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Purpose: Anaplastic lymphoma kinase (ALK) aberrations have been identified in pediatric-type infant gliomas, but their occurrence across age groups, functional effects, and treatment response has not been broadly established. Experimental Design: We performed a comprehensive analysis of ALK expression and genomic aberrations in both newly generated and retrospective data from 371 glioblastomas (156 adult, 205 infant/pediatric, and 10 congenital) with in vitro and in vivo validation of aberrations. Results: ALK aberrations at the protein or genomic level were detected in 12% of gliomas (45/371) in a wide age range (0–80 years). Recurrent as well as novel ALK fusions (LRRFIP1–ALK, DCTN1–ALK, PRKD3–ALK) were present in 50% (5/10) of congenital/infant, 1.4% (3/205) of pediatric, and 1.9% (3/156) of adult GBMs. ALK fusions were present as the only candidate driver in congenital/infant GBMs and were sometimes focally amplified. In contrast, adult ALK fusions co-occurred with other oncogenic drivers. No activating ALK mutations were identified in any age group. Novel and recurrent ALK rearrangements promoted STAT3 and ERK1/2 pathways and transformation in vitro and in vivo. ALK-fused GBM cellular and mouse models were responsive to ALK inhibitors, including in patient cells derived from a congenital GBM. Relevant to the treatment of infant gliomas, we showed that ALK protein appears minimally expressed in the forebrain at perinatal stages, and no gross effects on perinatal brain development were seen in pregnant mice treated with the ALK inhibitor ceritinib. Conclusions: These findings support use of brain-penetrant ALK inhibitors in clinical trials across infant, pediatric, and adult GBMs.

Background: Breast cancer brain metastasis (BCBM) represents an area of unmet clinical need. We previously demonstrated the genomic differences between primary BC and BCBM. In this study we examined the genomic differences between BCBM by BC subtypes and the potential actionable targets that might be taken forward for each subtype in clinical trials. Material and Methods: A total of 761 BCBMs were analyzed by comprehensive genomic profiling (CGP) for alterations in up to 395 genes (Foundation Medicine, USA). The samples were classified by immunohistochemistry (IHC) of the BCBM to ER+ (ER+/HER2-, ER+/HER2+), ER-/HER2+ and ER-/HER2-. Homologous recombination deficiency (HRD-gLOH; cutoff 16%), tumour mutational burden (TMB; cutoff 10 mutations/Mb), microsatellite instability (MSI) and PD-L1 prevalence and expression by IHC using the VENTANA SP142 assay (Immune Cell Score [IC] cutoff 1%) were also investigated. Results: For all BC subtypes the most enriched gene alterations in BCBM (>20% prevalence) were: TP53, MYC and PIK3CA. ESR1 and ERBB2 were more prevalent in ER+ and HER2+ tumours respectively, with ESR1 alterations significantly enriched in ER+/HER2- BCBMs (p< 0.0001). Frequently altered genes by BCBM subtype were: ER+/HER2-: CDH1 (8%) and BRCA2 (7%); ER+/HER2+: PIK3C2B (11%), MDM4 (11%), TBX3 (9%) and AKT2 (8%); ER-/HER2+: LYN (9%). Significantly enriched genes (p< 0.01) by BCBM subtype were: HER2+: CDK12 (15%); ER-/HER2-: BRCA1 (14%), CCND3 (9%), VEGFA, JAK2 (8% each) and the immune checkpoint inhibition (ICPI) biomarkers PDCD1LG2 (PDL2), CD274 (PDL1) (7% each). HRD-gLOH was high in ER+/HER2-: (43%) and ER-/HER2-: (70%). TMB and MSI were present in 10%-21% and 1-3% of BCBM respectively, whereas PDL1 protein expression by subtypes was: ER+/HER2- 23%, ER+/HER2+ 27%, ER-/HER2+ 57% , ER-/HER2- 48%. Table 1 summarizes the proportion of BCBMs with actionable alterations and selected clinical trials in BCBM. Conclusion: Clinically-relevant genomic alterations were identified across all BCBM subtypes. High HRD-gLOH, TMB and MSI were observed across all the BCBMs subtypes whereas PDL1/PDL2 alterations were a distinctive feature of ER-/HER2- BCBM. These data highlight the need to assess the genomic landscape of BCBM to enable rational treatment decisions with targeted agents, as well as to enable enrichment of relevant populations within clinical trials. Table 1. Selected studies with targeted therapy and/or immunotherapy in breast cancer brain metastases. The percentage (%) of actionable alterations by ER and HER2 status is summarized. ND: Not Detectable. Citation Format: Athina Giannoudis, Ethan Sokol, Shakti Ramkissoon, Talvinder Bhogal, Jeff Ross, Kimberly McGregor, Evangelia Razis, Rupert Bartsch, Richard S. Huang, Carlo Palmieri. Targetable alterations and genomic signatures within breast cancer brain metastases: Data from comprehensive genomic profiling of 761 breast cancer brain metastases [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P5-14-10.

