MAP2K1 Mutations Research Articles

The Genomic Landscape of Low-Grade Serous Ovarian/Peritoneal Carcinoma (LGSOC) and its Impact on Clinical Outcome (LBA 7)

The Genomic Landscape of Low-Grade Serous Ovarian/Peritoneal Carcinoma (LGSOC) and its Impact on Clinical Outcome (LBA 7)

Pediatric-Type Indolent B-Cell Lymphomas With Overlapping Clinical, Pathologic, and Genetic Features.

Pediatric-type follicular lymphoma (PTFL) and pediatric nodal marginal zone lymphoma (PNMZL) are rare pediatric-type indolent B-cell lymphomas (PedIBCL) that differ clinicopathologically from their adult counterparts. Accurate diagnosis is important to avoid overtreatment but is often challenging. The mutational landscape of PTFL is known and may aid diagnosis, but the genetic features of PNMZL are not well understood. We analyzed 21 cases of PedIBCL according to their clinicopathologic findings and classified them into PTFL (n=11), PNMZL (n=2), and “mixed type” tumors (n=8) showing ambiguous histology. We also analyzed 2 cases of adult B-cell lymphomas showing features of PedIBCL. Targeted sequencing of 121 lymphoma-related genes was performed. The median age of PedIBCL patients was 16 years (range: 3 to 47), and all but 1 PTFL patient were male. All patients presented with limited-stage disease, and only 1 relapsed. There were no significant differences in clinical features among the 3 PedIBCL groups. The most frequently mutated genes were MAP2K1, TNFRSF14, KMT2C, IRF8, and NOTCH2. The genetic features of all groups were similar to the established mutational landscape of PTFL. The 2 adult B-cell lymphomas cases also had MAP2K1, TNFRSF14, and IRF8 mutations, but the clinical features were not typical of PedIBCL. In summary, this study demonstrated that PTFL and PNMZL are similar diseases with overlapping clinical, pathologic, and genetic features; mixed type tumors can also occur. Atypical adult cases with similar histologic features were also observed. Therefore, the disease spectrum of PedIBCL may be much broader than is currently believed.

Adult Langerhans cell histiocytosis with thyroid gland involvement: clinical presentation, genomic analysis, and outcome.

The present study aims to evaluate the characteristics and treatment outcomes of adult Langerhans cell histiocytosis (LCH) patients with thyroid involvement. We retrospectively described the clinical, biological, and genomic characteristics of a series of 36 LCH patients with thyroid involvement in our center between January 2001 and December 2021. At the time of diagnosis, only one patient was classified as having single-system LCH, and 35 patients were classified as having multisystem (MS) LCH. Three patients had coexisting papillary thyroid carcinoma. Patients with thyroid gland involvement had higher frequencies of pituitary (88.6% vs. 53.4%, P < 0.001), liver (45.7% vs. 20.7%, P = 0.003), and lymph node (54.3% vs. 31.6%, P = 0.012) involvement and a lower frequency of bone (45.7% vs. 72.0%, P = 0.003) involvement than patients without thyroid gland involvement. Sixteen patients had abnormal thyroid function, including nine patients with primary hypothyroidism, one patient with central hypothyroidism, and six patients with subclinical hypothyroidism. BRAFV600E, BRAF N486_P490, and MAP2K1 mutations were detected in 14.3%, 57.1%, and 7.1% of patients, respectively. After a 43-month median follow-up, none of the patients died, and 15 patients experienced reactivation. The median event-free survival was 37.5months. Two of 6 patients with subclinical hypothyroidism had normal thyroid function, and 12 patients still had hypothyroidism after treatment. As the largest adult LCH cohort with thyroid gland involvement to date, we found that patients with thyroid gland involvement had different clinical characteristics, genetic profiles, and outcomes than patients without thyroid gland involvement.

QS17. Endothelial MAP2K1 Mutation Causes Abnormal Vascular Development in Inducible Mouse Strain

Purpose: Arteriovenous malformation (AVM) is defined by abnormal connections between arteries and veins instead of a normal capillary bed. Extracranial AVM is caused by an activating MAP2K1 mutation in endothelial cells. The downstream events and underlying mechanism that result in AVM are not known. The purpose of this study was to describe how the MAP2K1 mutation alters endothelial cell function. Methods: ROSA-GT-Map2k1-K57N+/- mice were crossed with Tg(Cdh5-CreER+/-; ROSA-GT-mTmG+/+) mice to produce offspring that would express mutant Map2k1 mRNA in endothelial cells after administration of tamoxifen. 9-day-old mouse brains were dissected and examined for phenotypic changes. Cerebral endothelial cells were isolated using anti-CD31 magnetic Dynabeads (Invitrogen). ddPCR was performed to calculate recombination frequency of the mutant allele. Bulk RNA sequencing was performed, and gene expression was compared using DESeq2. Genes with an adjusted p-value < 0.05 and absolute log2 fold change > 1 were called as differentially expressed genes. Significantly differentially expressed genes were clustered by their gene ontology. Results: Mutant mouse brains had abnormal vascular development with diffuse punctate vascular lesions. Confocal imaging identified vascular sacculations. The control mouse brain had normal vascular development and lacked these lesions. Mutant allele recombination frequency was 27% in the mutants and 0.1% in the control. Map2k1 expression was higher in mutant endothelial cells (log2FoldChange 2.18). 581 genes were significantly differentially expressed. The gene ontology pathways impacted most were (1) positive regulation of cell migration (GO:0030335) and (2) cell adhesion (GO:0007155). Conclusion: Expression of mutant Map2k1 in mouse endothelial cells results in abnormal cerebral vascular development. The transcriptome of these mutant cells shows significant changes in pathways affecting cell migration and cell adhesion. This inducible mouse strain is a promising animal model to study how the endothelial MAP2K1 mutation causes AVM and to test pharmacotherapy (MEK inhibitors) to treat AVM.

