MYD88 L265P Research Articles

Lymphoplasmacytic lymphoma with IgG-κ paraproteinemia presenting as a hepatic bulky mass

Lymphoplasmacytic lymphoma (LPL) usually involves bone marrow (BM) and sometimes lymph nodes and spleen. LPL presenting as a hepatic mass lesion is extremely rare, with only one case reported in the English literature. A 70-year-old Japanese female presented to us with a right hypochondriac mass with tenderness. Computed tomography (CT) revealed a 14 cm-sized bulky hepatic mass. Laboratory findings showed a normal white blood cell count of 4.1×109/L with 4% plasmacytoid lymphocytes; normocytic anemia, Hb 9.4 g/dL; high soluble IL-2 receptor level, 2,290 U/mL; and elevated IgG, 10,306 mg/dL. Furthermore, IgG-κ monoclonal protein was detected. 18F-fluorodeoxyglucose-positron emission tomography/CT revealed abnormal uptake in the liver mass; left supraclavicular, parasternal, abdominal, and left inguinal lymph nodes; and bilateral lung bases. Magnetic resonance imaging showed no bone lesions. BM aspiration and liver biopsy showed predominant infiltration of small lymphocytes admixed with plasmacytoid lymphocytes and plasma cells. In the liver specimen, lymphoepithelial lesions were not observed. The small lymphocytes were positive for CD20, CD79a, and bcl-2, and negative for CD5, CD10, cyclin D1, and IRTA1; plasma cells in BM were positive for CD19, CD45, IgG, and κ-chain, and negative for CD20, and CD56. MYD88 L265P mutation, reported in approximately 40% of non-IgM LPL cases, was not detected in the liver specimen and BM cells. The frequency is lower than that of typical IgM LPL. These findings led us to a diagnosis of LPL with IgG-κ paraproteinemia. The patient underwent four courses of R-CHOP and two courses of Bendamustine-R. Partial remission was achieved.

Immunoglobulin G Kappa Lymphoplasmacytic Lymphoma with Associated Al Amyloidosis: A Rare Combination!

Lymphoplasmacytic lymphoma (LPL) is a low-grade B-cell neoplasm, composed of small B-lymphocytes, plasmacytoid lymphocytes, and plasma cells involving bone marrow and sometimes lymph nodes or spleen and forms 1% of all non-Hodgkin lymphomas. LPL with bone marrow involvement and an Immunoglobulin M (IgM) monoclonal gammopathy of any concentration is designated as Waldenström macroglobulinemia (WM). Due to their frequent co-occurrence, the terms LPL and WM are often used interchangeably in clinical practice. However, although the clinical diagnosis of WM is restricted to cases with an IgM monoclonal protein, the World Health Organization recognizes that LPL can rarely present with paraproteins of immunoglobulin G (IgG) or immunoglobulin A (IgA). LPL associated with non IgM gammopathies are extremely rare and form >5% of all LPLs. We hereby present the case of a 65-year-old female with no previous co morbidities who presented with nonspecific complaints of weight loss, easy fatigability, and night sweats. Examination was within the normal limits except for the presence of subcentimetric axillary lymphadenopathy. She was found to have anemia with an altered A/G ratio M spike of 2.10 g/which was IgG kappa on immunofixation electrophoresis. The bone marrow revealed a mixture of lymphocytes, lymphoplasmacytoid cells, and plasma cells along with the presence of amyloid deposits. MYD88 L265P mutation done on peripheral blood was positive, and based on these, she was diagnosed with a case of IgG Kappa LPL with associated amyloidosis.

MYD88 Gene Polimorphism in Patients With Chronic Lenfoid Leukemia

Leukemia is a type of cancer that manifests itself with the excessive proliferation of blood cells, especially white blood cells, and affects the blood production system in the body. Leukemias are classified as acute or chronic and according to the extent and development of the tumor. In chronic lymphocyte leukemia (CLL), cancer cells are found in the bone marrow, blood, and lymph nodes. Therefore, in this study, the effects of polymorphisms in the MYD88 gene of CLL patients were investigated. For this purpose, 100 patients diagnosed with CLL and 70 healthy individuals without cancer history were included in the study as the control group. DNA was isolated from CLL patients and control groups. Polymorphisms in the MYD88 (L265p T/C) promoter regions were studied by RT-PCR. Genotype and allele frequencies were calculated directly. Statistical significance of genotypic distributions between CLL patients and control groups was evaluated with Z Ratios test and Fisher's Exact test.For the MYD88 L265p T/C region in 100 CLL patients, 95 wild type TT, 3 TC and 5 mutant type CC genotypes were detected. No statistically significant difference was found between CLL patients and control groups in the MYD88 L265p T/C region (p 0.05). According to the allele frequencies of the region, it was determined as 0.965 and 0.065 for T and C, respectively. According to Fisher's Exact results, it was calculated as 0.078 for TT, 0.078 for TC and 0.269 for Mutant, respectively. The MYD88 genotype frequencies of the patients included in the study do not appear to be statistically correlated with CLL patients. The research we have done as a result of our study will shed light on the studies with different SNPs to be made with the increase in the number of CLL patients..

Lymphoplasmacytic lymphoma: a clinicopathological and prognostic analysis of 27 cases

Objective: To study the clinical manifestations, pathologic features, diagnosis and differential diagnosis, treatment and prognosis of lymphoplasmacytic lymphoma/Waldenström's macroglobulinemia (LPL/WM). Methods: Twenty-seven cases of LPL from January 2016 to December 2020 at Guangdong Provincial People's Hospital were collected. The clinical data, histomorphology, immunophenotype, MYD88 L265P mutation, treatment and prognosis were analyzed retrospectively. Results: There were 19 males and 8 female patients, with median age of 63 years. The most common initial symptoms were fatigue related to anemia. Bone marrow was involved in all cases, lymphadenopathy was seen in 11 cases and splenomegaly in 10 cases. Monoclonal IgM type protein was detected in 25 cases, meeting the diagnostic criteria of WM. Microscopically, bone marrow and lymph nodes were infiltrated by small lymphocytes, plasmacytoid lymphocytes or plasma cells. The cells expressed pan B-cell markers and showed immunoglobulin light chain restriction. There was no expression of CD5, and low expression of CD23 and CD10; Ki-67 index was usually low. The positive rate of MYD88 L265P mutation was 73.9% (17/23). Most of the patients were treated with rituximab combined with alkylating agents, nucleoside analogues or immunomodulators, and the few patients with relapse or progression were treated with Ibutinib. During the 3-168 months' follow-up period, recurrence or progression were seen in nine cases. Thrombocytopenia, elevated β2-microglobulin and high-risk group were associated with recurrence or progression of the disease (P<0.05). The overall survival (OS) and progression-free survival (PFS) of the high-risk patients were significantly lower than those of the low-medium risk patients (P<0.05). Conclusions: LPL/WM is an exclusive diagnosis; the detection of MYD88 L265P mutation has high diagnostic value, but it is not specific. These cases should be assessed comprehensively for their clinical manifestation, serum IgM protein level and immunophenotype. The overall prognosis of LPL/WM is good, but there are still a small number of high-risk patients with rapid progress, and so the symptomatic patients should be diagnosed accurately and treated in a timely manner.

