A Retrospective Genomic Landscape of 661 Young Adult Glioblastomas Diagnosed Using 2016 WHO Guidelines for Central Nervous System Tumors.

The authors present a cohort of 661 young adult glioblastomas diagnosed using 2016 WHO World Health Organization Classification of Tumors of the Central Nervous System, utilizing comprehensive genomic profiling (CGP) to explore their genomic landscape and assess their relationship to currently defined disease entities. This analysis explored variants with evidence of pathogenic function, common copy number variants (CNVs), and several novel fusion events not described in literature. Tumor mutational burden (TMB) mutational signatures, anatomic location, and tumor recurrence are further explored. Using data collected from CGP, unsupervised machine-learning techniques were leveraged to identify 10 genomic classes in previously assigned young adult glioblastomas. The authors relate these molecular classes to current World Health Organization guidelines and reference current literature to give therapeutic and prognostic descriptions where possible.

Genomic Profiling Reveals Differences in Primary Central Nervous System Lymphoma and Large B-Cell Lymphoma, With Subtyping Suggesting Sensitivity to BTK Inhibition.

BACKGROUND: B-cell primary central nervous system (CNS) lymphoma (PCL) is diffuse large B-cell lymphoma (DLBCL) confined to the CNS. Less than 50% of patients with PCL achieve complete remission with current therapies. We describe the findings from comprehensive genomic profiling (CGP) of a cohort of 69 patients with PCL, 36 cases of secondary CNS lymphoma (SCL), and 969 cases of DLBCL to highlight their differences and characterize the PCL cohort. In addition, we highlight the differences in frequency of germinal center B-cell like (GCB) and non-GCB subtypes and molecular subtypes, particularly MCD and EZH subtypes, between PCL and DLBCL. MATERIALS AND METHODS: Sixty-nine cases of B-cell PCL, 36 cases of secondary CNS lymphoma (SCL), and 969 cases of DLBCL were evaluated by CGP of 405 genes via DNAseq and 265 genes via RNAseq for fusions (FoundationOne Heme). Tumor mutational burden (TMB) was calculated from 1.23 Mb of sequenced DNA. RESULTS: Genomic alterations with significant differences between PCL and DLBCL included MYD88, ETV6, PIM1, PRDM1, CXCR4, TP53, and CREBBP, while only MYD88 was significantly different between SCL and DLBCL. PCL cases were significantly enriched for the MCD molecular subtypes, which have an excellent response to BTKi. We report a patient with a durable complete response to BTKi consistent with their genomic profile. EBV status, CD274 amplification, and TMB status suggest that 38% of PCL patients may benefit from ICPI; however further study is warranted. CONCLUSION: CGP of PCLs reveals biomarkers, genomic alterations, and molecular classifications predictive of BTKi efficacy and potential ICPI efficacy. Given the limitations of standard of care for PCL, CGP is critical to identify potential therapeutic approaches for patients in this rare form of lymphoma.

A Case Series of Metastatic Malignant Gastrointestinal Neuroectodermal Tumors and Comprehensive Genomic Profiling Analysis of 20 Cases.

Malignant gastrointestinal neuroectodermal tumor (GNET) is an ultra-rare soft tissue sarcoma, therefore often misdiagnosed and has no available standard treatment. Here, we report 3 cases of metastatic GNET with variable clinical courses. Our small case series as well as extensive literature review, further support that GNET is a spectrum of diseases with variable inherent biology and prognosis. Surgical management in the setting of recurrent/metastatic disease may be appropriate for GNET with indolent nature. Response to systemic treatments including chemotherapy and targeted treatments is variable, likely related to heterogenous biology as well. Furthermore, we retrospectively identified 20 additional GNET cases from Foundation Medicine's genomic database and expanded on their clinicopathological and genomic features. Comprehensive genomic profiling (CGP) with DNA and RNA sequencing of this cohort, in the course of clinical care, demonstrated recurrent EWSR1 chromosomal rearrangements and a sparsity of additional recurrent or driver genomic alterations. All cases had low tumor mutational burden (TMB) and were microsatellite stable.

Comprehensive genomic profiling of histologic subtypes of urethral carcinomas.

