Implementation of a High-Accuracy Targeted Gene Expression Panel for Clinical Care.

This study describes the validation of a clinical RNA expression panel with evaluation of concordance between gene copy gain by a next-generation sequencing (NGS) assay and high gene expression by an RNA expression panel. The RNA Salah Targeted Expression Panel (RNA STEP) was designed with input from oncologists to include 204 genes with utility for clinical trial prescreening and therapy selection. RNA STEP was validated with the nanoString platform using remnant formalin-fixed, paraffin-embedded-derived RNA from 102 patients previously tested with a validated clinical NGS panel. The repeatability, reproducibility, and concordance of RNA STEP results with NGS results were evaluated. RNA STEP demonstrated high repeatability and reproducibility, with excellent correlation (r > 0.97, P < 0.0001) for all comparisons. Comparison of RNA STEP high gene expression (log2 ratio ≥ 2) versus NGS DNA-based gene copy number gain (copies ≥ 5) for 38 mutually covered genes revealed an accuracy of 93.0% with a positive percentage agreement of 69.4% and negative percentage agreement of 93.8%. Moderate correlation was observed between platforms (r = 0.53, P < 0.0001). Concordance between high gene expression and gene copy number gain varied by specific gene, and some genes had higher accuracy between assays. Clinical implementation of RNA STEP provides gene expression data complementary to NGS and offers a tool for prescreening patients for clinical trials.

Molecular characterization and survival analysis of a cohort of glioblastoma, IDH-wildtype.

Glioblastoma, IDH-wild type, the most common malignant primary central nervous system tumor, represents a formidable challenge in clinical management due to its poor prognosis and limited therapeutic responses. With an evolving understanding of its underlying biology, there is an urgent need to identify prognostic molecular groups that can be subject to targeted therapy. This study established a cohort of 124 sequential glioblastomas from a tertiary hospital and aimed to find correlations between molecular features and survival outcomes. Comprehensive molecular characterization of the cohort revealed prevalent alterations as previously described, such as TERT promoter mutations and involvement of the PI3K-Akt-mTOR, CK4/6-CDKN2A/B-RB1, and p14ARF-MDM2-MDM4-p53 pathways. MGMT promoter methylation is a significant predictor of improved overall survival, aligned with previous data. Conversely, age showed a marginal association with higher mortality. Multivariate analysis to account for the effect of MGMT promoter methylation and age showed that, in contrast to other published series, this cohort demonstrated improved survival for tumors harboring PTEN mutations, and that there was no observed difference for most other molecular alterations, including EGFR amplification, RB1 loss, or the coexistence of EGFR amplification and deletion/exon skipping (EGFRvIII). Despite limitations in sample size, this study contributes data to the molecular landscape of glioblastomas, prompting further investigations to examine these findings more closely in larger cohorts.

Detection of Constitutional Structural Variants by Optical Genome Mapping: A Multisite Study of Postnatal Samples.

Optical genome mapping is a high-resolution technology that can detect all types of structural variations in the genome. This second phase of a multisite study compares the performance of optical genome mapping and current standard-of-care methods for diagnostic testing of individuals with constitutional disorders, including neurodevelopmental impairments and congenital anomalies. Among the 627 analyses in phase 2, 405 were of retrospective samples supplied by five diagnostic centers in the United States and 94 were prospective samples collected over 18 months by two diagnostic centers (June 2021 to October 2022). Additional samples represented a family cohort to determine inheritance (n = 119) and controls (n = 9). Full concordance of results between optical genome mapping and one or more standard-of-care diagnostic tests was 98.6% (618/627), with partial concordance in an additional 1.1% (7/627).

EGFR and ERBB2 exon 20 insertion/duplication in advanced non-small cell lung cancer: genomic profiling and clinicopathologic features.