GEN-7 Liquid biopsy in brain tumor patients -The present and future-

We have previously published liquid biopsy for the diagnosis of brain tumors including PCNSL (JCO Precision Oncology, 2019; Leukemia and Lymphoma, 2019) and diffuse midline gliomas (DMG) (Diagnostics, 2021). We used the Maxwell RSC cfDNA extraction kit to extract circulating tumor DNA (ctDNA from) 1 milliliter of cerebrospinal fluid (CSF), and droplet digital PCR to detect MYD88 L265P mutations in PCNSL and H3F3A K27M mutations in DMG. From our initial experience, we were able to detect a high rate of MYD88 mutations in PCNSL, but not H3F3A mutations in DMG. We also observed that higher concentrations of ctDNA were obtained when prompt centrifugation and storage were done after obtaining CSF. Application of liquid biopsy to early detection of relapse and monitoring of treatment relapse are highly anticipated. In cases of PCNSL, we perform liquid biopsy when relapse is suspected on post-contrast MRI. However interestingly, the rate of MYD88 mutations detected is lower than that of newly-diagnosed cases. We would also like to share our experience performing liquid biopsy in conjunction with CSF cytology in brain tumor patients with evidence of leptomeningeal disease. From our initial experience, we would like to discuss the present limitations and future prospects of liquid biopsy in brain tumor patients.

ML-7 Liquid biopsy for MYD88 mutation in cerebrospinal fluid in patients with suspected primary CNS lymphoma

Background: Treatment intervention for central nervous system lymphoma (CNSL) requires pathological diagnosis by surgical biopsy. However, there are some cases in which the risk of surgery is high due to age, comorbidities, localization of lesions, etc. We are developing a CNSL diagnostic method based on the detection of MYD88 L265P mutation by digital PCR (dPCR) using CSF-DNA, and a high accuracy with a sensitivity of 92.9% and a specificity of 100% has been reported. Here, we report two cases with suspected brain stem CNSL, whose treatment strategy was determined by integrated clinico-laboratory information including neurological presentations, imaging, and the result of liquid biopsy. Result: Case 1. A 63-year-old woman visited our hospital with a complaint of right hemiplegia, which deteriorated in two months. MR images revealed a contrast-enhancing lesion in the left midbrain-ventral pons, suggesting CNSL. Biopsy was not considered because of its location, while dPCR using CSF-DNA showed a cluster of MYD88 mutation signals. Based on these work-ups, she was treated with high-dose methotrexate-based chemotherapy, resulting in a complete response with marked improvement of symptoms. Case 2. An 83-year-old man was referred for a history of diplopia and ataxic gait lasting for a month. MR images revealed an invasive lesion on his right midbrain-dorsal pons. Biopsy was declined due to the location, and liquid biopsy using CSF-DNA was performed to assist the diagnosis. In the first test, the CSF-DNA yield was too insufficient to determine the mutation signal by dPCR. The second dPCR using sufficient amount of CSF-DNA resulted in the Target/Total value of 0.049% which was lower than the threshold, suggesting the absence of MYD88 mutation. The patient underwent radiation therapy accordingly.Conclusions: CSF MYD88 mutation analysis by dPCR may have clinical utility and requires sufficient amount of CSF-DNA for exclusion of noise signals.

CD5-Negative, CD10-Negative Low-Grade B-Cell Lymphoproliferative Disorders of the Spleen.

CD5-negative, CD10-negative low-grade B-cell lymphoproliferative disorders (CD5-CD10-LPD) of the spleen comprise a fascinating group of indolent, neoplastic, mature B-cell proliferations that are essential to accurately identify but can be difficult to diagnose. They comprise the majority of B-cell LPDs primary to the spleen, commonly presenting with splenomegaly and co-involvement of peripheral blood and bone marrow, but with little to no involvement of lymph nodes. Splenic marginal zone lymphoma is one of the prototypical, best studied, and most frequently encountered CD5-CD10-LPD of the spleen and typically involves white pulp. In contrast, hairy cell leukemia, another well-studied CD5-CD10-LPD of the spleen, involves red pulp, as do the two less common entities comprising so-called splenic B-cell lymphoma/leukemia unclassifiable: splenic diffuse red pulp small B-cell lymphoma and hairy cell leukemia variant. Although not always encountered in the spleen, lymphoplasmacytic lymphoma, a B-cell lymphoproliferative disorder consisting of a dual population of both clonal B-cells and plasma cells and the frequent presence of the MYD88 L265P mutation, is another CD5-CD10-LPD that can be seen in the spleen. Distinction of these different entities is possible through careful evaluation of morphologic, immunophenotypic, cytogenetic, and molecular features, as well as peripheral blood and bone marrow specimens. A firm understanding of this group of low-grade B-cell lymphoproliferative disorders is necessary for accurate diagnosis leading to optimal patient management.

Abstract P073: Pharmacological inhibition of IRAK4 with CA-4948 is beneficial in marginal zone lymphoma models with secondary resistance to PI3K and BTK inhibitors

Abstract Background. Marginal zone lymphoma (MZL) is a heterogeneous B-cell malignancy for which no standard treatment exists. B-cell receptor (BCR) signaling dysregulation is a hallmark of MZL and can mimic antigen dependent BCR activation. However, single treatment with BCR signaling inhibitors shows limited complete response rate and relapsing patients. IRAK4 is a protein kinase downstream the Toll-like receptor signaling. CA-4948 is the first-in-class inhibitor of IRAK4 and it is in phase 1/2 studies for patients with relapsed/refractory clinical studies with favorable activity in lymphomas and myeloid neoplasms. Here, we assessed IRAK4 inhibition with CA-4948 against a panel of MZL cell lines in which we have developed secondary resistance to inhibitors of PI3K (idelalisib, copanlisib) or BTK (ibrutinib). Methods. Models with secondary resistance were developed by continuous exposing cell lines derived from splenic MZL (Karpas 1718 and VL51) to high doses of drugs for long period of times. Resistant and parental lines were exposed to increasing doses of CA-4948, alone or in combination with compounds they are resistant to. Cell viability was tested by MTT assay (72h treatment). Synergy of combinations was evaluated according to the Chou-Talalay combination index (CI), as well as potency and efficacy, estimated according to the MuSyC algorithm. Results. The parental Karpas1718, bearing a MYD88 L265P mutation, had an IC50 of 3.29 µM for CA-4948 as single agent, while the parental VL51 and all resistant models presented IC50 values in the range of 19-35 µM. IRAK4 inhibition with CA-4948 had strong benefit when combined to downstream BCR inhibitors. In resistant cells, CA-4948 was strongly synergistic with idelalisib, ibrutinib and, at lesser extent, with copanlisib. CA-4948 in combination with ibrutinib was strongly synergistic especially in the VL51 ibrutinib resistant model compared to the parental one. CA-4948 (from 1 to 5 µM) recovered sensitivity to ibrutinib at IC50 values close to the parental condition. Similarly, CA-4948 in combination with idelalisib was also synergistic and increased sensitivity to idelalisib in idelalisib resistant VL51. Less benefit was seen adding CA-4948 to copanlisib: the combination was synergistic in VL51 parental cell, but with only weak efficacy. In the Karpas1718, CA-4948 was synergistic either in idelalisib resistant or in parental, with improved benefit in the latter, in which synergistic effect was observed when combined with both idelalisib and ibrutinib. The synergistic activity of CA-4948 was mainly due to improved efficacy rather than potency of the combinatorial partners. Conclusions. Our results in MZL cell lines show that the IRAK4 inhibitor CA-4948 is synergistic with small molecules targeting BCR signaling, especially with idelalisib and ibrutinib. Data also suggest that concentrations of CA-4948 comparable to those achieved in patients might overcome or reduce secondary resistance to the tested tyrosine kinase inhibitors. Citation Format: Francesca Guidetti, Alberto J. Arribas, Filippo Spriano, Laura Barnabei, Reinhard von Roemeling, Elizabeth Martinez, Emanuele Zucca, Francesco Bertoni. Pharmacological inhibition of IRAK4 with CA-4948 is beneficial in marginal zone lymphoma models with secondary resistance to PI3K and BTK inhibitors [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P073.