BACKGROUND: Carcinoma of the urethra (UrthCa) is an uncommon Genitourinary (GU) malignancy that can progress to advanced metastatic disease. METHODS: One hundred twenty-seven metastatic UrthCa underwent hybrid capture-based comprehensive genomic profiling to evaluate all classes of genomic alterations (GA). Tumor mutational burden was determined on up to 1.1 Mbp of sequenced DNA, and microsatellite instability was determined on 114 loci. PD-L1 expression was determined by IHC (Dako 22C3). RESULTS: Forty-nine (39%) urothelial (UrthUC), 31 (24%) squamous (UrthSCC), 24 (19%) adenocarcinomas NOS (UrthAC), and 12 (9%) clear cell (UrthCC) were evaluated. UrthUC and UrthSCC are more common in men; UrthAC and UrthCC are more common in women. Ages were similar in all 4 groups. GA in PIK3CA were the most frequent potentially targetable GA; mTOR pathway GA in PTEN were also identified. GA in other potentially targetable genes were also identified including ERBB2 (6% in UrthUC, 3% in UrthSCC, and 12% in UrthAC), FGFR1-3 (3% in UrthSCC), BRAF (3% in UrthAC), PTCH1 (8% in UrthCC), and MET (8% in UrthCC). Possibly reflecting their higher GA/tumor status, potential for immunotherapy benefit associated with higher tumor mutational burden and PD-L1 staining levels were seen in UrthUC and UrthSCC compared to UrthAC and UrthCC. Microsatellite instability high status was absent throughout. CONCLUSIONS: Comprehensive genomic profiling reveals GA that may be predictive of both targeted and immunotherapy benefit in patients with advanced UrthCa and that could potentially be used in future adjuvant, neoadjuvant, and metastatic disease trials.

Landscape of Biomarkers in Non-small Cell Lung Cancer Using Comprehensive Genomic Profiling and PD-L1 Immunohistochemistry.

Comprehensive genomic profiling (CGP) and immunohistochemistry (IHC) are important biomarker tools used for patients with non-small cell lung cancer (NSCLC) given the expanding number of standard-of-care therapies that require companion diagnostic testing. We examined 9450 NSCLC real-world patient samples that underwent both CGP and programmed death-ligand 1 (PD-L1) IHC to understand the biomarker landscape in this patient cohort. By assessing National Comprehensive Cancer Network (NCCN)-recommended biomarkers including genomic alterations, tumor mutational burden (≥10 mutations/Mb cut-off), and PD-L1 expression (Tumor Proportion Score (TPS) ≥ 50% cut-off), we show that CGP + PD-L1 IHC yielded potentially actionable results for 70.5% of the 9,450 patients with NSCLC. Among the remaining 29.5% (2,789/9,450) of patients, 86.7% (2,419/2,789) were potentially eligible for another biomarker-associated therapy and/or clinical trial based on their genomic profile. In addition, in the PD-L1TPS≥50% disease subset, BRAF mutations, MET mutations, MET amplifications, and KRAS mutations were significantly enriched; and in the PD-L1TPS<50%, EGFR mutations, ERBB2 mutations, STK11 mutations, and KEAP1 mutations were enriched. These findings highlight the improved clinical utility of combining CGP with IHC to expand the biomarker-guided therapeutic options available for patients with NSCLC, relative to single biomarker testing alone.

Clinicopathologic and Genomic Characterization of PD-L1 Positive Urothelial Carcinomas.

INTRODUCTION: Pembrolizumab was approved with an accompanying companion diagnostic (CDx) assay (PD-L1 DAKO 22C3) for urothelial carcinoma (UC). In this study, we further characterize the clinicopathologic and genomic features of UC that are programmed death-ligand 1 (PD-L1) positive. MATERIALS AND METHODS: The cohort of this study consisted of a total of 528 consecutive UC patients with PD-L1 immunohistochemistry (IHC) and comprehensive genomic profiling (CGP). All PD-L1 IHC testing was performed using the DAKO 22C3 CDx assay for UC. PD-L1 positivity was determined at a combined positive score ≥ 10. RESULTS: A total of 44.5% (235/528) patients with UC were PD-L1positive . A lower PD-L1 positivity rate was detected in primary (42.3%, 148/350) versus metastatic sites (48.9%, 87/178). PD-L1 positivity was dependent on the location of the metastatic sites. CGP revealed PD-L1positive patients had more frequent genomic alterations (GAs) in TP53 (p = .006) and RB1 (p = .003) and less frequent GAs in FGFR3 (p = .001) and MTAP (p = .028). The APOBEC mutational signature and tumor mutational burden (TMB)-high were more common in PD-L1positive patients. By testing patients with UC with CGP, in addition to PD-L1 IHC, an additional 97 patients (18.4%) in the total cohort were eligible for immunotherapy based on TMB status. CONCLUSION: PD-L1positive and PD-L1negative urothelial carcinomas are genomically different. Also, our study provides the framework for future clinical investigation with regard to specimen site selection for PD-L1 testing as well as candidate biomarker genomic alterations that may predict for better response or lack of response to immune checkpoint inhibitors. IMPLICATIONS FOR PRACTICE: In this study, a higher prevalence of TP53 and RB1 alterations and APOBEC mutational signatures in the PD-L1positive urothelial carcinoma disease subset and enrichment of FGFR3 alterations in the PD-L1negative disease subset were found. These data provide the basis for future investigation into the role of these genomic changes as positive and negative predictors of immunotherapy response. Also, differences wer seen in PD-L1 positivity based on the collection site of the sample, which can provide a framework for future clinical trial design and could influence sample selection for PD-L1 testing in patients with urothelial carcinoma when multiple samples are available.