96. Activating MAP2K1 Mutation in Zebrafish Endothelial Cells Causes Arteriovenous Shunts

Purpose: Arteriovenous malformation (AVM) is a sporadic vascular malformation defined by a nidus of irregular blood vessels connecting arteries to veins instead of a normal capillary bed. Somatic activating mutations in MAP2K1 cause extracranial AVM. The purpose of this study was to create an AVM animal model using zebrafish. Methods: Single-cell stage casper;Tg(gata1:dsred) zebrafish embryos were injected with 1 nL of transgenesis mixture: 100 ng/μL of either (1) Control (pTol2-Fli:GFP) or (2) Mutant (pTol2-Fli:GFP-kdrl:MAP2K1K57N) plasmid DNA + 40 ng/μL Tol2 transposase mRNA. Erythrocytes in these fish express the fluorescent protein dsRed. Embryos were anesthetized with 0.4 mg/mL Tricaine, embedded in 1% low-melt agarose and then imaged using a Zeiss LSM 800 Confocal Microscope or a Nikon SMZ18 Fluorescence Microscope. Confocal images were obtained using Zen Blue version 2.5. Embryos were imaged 72 hours post-fertilization. Blood flow was visualized using dsRed fluorescence. Results: Injection of MAP2K1K57N plasmid resulted in abnormal arteriovenous shunts (58/96 transgenic embryos, 60%). The phenotype consisted of either (1) a proximal shunt with blood flowing through a direct connection between proximal aortic vessels to the common cardinal vein and immediately back into the heart (39/58 embryos, 67%) or (2) a mid-trunk shunt between the dorsal artery and posterior cardinal vein (19/58 embryos, 33%). Shunts were not present in control zebrafish (n=65). Endothelial cells at the site of the shunt expressed high levels of the marker transgene confirming shunts contained mutant endothelial cells. Conclusion: Zebrafish endothelial cells expressing mutant MAP2K1 form abnormal arteriovenous shunts. The phenotype recapitulates extracranial human AVM. Mutant MAP2K1 zebrafish are a promising animal model for testing pharmacotherapy to treat AVM.

Abstract 90: STAT3-driven MAPK activation represents a therapeutic vulnerability in ASCL1 high SCLC

Abstract Background Small cell lung cancer (SCLC) is an aggressive high grade neuroendocrine tumor (Hann et al. 2019; Bernhardt et al. 2016). Whilst MAPK mutations can be found in roughly 30% of human cancers (Schubbert et al. 2007) including non-small cell lung cancer (NSCLC), genomic and proteomic analyses have indicated suppression of MAPK pathway activity in SCLC (Cerami et al. 2012; Gao et al. 2013). Previous attempts to determine whether this might be therapeutically important (Ravi et al, 1998; Cristea et al. 2016/2020) have had conflicting conclusions. SCLC has recently been defined by the relative expression of four major transcriptional regulators (ASCL1, NEUROD1, POU2F3, YAP1) (Rudin et al., 2019). In this study we aimed to elucidate the effect of MAPK activation in these different SCLC subtypes and explore its therapeutic vulnerability. Results ASCL1-, NEUROD1- and POU2F3- driven SCLC cell lines were transduced with a doxycycline-inducible vector for expression of MEKDDS217D/S221D (MEK1). Activation through MEK1 in ASCL1-driven SCLC cell lines resulted in a significant decrease in cell growth over 9 days. This was associated with G2 cell-cycle arrest and senescence. Expression of MEK1 in the cells of other SCLC subtypes and NSCLC failed to show any appreciable changes in cell growth. Remarkably, athymic mice injected with a MEK1 expressing ASCL1-driven cell line showed significantly slower tumor formation and longer survival than when MEK1 was not expressed. Notably, we observed the opposite when MEK1 expressing NEUROD1-driven cells were injected into athymic mice. We observed strong upregulation of negative feedback regulators DUSP6 and SPRY2 upon MAPK activation which has previously been described in solid tumors and pre-B ALL (Courtois-Cox et al 2006; Shojaee et al. 2015). Interestingly, phosphokinase array demonstrated that, almost exclusively, STAT3 through phosphorylation at S727 was strongly upregulated in the ASCL1-driven subtype after MEK1 expression. This prompted us to look at other regulators of the JAK-STAT pathway and found an increase in phosphorylation of the inhibitory phosphatase PTPN6 (SHP1) in the ASCL1-driven subtype and no STAT5 phosphorylation in any of the subtypes. We next examined whether these cells were sensitive to STAT3 inhibition. Upon treatment with a STAT3 inhibitor, stattic, ASCL1-driven SCLC cells reached their IC50 after 3-5 days in comparison to 9 days for other SCLC subtypes. MEK inhibition through PD0325901 rescued growth inhibition upon MAPK activation and was further associated with a decrease in DUSP6, SPRY2 and pS727 STAT3. JAK1/2 inhibition through ruxolitinib had no effect on cell viability. Summary These findings suggest that ASCL1-driven SCLC in vitro and in vivo is sensitive to activation of MAPK signaling. Our data provides additional understanding of SCLC biology and its complex signaling networks and potential subtype-specific drug susceptibilities. Citation Format: Rebecca Caeser, Christopher Hulton, Emily Costa, Vidushi Durani, Megan Little, Xiaoping Chen, Sam E. Tischfield, Marina Asher, Faruk Erdem Kombak, Shweta S. Chavan, Nisargbhai S. Shah, Metamia Ciampricotti, Elisa de Stanchina, John T. Poirier, Charles M. Rudin, Triparna Sen. STAT3-driven MAPK activation represents a therapeutic vulnerability in ASCL1 high SCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 90.