Supplementary Figure 3: PPP1CB-ALK fusion is oncogenic and sensitive to targeted therapy. A. Immunoblot analysis of ALK downstream signaling proteins in cortical mNSC (CTX#6) and brainstem mNSC (BS#3) expressing PPP1CB-ALK. Cells were treated for 4 hours with ceritinib and lorlatinib at the indicated concentrations.B. Left panel: Soft agar colony-forming assay using stable mNSC PPP1CB-ALK demonstrated colony formation with dose-dependent inhibition by ceritinib. mNSC KRAS did not respond to ceritinib validating the drug specificity. Right panel: Quantification of colony formation under targeted drug inhibition using CellProfiler. Values represent colony counts relative to the untreated group ± s.d. The mean of three independent replicates is shown. Significance between treatments determined by the Mann-Whitney test. *P < 0.05, **P < 0.01.C. Cell viability of ceritinib-treated PPP1CB-ALK NIH-3T3 relative to eGFP-positive cells by CellTiterGlo.D. Relative viability of the ALK wild-type GBM line (BT164) and BT1857 lorlatinib-treated cells versus DMSO control.E. Relative viability of BT164 and BT1857 ceritinib-treated cells versus DMSO control.F. Relative viability of BT164 and BT1857 cells treated with an EGFR tyrosine kinase inhibitor neratinib.G. Pharmacodynamic analysis of NIH 3T3-PPP1CB-ALK s.c. tumors treated with vehicle or ceritinib at 30mg/kg for 5 days. Representative H&E stain, ALK, pSTAT3, pAKT S473 and Ki-67 IHC.H. Quantification of Ki-67-positive cells in vehicle and ceritinib-treated tumors using CellProfiler software. Error bars show standard error of the mean. *P < 0.05.I. Quantification of pSTAT3-positive cells in vehicle and ceritinib-treated tumors using CellProfiler software. Error bars show standard error of the mean. *P < 0.05.J. Tumor growth following subcutaneous implantation of PPP1CB-ALK and eGFP-expressing fibroblasts.K. Kaplan-Meier survival curves of SCID mice injected subcutaneously with mNSC CTX-PPP1CB-ALK (red, n = 5) or mNSC CTX-eGFP (black, n = 4) and treated with ceritinib at 30mg/kg/d for 15 days. Mice were euthanized at endpoint (tumor volume of 2000mm3). The grey area represents the treatment period.L. Vehicle or ceritinib-treated mouse weights at start and end of treatment.

Supplementary Figure 5: Single-cell RNA Sequencing and IHC examination of ALK in human neural development and immature neural cells. A. 2D representation of human brain cell subtypes (48 samples) during cortex development using a single cell RNA-seq atlas (cortex-dev.cells.ucsc.edu) shows that ALK transcript appears to be enriched in a certain cell type. Color-coded by ALK gene expression (beige to red) and age in weeks (rainbow).B. ALK-stained sagittal section of normal human cerebellum at 15 postconceptional weeks (pcw).C. ALK-stained sagittal section of human fetal neural stem cell pellets at 13 postconceptional weeks (pcw).D. ALK-stained sagittal section of human fetal neural stem cell pellets at 15 postconceptional weeks (pcw).