A pan-cancer analysis of PD-L1 immunohistochemistry and gene amplification, tumor mutation burden and microsatellite instability in 48,782 cases.

PD-L1 immunohistochemistry (IHC) currently has the most Food and Drug Administration (FDA) approvals as a companion diagnostic (CDx) for immunotherapies in specific tumor types; however, multiple other immunotherapy biomarkers exist. We performed this study to examine and report the prevalence of PD-L1 expression in a wide variety of tumor types and examine its relationship to microsatellite instability (MSI), tumor mutational burden (TMB), and CD274 (PD-L1) gene amplification. We performed a retrospective analysis of all cases in which both PD-L1 IHC (using the DAKO 22C3 IHC assay with either tumor proportion score (TPS) or combined positive score (CPS); or the VENTANA SP142 assay with infiltrating immune cell score (IC)) and comprehensive genomic profiling (CGP) were tested at Foundation Medicine between January 2016 and November 2019. Of note, PD-L1 positivity is defined per the CDx indication and tumor proportion score (TPS ≥ 1) for indications without a CDx claim; and TMB positivity is defined as ≥10 mutations/Mb. A total of 48,782 cases were tested for PD-L1 IHC and CGP. Immune cell expression of PD-L1 was more frequently identified than tumor cell expression of PD-L1. We saw a high correlation between PD-L1 expression and CD274 gene amplification (p < 0.0001), MSI and TMB (p < 0.0001), and PD-L1 and TMB (p < 0.0001). In addition, the combination of PD-L1 and TMB identified four unique disease subsets PD-L1-/TMB-, PD-L1+/TMB-, PD-L1-/TMB+, and PD-L1+/TMB+ with varying prevalence dependent on tumor type. Lastly, 50.3% (24527/48782) of the overall cohort was positive for at least one of the CDx or exploratory biomarkers described above. This is the largest pan-cancer analysis of relevant biomarkers associated with response to checkpoint inhibitors to date, including more than 48,000 cases. Additional clinical trials with treatment outcome data in individual tumor types are needed to determine whether the double positive PD-L1+/TMB+ disease subset would respond best to immunotherapy.

Reduced MFAP5 expression in stroma of gallbladder adenocarcinoma and its potential diagnostic utility.

The diagnosis of invasive adenocarcinoma of the gallbladder can sometimes be challenging. The presence of true desmoplastic reaction facilitates the diagnosis of invasion. However, desmoplasia-like changes can be observed in benign gallbladder conditions, and recognition of desmoplasia may be challenging based on morphology. In this study, we tested the expression pattern of microfibril-associated protein 5 (MFAP5), a promising immunohistochemical marker for desmoplasia, in benign gallbladders with desmoplasia-like reaction and gallbladders with invasive adenocarcinoma. We also evaluated the diagnostic utility of MFAP5 in challenging cases with an interobserver agreement study. The results showed that all benign cases retained intact/positive MFAP5 staining pattern in periglandular connective tissue, whereas 79.3% (23 out of 29) of cases of adenocarcinomas demonstrated diffuse and complete loss of MFAP5 staining in the tumor stroma. Interobserver agreement was improved by 2.66 times when images of MFAP5 immunohistochemistry were provided. In conclusion, MFAP5 expression is downregulated in the desmoplastic stroma of gallbladder adenocarcinoma and may provide a useful diagnostic marker in difficult cases.

Genomic Profiling of Circulating Tumor DNA From Cerebrospinal Fluid to Guide Clinical Decision Making for Patients With Primary and Metastatic Brain Tumors.