MYD88 L265P Augments Proximal B-Cell Receptor Signaling in Large B-Cell Lymphomas Via an Interaction with DOCK8

Diffuse large B-cell lymphoma (DLBCL) is a genetically heterogeneous disease comprised of five subtypes including a subset of poor-prognosis activated B cell (ABC)-enriched tumors with frequent MYD88L265P mutations, often in association with CD79B alterations (Cluster 5 DLBCLs) (Nat. Med. 2018; 24:679-690). Primary central nervous system lymphomas (PCNSLs) and primary testicular lymphomas (PTLs) have similar genetic signatures including recurrent MYD88L265P mutations and concurrent CD79B alterations (Blood 2016; 127: 869-81). These findings prompted us to evaluate a potential role for MYD88L265P in proximal B-cell receptor (BCR) signaling, in addition to its defined function as an intermediary in the Toll-Like Receptor (TLR) pathway and downstream NF-kB activation.In previous studies by Jabara et al., wild-type (WT) MYD88 was found to be constitutively associated with the DOCK8 adapter and the PYK2 tyrosine kinase in normal B-cells (Nat. Immunol. 2012; 13:612-20). In this setting, physiologic ligation of TLR9 with CpG oligodeoxynucleotides (CpG) induced PYK2-mediated phosphorylation of DOCK8, recruitment of Src kinases, including LYN, and downstream activation of the proximal BCR pathway member, spleen tyrosine kinase (SYK) (Nat. Immunol. 2012; 13:612-20). We postulated that mutated MYD88L265P might similarly augment proximal BCR signaling in DLBCLs in the absence of physiologic (CpG-induced) TLR9 signaling. Using three DLBCL cell lines (OCI-Ly1, SU-DHL4 and OCI-Ly7) with intact BCR signaling and WT endogenous MYD88 and CD79B, we first established that physiologic CpG activation of TLR signaling induced the phosphorylation of PYK2 and the proximal BCR signaling components, SYK and Bruton's tyrosine kinase (BTK). Thereafter, we genetically engineered these three DLBCL cell lines to express MYD88 L265P or MYD88 WT, alone or in association with CD79B Y196F. In all three cell lines, the co-expression of MYD88 L265P and CD79B Y196F significantly increased magnitude and duration of SYK and BTK phosphorylation following BCR crosslinking. These findings highlight the likely role of MYD88L265P in CD79BY196F-associated proximal BCR signaling in DLBCL.To elucidate the potential role of the DOCK8 adapter in MYD88 L265P-augmented BCR signaling, we first assessed the colocalization of MYD88 WT or MYD88 L265P with DOCK8 in the same three genetically engineered DLBCL cell lines using proximity ligation assays (PLA), which detect protein-protein interactions at less than 40 nm in situ. In each of these cell lines, we detected significantly increased co-localized MYD88 L265P/DOCK8 signals in comparison to MYD88 WT/DOCK8 signals (p<.0001, all). Additionally, there were significantly increased co-localized DOCK8/LYN signals in DLBCL cell lines that expressed MYD88 L265P rather than MYD88 WT (p<.0001, all). These data provide the first direct evidence of an enhanced association between MYD88 L265P, DOCK8 and LYN in BCR-dependent DLBCLs and a basis for enhanced BCR signaling in primary tumors with concurrent MYD88L265P and CD79B genetic alterations.We next analyzed the consequences of MYD88 L265P-associated, DOCK8-dependent increased proximal BCR signaling by depleting DOCK8 in BCR-dependent DLBCL cells with endogenous MYD88L265P/CD79BY196F alterations (HBL1 and TMD8) or endogenous unmutated MYD88 WT/CD79B WT (OCI-Ly1 and SU-DHL4). ShRNA-mediated DOCK8 knockdown (KD) significantly decreased BCR-mediated phosphorylation of SYK and BTK in MYD88L265P/CD79BY196F DLBCL cell lines but not in lines with MYD88 WT/CD79B WT, highlighting the specific role of DOCK8 in MYD88 L265P-associated proximal BCR signaling. Of great interest, DOCK8 KD selectively decreased the proliferation of MYD88L265P/CD79BY196F, but not MYD88WT/CD79BWT, DLBCLs (p<.004, HBL1 and p<.009, TMD8; p = non sig., OCI-Ly1 and SU-DHL4). Additionally, DOCK8 KD significantly increased the efficacy of chemical PI3Kα/δ (copanlisib) and BTK (ibrutinib) inhibition in MYD88L265P/CD79BY196F DLBCLs (HBL1 and TMD8). Taken together, these data identify DOCK8 as an intermediary in MYD88L265P-driven proximal BCR signaling and a possible treatment target in LBCLs with co-occurring MYD88L265P/CD79BY196F mutations. DisclosuresShipp: AstraZeneca: Consultancy, Research Funding; Immunitas Therapeutics: Consultancy; Bristol Myers Squibb: Research Funding; Merck: Research Funding; Bayer: Other: Institution: Research Grant/Funding; Abbvie: Other: Institution: Research Grant/Funding.