Purpose:Anaplastic lymphoma kinase (ALK) aberrations have been identified in pediatric-type infant gliomas, but their occurrence across age groups, functional effects, and treatment response has not been broadly established.Experimental Design:We performed a comprehensive analysis of ALK expression and genomic aberrations in both newly generated and retrospective data from 371 glioblastomas (156 adult, 205 infant/pediatric, and 10 congenital) with in vitro and in vivo validation of aberrations.Results:ALK aberrations at the protein or genomic level were detected in 12% of gliomas (45/371) in a wide age range (0–80 years). Recurrent as well as novel ALK fusions (LRRFIP1–ALK, DCTN1–ALK, PRKD3–ALK) were present in 50% (5/10) of congenital/infant, 1.4% (3/205) of pediatric, and 1.9% (3/156) of adult GBMs. ALK fusions were present as the only candidate driver in congenital/infant GBMs and were sometimes focally amplified. In contrast, adult ALK fusions co-occurred with other oncogenic drivers. No activating ALK mutations were identified in any age group. Novel and recurrent ALK rearrangements promoted STAT3 and ERK1/2 pathways and transformation in vitro and in vivo. ALK-fused GBM cellular and mouse models were responsive to ALK inhibitors, including in patient cells derived from a congenital GBM. Relevant to the treatment of infant gliomas, we showed that ALK protein appears minimally expressed in the forebrain at perinatal stages, and no gross effects on perinatal brain development were seen in pregnant mice treated with the ALK inhibitor ceritinib.Conclusions:These findings support use of brain-penetrant ALK inhibitors in clinical trials across infant, pediatric, and adult GBMs.

Supplementary Figure 6: Single cell RNA sequencing comparison of ALK expression and other infant/pediatric glioma kinase drivers in developing human and mouse brain. A. 2D representation of human brain cell subtypes (48 samples) during cortex development using a single cell RNA-seq atlas (http://cells.ucsc.edu/?ds=cortex-dev) shows that ALK positive cells make up less than 1% of all cells in the dataset and aren't localized within a specific cluster.B. Cluster-level summary of the human scRNA dataset shows that ALK expression (TPM) is decreased compared to EGFR, NTRK2, and NTRK3, and ALK positive cells make up a much lower percentage of each cluster compared to those 3 genes.C. 2D representation of developmental mouse forebrain (E12.5 – P6) in a single cell RNA-seq atlas (http://cc-shiny-01.functionalgenomics.ca/braindex/clusters) shows similar results as seen in the human scRNA Nowakowski dataset.D. Cluster-level summary of the mouse scRNA dataset corroborates results seen in the Nowkowski scRNA dataset; that Alk expression (log-normalized) is decreased, and less prevalent than EGFR, NTRK2, and NTRK3.E. 2D representation of RNA sequencing data for NTRK1-3 genes in the Nowakowski developing human cortex dataset. Compared to ALK expression, the NTRK genes have increased expression levels, increased prevalence in the dataset, and show increased localization to specific cluster.F. Similar to (E), the 2D representation of Ntrk genes in the developing mouse scRNA dataset shows increased expression and prevalence compared to Alk.

Supplementary Figure 2: T1 post-contrast axial and sagittal images of ALK-fused and non ALK-altered GBMs and ALK staining in ALK-fused congenital GBMs. A. Example images from patients with ALK aberrations (76, 89, 116, 197)B. In contrast to those without ALK aberrations (right columns). Contrast is gadolinium for those where applicable and show enhancement within both the tumor categories.C. Representative H&E and ALK IHCs of congenital GBM harboring an ALK fusion.

Supplementary Figure 4: Characterization of the lung cancer recurrent DCTN1-ALK fusion identified in cGBM (case ALK.232). A. Integrative Genomic Viewer (IGV) screenshot of the DCTN1-ALK variant.B. Predicted sequence of DCTN1-ALK fusion (DCTN1 exon 1-26, ALK exon 20-29 conserved).C. PDX DCTN1-ALK sub-renal capsule engraftment confimed by IHC.C. Relative viability of the ALK wild-type GBM line (BT164) and BT1857 ceritinib-treated cells versus DMSO control.D. Relative viability of BT164 and BT1857 lorlatinib-treated cells versus DMSO control.E. Relative viability of BT164 and BT1857 cells treated with an EGFR tyrosine kinase inhibitor neratinib.F. PDX DCTN1-ALK sub-renal capsule engraftment confirmed by IHC.

Supplementary Figure 1: Characterization of the novel LRRFIP1-ALK fusion (case ALK.223) A. Visualization of the read coverage at the fusion sites in ALK and LRRFIP1 genes using Integrative Genomic Viewer.B. Pile-up plot generated by SvABA using whole-genome sequencing data shows the breakpoint coordinates.C. Copy number ratio profile by WGS showing no somatic copy number alterations anywhere in the genome. The x-axis represents the chromosomes.D. Copy number ratio profile on chromosome 2 showing no copy number change of ALK and LRRFIP1 genes.E. Two RT-PCR products of 186bp (variant 1) and 189bp (variant 3) were detected by agarose gel electrophoresis with total RNA from tumor specimen ALK.223.F. Primer sequences of ALK-LRRFIP1 (variants 1 and 2) and LRRFIP1-ALK (variant 3) and the predicted DNA product size.G. Western Blot analysis of ALK signaling pathway in subcutaneous tumors treated with vehicle or lorlatinib. Normal brain was used as control.H. Representative 40x images and CellProfiler quantification of Cleaved Caspase 3- and Ki-67- stained subcutaneous tumors.