Despite advances in systemic therapies for solid tumors, the development of brain metastases remains a significant contributor to overall cancer mortality and requires improved methods for diagnosing and treating these lesions. Similarly, the prognosis for malignant primary brain tumors remains poor with little improvement in overall survival over the last several decades. In both primary and metastatic central nervous system (CNS) tumors, the challenge from a clinical perspective centers on detecting CNS dissemination early and understanding how CNS lesions differ from the primary tumor, in order to determine potential treatment strategies. Acquiring tissue from CNS tumors has historically been accomplished through invasive neurosurgical procedures, which restricts the number of patients to those who can safely undergo a surgical procedure, and for which such interventions will add meaningful value to the care of the patient. In this review we discuss the potential of analyzing cell free DNA shed from tumor cells that is contained within the cerebrospinal fluid (CSF) as a sensitive and minimally invasive method to detect and characterize primary and metastatic tumors in the CNS.

Molecular profiling of mesonephric and mesonephric-like carcinomas of cervical, endometrial and ovarian origin.

Mesonephric carcinoma is a rare cancer that most often arises within the cervix, and less frequently, in the ovary and endometrium. A retrospective search of our CLIA-certified and CAP-accredited reference molecular laboratory database (Foundation Medicine, Inc.) identified 20 mesonephric or mesonephric-like, cervical (n = 10), endometrial (n = 5), ovarian (n = 4) or peri-bladder (n = 1) carcinomas that had undergone comprehensive genomic profiling via next generation sequencing. Activating KRAS mutations were present in 90%, 18 of 20 cases, including G12V (n = 7), G12D (n = 6), G12A (n = 3) and G12C (n = 2). Other recurrent alterations were identified in ARID1A (25%), PIK3CA (20%), CTNNB1 (15%), TP53 (10%), MLL2 (10%) and CDKN2A (10%). One KRAS wild-type case had a GATA3 mutation as the sole alteration, while the second KRAS wild-type case had an EGFR exon 20 insertion D770_N771insSVD alteration. All tumors were negative for HPV DNA, microsatellite instability, high tumor mutational burden and homologous recombination deficiency. A circulating tumor DNA (ctDNA) liquid biopsy from peripheral blood, which was performed 6 years after original solid tumor resection in one patient with suspected lung metastasis, revealed concordance of KRAS alteration, gains of chromosomes 1q, 2, 10, 12 and 20, plus new TP53 alterations in the liquid biopsy compared to the original sample. KRAS G12 mutation is major driver of mesonephric and mesonephric-like carcinomas, with less frequent contribution by ARID1A and PIK3CA pathways in tumors of non-cervical origin. ctDNA liquid biopsy may be useful in detecting mutations in recurrent or metastatic patients, who may potentially be eligible for trials against emerging targeted therapies.

Biomarkers in Breast Cancer: An Integrated Analysis of Comprehensive Genomic Profiling and PD-L1 Immunohistochemistry Biomarkers in 312 Patients with Breast Cancer.

BACKGROUND: We examined the current biomarker landscape in breast cancer when programmed death-ligand 1 (PD-L1) testing is integrated with comprehensive genomic profiling (CGP). MATERIAL AND METHODS: We analyzed data from samples of 312 consecutive patients with breast carcinoma tested with both CGP and PD-L1 (SP142) immunohistochemistry (IHC) during routine clinical care. These samples were stratified into hormone receptor positive (HR+)/human epidermal growth factor receptor negative (HER2-; n = 159), HER2-positive (n = 32), and triple-negative breast cancer (TNBC) cohorts (n = 121). RESULTS: We found that in the TNBC cohort, 43% (52/121) were immunocyte PD-L1-positive, and in the HR+/HER2- cohort, 30% (48/159) had PIK3CA companion diagnostics mutations, and hence were potentially eligible for atezolizumab plus nab-paclitaxel or alpelisib plus fulvestrant, respectively. Of the remaining 212 patients, 10.4% (22/212) had a BRCA1/2 mutation, which, if confirmed by germline testing, would allow olaparib plus talazoparib therapy. Of the remaining 190 patients, 169 (88.9%) were positive for another therapy-associated marker or a marker that would potentially qualify the patient for a clinical trial. In addition, we examined the relationship between immunocyte PD-L1 positivity and different tumor mutation burden (TMB) cutoffs and found that when a TMB cutoff of ≥9 mutations per Mb was applied (cutoff determined based on prior publication), 11.6% (14/121) patients were TMB ≥9 mutations/Mb and of these, TMB ≥9 mutations per Mb, 71.4% (10/14) were also positive for PD-L1 IHC. CONCLUSION: Our integrated PD-L1 and CGP methodology identified 32% of the tested patients as potentially eligible for at least one of the two new Food and Drug Administration approved therapies, atezolizumab or alpelisib, and an additional 61.2% (191/312) had other biomarker-guided potential therapeutic options. IMPLICATIONS FOR PRACTICE: This integrated programmed death-ligand 1 immunohistochemistry and comprehensive genomic profiling methodology identified 32% of the tested patients as eligible for at least one of the two new Food and Drug Administration-approved therapies, atezolizumab or alpelisib, and an additional 61.2% (191/312) had other biomarker-guided potential therapeutic options. These findings suggest new research opportunities to evaluate the predictive utility of other commonly seen PIK3CA mutations in hormone receptor-positive breast cancers and to standardize tumor mutation burden cutoffs to evaluate its potentially predictive role in triple-negative breast cancer.