Single Cell Analysis of MYD88 L265P and MYD88 WT Waldenström Macroglobulinemia Patients

Waldenström macroglobulinemia (WM) is a low-grade B- cell lymphoproliferative disorder characterized by bone marrow (BM) infiltration by small lymphoplasmacytic lymphoma (LPL) cells that secrete monoclonal IgM immunoglobulin. Our understanding of the pathogenesis of WM has improved significantly by the discovery that the vast majority of patients with WM harbor the somatic mutation L265P in MYD88 gene on their clonal cells. However, the small fraction of patients (3-10%) that lack mutations in MYD88 is characterized by a different clinical course, with an increased risk of disease transformation and probably shorter overall survival. WM is a clonal disease but there may be significant heterogeneity within the clone, which may be associated with clinical outcomes, however, this has not been extensively explored. Recent advances in the area of single-cell RNA sequencing (scRNA-Seq) technologies have improved so that it is feasible to sequence and analyze thousands of cells per tumor delivering significant insights into a tumor's cellular heterogeneity and the biological features that distinguish different cell populations.To better understand the differences occurring in the B cell composition between WM patients harboring the MYD88 L265Pmutation and those with MYD88 WT respectively, we performed single-cell RNA sequencing on CD19+ sorted cells from BM aspirates of these patients. Both patients were wild type to CXCR4 mutations. Overall, we characterized 8,517 cells from one MYD88 L265P WM patient (3,116 cells), one MYD88 WT patient (3,348 cells) and one healthy donor as control (2,053 cells). Clustering of these cells based on their gene expression profile, revealed that the majority of the cells form a large cluster corresponding to B cells (naïve, immature and mature;6852 cells) while the rest of the cells were classified into four broad cell types ranging from pro-B cells (1243 cells) to only few cells belonging to mature populations engaged in the immune response such as plasma cells (190 cells), T cells (124 cells) and monocytes (53 cells) indicating a high purity of the samples (≥95%) (Fig 1A). Clonality assessment showed high lambda light chain expressing populations in both patients and as expected an evenly distributed polyclonal population in the healthy donor.To investigate the relative composition of the different cell types between the two genetically distinct WM patients (MYD88 L265P vs MYD88 WT) we searched for populations that were enriched in the B cell repertoire compared to the normal BM. From our data we observe a significant enrichment of pro-B cells in the MYD88 WT patient which is almost completely missing in the MYD88 L265P patient and an enhanced naïve B cell population in MYD88 L265P patient compared to both MYD88 WT patient and the healthy donor (Fig 1B, C). In addition, immature B cells are 3-fold higher in WM patients compared to the healthy donor. Differential gene expression analysis in B cells showed that among both patients several genes are highly expressed (>1.5 log2 fold-change) compared to the healthy donor such as CD52, FCMR, CD74, CD37 and HLA-DPA1. Genes that were highly upregulated in the MYD88 mutant compared to the wild-type WM patient include CXCR4, TXNIP, ID3, LLT1, MTRNR2L8, ZFP36L2 and ZNF331. On the other hand, genes that are highly upregulated in the wild-type phenotype compared to the mutant phenotype include MS4A1, CD79A, CD82, CD1C, KIAA0040, CD72, EGR1, SYK, and LGALS1 (Fig 1D, E). Distinct pathways enriched by genes differentially expressed by the MYD88 mutant B cell population include response to corticosteroid and hypoxia whereas in the MYD88 wild type patient unique processes include B cell activation and receptor signaling pathways, immune response and NFκB signaling.In summary, our results show that there is a distinctive transcriptional profile in the B cells from the MYD88 WT patient where most differentially expressed genes do not overlap with the gene expression of the B cells from the MYD88 L265P patient, which may provide further mechanistic insights into the differences in biology and clinical presentation in patients with MYD88 WT versus those harboring MYD88 L265P. Further evaluation in additional patient samples is ongoing to gain additional insights in pathways that may be involved in disease transformation and response to therapy with targeted therapies, either targeting CD20 or BCR pathway. [Display omitted] DisclosuresGavriatopoulou: Genesis: Honoraria; Sanofi: Honoraria; Karyopharm: Honoraria; Takeda: Honoraria; GSK: Honoraria; Janssen: Honoraria; Amgen: Honoraria. Terpos: Genesis: Consultancy, Honoraria, Research Funding; GSK: Honoraria, Research Funding; Janssen-Cilag: Consultancy, Honoraria, Research Funding; Novartis: Honoraria; Takeda: Consultancy, Honoraria, Research Funding; Sanofi: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; BMS: Honoraria; Amgen: Consultancy, Honoraria, Research Funding. Dimopoulos: BMS: Honoraria; Takeda: Honoraria; Beigene: Honoraria; Janssen: Honoraria; Amgen: Honoraria. Kastritis: Amgen: Consultancy, Honoraria, Research Funding; Genesis Pharma: Honoraria; Takeda: Honoraria; Janssen: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria, Research Funding.

Prognostic Impact of MYD88 L265P Mutation By Droplet Digital PCR in IgM MGUS and Smoldering Waldenström Macroglobulinemia

BACKGROUND: MYD88 L265P mutation is highly prevalent in IgM monoclonal gammopathy of undetermined significance (MGUS), smoldering Waldenström macroglobulinemia (SWM) and symptomatic WM. Allele-specific PCR (AS-PCR) has been used routinely to assess MYD88 mutation; however, with the advent of more precise high-throughput technologies such as droplet digital PCR (ddPCR), absolute quantification can be achieved. There is no data regarding ddPCR applicability in asymptomatic IgM monoclonal gammopathies or as a prognostic biomarker. Here, we aimed to compare MYD88 quantification by ddPCR with clinical and laboratory features and to analyze the prognostic impact in a series of patients (pts) with IgM MGUS and SWM.METHODS: We analyzed bone marrow (BM) and peripheral blood (PB) samples stored from pts diagnosed with IgM MGUS and SWM at our institution from 1980 to 2020. DNA extraction methods followed manufacturer instructions (Qiagen) to obtain genomic DNA from unsorted BM samples and cell-free DNA (cfDNA) from PB. MYD88 L265P mutation was quantified by ddPCR using a Bio-Rad commercial assay (HEX-labeled wild-type allele; FAM-labeled mutant allele). We used OCI-Ly3 DLBCL ABC cell line, homozygous for MYD88 L265P, as a positive control. ddPCR was performed following Bio-Rad technical specifications using the QX200 droplet reader. Data was analyzed using QuantaSoft v.1.0 software (Bio-Rad). Absolute quantification of the mutation was expressed as percentage of fractional abundance. For survival analysis, we used a competing risk analysis to evaluate the prognostic impact of MYD88 mutation on progression to symptomatic WM.RESULTS: A total of 217 unsorted samples were analyzed (187 BM and 30 PB). Genomic DNA from unsorted BM samples was extracted from pts diagnosed with IgM MGUS (46%), SWM (44%), and symptomatic WM (10%). cfDNA was obtained from a subgroup of pts with IgM MGUS (52%) and SWM (48%). Median age at diagnosis was 68 (range 61 to 76). AS-PCR could detect the mutation in 22 (31%) pts with IgM MGUS and 49 (75%) with SWM. ddPCR improved precision detection up to 48 (55%) pts with IgM MGUS and 68 (83%) with SWM. All pts with symptomatic WM harbored the MYD88 mutation, as identified by both techniques. Median absolute quantification from BM was 2.3% and 7% for pts with IgM MGUS and SWM, respectively (p<0.001).Pearson correlation coefficients comparing BM MYD88 mutation quantification by ddPCR with serum M-protein size, IgM concentration, BM lymphoplasmacytic infiltration rate and BM CD19+ cells were 0.3, 0.4, 0.6, and 0.9 (p<0.0004), respectively. Similar coefficients were observed in symptomatic WM regarding BM infiltration rate (0.6; p=0.001) and BM CD19+ cells (0.9; p<0.0001). Spearman correlation coefficients comparing cfDNA MYD88 mutation quantification with BM lymphoplasmacytic infiltration rate and BM CD19+ cells were 0.4 and 0.5 (p<0.008), respectively. Agreement regarding MYD88 mutation detection by ddPCR in BM DNA and cfDNA samples was 82% (Cohen kappa index 0.6).With a median overall survival of 13 years in pts with IgM MGUS and SWM, 13% of them progressed to symptomatic WM while 22% died without progression. Cox univariate analysis using continuous values for MYD88 quantification (p=0.004), serum IgM (p<0.001), BM lymphoplasmacytic infiltration (p<0.001), and serum albumin (p=0.04) were significant. X-tile software was used to find the optimal cutoff point of MYD88 quantification as a biomarker. 4.5% was established for pts with IgM MGUS while 25% for SWM. Using the Fine and Gray regression model in a competing risk analysis taking death without progression as a competing event, higher MYD88 mutation burden negatively impacted the risk of progression of IgM MGUS (SHR 4.6; p=0.003) and SWM (SHR 6; p<0.001) (Figure 1).CONCLUSION: Quantification of the MYD88 L265P mutation by ddPCR has higher precision and sensitivity compared to AS-PCR; thus ddPCR could be used as a potential new and useful biomarker. MYD88 tumor burden correlated with well-known laboratory parameters used for diagnosis and risk stratification, whether using genomic DNA from unsorted BM samples or cfDNA. Risk of progression was higher in patients harboring an increased mutant allele burden. This is the first report showing the prognostic impact of MYD88 quantification in a series of patients with asymptomatic IgM gammopathy and long-term follow up. [Display omitted] DisclosuresCibeira: Akcea: Honoraria, Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees. Bladé Creixenti: Janssen, Celgene, Takeda, Amgen and Oncopeptides: Honoraria. Rosinol: Janssen, Celgene, Amgen and Takeda: Honoraria. Fernandez de Larrea: BMS: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Takeda: Honoraria, Research Funding; GSK: Honoraria; Sanofi: Consultancy; Janssen: Consultancy, Honoraria, Research Funding.