Background: Exon 20 (ex20) in-frame insertions or duplications (ins/dup) in epidermal growth factor receptor (EGFR) and its analog erb-b2 receptor tyrosine kinase 2 (ERBB2) are each detected in 1.5% of non-small cell lung cancer (NSCLC). Unlike EGFR p.L858R or ex19 deletions, ex20 ins/dup is associated with de novo resistance to classic EGFR inhibitors, lack of response to immune checkpoint inhibitors, and poor prognosis. US Food and Drug Administration has approved mobocertinib and amivantamab for targeting tumors with this aberration, but the number of comprehensive studies on ex20 ins/dup NSCLC is limited. We identified 18 cases of NSCLCs with EGFR/ERBB2 ex20 ins/dup and correlated the findings with clinical and morphologic information including programed death-ligand 1 (PD-L1) expression. Methods: A total of 536 NSCLC cases tested at our institution between 2014 and 2023 were reviewed. A custom-designed 214-gene next-generation sequencing panel was used for detecting DNA variants, and the FusionPlex CTL panel (ArcherDx) was used for the detection of fusion transcripts from formalin-fixed, paraffin-embedded tissue. Immunohistochemistry (IHC)for PD-L1 was performed using 22C3 or E1L3N clones. Results: Nine EGFR and nine ERBB2 ex20 ins/dup variants were identified from an equal number of men and women, 14 were non- or light smokers, and 15 had stage IV disease. All 18 cases were adenocarcinomas. Seven of the 11 cases with available primary tumors had acinar predominant pattern, two had lepidic predominant pattern, and the remainder had papillary (one case) and mucinous (one case) patterns. Ex20 ins/dup variants were heterogenous in-frame one to four amino acids spanning A767-V774 in EGFR and Y772-P780 in ERBB2 and were clustered in the loop following the C-helix and α C-helix. Twelve cases (67%) had co-existing TP53 variants. Copy number variation in CDK4 amplification was identified in one case. No fusion or microsatellite instability was identified in any case. PD-L1 was positive in two cases, low positive in four cases, and negative in 11 cases. Conclusions: NSCLCs harboring EGFR/ERBB2 ex20 ins/dup are rare and tend to be acinar predominant, negative for PD-L1, more frequent in non- or light smokers, and mutually exclusive with other driver mutations in NSCLC. The correlation of different EGFR/ERBB2 ex20 ins/dup variants and co-existing mutations with response to targeted therapy and the possibility of developing resistant mutations after mobocertinib treatment warrants further investigation.

RNA sequencing steps toward the first line.

DNA is the sequence that codes for proteins. Messenger RNA is transcribed from the DNA sequence of genes and translated into protein. It can be difficult to predict how a change in the DNA sequence will affect messenger RNA and protein quantity and quality. DNA translocation changes can cause the joining of sequences from two different genes or different parts of the same gene. DNA sequencing is often used clinically to predict how DNA changes might affect proteins. Alternatively, RNA sequencing can be used as a more direct measure of the effect of DNA changes on the protein products. This sequencing is important for identifying changes in cancer that may indicate response to targeted therapy, prognosis, or diagnosis.

Multisite Assessment of Optical Genome Mapping for Analysis of Structural Variants in Constitutional Postnatal Cases.

This study compares optical genome mapping (OGM) performed at multiple sites with current standard-of-care (SOC) methods used in clinical cytogenetics. This study included 50 negative controls and 359 samples from individuals (patients) with suspected genetic conditions referred for cytogenetic testing. OGM was performed using the Saphyr system and Bionano Access software version 1.7. Structural variants, including copy number variants, aneuploidy, and regions of homozygosity, were detected and classified according to American College of Medical Genetics and Genomics guidelines. Repeated expansions in FMR1 and contractions in facioscapulohumeral dystrophy 1 were also analyzed. OGM results were compared with SOC for technical concordance, clinical classification concordance, intrasite and intersite reproducibility, and ability to provide additional, clinically relevant information. Across five testing sites, 98.8% (404/409) of samples yielded successful OGM data for analysis and interpretation. Overall, technical concordance for OGM to detect previously reported SOC results was 99.5% (399/401). The blinded analysis and variant classification agreement between SOC and OGM was 97.6% (364/373). Replicate analysis of 130 structural variations was 100% concordant. On the basis of this demonstration of the analytic validity and clinical utility of OGM by this multisite assessment, the authors recommend this technology as an alternative to existing SOC tests for rapid detection and diagnosis in postnatal constitutional disorders.