Vulvar Squamous Cell Carcinoma: Comprehensive Genomic Profiling of HPV+ Versus HPV- Forms Reveals Distinct Sets of Potentially Actionable Molecular Targets.

PURPOSE: Vulvar squamous cell carcinoma (vSCC) encompasses two predominant variants: one associated with detectable high-risk strains of human papillomavirus (hrHPV) and a second form often occurring in the context of chronic dermatitis in postmenopausal women. Genomic assessment of a large-scale cohort of patients with aggressive vSCC may identify distinct mutational signatures. MATERIALS AND METHODS: Tumor samples from a total of 280 patients with vSCC underwent hybridization capture with analysis of up to 406 cancer-related genes. Human papillomavirus (HPV) sequences were detected by de novo assembly of nonhuman sequencing reads and aligned to the RefSeq database. Immunohistochemistry for programmed death-ligand 1 (PD-L1) was assessed. RESULTS: One hundred two of 280 vSCCs (36%) contained hrHPV sequences, predominantly HPV 16 (88%). The HPV-positive (HPV+) group was significantly younger (median age, 59 v 64 years; P = .001). Compared with HPV-negative (HPV-) vSCCs, HPV+ tumors showed more frequent pathogenic alterations in PIK3CA (31% v 16%; P = .004), PTEN (14% v 2%; P < .0001), EP300 (14% v 1%; P < .0001), STK11 (14% v 1%; P < .0001), AR (5% v 0%; P = .006), and FBXW7 (10% v 3%; P = .03). In contrast, HPV- vSCCs showed more alterations in TP53 (83% v 6%; P < .0001), TERTp (71% v 9%; P < .0001), CDKN2A (55% v 2%; P < .0001), CCND1 amplification (22% v 2%; P < .0001), FAT1 (25% v 4%; P < .0001), NOTCH1 (19% v 6%; P = .002), and EGFR amplification (11% v 0%; P < .0001), as well as a higher rate of 9p24.1 (PDL1/PDL2) amplification (5% v 1%) and PD-L1 immunohistochemistry high-positive tumor staining (33% v 9%; P = .04). CONCLUSION: Comprehensive molecular profiles of vSCC vary considerably with hrHPV status and may inform patient selection into clinical trials. Sixty-one percent of HPV+ vSCCs had a pathogenic alteration in the PI3K/mTOR pathway, whereas HPV- vSCCs showed alterations in TP53, TERTp, CDKN2A, CCND1, and EGFR, and biomarkers associated with responsiveness to immunotherapy.

CYLD-mutant cylindroma-like basaloid carcinoma of the anus: a genetically and morphologically distinct class of HPV-related anal carcinoma.

