotic activity and post-surgical residual volume can be combined to evaluate the prognosis for patients with resected astrocytomas IDH-mutant. Patients with < 6 mitoses per 3 mm2 and a residual volume < 1 cm3 were the best candidates for observational follow-up.

ROS1 Alterations as a Potential Driver of Gliomas in Infant, Pediatric, and Adult Patients

Gliomas harboring oncogenic ROS1 alterations are uncommon and primarily described in infants. Our goal was to characterize the clinicopathological features and molecular signatures of the full spectrum of ROS1 fusion–positive gliomas across all age groups. Through a retrospective multi-institutional collaboration, we report a collection of unpublished ROS1 fusion gliomas along with the characterization and meta-analysis of new and published cases. A cohort of 32 new and 58 published cases was divided into the following 3 age groups: 19 infants, 40 pediatric patients, and 31 adults with gliomas. Tumors in infants and adults showed uniformly high-grade morphology; however, tumors in pediatric patients exhibited diverse histologic features. The GOPC::ROS1 fusion was prevalent (61/79, 77%) across all age groups, and 10 other partner genes were identified. Adult tumors showed recurrent genomic alterations characteristic of IDH wild-type glioblastoma, including the +7/−10/CDKN2A deletion; amplification of CDK4, MDM2, and PDGFRA genes; and mutations involving TERTp, TP53, PIK3R1, PIK3CA, PTEN, and NF1 genes. Infant tumors showed few genomic alterations, whereas pediatric tumors showed moderate genomic complexity. The outcomes were significantly poorer in adult patients. Although not statistically significant, tumors in infant and pediatric patients with high-grade histology and in hemispheric locations appeared more aggressive than tumors with lower grade histology or those in nonhemispheric locations. In conclusion, this study is the largest to date to characterize the clinicopathological and molecular signatures of ROS1 fusion–positive gliomas from infant, pediatric, and adult patients. We conclude that ROS1 likely acts as a driver in infant and pediatric gliomas and as a driver or codriver in adult gliomas. Integrated comprehensive clinical testing might be helpful in identifying such patients for possible targeted therapy.

A comprehensive genomic study of 390 H3F3A-mutant pediatric and adult diffuse high-grade gliomas, CNS WHO grade 4.

Malignant brain tumors, known as H3K27-altered diffuse midline glioma (DMG) and H3G34-mutant diffuse hemispheric glioma (DHG), can affect individuals of all ages and are classified as CNS WHO grade 4. We comprehensively characterized 390 H3F3A-mutant diffuse gliomas (201 females, 189 males) arising in pediatric patients (under 20years old) and adults (20years and older) evaluated by the CGP program at Foundation Medicine between 2013 and 2020. We assessed information from pathology reports, histopathology review, and clinical data. The cohort included 304 H3K27M-mutant DMG (156 females, 148 males) and 86 H3G34-mutant DHG (45 females, 41 males). Median patient age was 20years (1-74years). The frequency of H3K27M-mutant DMG was similar in both pediatric and adult patients in our cohort-48.6% of the patients were over 20years old, 31.5% over 30, and 18% over 40 at initial diagnosis. FGFR1 hotspot point mutations (N546K and K656E) were exclusively identified in H3K27M-mutant DMG tumors (64/304, 21%; p = 0.0001); these tend tooccur in older patients (median age: 32.5years) and mainly arose in the diencephalon. H3K27M-mutant DMG had higher rates of mutations in NF1 (31.0 vs 8.1%; p = 0.0001) and PIK3CA/PIK3R1 (27.9% vs 15.1%; p = 0.016) compared to H3G34-mutant DHG. However, H3G34-mutant DHG had higher rates of targetable alterations in cell-cycle pathway genes (CDK4 and CDK6 amplification; CDKN2A/B deletion) (27.0 vs 9.0%). Potentially targetable PDGFRA alterations were identified in ~ 20% of both H3G34-mutant DHG and H3K27M-mutant DMG. Overall, in the present study H3K27M-mutant DMG occurred at similar rates in both adult and patient patients. Through our analysis, we were able to identify molecular features characteristic of DMG and DHG. By identifying the recurrent co-mutations including actionable FGFR1 point mutations found in nearly one-third of H3K27M-mutant DMG in young adults, our findings can inform clinical translational studies, patient diagnosis, and clinical trial design.