The Genetics of Early-Stage Melanoma in a Veteran Population.

To improve understanding of the genetic signature of early-stage melanomas in Veterans, hotspot mutation profiling using next-generation sequencing (NGS) was performed on melanoma tissue samples from patients at the Iowa City Veterans Affairs Medical Center (VAMC). Genetic analysis identified BRAF (36.3%), TP53 (25.9%), NRAS (19.3%), CDKN2A (11.1%), KIT (8.1%), and BAP1 (7.4%) mutations with the highest prevalence. Although common variants in BRAF were detected at lower rates than what is reported for the general population, 55.6% of cases showed activating mutations in the RAS/RAF pathways. Variants in TP53 and KIT were detected at higher rates than in the general population. Veterans with prior history of melanoma were at significantly higher odds of having TP53 mutation (OR = 2.67, p = 0.04). This suggests that TP53 may be a marker for recurrent melanoma and possibly alternative exposures in the military population. This study provides new information regarding the genetics of melanoma in a Veteran population and early-stage melanomas, highlighting risk factors unique to this population and contributing to the conversation about preventing melanoma deaths in US Military personnel.

Pharmacological ascorbate improves the response to platinum-based chemotherapy in advanced stage non-small cell lung cancer.

PURPOSE: Platinum-based chemotherapy with or without immunotherapy is the mainstay of treatment for advanced stage non-small cell lung cancer (NSCLC) lacking a molecular driver alteration. Pre-clinical studies have reported that pharmacological ascorbate (P-AscH-) enhances NSCLC response to platinum-based therapy. We conducted a phase II clinical trial combining P-AscH- with carboplatin-paclitaxel chemotherapy. EXPERIMENTAL DESIGN: Chemotherapy naïve advanced stage NSCLC patients received 75 g ascorbate twice per week intravenously with carboplatin and paclitaxel every three weeks for four cycles. The primary endpoint was to improve tumor response per Response Evaluation Criteria in Solid Tumors (RECIST) v1.1 compared to the historical control of 20%. The trial was conducted as an optimal Simon's two-stage design. Blood samples were collected for exploratory analyses. RESULTS: The study enrolled 38 patients and met its primary endpoint with an objective response rate of 34.2% (p = 0.03). All were confirmed partial responses (cPR). The disease control rate was 84.2% (stable disease + cPR). Median progression-free and overall survival were 5.7 months and 12.8 months, respectively. Treatment-related adverse events (TRAE) included one grade 5 (neutropenic fever) and five grade 4 events (cytopenias). Cytokine and chemokine data suggest that the combination elicits an immune response. Immunophenotyping of peripheral blood mononuclear cells demonstrated an increase in effector CD8 T-cells in patients with a progression-free survival (PFS) ≥ 6 months. CONCLUSIONS: The addition of P-AscH- to platinum-based chemotherapy improved tumor response in advanced stage NSCLC. P-AscH- appears to alter the host immune response and needs further investigation as a potential adjuvant to immunotherapy.

Genomic Profiling of Metastatic Uveal Melanoma Shows Frequent Coexisting BAP1 or SF3B1 and GNAQ/GNA11 Mutations and Correlation With Prognosis.

OBJECTIVES: To identify therapeutic targets and correlate with clinical outcomes from mutation profiling of metastatic uveal melanoma (UM) using next-generation sequencing (NGS). METHODS: Melanoma cases that were tested using DNA-based NGS panels of 25 and/or 214 genes were evaluated retrospectively (263 cases) and identified 27 UM cases. BAP1 expression was examined by immunohistochemistry. RESULTS: Mutations in GNA11 (14) and GNAQ (12) were found in 96% (n = 27) of cases of UM, and most had coexisting BAP1 (17) or SF3B1 (4) mutations. Coexisting GNAQ/11-SF3B1 mutations correlated with a longer average time to first metastasis compared with GNAQ/11-BAP1 mutations (99.7 vs 38.5 months, P = .047). Three patients with BAP1 mutations received trametinib; two are still alive (15 months; 23 months), and one died (32 months). In non-UMs, only 4.2% (n = 236) had BAP1 and 3.8% had SF3B1 mutations; none had coexisting GNAQ/11 mutations. CONCLUSIONS: Coexisting BAP1/SF3B1 and GNAQ/11 mutations were unique to UM. SF3B1 mutations were reported to be UM-specific in melanoma and associated with rare/no metastasis. The finding of mutated SF3B1 in 14.8% (n = 27) of UMs suggests its role should be further evaluated. The correlation of BAP1/SF3B1 mutation with survival also warrants investigation.