Rare reports of anal carcinoma (AC) describe histologic resemblance to cutaneous cylindroma, but mutations in the tumor suppressor CYLD, the gene responsible for familial and sporadic cylindromas, have not been systematically investigated in AC. Here, we investigate CYLD-mutant AC, focusing on molecular correlates of distinct histopathology. Comprehensive genomic profiling (hybrid-capture-based DNA sequencing) was performed on 574 ACs, of which 75 unique cases (13%) harbored a CYLD mutation. Clinical data, pathology reports, and histopathology were reviewed for each CYLD-mutant case. The spectrum of CYLD mutations included truncating (n = 50; 67%), homozygous deletion (n = 10; 13%), missense (n = 16; 21%), and splice-site (n = 3; 4%) events. Compared with CYLD-wildtype AC (n = 499), CYLD-mutant ACs were significantly enriched for females (88% vs. 67%, p = 0.0001), slightly younger (median age 59 vs. 61 years, p = 0.047), and included near-universal detection of high-risk HPV sequences (97% vs. 88%, p = 0.014), predominantly HPV16 (96%). The CYLD-mutant cohort also showed significantly lower tumor mutational burden (TMB; median 2.6 vs. 5.2 mut/Mb, p < 0.00001) and less frequent alterations in PIK3CA (13% vs. 31%, p = 0.0015). On histopathologic examination, 73% of CYLD-mutant AC (55/75 cases) showed a striking cylindroma-like histomorphology, composed of aggregates of basaloid cells surrounded by thickened basement membranes and containing characteristic hyaline globules, while only 8% of CYLD-wildtype tumors (n = 34/409) contained cylindroma-like hyaline globules (p < 0.0001). CYLD-mutant carcinomas with cylindroma-like histomorphology (n = 55) showed significantly lower TMB compared with CYLD-mutant cases showing basaloid histology without the distinctive hyaline globules (n = 14) (median 1.7 vs. 4.4 mut/Mb, p = 0.0058). Only five CYLD-mutant cases (7%) showed nonbasaloid conventional squamous cell carcinoma histology (median TMB = 5.2 mut/Mb), and a single CYLD-mutant case showed transitional cell carcinoma-like histology. Within our cohort of ACs, CYLD mutations characterize a surprisingly large subset (13%), with distinct clinical and genomic features and, predominantly, a striking cylindroma-like histopathology, representing a genotype-phenotype correlation which may assist in classification of AC.

Genomic profiling of BCOR-rearranged uterine sarcomas reveals novel gene fusion partners, frequent CDK4 amplification and CDKN2A loss.

OBJECTIVE: Genomic alterations of BCOR via ZC3H7B-BCOR fusion or BCOR internal tandem duplication (ITD) define a subset of endometrial stromal sarcoma (ESS). The goals of this study were to: 1) determine the molecular landscape of BCOR-rearranged ESS, 2) to identify novel BCOR fusion gene partners in ESS and their associated clinicopathological characteristics, and 3) to potentially unravel targetable genomic alterations in BCOR-mutated ESS. METHODS: A retrospective database search of a CLIA-certified molecular laboratory was performed for uterine sarcomas that contained BCOR rearrangements or BCOR ITD. The cases were previously assayed by comprehensive genomic profiling via both DNA- and RNA-based targeted next generation sequencing during the course of clinical care. Clinicopathological and genomic data was centrally re-reviewed. RESULTS: We identify largest cohort of BCOR-rearranged ESS to date (n = 40), which included 31 cases with canonical ZC3H7B-BCOR fusion as well as 8 cases with novel BCOR gene rearrangement partners, such as BCOR-L3MBTL2, EP300-BCOR, BCOR-NUTM2G, BCOR-RALGPS1, BCOR-MAP7D2, RGAG1-BCOR, ING3-BCOR, BCOR-NUGGC, KMT2D-BCOR, CREBBP-BCOR and 1 case with BCOR internal rearrangement. Re-review of cases with novel rearrangements demonstrated sarcomas with spindle, epithelioid or small round cell components and frequent myxoid stromal change. Comprehensive genomic profiling revealed high frequency of CDK4 and MDM2 amplification in 38% and 45% of BCOR-rearranged cases, respectively, and homozygous deletion of CDKN2A, which encodes an inhibitor of CDK4 in 28% of cases. Notably, CDK4 and MDM2 amplification was absent in all cases from 15 different ESS cases harboring BCOR ITD. CONCLUSIONS: Alterations of CDK4 pathway members, for which targeted therapy is clinically available (i.e. palbociclib), via CDK4 amplification or CDKN2A loss, contributes to the pathogenesis of BCOR-rearranged uterine sarcomas, which may have therapeutic implications.

Treatment of Pediatric Glioblastoma with Combination Olaparib and Temozolomide Demonstrates 2-Year Durable Response.