Clinical characteristics and treatment outcomes in patients with histiocytic neoplasms harboring class 3 MAP2K1 mutations, including treatment with the ERK inhibitor ulixertinib.

e19081 Background: Histiocytic neoplasms (HN) are clonal myeloid disorders with diverse clinical phenotypes. HN nearly invariably harbor mutations of the mitogen activated protein kinase (MAPK) pathway, including the BRAFV600E mutation in HN subtypes that are responsive to BRAF inhibition. More recently characterized, the second most frequently mutated gene driving HN is MAP2K1, with broad responsiveness to MEK inhibition reported. The most common MAP2K1 variant observed in our cohort (n=300 patients) is the exon 3 p.E102_I103 in-frame deletion, among the Class 3 MAP2K1 mutants predicted to be resistant to allosteric MEK inhibition. We present clinical and treatment characteristics of HN patients with Class 3 MAP2K1 mutations. Methods: Patients with HN and exon 3 p.E102_I103del or similar mutations identified by tumor sequencing were included. Sites of disease were captured. First- and later-line treatments were categorized as observation, chemotherapy (vinblastine, cytarabine, cladribine, methotrexate), immune modulation (anakinra or interferon), MEK inhibition (trametinib or cobimetinib), or ERK inhibition (ulixertinib). Clinical and radiologic responses were captured as partial response (PR), complete response (CR), or progressive disease (PD). PD includes relapse following PR or CR. Results: 16 patients were identified. 8 (50%) were female, and median age at HN diagnosis was 31 (range 22-58). 10 patients had Langerhans cell histiocytosis (LCH), 4 had Erdheim-Chester disease (ECD), 2 had mixed histiocytosis. Sites of HN were bone (16; 100%), lymph node (8; 50%), brain (8; 50%), skin/subcutaneous (4; 25%), retroperitoneum (3; 19%), cardiovascular (3; 19%), abdomen (2; 13%), reproductive (1; 6%) and other sites (5; 31%). Mutations identified were MAP2K1 p.E102_103del (13; 81%), p.L101_103delinsF (1; 6%), p.P105_I107delinsL (1; 6%), and p.I103_A106del (1; 6%). 2 (13%) patients had spontaneous regression of disease and were observed; 3 (19%) patients had CR to first-line chemotherapy. 3 (19%) patients have had CR/PR to first-line MEK inhibition. 8 (50%) patients had PD following chemotherapy and/or immune modulation; of those, 1 was lost to follow-up, 4 had CR/PR to MEK inhibition; however, 3 had PD despite MEK inhibition. These three patients and one treatment-naïve patient were treated with an oral ERK1/2 inhibitor, ulixertinib, on prospective protocols. 3 of 4 had a clinical or radiologic PR (1) or CR (2). Conclusions: Histiocytic neoplasms with Class 3 MAP2K1 mutations represent a diverse spectrum of disease characterized by frequent bone, nodal and neurologic involvement, by frequent resistance to chemotherapy. This entity is resistant to MEK inhibition in some patients, a phenomenon previously undocumented, and responsive to ERK inhibition, which may be a promising therapeutic approach to HN.

Progress in the Treatment of Adult Langerhans Cell Histiocytosis --Review

Langerhans cell histiocytosis (LCH) is a rare proliferative disease dominated by the proliferation of Langerhans cells, which is inflammatory myeloid neoplasms. Its clinical manifestations are variable, occurring at any age and at any site, and it is rarer in adults than in children. The gold standard for diagnosis is histopathological biopsy. Due to the rarity of adult LCH and the heterogeneity of this disease, treatment of adult LCH should be developed according to the extent of the disease and risk stratification. With the discovery of MAPK, PI3K and c-KIT signaling pathway activation, especially BRAF V600E and MAP2K1 mutations, targeted therapy has become a hot spot for therapeutic research. Meanwhile, the discovery of high expression of M2-polarized macrophages and Foxp3+ regulatory T cells (Treg) in LCH has provided an important basis for the immunotherapy. In this article, we will focus on reviewing the latest research progress in the treatment of adult LCH in recent years, and provide a reference for clinical research on the treatment of adult LCH patients.