Treatment of Relapsed/Refractory Waldenström Macroglobulinemia Patients: Final Clinical and Molecular Results of the Phase II Brb (Bendamustine, Rituximab and Bortezomib) Trial of the Fondazione Italiana Linfomi (FIL)

Treatment of Relapsed/Refractory Waldenström Macroglobulinemia Patients: Final Clinical and Molecular Results of the Phase II Brb (Bendamustine, Rituximab and Bortezomib) Trial of the Fondazione Italiana Linfomi (FIL)

New Anti-Cancer Drug Compounds to Treat Mutated Waldenstrom Myd88 L265P

Waldenstrom Macroglobulinemia (WM) is a non-Hodgkin lymphoma, often associated with production of monoclonal IgM in a large amount. The increased level of IgM leads to the increased level of blood viscosity, potentially causing spontaneous bleeding, headaches, vertigo and could lead to stroke and coma. WM is a rare disease, affecting around 3 cases per million per year in the USA. Different chromosomal abnormalities can be cause of WM, but the most common mutation detected in WM is the mutation L265P on the Myd88 protein, a downstream regulator of TLR4 pathway. While today there are many treatment options to manage WM, including plasmapheresis, monoclonal (rituximab) and immunomodulating drugs (halidomide), cytokines, mTOR inhibitors (everolimus), Burton tyrosine kinase (Ibrutinib), there isn't a single effective enough treatment for WM that are widely used.In this study, MWCL-1 cells (adult WM), expressing the Myd88 L265P mutation, were used to test different drugs to monitor their effect in cell proliferation and apoptosis activities, in comparison to midostaurin (an inhibitor of FLT3), for 24 hours treatment. MWCL-1 cells were cultured in RPMI1640, in presence of 2, 10 and 25 % of fetal bovine serum (FBS). 25 % FBS was tested to be as close as possible with the level of serum present in human blood. Adipose stromal cells (ASC) were used as a reference control, in absence of serum. Compounds A (targeting TLR pathway, through IRAK-4) was used at various concentrations (0.05 to 100 µM), over 24h of treatment, in combination or not with compound B (targeting the DNA). Compound A has a high affinity for IRAK4, downstream kinase of TLR pathway. TLR pathway activation in leukemia, through Myd88 and IRAK4, is involved in the pro-inflammation response but it is also inducing proliferation and cell survival. Compound B is an inhibitor of the poly (ADP-ribose) polymerase (PARP), which are enzymes that are involved in DNA transcription, cell cycle regulation and DNA repair.In the ASC, the caspase activity of compound A was not different from the control, but the number of cells was lower at 50 and 100 µM, indicating that the compound A was delaying the cell growth. The number of MWCL-1 cells decreased only at higher compound A concentration (50 and 100 µM), during a 24h treatment, at 2, 10 and 25 % FBS. When the cells were treated with the compound A, the level of caspase 3/7 activity was elevated only at the highest concentration (50 and 100 µM), which is consistent with the decreased number of cells, at 2, 10 and 25 % FBS. MWCL-1 were more resistant to the anti-cancer effect of the compound A at 25 % FBS, indicating that it could work on patients because 25 % FBS is close to the patient's serum level. Midostaurin, from 0.05 to 100 µM, was reducing the number of cells and increasing the caspase 3/7 activity in the cells, in a dose response manner, at 2, 10 and 25 % FBS. However, data suggest that at the highest midostaurin dose (100 µM), compound A was more efficient than midostaurin (100 µM). Compound B alone, at 10 µM, had no effect on the MWCL-1 cell number and on the caspase 3/7 activity, for the 2, 10 and 25 % FBS. However, a synergistic effect was demonstrated when compound B (10 µM) was combined with compound A at 50 and 100 µM. The combination decreased the number of MWCL-1 cells and increased the caspase activity 3/7, more so than it was compared to the single compound studies. Our data suggest that it would be possible to lower the posology of the compound A when it will be given to the patients, in combination with the compound B.In conclusion, the data showed compound A alone or combined with the compound B decreased the level of proliferation and increased the level of apoptosis of MWCL-1 cells, cultured with 2 to 25 % of FBS. The use of compound A at high concentration to reduce the number of MWCL-1 reflect the difficulties to treat WM patient with the existing approved drugs. Further studies will be necessary to understand more the molecular mechanism affected by the compound A and/or B in the TLR-IRAK4 pathway, and to study their effect in vivo. DisclosuresOliva: Emmaus Lifesciences, Inc.: Current Employment. Villanueva: Emmaus Lifesciences, Inc.: Current Employment. Ochiai: Emmaus Lifesciences, Inc.: Ended employment in the past 24 months. Niihara: Emmaus Lifesciences, Inc.: Current Employment.