For pediatric patients with high-grade gliomas, standard-of-care treatment includes surgery, chemotherapy, and radiation therapy; however, most patients ultimately succumb to their disease. With advances in genomic characterization of pediatric high-grade gliomas, the use of targeted therapies in combination with current treatment modalities offer the potential to improve survival in this patient population. In this report, we present the case of a 3-year-old girl with glioblastoma who continues to experience an exceptional and durable response (>2 years) to the poly (ADP-ribose) polymerase (PARP) inhibitor olaparib. Our patient presented with persistent and progressive seizure activity that upon workup was the result of a large heterogeneously enhancing, mixed cystic and solid mass in the left frontal-parietal-temporal region. Histopathologic analysis of resected tumor tissue confirmed the diagnosis of glioblastoma, and comprehensive genomic profiling demonstrated absence of any BRAF or H3F3A mutations. Genomic profiling, however, did reveal a probable germline heterozygous BRCA2 Lys3326Ter (K3226*) nonsense variant. After debulking surgery, the patient received standard-of-care treatment with radiation and temozolomide. Nine months later the PARP inhibitor olaparib was administered in combination with temozolomide for 16 cycles. This regimen was well tolerated by the patient and serial imaging showed reduction in tumor size. Since completion of the regimen, the patient remains neurologically intact with no evidence of tumor recurrence. To our knowledge, this represents the first case of a pediatric glioblastoma that maintains a durable response to a therapeutic strategy that included the PARP inhibitor olaparib and more generally highlights the potential clinical utility of incorporating these agents into the treatment of pediatric high-grade gliomas. KEY POINTS: Germline mutations detected in pediatric gliomas may represent a cancer predisposition syndrome. Integrating molecular testing into routine clinical care for pediatric patients with glioma is critical to identify therapeutic targets and patients with a cancer predisposition syndrome. Patients with glioma with defects in DNA repair pathway components (e.g., BRCA1/2) may show increased responsiveness to poly (ADP-ribose) polymerase (PARP) inhibitors. Combining PARP inhibitors with temozolomide (standard-of-care treatment) revealed no adverse events or toxicities over the course of 18 months.

Comprehensive Assessment of Immuno-oncology Biomarkers in Adenocarcinoma, Urothelial Carcinoma, and Squamous-cell Carcinoma of the Bladder.

BACKGROUND: In patients with rare histologies of bladder cancer, including adenocarcinoma of the bladder (ACB) and squamous-cell carcinoma (SCC), there are limited standard therapy options, defining an unmet medical need. OBJECTIVE: In this comparative comprehensive genomic profiling (CGP) study, genomic alterations (GAs), and immuno-oncology (IO) biomarkers have been analyzed. DESIGN, SETTING, AND PARTICIPANTS: Within the Foundation Medicine database, 143 cases with centrally reviewed pure ACB, 2142 with pure urothelial carcinoma (UC), and 83 with pure SCC were subjected to CGP. All patients developed advanced disease following a primary diagnosis of bladder cancer. INTERVENTION: CGP using a hybrid capture-based assay and immunohistochemistry (IHC). OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Tumor mutational burden (TMB) was determined on 1.1 Mbp of sequenced DNA, and microsatellite instability (MSI) was determined on 114 loci. Programmed cell-death ligand-1 (PD-L1) expression was determined by IHC (Ventana SP-142 assay), with >1% tumor cells (TCs) or tumor-infiltrating lymphocytes (TILs) scoring positive. RESULTS AND LIMITATIONS: Pure ACB patients were younger and more often female than pure UC and pure SCC patients. UC and SCC had a significantly higher median TMB than ACB (p < 0.001). Rare CD274 (PD-L1) amplification cases were more frequently seen in SCC than in UC (5% vs 1%), and were not seen in ACB. MSI high status was very uncommon in all tumor types (0-1%). The frequencies of PD-L1 expression in both TCs and TILs was higher in UC and SCC (both 30%) than in ACB (18%). The results are limited by their retrospective nature and lack of clinical data annotation. CONCLUSIONS: Deep sequencing revealed significant differences in IO biomarkers among the three major subtypes of bladder carcinomas. UC and SCC revealed higher frequencies of PD-L1 expression and higher TMB than ACB, and SCC has the highest frequency of CD274 amplification. The presence of pure SCC features should not disqualify patients for inclusion in IO trials. PATIENT SUMMARY: Tumor samples from patients diagnosed with advanced pure adenocarcinoma of the bladder (ACB) or pure squamous-cell carcinoma (SCC) have been analyzed in terms of frequency of putative immunotherapy biomarkers. The results indicated that pure SCC of the bladder was characterized by genomic features that portend similar response possibilities to immunotherapy compared with the classical pure urothelial carcinoma. Conversely, for pure ACB there might be different therapeutic opportunities, such as targeted therapies against peculiar genomic alterations in selected patients.

Comprehensive Genomic Landscapes in Early and Later Onset Colorectal Cancer.