Updated clinical outcomes from ULI-EAP-100, an intermediate expanded access program for ulixertinib (BVD-523).

e15101 Background: Ulixertinib is a first-in-class selective small molecule inhibitor of ERK1/2 with demonstrated clinical efficacy in patients with solid tumors harboring MAPK pathway alterations in BRAF and NRAS. Emerging clinical data suggest improved efficacy for certain patient populations when MEK/ERK inhibitors are combined with inhibitors of autophagy (hydroxychloroquine), CDK4/6 (palbociclib), or EGFR/BRAF (cetuximab/encorafenib). Methods: ULI-EAP-100 (NCT04566393) is an ongoing intermediate expanded access protocol (EAP) that offers ulixertinib to US patients with advanced solid tumors harboring MAPK pathway alterations including but not limited to KRAS, BRAF, NRAS, MEK1/2, and ERK1/2, who have exhausted or not tolerated standard-of-care. Combinations with approved agents are permitted at the discretion of the treating physician and approval. Clinical data is collected and structured in a 21 CFR Part 11-compliant database according to XCELSIOR (NCT03793088), a real-world observational registry. Results: As of February 1st, 2022, 48 patients (pts) across 17 sites have been treated with ulixertinib under ULI-EAP-100 including pts with colorectal cancer (CRC) (21), melanoma (8), pancreatic ductal adenocarcinoma (PDAC) (6), lung cancer (5), cholangiocarcinoma (4), glioblastoma (2), and appendiceal cancer (2). Qualifying alterations were mutations in KRAS (21 pts), BRAF (13), NRAS (3), MAP2K1 (1), multiple genes (7), and overexpression of MAPK1 (1). Regimens included ulixertinib monotherapy (28 pts), ulixertinib + hydroxychloroquine (12), ulixertinib + encorafenib + EGFRi (cetuximab, panitumumab) (5), ulixertinib + palbociclib (2), and ulixertinib + encorafenib + atezolizumab (1). Thirty-nine (39) pts have discontinued ulixertinib for reasons including progression (25), AEs (9), and patient choice including hospice (5). Most common AEs (any grade/relatedness) were acneiform rash (29%), fatigue (25%), diarrhea (19%), abdominal pain (15%), and anorexia (15%). Grade 3/4 SAEs occurred in 15 pts (31%). Nine pts (19%) expired while on protocol, all attributed to disease progression. Four pts attained clinical benefit including BRAF V600E PDAC and glioblastoma pts on monotherapy for 413 and 128 days, respectively. A BRAF D594G melanoma pt’s tumor rapidly shrunk with monotherapy permitting surgery and rendering pt with no evidence of disease and a BRAF V600E CRC pt experienced a complete response following ulixertinib + encorafenib + cetuximab combination therapy. Conclusions: ULI-EAP-100 is a unique EAP permitting monotherapy and personalized combination regimens at treating physicians’ discretion. Safety of ulixertinib in this program is consistent with previous experience and all grade 5 AEs were attributed to disease progression. Multiple patients who exhausted prior therapies gained significant clinical benefit from ulixertinib under ULI-EAP-100. Clinical trial information: NCT04566393.

Natural distribution of genomic alterations correlated with the resistance to KRASG12C inhibitor in Chinese colorectal cancer.

e15550 Background: KRAS inhibitors exhibited promising efficacy as systemic therapy in KRAS G12C-mutant colorectal cancers (CRC) lately. Recent studies have revealed a number of genomic alterations related to acquired resistance to this type of drugs. The discovered resistant gene mutations include: 1. Novel acquired secondary KRAS mutations within the adagrasib-binding pocket such as Y96C, H95D/Q/R, R68S, 2. Activating mutations in genes ( KRAS, BRAF, MAP2K1) MAPK signaling pathway, 3. Oncogenic fusions involving ALK, RET, BRAF, RAF1, and FGFR3, 4. Gene amplifications in KRAS, MET, MYC. Although found as acquired resistance mechanism to KRAS inhibitors, these genomic alterations might be primary or secondary in the setting of other type of therapies. Characterizing the natural distribution of these genomic alterations in cancer patients would contribute to clarifying the application potential of KRAS inhibitors. We herein performed a large-sample size analysis in Chinese colorectal cancer. Methods: A total of 9385 early-stage, late-stage, treated or treatment-naive CRC were included in this analysis. Calling of single nucleotide variants (SNV), copy number variants (CNV), insertion/deletions (indels), fusions and evaluation of tumor mutational burden (TMB) were performed using a wide panel next-generation sequencing (NGS) testing. Results: 43.4% (4069/9385) cases were KRAS-mutant in this cohort. KRAS G12C account for 5.8% (236/4069) in all KRAS-mutant CRC, and 15.7% (37/236) were harbored the above resistance genomic alterations. Fifteen KRAS G12C-mutant CRC were detected to have activating MAPK mutations, including G12D/V (n = 6), G13D (n = 3), Q61H (n = 6), NRAS Q61K (n = 3), MAP2K1 K57T (n = 3), and BRAF V600E (n = 4). Ten KRAS G12C-mutant CRC were detected to have oncogenic fusions, including TGFBR2- CACNA1D (n = 3), TPM4- ALK (n = 1), FGFR3- TACC3 (n = 1), ETV6- SNRPC (n = 1), KAZN- PAX7 (n = 1), NRG1- TTC12 (n = 1), TCF7L2- EWSR1 (n = 1) and ERBB2 arrangement (n = 2). It is noteworthy that a single case was with 5 gene arrangement events in 17q ( ERBB2- ZPBP2, IKZF3- ERBB2, PGAP3- ERBB2, RARA- ERBB2, RARA- GRB7). Thirteen KRAS G12C-mutant CRC were detected to have amplifications in KRAS (n = 5), MYC (n = 5) and MET (n = 4) genes. TP53 (81%), APC (65%) and PIK3CA (32%) were the utmost frequent somatic mutational genes in this subgroup of CRC. In addition, all cases were microsatellite stable (MSS). Tumors harboring fusion variants showed relatively high TMB level in this cohort. Conclusions: Our analysis results indicate about 15% KRAS G12C-mutant Chinese CRC were probable resistant to KRASG12C inhibitor. One-third of these patients had co-existed oncogenic fusions and higher TMB levels, suggesting possibilities for other therapeutic choices.