Additional information tables 1 to 8.

This file contains supplemental tables S1-S7. The data contained are as follows: (Table S1) Gene lists on the FoundationOne assay, (Table S2) Sample counts per disease, (Table S3) Foundation Medicine sample counts compared to TCGA sample counts, (Table S4) Genes with altered frequencies between local and metastatic disease, (Table S5) Merged MutationAssessor and PolyPhen-2 outputs, (Table S6) Enrichment of NOTCH1 alterations across diseases, (Table S7) Curated list of tumor suppressor genes on the FoundationOne assay.

Long tail distribution of the top 50 altered genes in pediatric sarcoma (S1); Long tail distribution of the top 50 altered genes in pediatric extracranial embryonal tumors (S2); Long tail distribution of the top 50 altered genes in pediatric brain tumors (S3); Long tail distribution of the top 50 altered genes in pediatric heme malignancies (S4); Long tail distribution of the top 50 altered genes in pediatric carcinomas (S5); Long tail distribution of the top 50 altered genes in pediatric gonadal tumors (S6); Known fusions identified in the pediatric cohort (S7); Tileplots showing the pattern of co-occurring alterations with novel MLL3 mutations (S8); Tileplot showing the pattern of co-occurring alterations with a novel PRSS1 mutation (S9); Tileplot showing the pattern of co-occurring alterations with novel DKC1 deletions (S10).

Gene lists for Version 1 of the FoundationOne Assay (S1); Gene lists for Version 2 of the FoundationOne Assay (S2); Gene lists for Version 3 of the FoundationOne Assay (S3); Gene lists for Version 4 of the FoundationOne Assay (S4); Gene lists for Version 5 of the FoundationOne Assay (S5); Distribution of samples across different FoundationOne assays (S6); Filtered variants of unknown significance (S7).

Genomic profiling is widely predicted to become a standard of care in clinical oncology, but more effective data sharing to accelerate progress in precision medicine will be required. Here, we describe cancer-associated genomic profiles from 18,004 unique adult cancers. The dataset was composed of 162 tumor subtypes including multiple rare and uncommon tumors. Comparison of alteration frequencies to The Cancer Genome Atlas identified some differences and suggested an enrichment of treatment-refractory samples in breast and lung cancer cohorts. To illustrate novelty within the dataset, we surveyed the genomic landscape of rare diseases and identified an increased frequency of NOTCH1 alterations in adenoid cystic carcinomas compared with previous studies. Analysis of tumor suppressor gene patterns revealed disease specificity for certain genes but broad inactivation of others. We identified multiple potentially druggable, novel and known kinase fusions in diseases beyond those in which they are currently recognized. Analysis of variants of unknown significance identified an enrichment of SMAD4 alterations in colon cancer and other rare alterations predicted to have functional impact. Analysis of established, clinically relevant alterations highlighted the spectrum of molecular changes for which testing is currently recommended, as well as opportunities for expansion of indications for use of approved targeted therapies. Overall, this dataset presents a new resource with which to investigate rare alterations and diseases, validate clinical relevance, and identify novel therapeutic targets. Cancer Res; 77(9); 2464–75. ©2017 AACR.

Purpose: Isocitrate dehydrogenase (IDH) mutations in glioma patients confer longer survival and may guide treatment decision making. We aimed to predict the IDH status of gliomas from MR imaging by applying a residual convolutional neural network to preoperative radiographic data.Experimental Design: Preoperative imaging was acquired for 201 patients from the Hospital of University of Pennsylvania (HUP), 157 patients from Brigham and Women's Hospital (BWH), and 138 patients from The Cancer Imaging Archive (TCIA) and divided into training, validation, and testing sets. We trained a residual convolutional neural network for each MR sequence (FLAIR, T2, T1 precontrast, and T1 postcontrast) and built a predictive model from the outputs. To increase the size of the training set and prevent overfitting, we augmented the training set images by introducing random rotations, translations, flips, shearing, and zooming.Results: With our neural network model, we achieved IDH prediction accuracies of 82.8% (AUC = 0.90), 83.0% (AUC = 0.93), and 85.7% (AUC = 0.94) within training, validation, and testing sets, respectively. When age at diagnosis was incorporated into the model, the training, validation, and testing accuracies increased to 87.3% (AUC = 0.93), 87.6% (AUC = 0.95), and 89.1% (AUC = 0.95), respectively.Conclusions: We developed a deep learning technique to noninvasively predict IDH genotype in grade II–IV glioma using conventional MR imaging using a multi-institutional data set. Clin Cancer Res; 24(5); 1073–81. ©2017 AACR.