PURPOSE: The incidence rates of colorectal cancers are increasing in young adults. The objective of this study was to investigate genomic differences between tumor samples collected from younger and older patients with colorectal cancer. EXPERIMENTAL DESIGN: DNA was extracted from 18,218 clinical specimens, followed by hybridization capture of 3,769 exons from 403 cancer-related genes and 47 introns of 19 genes commonly rearranged in cancer. Genomic alterations (GA) were determined, and association with patient age and microsatellite stable/microsatellite instability high (MSS/MSI-H) status established. RESULTS: Overall genomic alteration rates in the younger (<40) and older (≥50) cohorts were similar in the majority of the genes analyzed. Gene alteration rates in the microsatellite stable (MSS) younger and older cohorts were largely similar, with several notable differences. In particular, TP53 (FDR < 0.01) and CTNNB1 (FDR = 0.01) alterations were more common in younger patients with colorectal cancer, and APC (FDR < 0.01), KRAS (FDR < 0.01), BRAF (FDR < 0.01), and FAM123B (FDR < 0.01) were more commonly altered in older patients with colorectal cancer. In the MSI-H cohort, the majority of genes showed similar rate of alterations in all age groups, but with significant differences seen in APC (FDR < 0.01), BRAF (FDR < 0.01), and KRAS (FDR < 0.01). CONCLUSIONS: Tumors from younger and older patients with colorectal cancer demonstrated similar overall rates of genomic alteration. However, differences were noted in several genes relevant to biology and response to therapy. Further study will need to be conducted to determine whether the differences in gene alteration rates can be leveraged to provide personalized therapies for young patients with early-onset sporadic colorectal cancer.

Genomic Features of Metastatic Testicular Sex Cord Stromal Tumors.

BACKGROUND: Metastatic testicular sex cord stromal tumors of the testis (MSCSTs) comprise an extremely uncommon form of genitourinary malignancy. OBJECTIVE: To perform comprehensive genomic profiling (CGP) to enable the search for potential therapy targets. DESIGN, SETTING, AND PARTICIPANTS: Ten patients with testicular Leydig cell tumors (LCTs), six with Sertoli cell tumors (SCTs), and three with undifferentiated sex cord stromal tumors (USCSTs) and a comparison group of 366 patients with ovarian sex cord stromal tumors (SCSTs) underwent hybrid-capture-based CGP to evaluate all classes of genomic alterations (GAs). The tumor mutational burden (TMB) was determined on 1.1 Mbp of sequenced DNA, and microsatellite instability (MSI) was determined on 114 loci. INTERVENTION: CGP on tumor samples. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Descriptive analyses and differences between histological subgroups were reported. RESULTS AND LIMITATIONS: In these patients, all of whom had metastatic disease at the time of sequencing, the primary testis tumor was sequenced in six (32%) patients and a metastatic site in 13 (68%) patients. The overall frequencies of GAs were similar in LCTs, SCTs, and USCSTs, ranging from 3.0 to 3.5 GAs/tumor. The most frequent untargetable GAs included CTNNB1 and CDKN2A/B, both ranging from 20% to 33% of cases. Targetable GAs were uncommon in all MSCST subgroups, but several tumors showed potential for cell-cycle inhibitors (CDK4 in LCTs), mTOR inhibitors (RICTOR, NF2, and PTEN in all three tumor types), hedgehog inhibitors (PTCH1 in LCTs), and poly(ADP-ribose) polymerase inhibitors (BAP1 in SCTs). No MSI-high status was identified. The TMB was also low in all MSCST groups, and tumors featuring a TMB of ≥10 mutations/Mb were not identified. GA findings from ovarian SCSTs largely recapitulated those from MSCSTs. A lack of clinical outcome correlation is a limitation of the present analyses. CONCLUSIONS: Rare cases of testicular MSCSTs have GAs linked to potential targeted therapy benefits on CGP. In contrast, the lack of MSI-high status and an overall low TMB indicate a likely lack of benefit for immunotherapies. PATIENT SUMMARY: Genomic profiling can guide clinical research and disclose therapeutic opportunities for patients with rare testicular cancers for which standard therapies are lacking.

Genomic Landscape of Adult and Pediatric BCR-ABL1-Like B-Lymphoblastic Leukemia Using Parallel DNA and RNA Sequencing.

BCR-ABL1-like B-Acute Lymphoblastic Leukemia (B-ALL) is a subset of B-ALL with a poor prognosis that is found in all age groups. Definitive identification of these patients is difficult in routine clinical practice as gene expression profiling, the gold standard test, is not widely available. Comprehensive genomic profiling performed on 450 patients with extensive fusion profiling revealed a wide range of genomic alterations which were consistent with a classification of BCR-ABL1-like B-ALL in 29% of cases. This manuscript highlights a clinically available alternative method for identifying a large subset of patients with BCR-ABL1-like B-ALL.