Abstract 402: High Prevalence Of Ras/mapk Pathway Mutations In Cell-free Dna Of Extracranial Arteriovenous Malformation: A New Method To Detect The Genotype

Purpose: Extracranial Arteriovenous Malformations(AVMs) were primarily caused by somatic mutations in KRAS and MAP2K1 genes. Targeted chemotherapies are emerging but require molecular diagnosis. Since the AVMs were high flow and high pressure. It is hard to acquire the AVM lesion tissue where the mutant variants were only detected in. Few were detected in Cell-free DNA(CfDNA). Since absolute ethanol embolism is one of the effective methods for treating AVM. We hypothesized that CfDNA of post-embolism could provide the genotype of patients with AVMs. Methods: Peripheral Blood, lesion tissue specimens under the guidance of digital subtraction angiography(DSA), CfDNAs isolated from plasma of 40 patients underwent interventional embolism(before and after as T1(1h, setting first injecting ethanol moment as T0 )). 4 patients ‘s CfDNAs were collaborated in 8h(T8), 16h(T16), 24h(T24), 48h(T48), and 72h(T72) after T0. All the specimen were sequenced by a targeted NGS panel of vascular anomalies Results: Variants were detected in tissue and CfDNA but none in peripheral blood. The prevalence for tissue, CfDNA of pre-embolism and post-embolism(1h) were 73%(27/37), 27.5%(11/40), and 90%(36/40), respectively. KRAS(p.Gln61, p.Gly12) and MAP2K1(p.Gln56, p.Lys57) were the mutant hot spots. Novel mutations in BRAF, RASA1, KRAS, and MAP2K1 were also detected in tissue and cfDNA. As for specificity, area under the ROC curve were 0.8125(P&lt;0.0001), 0.6375(P=0.0343), and 0.9500(P&lt;0.0001). For the variants allele frequency(VAF) of CfDNA, the VAF reached a surge at T1, then a second surge at T24. Conclusion: AVMs were caused by mutations in RAS/MAPK pathway where cfDNA of post-embolism(T1) of absolute ethanol interventional therapy could provide the accurate genotype of the patients in sensitivity and specificity. VAF detected in T24 could provide information related to the natural metabolism such as the indications or prognosis of targeted chemotherapy. Overall, CfDNA of post-embolism provides us a new method for approaching the precise medicine of AVM.

The Morpho-Molecular Landscape of Spitz Neoplasms.

Spitz neoplasms are a heterogeneous group of melanocytic proliferations with a great variability in the histological characteristics and in the biological behavior. Thanks to recent discoveries, the morpho-molecular landscape of Spitz lineage is becoming clearer, with the identification of subtypes with recurrent features thus providing the basis for a more solid and precise tumor classification. Indeed, specific mutually exclusive driver molecular events, namely HRAS or MAP2K1 mutations, copy number gains of 11p, and fusions involving ALK, ROS, NTRK1, NTRK2, NTRK3, MET, RET, MAP3K8, and BRAF genes, correlate with distinctive histological features. The accumulation of further molecular aberrations, instead, promotes the increasing malignant transformation of Spitz neoplasms. Thus, the detection of a driver genetic alteration can be achieved using the appropriate diagnostic tests chosen according to the histological characteristics of the lesion. This allows the recognition of subtypes with aggressive behavior requiring further molecular investigations. This review provides an update on the morpho-molecular correlations in Spitz neoplasms.

Deciphering Genetic Alterations of Hairy Cell Leukemia and Hairy Cell Leukemia-like Disorders in 98 Patients.