Monocytes Exposed to Immune Complexes Reduce pDC Type 1 Interferon Response to Vidutolimod.

Vidutolimod, also known as CMP-001, is a virus-like particle composed of the Qß bacteriophage coat protein encasing a TLR9 agonist. Vidutolimod injected intratumorally is showing promise in early phase clinical trials based on its ability to alter the tumor microenvironment and induce an anti-tumor immune response. We previously demonstrated that the in vivo efficacy of vidutolimod is dependent on the presence of anti-Qß antibodies that enhance opsonization and uptake of vidutolimod by TLR9-expressing plasmacytoid dendritic cells (pDCs). Here, we evaluated the effect of immune complexes, including anti-Qß-coated vidutolimod, on induction of Type 1 Interferon production by peripheral blood mononuclear cells in response to vidutolimod and soluble TLR9 agonists. Immune complexes, including but not limited to anti-Qß-coated vidutolimod, indirectly suppressed TLR9-mediated Type 1 Interferon production by pDCs in a monocyte-dependent manner. These findings indicate that anti-Qß-coated vidutolimod has effects in addition to those mediated by TLR9 that could have important clinical implications for understanding the mechanism of action of this exciting new approach to in situ immunization and cancer immunotherapy.

PTCH1-GLI1 Fusion-Positive Ovarian Tumor: Report of a Unique Case With Response to Tyrosine Kinase Inhibitor Pazopanib.

Recurrent GLI1 gene fusions have been recently described in a subset of soft tissue tumors showing a distinct monotonous epithelioid morphology with a rich capillary network and frequent S100 protein expression. Three different fusion partners-ACTB, MALAT1, and PTCH1-have been reported with the PTCH1-GLI1 fusion from 2 patients only, both with head and neck tumors. Herein, we report for the first time a PTCH1-GLI1 fusion in a primary ovarian tumor from a female patient aged 54 years who presented with a 21-cm right ovarian mass and mesenteric metastasis. The tumor was diagnosed as "favor malignant melanoma" based on histologic examination and extensive immunohistochemistry studies. The patient received 4 cycles of pembrolizumab and 2 cycles of trabectedin but developed multiple metastases. A next-generation sequencing-based assay detected a PTCH1-GLI1 fusion, which led to a revised pathologic diagnosis and a change of the patient's management. The patient was switched to the tyrosine kinase inhibitor (TKI) pazopanib to target the sonic hedgehog pathway. Her disease was stable 49 months post TKI therapy. Our case report is the first to show that a tumor with GLI1 oncogenic activation was sensitive to a TKI. The morphologic and immunohistochemistry similarities of our patient's tumor to other recently described tumors harboring GLI1 fusions suggest that these tumors may all belong to the same entity of GLI1 fusion-positive neoplasms and may be treated similarly.

Validation of Optical Genome Mapping for the Molecular Diagnosis of Facioscapulohumeral Muscular Dystrophy.

The molecular diagnosis of facioscapulohumeral muscular dystrophy (FSHD) relies on detecting contractions of the unique D4Z4 repeat array at the chromosome 4q35 locus in the presence of a permissive 4q35A haplotype. Long, intact DNA molecules are required for accurate sizing of D4Z4 repeats. We validated the use of optical genome mapping to determine size and haplotype of D4Z4 alleles for FSHD analysis. The cohort included 36 unique DNA specimens from fresh blood samples or archived agarose plugs. High-molecular- weight DNA underwent sequence-specific labeling followed by separation and image analysis with data collection on the Saphyr system. D4Z4 allele sizes were calculated and haplotypes determined from the labeling patterns. Each specimen had previous diagnostic testing using restriction enzyme digests with EcoRI, EcoRI/BlnI, XapI, or HindIII, followed by pulsed field gel electrophoresis and Southern blot analysis with appropriate probes. Optical genome mapping detected 4q35 and 10q26 alleles ranging from 1 to 79 D4Z4 repeats and showed strong correlation with Southern blot allele sizing (R2 = 0.95) and haplotyping (133 of 134; 99.4% haplotype match). Analysis of inter-assay and intra-assay runs showed high reproducibility (0.03 to 0.94 %CV). Subsequent optical genome mapping for routine clinical testing from 315 clinical FSHD cases compared favorably with historical result trends. Optical genome mapping is an accurate and highly reproducible method for chromosomal abnormalities associated with FSHD.