Supplementary Data

Supplemental Figure: Mean plasma concentrations of vandetanib.

Supplemental Tables 1-3. Supplementary Table 1: Correlations in the vandetanib/RT/TMZ arm between best radiographic responses with pre-treatment and on-treatment changes in blood biomarkers. Supplementary Table 2: Correlations in the vandetanib/RT/TMZ arm between overall survival with pre-treatment and on-treatment changes in blood biomarkers. Supplementary Table 3: Comparison of PFS or OS by log-rank p-value based on tissue biomarker analysis

This file contains: cellularity distribution, exon coverage distribution, variant filtering flow chart, detailed disease distributions, FMvTCGA long tail plots by local/metastatic disease, local disease versus plural fluid in FM lung adeno, adenoid cystic carcinoma tile plot, summary of known fusions observed in novel diseases, ERBB2 tile plot in cervical cancers, TCGA MET CN to expression correlation, tile plot of AKT1 E17K colorectal tumors, and distribution of alterations in tumor suppressor genes.

Details for variant processing, analysis of TCGA data, and the FM mutation hotspot caller.

Pediatric cancers are generally characterized by low mutational burden and few recurrently mutated genes. Recent studies suggest that genomic alterations may help guide treatment decisions and clinical trial selection. Here, we describe genomic profiles from 1,215 pediatric tumors representing sarcomas, extracranial embryonal tumors, brain tumors, hematologic malignancies, carcinomas, and gonadal tumors. Comparable published datasets identified similar frequencies of clinically relevant alterations, validating this dataset as biologically relevant. We identified novel ALK fusions in a neuroblastoma (BEND5–ALK) and an astrocytoma (PPP1CB–ALK), novel BRAF fusions in an astrocytoma (BCAS1–BRAF) and a ganglioglioma (TMEM106B–BRAF), and a novel PAX3–GLI2 fusion in a rhabdomyosarcoma. Previously characterized ALK, NTRK1, and PAX3 fusions were observed in unexpected malignancies, challenging the "disease-specific" alterations paradigm. Finally, we identified recurrent variants of unknown significance in MLL3 and PRSS1 predicted to have functional impact. Data from these 1,215 tumors are publicly available for discovery and validation. Cancer Res; 77(2); 509–19. ©2017 AACR.