Simple SummaryThe diagnosis of hairy cell leukemia (cHCL) and HCL-like disorders, including the variant form of HCL (vHCL) and splenic diffuse red pulp lymphoma (SDRPL) can be challenging, particularly in complex situations. The integration of all data, including molecular data, is essential for distinguishing the different entities. The BRAFV600E mutation is identified in most cHCL cases, whereas it is absent in vHCL and SDRPL. MAP2K1 mutations are observed in half of vHCL cases and in cHCL BRAFWT and they are associated with a worse prognosis. The interest in deep sequencing for the diagnosis and prognosis of hairy cell leukemia and HCL-like disorders is essential. Some KLF2 genetic alterations have been localized on the AID consensus motif, suggesting an AID-induced mutation mechanism. KLF2 is the second most altered gene in HCL, and mutations must be investigated to confirm whether AID could be responsible for the genetic alterations in this gene. Clonal evolution can be observed in half of the cases.Hairy cell leukemia (cHCL) patients have, in most cases, a specific clinical and biological presentation with splenomegaly, anemia, leukopenia, neutropenia, monocytopenia and/or thrombocytopenia, identification of hairy cells that express CD103, CD123, CD25, CD11c and identification of the V600E mutation in the B-Raf proto-oncogene (BRAF) in 90% of cases. Monocytopenia is absent in vHCL and SDRPL patients and the abnormal cells do not express CD25 or CD123 and do not present the BRAFV600E mutation. Ten percent of cHCL patients are BRAFWT and the distinction between cHCL and HCL-like disorders including the variant form of HCL (vHCL) and splenic diffuse red pulp lymphoma (SDRPL) can be challenging. We performed deep sequencing in a large cohort of 84 cHCL and 16 HCL-like disorders to improve insights into the pathogenesis of the diseases. BRAF mutations were detected in 76/82 patients of cHCL (93%) and additional mutations were identified in Krüppel-like Factor 2 (KLF2) in 19 patients (23%) or CDKN1B in 6 patients (7.5%). Some KLF2 genetic alterations were localized on the cytidine deaminase (AID) consensus motif, suggesting AID-induced mutations. When analyzing sequential samples, a clonal evolution was identified in half of the cHCL patients (6/12 pts). Among the 16 patients with HCL-like disorders, we observed an enrichment of MAP2K1 mutations in vHCL/SDRPL (3/5 pts) and genes involved in the epigenetic regulation (KDM6A, EZH2, CREBBP, ARID1A) (3/5 pts). Furthermore, MAP2K1 mutations were associated with a bad prognosis and a shorter time to next treatment (TTNT) and progression-free survival (PFS), independently of the HCL classification.

Attempting to Solve the Pigmented Epithelioid Melanocytoma (PEM) Conundrum: PRKAR1A Inactivation Can Occur in Different Genetic Backgrounds (Common, Blue, and Spitz Subgroups) With Variation in Their Clinicopathologic Characteristics.

Pigmented epithelioid melanocytoma is a rare cutaneous melanocytic proliferation considered high-grade melanocytoma in the 2018 WHO Classification of Skin Tumors. Little has been reported about the associated genetic drivers in addition to BRAF and MAP2K1 mutations or PRKCA gene fusions. Here, we present a series of 21 cases of PRKAR1A -inactivated melanocytic tumors in which we could assess the associated genetic background. We identified 9 different driver genes related to the common, Spitz, blue nevi, and PRKC -fused groups. Nine cases were associated with a canonical BRAF p.V600E mutation, a hallmark of the common nevus group. They occurred mainly in young adults. All were combined (biphenotypic) cases with a variable proportion of compound nevus. The pigmented epithelioid melanocytoma component was made of thin fascicules or isolated epithelioid cells covered by a dense hyperpigmented melanophage background and was predominantly located in the upper dermis. One such case was malignant. Six cases were associated with Spitz-related genetic anomalies ranging from HRAS or MAP2K1 mutations to gene fusions involving MAP3K8 , MAP3K3 , and RET . They occurred mainly in children and young adults. Morphologically, they showed large confluent junctional nests in a hyperplastic epidermis and a fascicular dermal component of spindled and epithelioid melanocytes with a frequent wedged silhouette. Intravascular invasion was observed in 4/6 cases. Five cases were associated with canonical mutations of the blue nevus group with 4 CYSLTR2 p.L129Q and 1 GNAQ p.Q209L mutations. They were removed mainly in adults and showed a frequent junctional component with epidermal hyperplasia. The dermal component showed dense fascicules of spindled and epithelioid melanocytes predominating over melanophages. One case occurred in a PRKCA -fused tumor in an adolescent with classic morphologic features. These results could potentially shift the concept of PRKAR1A -inactivated melanocytoma, changing from a rather unified model to a more complex one, including genetic subgroup variations with clinical and morphologic specificities. The genetic background of PRKAR1A -inactivated melanocytic tumors should be systematically explored to better understand the extent and clinical behavior of these complex lesions.