Melanoma Brain Metastases in the Era of Targeted Therapy and Checkpoint Inhibitor Therapy.

Brain metastases commonly develop in melanoma and are associated with poor overall survival of about five to nine months. Fortunately, new therapies, including immune checkpoint inhibitors and BRAF/MEK inhibitors, have been developed. The aim of this study was to identify outcomes of different treatment strategies in patients with melanoma brain metastases in the era of checkpoint inhibitors. Patients with brain metastases secondary to melanoma were identified at a single institution. Univariate and multivariable analyses were performed to identify baseline and treatment factors, which correlated with progression-free and overall survival. A total of 209 patients with melanoma brain metastases were identified. The median overall survival of the cohort was 5.3 months. On multivariable analysis, the presence of non-cranial metastatic disease, poor performance status (ECOG 2-4), whole-brain radiation therapy, and older age at diagnosis of brain metastasis were associated with poorer overall survival. Craniotomy (HR 0.66, 95% CI 0.45-0.97) and treatment with a CTLA-4 checkpoint inhibitor (HR 0.55, 95% CI 0.32-0.94) were the only interventions associated with improved overall survival. Further studies with novel agents are needed to extend lifespan in patients with brain metastases in melanoma.

CLIA Laboratory Testing for Facioscapulohumeral Dystrophy: A Retrospective Analysis.

OBJECTIVE: To summarize facioscapulohumeral muscular dystrophy (FSHD) diagnostic testing results from the University of Iowa Molecular Pathology Laboratory. METHODS: All FSHD tests performed in the diagnostic laboratory from January 2015 to July 2019 were retrospectively reviewed. Testing was by restriction enzyme digestion and Southern blot analysis with sequencing of SMCHD1, if indicated. Cases were classified as FSHD1 (4q35 EcoRI size ≤40 kb; 1-10 D4Z4 repeats), FSHD2 (permissive 4q35A allele, D4Z4 hypomethylation, and pathogenic SMCHD1 variant), or non-FSHD1,2. We also noted cases with borderline EcoRI fragment size (41-43 kb; 11 D4Z4 repeats), cases that meet criteria for both FSHD1 and FSHD2, somatic mosaicism, and cases with hybrid alleles that add complexity to test interpretation. RESULTS: Of the 1,594 patients with FSHD tests included in the analysis, 703 (44.1%) were diagnosed with FSHD. Among these positive tests, 664 (94.5%) met criteria for FSHD1 and 39 (5.5%) met criteria for FSHD2. Of all 1,594 cases, 20 (1.3%) had a 4q35 allele of borderline size, 23 (1.5%) were somatic mosaics, and 328 (20.9%) had undergone translocation events. Considering only cases with at least 1 4q35A allele, D4Z4 repeat number differed significantly among groups: FSHD1 cases median 6.0 (interquartile range [IQR] 4-7) repeats, FSHD2 cases 15.0 (IQR 12-22) repeats, and non-FSHD1,2 cases 28.0 (IQR 19-40) repeats. CONCLUSION: FSHD1 accounts for 94.5% of genetically confirmed cases of FSHD. The data show a continuum of D4Z4 repeat numbers with FSHD1 samples having the fewest, FSHD2 an intermediate number, and non-FSHD1,2 the most.

Multivariable Analysis of 169 Cases of Advanced Cutaneous Melanoma to Evaluate Antibiotic Exposure as Predictor of Survival to Anti-PD-1 Based Immunotherapies.