High Frequency of CNS Involvement in Transformed Waldenström Macroglobulinemia

INTRODUCTIONCentral nervous system (CNS) relapse is a challenging situation in diffuse large B-cell lymphoma (DLBCL). High CNS-International Prognostic Index (IPI), activated B-cell (ABC) subtype and MYD88 L265P mutation, features often found in transformed Waldenström macroglobulinemia (WM), are associated with a higher risk for developing CNS relapse. This study was aimed to describe CNS involvement in a large cohort of transformed WM.METHODSThis international multicenter retrospective study included patients with a diagnosis of WM and a concurrent or sequential histological diagnosis of DLBCL. CNS disease was diagnosed by detection of DLBCL cells in the cerebrospinal fluid and/or by brain biopsy. Patients with CNS involvement by lymphoplasmacytic cells (Bing-Neel syndrome) were excluded. Of 254 identified patients with a diagnosis of histological transformation (HT) between 1988 and 2020, 19 were excluded due to lack of data on extranodal involvement. The first part of the analysis focused on baseline CNS involvement. Clinicobiological characteristics were compared between groups using Chi-square or Fisher's exact tests or Mann Whitney tests as appropriate. We analyzed CNS recurrence in the second part of the study. Forty-eight additional patients were excluded due to baseline CNS involvement (n = 25), absence of treatment at HT (n = 14) and lack of details on follow-up (n = 9). Cumulative incidence of CNS relapse was analyzed using competing-risk models that accounted for other events like systemic relapse or death from any cause, reporting sub-hazard ratio (SHR).RESULTSBaseline CNS involvement was present in 25 patients (11%) with transformed WM, including 10 (4%) with parenchymal disease, 10 with leptomeningeal, 4 (2%) with both, and 1 with unspecified CNS involvement. Characteristics associated with baseline CNS involvement were performance status 2-4 (P=0.03) and ≥2 extranodal sites (P=0.02). Median survival after HT was 1 year [0.7-2.5], comparable to the one of patients without CNS disease (1.8 year [1.2-2.6], P=0.74). We observed no difference in survival based on isolated CNS involvement (n = 10) compared to CNS and systemic involvement (n = 15) (P=0.94).Twenty-three CNS relapses occurred (12%). The 2-year and 3-year rates of CNS relapse were 9% (95% CI, 6-14) and 11% (95% CI, 7-16) (Figure 1). The median time to relapse in the CNS was 11 months (95% CI, 7-25). Thirteen CNS recurrences (57%) occurred during the first year of follow-up. Seventy percent were isolated CNS relapses. The location was leptomeningeal in 43% of cases, parenchymal in 35%, both in 17%, and unspecified in 4%. According to CNS-IPI risk groups (data available for 20 patients), 9 patients (45%) belonged to the high-risk group, 10 (50%) to the intermediate-risk group and 1 (5%) to the low-risk group. Prior to CNS relapse, 87% of patients had received rituximab, and 39% had received CNS prophylaxis (30% intrathecal chemotherapy, 4% high-dose methotrexate (HD-MTX), and 4% both). After CNS recurrence, 96% of patients received salvage treatment: combination of HD-MTX and HD-cytarabine (48%), HD-MTX alone (30%), or HD-cytarabine alone (9%). Four patients underwent consolidative autologous stem cell transplantation. The median survival after CNS relapse was 5.6 months. Factors associated with 3-year cumulative incidence of CNS recurrence in univariate analysis were involvement of kidney/adrenal glands (HR, 4.4; P=0.01) and MYD88 L265P mutation (P=0.01) (Figure 2A and B). Of note, among 74 patients (over 187, 40%) with data available for MYD88 mutation status, 11 CNS relapses occurred in patients with MYD88 L265P mutation (n = 54, 20%) whereas no relapse were observed in MYD88 WT cohort (n = 20). A trend toward higher risk of CNS relapse for ≥2 extranodal sites (HR, 2.3, 95% CI 0.98-5.3; P=0.06) was observed. Cumulative incidence according to CNS-IPI risk groups (0% in the low-risk, 9% in the intermediate-risk and 14% in the high-risk group) was not statistically significant (P=0.47).CONCLUSIONCNS involvement occurs frequently in transformed WM. Rate of CNS relapse seems similar to DLBCL patients belonged to the CNS-IPI high-risk group. Special attention should be paid to patients with kidney/adrenal involvement and MYD88 L265P mutation. [Display omitted] DisclosuresVos: Sanofi: Membership on an entity's Board of Directors or advisory committees; Celgene: Other: Travel reimbursement. Treon: X4: Research Funding; Janssen: Consultancy, Research Funding; Dana Farber Cancer Institute: Current Employment; BMS: Consultancy, Research Funding; Self: Patents & Royalties: Holder of multiple patents related to testing and treatment of MYD88 and CXCR4 mutated B-cell malignancies; AbbVie: Consultancy, Research Funding; BeiGene: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding. Dimopoulos: Janssen: Honoraria; Beigene: Honoraria; Amgen: Honoraria; BMS: Honoraria; Takeda: Honoraria. Kapoor: Cellectar: Consultancy; Karyopharm: Consultancy; BeiGene: Consultancy; Pharmacyclics: Consultancy; Sanofi: Consultancy; Amgen: Research Funding; Ichnos Sciences: Research Funding; Regeneron Pharmaceuticals: Research Funding; Glaxo SmithKline: Research Funding; Karyopharm: Research Funding; Sanofi: Research Funding; Takeda: Research Funding; AbbVie: Research Funding. Castillo: Abbvie: Consultancy, Research Funding; BeiGene: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding; Janssen: Consultancy; Roche: Consultancy; TG Therapeutics: Research Funding.

The Contribution of CD148, CD180 and CD200 Combination in the Diagnosis of Chronic B-Cell Lymphoproliferative Disorders