Abstract PD15-03: Overlapping molecular features (proliferation, immune signatures andTP53mutations) associated with palbociclib resistance inER+HER2- primary breast cancer

Abstract Background: Cyclin-dependent kinase 4/6 (CDK4/6) inhibitors in combination with anti-hormone therapy are highly effective treatments for oestrogen receptor positive (ER+) and HER2 negative (HER2-) advanced breast cancer. Pre-clinical and clinical studies have reported mechanisms of resistance to CDK4/6 inhibitors to include interferon signalling, high CCNE1 expression and loss of RB1 expression. The PALLET phase II randomized neoadjuvant trial of letrozole (LET) ± palbociclib (PALBO) in postmenopausal ER+HER2- primary breast cancer showed that suppression of proliferation as measured by Ki67 was significantly greater with addition of PALBO to LET but did not result in all patients achieving complete cell-cycle arrest, indicating intrinsic resistance in some patients. We report phenotypes/genotypes associated with that resistance. Methods: In the PALLET trial, 307 patients were randomized to LET+PALBO (n=204) or LET (n=103) for 14wks. For the first 2wks of LET+PALBO patients were randomised to LET, PALBO or LET+PALBO. RNA-seq of baseline samples from consented patients was performed on fresh frozen biopsies for 224 patients (LET-only n=77; LET+PALBO n=147); whole exome sequencing was performed on 188 tumors and matched blood samples (LET-only n=61; LET+PALBO n=127). After 14wks of treatment, those patients with Ki67% &amp;lt; 2.7% were classified as having complete cell-cycle arrest (CCCA). Differentially expressed genes (DEGs) were identified between patients sensitive (CCCA) and resistant (non-CCCA) to treatments with or without PALBO at 14wks by DESeq2. Mutect2 and VarScan was used to identify somatic mutations and CNVkit was used to identify copy number alterations in whole exome sequencing (WES). Results: In LET+PALBO treated patients, higher expression of E2F targets, interferon gamma response and mTORC1 signalling genes were observed in baseline gene expression of non-CCCA patients at 14wks (FDR&amp;lt;0.05, GSEA). Similar results were also observed if using 2wk Ki67 data. In LET-only non-CCCA patients, higher expression of mTORC1 signalling and lower expression of oestrogen response genes (FDR&amp;lt;0.05, GSEA) were observed. Additional analysis of baseline gene expression for non-CCCA at 14wks LET+PALBO patients showed higher expression of immune checkpoint inhibition associated genes including IFNG, IDO1, PD-L1 (FDR&amp;lt;0.05, DESeq2), higher expression of genes expressed only in immune cells and two gene signatures related to interferon signalling and immune checkpoint blockade (FDR&amp;lt;0.05, GSEA). Somatic mutation analysis showed a trend for enrichment of mutations in TP53 for both LET-only and LET+PALBO non-CCCA patients (p=0.02 and p=0.06, respectively, Fisher-exact) and significant enrichment of MAP3K1 mutations in LET-only CCCA patients (p&amp;lt;0.05, Fisher-exact). TP53 mutations were also associated with higher proliferation, mTORC1 and immune related signatures (all p&amp;lt;0.01, Mann-Whitney). Change at 14wks Ki67 was significantly different (p=0.02, Mann-Whitney) between TP53wt and TP53MUT for LET-only patients (median WT = -92%, MUT = -66%, p=0.02, Mann-Whitney ) but not for LET+PALBO (median WT = -99% MUT = -95%, p=0.13, Mann-Whitney). No copy number alterations were significantly enriched in LET+PALBO non-CCCA patients. Conclusion: We observe, confirming previous studies, an association of CDK4/6 inhibitor resistance, high expression of CCNE1 and genes related to interferon gamma signalling. We show that there is an overlap between resistance mechanisms and TP53 mutations. However, ER+HER2- patients with TP53 mutations may still benefit from PALBO adding to suppression of proliferation compared to LET-only treatment. Citation Format: Eugene F Schuster, Hui Xiao, Maggie Cheang, Elena Lopez-Knowles, Lucy Kilburn, Viktoriya Korchina, Sejal Salvi, Samuel A Jacobs, Melanie Finnigan, David A Wheeler, Shannon Puhalla, Donna Muzny, Harsha Doddapaneni, Katherine Pogue-Geile, Yuan Liu, Judith Bliss, Stephen Johnston, Mitch Dowsett, Mothaffar Rimawi, On behalf of the PALLET Trialists. Overlapping molecular features (proliferation, immune signatures andTP53mutations) associated with palbociclib resistance inER+HER2- primary 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 PD15-03.

Mucinous Histology Is Associated with Resistance to Anti-EGFR Therapy in Patients with Left-Sided RAS/BRAF Wild-Type Metastatic Colorectal Cancer.

BackgroundLimited studies have suggested that mucinous histology is associated an attenuated response to anti-epidermal growth factor receptor (EGFR) therapy.MethodsWe conducted a single-institution, retrospective study to review the anti-EGFR response and the molecular profile of patients with left-sided microsatellite stable RAS/BRAF wild-type mucinous metastatic colorectal cancer.ResultsIn comparison to nonmucinous population (n = 98), mucinous histology (n = 20) was associated with a younger age (48 vs 54, P = .02), wild-type APC (80% vs 15.3%, P < .0001), and wild-type TP53 (40% vs 8.2%, P = .001). Guanine nucleotide binding protein, alpha stimulating (GNAS) mutations were exclusively found in mucinous tumors (20% vs 0, P < .0001). Genomic alterations associated with resistance to anti-EGFR therapy, such as ERBB2 amplification, PIK3CA mutation, MAP2K1 mutation, and KRAS amplification, were identified in patients with left-sided RAS/BRAF wild-type mucinous metastatic colorectal cancer. Mucinous histology was not associated with a worse outcome than non-mucinous histology (34.3 vs 42.2 months, P = .85). However, patients with left-sided RAS/BARF wild-type mucinous colorectal cancer treated with first-line anti-EGFR therapy had significantly worse progression-free survival (4 vs 6.5 months, hazard ratio [HR] = 5.3, 95% confidence interval [CI] 1.3-21.7, P = .01) than patients treated with the first-line vascular endothelial growth factor A antibody, bevacizumab. Anti-EGFR therapy was associated with limited responses and a short PFS across all lines of therapy in 12 patients with left-sided RAS/BRAF wild-type mucinous colorectal cancer.ConclusionsMucinous histology is associated with diminished benefits from anti-EGFR therapy in patients with left-sided RAS/BRAF wild-type colorectal cancer. These patients should be considered for bevacizumab-based therapy in the first- and second-line settings.