Recently antibiotic exposure has been associated with worse outcomes in patients undergoing treatment with antibodies directed against programmed cell death protein-1 (PD-1). We reviewed data of 1264 patients enrolled at Melanoma Skin and Ocular Tissue Repositories at University of Iowa Hospitals and Clinic. Reviewed data included patient demographics, prior medical history, baseline hematologic and disease parameters and outcomes including progression-free survival (PFS) and overall survival (OS). Cox regression models were used to determine predictive markers. Overall, 169 patients with advanced cutaneous melanoma received anti-PD-1 based therapies. Median follow up was 18.46 (range 0.89 to 62.52) months. On multivariable analysis brain metastasis, higher absolute neutrophil count (ANC) and lower absolute lymphocyte count were associated with poorer PFS while brain and liver metastasis and lower albumin were associated with poorer OS. Prior antibiotics, radiation as well as age, gender, basal metabolic index (BMI), smoking status, BRAF mutation, line of therapy (first or latter), prior treatments (ipilimumab or BRAF inhibitors), hemoglobin, neutrophil-to-lymphocyte ratio, white blood cell, platelet and eosinophil counts were not associated with PFS or OS in multivariable analysis. Contrary to some prior studies BMI, radiation, and antibiotics were not associated with PFS or OS.

Exploration of PCORnet Data Resources for Assessing Use of Molecular-Guided Cancer Treatment.

PURPOSE: Examine the ability of PCORnet data resources to investigate molecular-guided cancer treatment. PATIENTS AND METHODS: Patients (N = 86,154) had single primary solid tumors (diagnosed 2013-2017) from hospital oncology registries linked to the PCORnet Common Data Model (CDM) at 11 medical institutions. Molecular and anatomic test procedures and oral and infused therapies were identified with Current Procedural Terminology (CPT) and Healthcare Common Procedure Coding System (HCPCS) codes, RxNorm Concept Unique Identifier, and National Drug Codes from CDM tables. Chart review (2 institutions, n = 213) for advanced colorectal cancer and Medicare claims linkages (7 institutions, n = 1,731) for breast cancer explored options for increasing electronic data capture. RESULTS: Molecular testing prevalence detected via analyte-specific molecular CPT/HCPCS codes was 5.5% (n = 4,784); for the nonspecific anatomic pathology codes, for which only some testing is performed to guide therapy selection, it was an additional 44.8% (n = 38,610). Molecular-guided therapy prevalence was 5% (n = 4,289). Testing and treatment were most common with stage IV disease and varied across cancer types and study institutions (testing, 0%-10.4%; treatment, 0.8%-8.4%). Therapy-concordant test results were found in charts for all 36 treated patients with colorectal cancer at the 2 institutions, 3 (8.3%) of whom received treatment outside the institution. Breast cancer Medicare claims linkage increased rates of identified testing from 62.7%-98.9% and treatment from 3.9%-8.2%. CONCLUSION: Although a minority of patients received molecular-guided therapies, the majority had testing that could guide cancer treatment. Claims data extended electronic data capture for therapies and test orders but often was uninformative for types of test ordered. Test results continue to require text data curation from narrative pathology reports.

Coactivation of NF-κB and Notch signaling is sufficient to induce B-cell transformation and enables B-myeloid conversion.

NF-κB and Notch signaling can be simultaneously activated in a variety of B-cell lymphomas. Patients with B-cell lymphoma occasionally develop clonally related myeloid tumors with poor prognosis. Whether concurrent activation of both pathways is sufficient to induce B-cell transformation and whether the signaling initiates B-myeloid conversion in a pathological context are largely unknown. Here, we provide genetic evidence that concurrent activation of NF-κB and Notch signaling in committed B cells is sufficient to induce B-cell lymphomatous transformation and primes common progenitor cells to convert to myeloid lineage through dedifferentiation, not transdifferentiation. Intriguingly, the converted myeloid cells can further transform, albeit at low frequency, into myeloid leukemia. Mechanistically, coactivation of NF-κB and Notch signaling endows committed B cells with the ability to self renew. Downregulation of BACH2, a lymphoma and myeloid gene suppressor, but not upregulation of CEBPα and/or downregulation of B-cell transcription factors, is an early event in both B-cell transformation and myeloid conversion. Interestingly, a DNA hypomethylating drug not only effectively eliminated the converted myeloid leukemia cells, but also restored the expression of green fluorescent protein, which had been lost in converted myeloid leukemia cells. Collectively, our results suggest that targeting NF-κB and Notch signaling will not only improve lymphoma treatment, but also prevent the lymphoma-to-myeloid tumor conversion. Importantly, DNA hypomethylating drugs might efficiently treat these converted myeloid neoplasms.