Introduction: B-cell immunophenotype could be swiftly assessed by flow cytometry on blood samples or bone marrow aspirate specimens. It provides crucial information later refined with histologic, genetic and molecular features to assert accurate diagnosis of chronic B-cell lymphoproliferative disorders (B-CLPD). Besides Matutes score we identified additional useful markers, i.e. CD148 and CD180 to classify mantle cell lymphoma (MCL) and marginal zone lymphoma (MZL), respectively. Furthermore, CD200 is known to be highly expressed in chronic lymphoid leukemia (CLL) while absent in MCL.Hypothesis: The determination of CD148, CD180 and CD200 expression on B-cells by flow cytometry on blood samples and/or bone marrow aspirates could be a potent tool to accurately identify B-CLPD. We postulated the existence of the following specific expression patterns in B-CLPD: CD148 dim/CD180 dim/CD200 bright for CLL, CD148 dim/CD180 dim/CD200 dim for lymphoplasmocytic lymphoma (LPL), CD148 bright/CD180 dim/CD200 neg/dim for MCL and CD148 dim/CD180 bright/CD200 dim for MZL .Methods: In a prospective study we investigated the expression of CD148/CD180/CD200 on B-cells from 673 patients at the time of B-CLPD diagnosis in our hospital from 2014 to 2020. We analyzed 440 blood and 233 bone marrow aspirate specimens using a BD FACSCanto II flow cytometry instrument. Based solely on CD148/CD180/CD200 specific expression patterns we postulated a diagnosis of CLL, LPL, MCL or MZL. These postulated diagnoses were later confronted to the final diagnoses when all histologic, genetic and molecular features were finalized. Sensitivity, specificity, positive and negative predictive values of the expression profiles were determined. In addition, to investigate the relative importance of these three CD markers we then normalized their mean fluorescence intensities (MFI) and applied several supervised machine learning algorithms including Logistic Regression, Random Forest and Light Gradient Boosting Machine (LightGBM).Results: Out of the 673 clinical samples the CD148/CD180/CD200 expression patterns classified 212 specimens as CLL/SLL (30.8%), 160 as LPL (23.8%), 76 as MCL (11.28%) and 169 as MZL (25%). These diagnosis hypotheses were retrospectively compared to the final diagnoses based on all histologic, genetic and molecular features These diagnosis hypotheses of CLL, LPL, MCL and MZL were consistent with the final diagnosis in 583 out of the 617 corresponding cases (94%) with high positive and negative predictive values. The characteristics of the diagnosis accuracy are detailed in the table below. HCL and FL were not further investigated as their immunophenotype usually do not overlap with those of other B-CLPD.Seventeen out of 617 patients (17/617, 5.3%) did not displayed a clear CD148/CD180/CD200 pattern: 9 LPL, 4 CLL and 4 MZL. In sixteen patients (16/617, 5.0%) the diagnosis hypothesis based on this strategy was not confirmed after completion of the exploration including karyotype, MYD88 L265P mutational status, CCND1 overexpression and pathology explorations.We next investigated the relative importance of these 3 markers. We focused on MFI values of CD148, CD180 and CD200 and three categorical “positive or negative” markers (CD5, CD23, FMC7) that were assembled into a composite marker. After Cox-box normalization of CD148, CD180 and CD200 MFIs, a set of supervised machine learning algorithms including Logistic Regression, Random Forest and Light Gradient Boosting Machine (LightGBM) were applied to the cohort of CLL, LPL, MCL and MZL.We established that the highest diagnosis weights were obtained for CD200 in CLL, CD200 and CD148 in MCL (negatively and positively, respectively), CD180 in MZL. In LPL, CD148, CD180 and CD200 had the highest weights using LightGBM and Random Forest algorithms, while Logistic Regression determined that CD5 and CD23 had the highest (negative) weights.In conclusion, the determination of CD148/CD180/CD200 surface expression patterns by flow cytometry, along with morphology, allowed to assert an accurate diagnosis hypothesis in CLL, MCL, LPL and MZL with high positive and negative predictive values. Machine learning algorithms allowed to measure the relative importance of these markers, that could be of great help in case of discordant expression of the main diagnosis markers. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

The Landscape of Immunoglobulin Heavy Chain Gene Repertoire in Lymphoplasmacytic Lymphoma / Waldenström Macroglobulinemia

Introduction Immunoglobulin heavy-chain variable genes (IGHV) is critical for the defining epitope binding affinityand B cell differentiation. Lymphoplasmacytic lymphoma/Waldenström macroglobulinemia (LPL/WM) is a heterogeneous diseasewhose role of IGHV usage remains unknown. Besides, the clinical relevance of IGHV repertoire for LPL/WM remain largely unexplored. The aim of our study is to explore the IGH repertoire of LPL/WM in by far the largest series, and to evaluate the correlation between IGH rearrangements and genetic aberrations and clinical characteristics of LPL/WM patients.Methods A total of 162 patients with a diagnosis of LPL/WM were included in this study. Polymerase chain reaction (PCR)amplification of IGHV-IGHD-IGHJ was performed on genomic DNA or cDNA samples using the IGH Somatic Hypermutation Assay v2.0 (Invivoscribe, Technologies, San Diego, US). Sequences were aligned to IMGT (http://www.imgt.org/IMGT_vquest/vquest) and IGBLAST (https://www.ncbi.nlm.nih.gov/igblast/) databases. IGH gene repertoires, mutation status, IGHV CDR3 characteristics, genetic aberrations, MYD88 mutation status and clinical characteristics were collected to evaluate the relevance.Results Productive IGHV-D-J rearrangements were obtained in 136 out of 162 patients (84.0%). The IGHV gene repertoire was remarkably biased in LPL/WM. IGHV3-23 (15.4%), IGHV4-34 (10.3%), IGHV3-7 (8.1%), IGHV3-30 (7.4%) and IGHV3-74 (7.4%) were significantly overrepresented in LPL/WM(Figure 1). Among the 134 IGHD data, the most frequent segment was IGHD3-10 (21/134, 15.7%), followed by IGHD6-13 (18/134, 13.4%) (Figure 2). Among the 134 IGHJ data, IGHJ4 segment was selected in more than half of these rearrangements (70/136; 51.5%), followed by IGHJ6 (23/136; 16.9%) and IGHJ5 (21/136; 15.4%) (Figure 3). Most of the cases were mutated (97.0%) using a 98% IGHV germline homology cutoff. IGHV3-30 was associated with long heavy chain CDR3, indicating the specific antigen selection in LPL/WM. Patients with IGHV3-7 were significantly more likely to harbor 6q deletion (p<0.001)(Figure 4) and abnormal karyotype (p=0.004)(Figure 5).The IGHV hypermutation rate in patients with MYD88 L265P mutation was significantly higher than in wild-type patients (7.3% vs5.6%, p=0.009). IGHV3-23 and IGHV3-74 segments were more frequently detected in MYD88 mutated LPL/WM patients (25.7% vs. 4.3%, p=0.025). IGHV3-7 and IGHV4-59 were represented more in MYD88 wildtype patients (30.4% vs. 8.9%，p=0.005). Moreover, Patients with IGHV4 especially IGHV4-34 had higher level of LDH. IGHV4 was a prognostic marker of shorter progression-free-survival (Figure 6).Conclusion LPL/WM appears to be composed ofdifferent subgroups based on the IGHV repertoire. The mutational status and the IGHV CDR3 length indicated the role for antigenselection in LPL/WM development. The presence of IGHV4 genes proved to be a potential risk factor associated with outcome which deserved further study.These results showed for the first time that IGHV repertoire had clinical relevance in LPL/WM. [Display omitted] DisclosuresWang: AbbVie: Consultancy; Astellas Pharma, Inc.: Research Funding.

Ethnic Disparities in Waldenström's Macroglobulinaemia in the United Kingdom - Analysis from the Wmuk Rory Morrison Registry