HGG-15. HIGH-GRADE GLIOMAS OCCURRING IN TEENAGERS AND YOUNG ADULT PATIENTS COMPRISE NOVEL MOLECULAR SUBGROUPS

Abstract High-grade gliomas (HGG) arise in any central nervous system location, in adults and children, with a poor prognosis. HGGs in teenagers and young adults (TYA) are understudied; this project aimed to characterise these tumours and identify potential therapeutic targets. HGG samples (n=195, FFPE/FF, 13-30 years) were collected, excluding well-characterised entities (histone/IDH-mutant). DNA methylation profiling (Illumina EPIC BeadArrays, brain tumour classifier (MNP v12.5 R package), n=195) classified cases against a reference cohort. Calibrated scores guided workflows to characterise mutational landscapes including RNA-based ArcherDx fusion panel (n=112), whole exome sequencing (n=84), and histological reviews in parallel. Well-characterised entities (n=50, including IDH-mutant tumours (n=18), PXA (n=12)) were excluded. Of the cases scoring &amp;gt;0.6, 64% classified as paediatric-type subgroups (pedHGG-RTK1A/B/C (30%), pedHGG-MYCN (10%), pedHGG-A/B (7%), pedHGG-RTK2A (6%)). 29% classified as subgroups more frequently seen in adults (GBM-MES (12%), GBM-RTK1/2 (3%)). 54% classified with a score &amp;lt;0.6. 38% cases were assigned to novel, recently identified, poorly-characterised subgroups with distinct methylation profiles and molecular features (HGG-B, HGG-E, ANTCON, GBM-CBM). Frequency comparisons to publicly available methylation data showed GBM-MES-ATYP tumours are TYA age-specific, in contrast to adult age-specific GBM-MES-TYP. Copy number profiling identified frequent changes across the cohort, including chromosomal gains (chr1q (54%), chr2 (22%), chr7 (41%)) and losses (chr10 (40%), chr13 (64%)). Focal amplifications included PDGFRA (12%), CDK4 (7%), MYCN/ID2 (3%), MYC (2%) and EGFR (0.7%) and the most common focal deletion was CDKN2A/p16 (12%). The most frequent glioma-associated somatic variants included TP53, PDGFRA, EGFR, and NF1. BCOR alterations were enriched compared to adult and paediatric reference cohorts. 10% cases showed a hypermutator phenotype, enriched in HGG-E. CDK4 amplifications were recurrent in GBM-RTK1 and GBM-CBM. Histology showed variable cytological and architectural features within novel subgroups. TYA HGG comprise novel subgroups with distinct methylation profiles and molecular characteristics, representing opportunities to refine future treatment.

Genomic landscape of CDK6 amplification and its potential prognostic value in adult-type diffuse gliomas.

e14038 Background: Glioma is commonly described as a disease of neuroglial cell proliferation. And the gene related to cell cycle pathway alter frequently in glioma including CDKN2A/B, CDK4 and CDK6. CDKN2A/B homozygous deletion is proved to be associated with worse survival in IDH-mut gliomas and meningiomas. However, the correlation between CDK4/6 amplification and the prognosis of glioma remains unclear. Herein, we explored the distribution of CDK4/6 alterations and whether they are the prognostic biomarker of glioma. Methods: A total of 90 patients with adult-type diffuse glioma were enrolled to this retrospective study from our center. Whole genome sequencing (WES) was performed to identify the mutations of the cohort. Results: CDK4 and CDK6 amplified in 17 (18.9%) and 12 (13.3%) patients, respectively. Four patients harbored both CDK4 and CDK6 amplification. The Cox multivariate regression revealed only CDK6 amplification was the independent factor (HR 4.0, 95% CI 1.1-15.2, p=0.041) of overall survival (OS). Interestingly, CDK6 amplification occurred easier in female and trended to be co-mutant with CDKN2A/B homozygous deletion. CDK6 amplification was found to be significantly associated with shorter OS ( p&lt;0.05) of glioblastoma (GBM). Although TERTp was not related to the OS of GBM according to TCGA, patients harboring both TERTp and CDK6 amplification had the significant worst OS ( p&lt;0.001). Conclusions: Our study compared the mutational characteristics of gliomas with CDK4/6 alterations and demonstrated the potential prognostic value of CDK6 amplification, and provide the new basis of precise diagnosis and treatment in gliomas.