Background: There is interest in exploring neoadjuvant chemotherapy + immune checkpoint inhibitor therapy in stage II-III ER+ breast cancer, but there is no information on correlation between PD-L1 expression and Oncotype DX Recurrence Score (RS) or histologic grade that currently inform patient selection for adjuvant/neoadjuvant chemotherapy. The goal of this study was to assess associations between PD-L1 protein expression, RS, tumor grade, and stromal tumor infiltrating lymphocyte (TIL) score in early stage ER+ cancers. Methods:. Formalin fixed surgical pathology blocks of 213 patients who had RS determination as routine care between 2012 and 2017 were retrieved from Yale Pathology. PD-L1 immunohistochemistry was performed with the SP142 assay by Foundation Medicine, cases with ≥1% tumor infiltrating immune cell positivity in the tumor area were considered PD-L1+. TIL scores were determined prospectively as part of this study by breast pathologists following the international TIL scoring guidelines. We compared PD-L1 expression positivity rates across RS (25) and TIL categories (30%), and tumor grade using Wilcoxon and Chi-square tests. Multivariate analysis was performed using logistic regression. Results: Patient characteristics are shown in the table below. PD-L1 results were available for 201, and TIL scores for 203 patients. Overall, 53% of cases were PD-L1+, but expression levels were low, among the positive cases only 14% had positivity ≥ 5%. PD-L1 expression was significantly higher among cases with RS>25 (78% PD-L1+, among these 19% had PD-L1+ ≥5%), compared to RS 5%) compared to grade 2 (49% PD-L1+) or 1 tumors (48% PD-L1+, all at 1% level). T2 and T3 tumors also had significantly more frequent PD-L1 expression (67% and 83%, respectively) compared to T1 cancers (48%). There was no correlation between PD-L1 expression and age, nodal status or histology. In multivariate analysis including age, grade, tumor size, histology, nodal status, TIL score and RS, only TIL and RS remained as independent predictors of PDL1 positivity. Conclusions: Approximately half of early stage ER+ breast cancers were PD-L1+ using the SP142 assay, but expression levels are low with only 14% showing ≥ 5% immune cell staining. PD-L1 expression is significantly more frequent and higher in larger tumors (T2, T3), grade 3 cancers, and in cancers with RS >25. PD-L1 expression also correlates with TIL score but not with histologic type, nodal status or age. These findings suggest that the most chemotherapy sensitive (grade 3, RS> 25), larger ER+ cancers may benefit from immunotherapy added to chemotherapy, similar to triple negative cancers. n (%)PD-L1 positivePD-L1 negativep-value (Chi-sq)Histology0.9IDC151 (71%)7567ILC53 (25%)2723other9 (4%)54Grade0.02174 (35%)33362110 (53%)5152329 (12%)236Tumor Size0.008T1145 (68%)6374T262 (29%)3919T3/T46 (3%)51Nodal Status0.9N0177 (84%)9077N135 (16%)1716Age (years)range 39-850.8≤5036 (17%)1815>50177 (83%)8979Recurrence Score0.003RS 2539 (18%)288TIL Count0.0000008 Citation Format: Kim RM Blenman, Malini Harigopal, Richard Huang, Emily Reisenbichler, Tao Qing, Eiman Ibrahim, Kamaljeet Singh, Shakti Ramkissoon, Roberts Mustimbo, Jeffrey Ross, Lajos Pusztai. PD-L1 protein expression in relation to Recurrence Score values in early stage ER+HER2- breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P2-08-03.

Understanding the genomic landscape of breast cancer brain metastases (BCBMs) is key to elucidating their cause and developing novel treatments. In this study, comprehensive genomic profiling was performed on 822 BCBMs, 11,988 local breast cancer (BC) biopsies and 15,516 non-central nervous system (N-CNS) metastases (all unpaired samples). Clinically-relevant genomic alterations were significantly enriched in BCBMs compared to local BCs and N-CNS metastases. Homologous recombination deficiency as measured by BRCA1/2alteration prevalence and loss-of-heterozygosity and immune checkpoint inhibitor (ICI) biomarkers [Tumour mutation burden (TMB)-High, Microsatellite instability (MSI)-High, PD-L1/L2)] were significantly more prevalent in BCBM than local BC and N-CNS. High PD-L1 protein expression was observed in ER-negative/HER2-negative BCBMs (48.3% vs 50.0% in local BCs, 21.4% in N-CNS). Collectively, our data highlights that a high proportion of BCBMs are potentially amenable to treatment with targeted therapeutic agents including PARP inhibitors and ICIs.

Background: Adult primary retroperitoneal sarcomas (RPSs) are a group of heterogeneous tumors with different histological subtypes. Comprehensive genomic profiling (CGP) analyses have recently provided significant insights into the biology of sarcomas by identifying genomic alterations (GAs) which could benefit from targeted therapies. Methods: RPS were evaluated by CGP using next-generation sequencing of up to 406 cancer-related genes. Tumor mutational burden (TMB) was determined on 0.83 to 1.14 mut/Mb of sequenced DNA. Finally, PD-L1 expression was determined. Results: Overall, 296 cases of primary RPS were analyzed. Liposarcoma (LPS) subtype had more GA/tumor than leiomyosarcoma (LMS) subtypes, with follicular dendritic cell sarcomas harboring the highest and synovial sarcomas the lowest. TP53 and Rb1 alterations were the highest in LMS, and CDK4/6 and MDM2 in LPS. However, both the TMB and targetable GA rates were low across subtypes. PD-L1 immunostaining was low positive in 21% and high positive in 5% of patients, respectively. Conclusions: CGP analysis revealed that potentially actionable genomic targets were rare in our cohort of RPS. Moreover, RPSs seem less likely to respond to immune checkpoint inhibitors based on putative biomarkers status. Nevertheless, genomic stratification according to histological subtypes led to description of GAs that can inform future clinical trials design.