Durable Clinical Benefit in Patients with Advanced Cutaneous Melanoma after Discontinuation of Anti-PD-1 Therapies Due to Immune-Related Adverse Events.

INTRODUCTION: Anti-PD-1 therapies, pembrolizumab and nivolumab, are currently the standard of care for treatment of patients with metastatic melanoma. Treatment is usually continued until toxicity or disease progression. Though these therapies are well tolerated, some patients discontinue them due to immune-related adverse events (irAE). Discontinuation of therapy brings challenges to their management due to limited treatment options and lack of long-term prognostic information for these patients. Herein, we reviewed patients at our institution to analyze their clinical outcomes. MATERIALS AND METHODS: Charts of 1264 consecutive patients enrolled between 8/1/2012 and 7/31/2017 at Melanoma Skin & Ocular Tissue Repositories at Holden Comprehensive Cancer Center at the University of Iowa Hospitals and Clinic were reviewed. Eligible patients were those who received single-agent anti-PD-1 therapy and subsequently discontinued it due to irAE. Reviewed data included patient demographics, prior medical history, baseline disease parameters, and outcomes. Kaplan-Meier survival analysis was done to determine progression-free survival (PFS) and overall survival (OS). RESULTS: Overall 169 patients with advanced, unresectable, or metastatic cutaneous melanoma received anti-PD-1 therapy of which 16 (9.5%) white, non-Hispanic patients with median age of 64.5 (range 35 to 81 years) discontinued treatment due to irAE. Fifteen patients received pembrolizumab and one received nivolumab. The median duration of treatment was 4.7 (range 0.7 to 11.5) months. Median follow-up was 30.3 (range 4.6 to 49.4) months. Median PFS was 24.6 months and median OS was not reached. Durable clinical benefit (time to progression or next treatment of more than 6 months from last treatment) was observed in 13 (81.2%) patients. At the time of analysis, 8 patients had progressed and 4 patients died (all-cause). DISCUSSION: Our results suggest that advanced melanoma patients discontinuing anti-PD-1 therapy due to irAE usually experience durable clinical benefit. However, caution is needed with these agents in patients with underlying autoimmune diseases.

BCR-NTRK2 fusion in a low-grade glioma with distinctive morphology and unexpected aggressive behavior.

A 52-yr-old man was found to have a 6.6-cm left frontotemporal mass. Biopsy revealed a low-grade astrocytic neoplasm with significant infiltration and an unusual morphologic appearance. Only rare mitotic figures were seen and the Ki-67 proliferative index was very low. Unexpectedly, the low-grade astrocytoma showed rapid progression within a short time, but subsequent resection showed similar histologic findings to the original biopsy with only slightly more mitoses and a marginally increased Ki-67 proliferative index. Molecular testing performed on the tumor showed no alterations in the IDH1, IDH2, EGFR, or BRAF genes by sequencing, intact 1p/19q by FISH, and a novel BCR-NTRK2 fusion transcript by reverse transcription and anchored multiplex PCR. The patient underwent standard-of-care therapy, both first and second line, for a high-grade glioma because of the aggressive behavior, but the glioma continued to progress despite treatment, and the patient died within 13.5 mo of the original diagnosis. At the time of diagnosis, the BCR-NTRK2 fusion transcript had not been described in solid tumors; however, a recent publication described this fusion transcript in two glioblastomas. Although no approved therapy was available for this patient, FDA approval has now been given for solid tumors with any NTRK gene family fusions. This unexpected molecular finding in a deceptively low-grade-appearing glioma supports the use of expanded molecular testing in gliomas and solid tumors, particularly in instances where targeted therapies are available.