Tumor board presentation of a woman with metastatic, hormone receptor‐positive, mismatch repair‐deficient endometrial cancer

Tumor board presentation of a woman with metastatic, hormone receptor‐positive, mismatch repair‐deficient endometrial cancer

Combined treatment with vemurafenib and cobimetinib in Langerhans cell histiocytosis and Erdheim-Chester disease overlap syndrome: A case Report

Histiocytoses are clonal disorders diseases derived from the monocyte-macrophage lineage. The Erdheim–Chester Disease (ECD) and Langerhans Cell Histiocytosis (LCH) may occur in association with overlapping clinical, histopathological and molecular features, harboring somatic MAP2K1 mutations in more than 50% of patients. BRAF and MEK inhibitors have shown to be efficacious in ECD and LCH, including responses in patients with CNS involvement. This case report describes a 59-year-old woman who presented with vemurafenib-refractory ECD/LCH overlap syndrome treated with vemurafenib/cobimetinib dual therapy, with rapidly progressing neurological involvement after its initiation. Although targeted therapy plays a crucial role in the treatment of histiocytosis, only anecdotal clinical cases treated with dual therapy have been reported in ECD or LCH and collaborative trials are needed to improve outcomes.

Spectrum of MAP3K1 mutations in breast cancer is luminal subtype-predominant and related to prognosis.

MAP3K1 is a MAPK family serine-threonine kinase that is frequently mutated in human cancer. The association between mutations in the MAP3K1 gene and the clinicopathological characteristics and prognosis of patients with breast cancer remain unclear in the Chinese population. Thus, the aim of the present retrospective study was to investigate the possible role and function of MAP3K1 in breast cancer. Data obtained from 412 consecutive patients with breast cancer were selected from Guangdong Provincial People's Hospital (GDPH) for analysis in the present study. Mutations were assessed using next-generation sequencing. The association between MAP3K1 mutations and clinicopathological features were analyzed and further compared with the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) cohort and data from The Cancer Genome Atlas (TCGA). In the GDPH cohort, a total of 45 mutations MAP3K1 were identified in 8.5% (n=35) of the 412 patients, compared with 9.7% (n=244) in METABRIC and 7.9% (n=88) in TCGA. The majority of the mutations identified in the in three cohorts were truncating mutations, followed by mis-sense mutations. Mutations in MAP3K1 were predominant in patients with the luminal A and B breast cancer subtypes in METABRIC datasets (P<0.001), although no significant differences were observed in the GDPH cohort (P=0.227). In the METABRIC cohort, patients with MAP3K1 mutations experienced a improved overall survival (OS) rate than patients without MAP3K1 mutations (P=0.006). In patient with hormone receptor (HR)+ breast cancer, a more significantly higher OS rate was observed in patients with MAP3K1 mutations (P<0.001). MAP3K1 expression was associated with OS in the HR+ subgroup. Moreover, the MAP3K1 methylation levels were reduced in primary breast cancer tissue, compared with normal tissue. Thus, the present findings identified MAP3K1 mutations in Chinese patients with breast cancer, and compared MAP3K1 mutations between the cohorts from Western and Eastern countries.

Characterization of bone homeostasis in individuals affected by cardio-facio-cutaneous syndrome.

Cardio-facio-cutaneous syndrome (CFCS) is a rare disorder characterized by distinctive craniofacial appearance, cardiac, neurologic, cutaneous, and musculoskeletal abnormalities. It is due to heterozygous mutations in BRAF, MAP2K1, MAP2K2, and KRAS genes, belonging to the RAS/MAPK pathway. The role of RAS signaling in bone homeostasis is highly recognized, but data on bone mineral density (BMD) in CFCS are lacking. In the present study we evaluated bone parameters, serum and urinary bone metabolites in 14 individuals with a molecularly confirmed diagnosis of CFCS. Bone assessment was performed through dual X-ray absorptiometry (DXA); height-adjusted results were compared to age- and sex-matched controls. Blood and urinary bone metabolites were also analyzed and compared to the reference range. Despite vitamin D supplementation and almost normal bone metabolism biomarkers, CFCS patients showed significantly decreased absolute values of DXA-assessed subtotal and lumbar BMD (p ≤ 0.05), compared to controls. BMD z-scores and t-scores (respectively collected for children and adults) were below the reference range in CFCS, while normal in healthy controls. These findings confirmed a reduction in BMD in CFCS and highlighted the importance of monitoring bone health in these affected individuals.