Introduction:. Breast cancer (BC) brain metastases (BM) are a growing clinical problem as survival for metastatic BC improves. Genomic analyses of BM are key to developing more targeted therapeutic interventions. In this study we explored the gene rearrangements of BM and compared these to a cohort of non-paired local breast cancers (BCs) and non-CNS metastasis (N-CNS). Methods:. We analyzed 822 BMs and compared them to 11,988 local, breast-biopsied BCs and 15,516 N-CNS (samples unpaired) profiled with comprehensive genomic profiling (CGP) (Foundation Medicine, Cambridge, MA, USA) examining all classes of alterations in at least 324 genes. The prevalence of individual gene rearrangements within the BM was compared to those within BC and N-CNS cases. Further analysis was undertaken by classifying BM, BC and N-CNS specimens with respect to their oestrogen receptor (ER) and human epidermal growth factor receptor 2 (HER2) status, available for a subset of samples. Results:. CGP identified 307 rearrangement events in BMs impacting 169 genes. Twenty genes exhibited recurrent rearrangement events with a prevalence greater than 0.5% in either the BM, BC or N-CNS. 10 of these 20 genes were significantly more prevalent within the BM specimens vs. BC and N-CNS (p The prevalence of CDK12 rearrangements within BM, BCs, and other N-CNS in different sub-types of BC.SubtypePrevalenceP-valueBM BCN-CNSBM vs. BCBM vs. . N-CNSAll3.53%. (29/822)0.86%. (103/11,988)0.68%. (105/15,516)3.55x10-91.85x10-11HER2+(irrespective of ER status)14.59%. (27/185)7.80%. (86/1,102)7.87%. (94/1,194)0.00460.0048HER2+, ER-15.29%. (13/85)10.53%. (6/57)1.96%. (1/51)0.460.02HER2+, ER+13.64%. (9/66)7.14%. (6/84)10.20%. (5/49)0.270.77HER2-, ER-0.67%. (2/298)0.17%. (1/585)0.69%. (2/291)0.261.00HER2-, ER+0.00%. (0/212)0.16%. (1/641)0.00%. (0/634)1.001.00 Citation Format: Talvinder Bhogal, Ethan Sokol, Shakti Ramkissoon, Athina Giannoudis, Kimberly McGregor, Allison Clark, Evangelia Razis, Rupert Bartsch, Richard Huang, Carlo Palmieri. Comprehensive analysis of the gene rearrangements within 822 breast cancer brain metastases reveals an enrichment of cyclin dependent kinase 12 rearrangements in HER2-positive brain metastases [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-07-04.

Background: NCSLC is the leading cause of cancer deaths in the United States (USA). The 5-year survival rate is 22.9% (7% in those with metastases). Personalized therapies have shown promise but a better understanding of the genomic landscape of the tumor may help further improve survival. Here, we present the genomic landscape and key features of NCSLC patients using a real-world clinico-genomics dataset. Methods: The ConcertAI Genome360TM NCSLC dataset is a deeply curated real-world dataset of 11349 patients from the USA who have undergone a next generation sequencing test. This dataset covers >2000 genes. Here we present the demographic, clinical and genomic characteristics of this cohort. The top 20 genes with pathogenic mutations were identified and co-mutational analysis was performed to identify the most significant genes that were positively and negatively correlated to these genes using chi-squared test with Bonferroni correction. Other clinically relevant biomarkers such as tumor mutation burden (TMB), microsatellite instability (MSI) and Programmed death-ligand 1 (PD-L1) status were also characterised. Results: The median age at diagnosis was 70 years with an equal gender distribution. Adenocarcinoma (69%) and squamous cell carcinoma (24%) were the two most common histologies. Most patients were diagnosed at stage IV (57%) or III (19%). Brain, bone, and liver were the most prevalent metastatic sites. 22% patients had high expression of PD-L1, 28% had high TMB and only ~1% patients had high MSI. The 20 most frequently mutated genes along with their top 3 most significant (p-value < 3e-5) positively and negatively correlated genes are listed in Table 1. Conclusion: This analysis provides a deeper understanding of the genomic landscape of NSCLC. The co-mutational analysis provides insights into pathways that are perturbed together providing opportunities to develop treatments to overcome such co-mutations. Table 1. Top 20 mutated genes and their associated positively/negatively correlated genes. Gene Prevalence (%) Positively Correlated Genes Negatively Correlated Genes TP53 65 SOX2, NFE2L2, FGFR1 KRAS, MDM2, STK11 KRAS 28 STK11, RBM10, ATM EGFR, KMT2D, SOX2 CDKN2A 19 TP53, CCND1, SOX2 RB1 EGFR 13 MDM2, CDK4, CCNE1 STK11, KEAP1, KMT2D STK11 12 KEAP1, KRAS, MYC SOX2, PIK3CA, KMT2D MTAP 9 CDKN2A RB1 PIK3CA 7 SOX2, TP53, NFE2L2 KRAS, STK11 KEAP1 6 STK11, KRAS, CDKN2A EGFR SMARCA4 6 STK11, KEAP1, APC EGFR NF1 6 TP53, RASA1, PTPN11 KRAS RBM10 5 KRAS, IL7R, ATM TP53 KMT2D 7 TP53, NFE2L2, NOTCH1 KRAS, EGFR, STK11 DNMT3A 6 TET2, ASXL1, CHEK2 - RB1 7 TP53, PTEN, CCNE1 MTAP, KRAS, CDKN2A ARID1A 7 CDKN2A, TP53, FAT1 EGFR ATM 4 KRAS, NKX2-1, NFKBIA TP53 PTEN 7 RB1, TP53, KMT2D KRAS BRAF 11 SETD2, CDK6 KRAS MET 7 CDK6, MDM2, ROS1 KRAS TERT 4 MITF, PTCH1, EP300 - Citation Format: neeraj kumar singh, Kamal S. Saini, Sangavai Chakkrapani, Eric Severson, Nahush Nagaraj, Shakti Ramkissoon, Isagani Chico, Ariel Aguilo, Rohini George, Laura Vidal, Smita Agrawal. Genomic landscape of patients with non-small cell lung cancer (NSCLC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 923.

Purpose: Anaplastic Lymphoma Kinase (ALK) aberrations have been identified in pediatric type infant gliomas, but their occurrence across age groups, functional effects, and treatment response have not been broadly established. Experimental Design: We performed a comprehensive analysis of ALK expression and genomic aberrations in both newly-generated and retrospective data from 371 glioblastomas (156 adult, 205 infant/pediatric and 10 congenital) with in vitro and in vivo validation of aberrations. Results: ALK aberrations at the protein or genomic level were detected in 12% of gliomas (45/371) in a wide age range (0-80 years). Recurrent as well as novel ALK fusions (LRRFIP1-ALK, DCTN1-ALK, PRKD3-ALK) were present in 50% (5/10) of congenital/infant, 1.4% (3/205) of pediatric, and 1.9% (3/156) of adult GBMs. ALK fusions were present as the only candidate driver in congenital/infant GBMs, and were sometimes focally amplified. In contrast, adult ALK fusions co-occurred with other oncogenic drivers. No activating ALK mutations were identified in any age group. Novel and recurrent ALK rearrangements promoted STAT3 and ERK1/2 pathways and transformation in vitro and in vivo. ALK-fused GBM cellular and mouse models were responsive to ALK inhibitors, including in patient cells derived from a congenital GBM. Relevant to treatment of infant gliomas, we showed that ALK protein appears minimally expressed in the forebrain at perinatal stages and no gross effects on perinatal brain development was seen in pregnant mice treated with the ALK inhibitor ceritinib. Conclusions: These findings support expanded evaluation of brain-penetrant ALK inhibitors in clinical trials across infant, pediatric, and adult GBMs. Citation Format: Anne-Florence Blandin, Ross Giglio, Maya Srikanth Graham, Guadalupe Garcia, Seth Malinowski, Jared K. Woods, Shakti Ramkissoon, Lori Ramkissoon, Frank Dubois, Kate Schoolcraft, Jessica W. Tsai, Dayle K. Wang, Robert Jones, Jayne Vogelzang, Kristine Pelton, Sarah Becker, Fiona Watkinson, Claire Sinai, Elizabeth Cohen, Matthew Booker, Michael Tolstorukov, Veerle Haemels, Liliana Goumnerova, Karen Wright, Mark Kieran, Katie Fehnel, David Reardon, Arnault Tauziede-Espariat, Rishi Lulla, Benjamin Carcamo, Stanley Chaleff, Alain Charest, Frederik De Smet, Azra H. Ligon, Adrian Dubuc, Melanie Pagès, Pascale Varlet, Patrick Wen, Brian Alexander, Susan Chi, Sanda Alexandrescu, Ralf Kittler, Robert Bachoo, Rameen Beroukhim, Pratiti Bandopadhayay, Keith L. Ligon. ALK amplification and rearrangements are recurrent targetable events in congenital and adult glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1201.