Introduction: Waldenström's Macroglobulinaemia (WM) is a rare indolent B-cell lymphoma characterised by lymphoplasmacytic infiltrate with an IgM paraproteinaemia. Approximately 400 patients (pts) are diagnosed annually in the United Kingdom (UK). National and regional registries provide important insights on the epidemiology and clinical features of this distinct uncommon disorder. Previous authors from the United States have reported significant differences in age at diagnosis and overall survival (OS) in WM between ethnic groups from Surveillance, Epidemiology and End Results data [1]. We aimed to determine baseline demographics, symptoms and survival outcomes across different ethnic groups in the UK.Methods: We retrospectively reviewed data from the WMUK Rory Morrison Registry, collating descriptive data of WM as well as non-IgM Lymphoplasmacytic lymphoma (LPL) cases from 20 centres across the UK, diagnosed between June 1978 and May 2021. Research ethics approval was obtained.Results: 732 pts with documented ethnicity were included here. Ethnicity was categorised in accordance with the UK National Census. Those other than the White cohort (662/732; 90%) were collectively termed ‘Ethnic Minorities’ (EM) (70/732; 9%): 39/70 (56%) Asian (13 Indian, 6 Chinese, 4 Pakistani, 16 other); 19/70 (27%) Mixed (1 White & Black African, 1 White & Asian, 1 Arab, 16 other); 12/70 (17%) Black (6 Caribbean, 3 African, 3 other).Baseline characteristics are displayed in table 1. EM presented at a significantly younger age compared to White cohort (60 vs 64 years, p=0.01) and had a significantly lower paraprotein at diagnosis (11 vs 18g/L, p=0.05). MYD88 L265P mutation tested in 250 pts (34%): positive in the majority 218/250 (87%) and notably less common in EM (p=0.09), most significantly in the Asian cohort compared to White (67% v 88%, p=0.01). The EM cohort had a smaller proportion of WM diagnoses compared to White cohort (77% vs 93%, p=0.02) with the Asian cohort accounting for the highest proportion of non-IgM LPL (15% vs 4%, p=0.002). Bone marrow trephine infiltration at diagnosis was lower in the EM cohort compared to the White cohort, although not statistically significant (25% vs 37%, respectively; p=0.10). IPSSWM scores were similar throughout.Approximately half of both the White & EM cohorts were asymptomatic at diagnosis. The most common presenting symptoms in EM were anaemia-related (10/34; 29%), B symptoms (6/34; 18%) and fatigue (4/34; 12%). 6/18 (33%) of Asians presented with cryoglobulinaemia, amyloid, nephropathy or acquired von Willebrand's Disease.Of EM, 51/70 (73%) had treatment, 35/51 (69%) within the first year from diagnosis with time to treatment 2 months (range 0-364). Time to next therapy was 14 months. Median lines of therapy were 2 (range 1-8). The most common indication for first line therapy was lymphoma-related (16/39; 41%) followed by paraprotein-related (peripheral neuropathy, hyperviscosity and autoimmune) (13/39; 33%). First line therapies included Bendamustine-Rituximab (BR; 8/49; 16%), Dexamethasone Rituximab Cyclophosphamide (DRC; 7/49; 14%) and R-CHOP (5/49; 10%) with smaller proportions receiving Rituximab monotherapy, Cladribine + Rituximab or Fludarabine + Cyclophosphamide + Rituximab (FCR) (3/49 (6.1% each). Two of 41 (5%) of EM had first treatment as a part of a clinical trial, compared to 16/156 (10%) of the White cohort (p=0.28). Response to first line therapy was similar between the White & EM cohorts (Table 1).For EM, at a median follow up of 7 years, 17 died (1/17; 6% disease related). Median OS was not reached (Figure 1). Estimated 5-year and 10- year OS was 91% (95% CI 96-80%) and 75% (95% CI 85-60%), respectively.Conclusions: This first report on ethnic disparities in WM and non-IgM LPL in the UK identified key differences in presentation of disease. Ethnic minorities present at a younger age, with a lower paraprotein. The Asian cohort had a greater proportion of non-IgM LPL cases and fewer MYD88 mutated cases. A small proportion of Ethnic Minorities have treatment on a clinical trial which may warrant further attention. Further analysis including associations with socioeconomic status, deprivation indices and comorbidities are ongoing.•Ailawadhi, Sikander et al. “Outcome disparities among ethnic subgroups of Waldenström's macroglobulinemia: a population-based study.” Oncology vol. 86,5-6 (2014): 253-62. doi:10.1159/000360992 DisclosuresMcCarthy: Janssen: Honoraria; Astra zenica pharmaceuticals: Honoraria; Amgen: Honoraria. Pratt: Amgen: Consultancy; Binding Site: Consultancy; BMS/Celgene: Consultancy; Gilead: Consultancy; Janssen: Consultancy; Takeda: Consultancy. El-Sharkawi: AbbVie: Honoraria, Other: Travel Support, Ad boards; AstraZeneca: Honoraria, Other: Ad boards; Janssen: Honoraria, Other: Ad boards; Roche: Honoraria; Takeda: Honoraria; Novartis: Other: Travel Support; ASTEX: Other: Ad boards; Beigene: Other: Ad boards; Kyowa Kirin: Other: Ad boards. Linton: Genmab: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Aptitude Health: Honoraria; Hartley Taylor: Honoraria; Celgene: Research Funding; BeiGene: Research Funding; University of Manchester: Current Employment. Gatto: Roche: Consultancy. D'Sa: Janssen Cilag: Honoraria, Research Funding; BeiGene: Honoraria, Research Funding; Sanofi: Honoraria.

Efficacy and Safety of Orelabrutinib in Relapsed/Refractory Waldenstrom's Macroglobulinemia Patients

BackgroundWaldenstrom's Macroglobulinemia (WM) is a B-cell disorder characterized primarily by bone marrow infiltration with lymphoplasmacytic cells, along with immunoglobulin M (IgM) monoclonal gammopathy. Bruton's tyrosine kinase (BTK) plays a key role in signaling pathways for the survival of WM clone, particular in patients harboring MYD88 L265P mutations. However, due to target selectivity issue, Clinical uses of early BTK inhibitors are still compromised with off-target activities to many other kinases besides BTK. Orelabrutinib is a novel, highly potent small molecule inhibitor of BTK with superior selectivity for B-cell malignancies and autoimmune diseases. Preliminary efficacy and safety data of ICP-CL-00105 in relapsed/refractory WM patients are presented here.MethodsICP-CL-00105 is a single arm, multiple centers, open label, phase 2 study in clinical and histopathological confirmed patients with R/R WM requiring treatment per IWWM-7. MYD88 and CXCR4 mutations were assessed in bone marrow samples at baseline. Orelabrutinib at a daily dose of 150mg was administered orally until disease progression or unacceptable toxicity. Blood samples for IgM were assessed at baseline and every cycle for 6 cycles and every 3 cycles thereafter by central lab. Responses were assessed in accordance with IWWM-6 and NCCN guidelines. The primary endpoint was major response rate (MRR) as assessed by IRC. Key secondary endpoints were MRR as assessed by investigator, overall response rate (ORR), duration of major response (DOMR), progression-free survival (PFS), OS, changes in IgM levels from baseline, improvement on hemoglobin levels and safety. Treatment-emergent adverse events (TEAEs) and treatment-related adverse events (TRAEs) were assessed according to NCI CTCAE v4.03.ResultsAs of June 1, 2021, for the 47 patients the median follow-up duration was 10.5 months. The median age was 63 years (range, 56-68 years), 40 patients (85.1%) were male. 87.2% of patients were at intermediate or high risk according to the Prognostic Scores (IPSS). The proportion of patients with MYD88 L265PCXCR4 wildtype was 83% at baseline. With a median duration of treatment of 9.2 months, MRR was 74.5% as assessed by investigator. ORR was 87.2% with 97.9% patients achieved disease control. The estimated 12-month DOMR were 89.5%. The estimated 12-month PFS and OS were 88.0% and 92.3%, respectively. The median PFS and median OS have not been reached. The MRR was higher in patients with MYD88 L265PCXCR4 wildtype (79.5%). The median IgM level was 30.3g/L at baseline. The decline in the serum IgM levels from baseline were observed with a median reduction by 79.0% (IQR: -89.4, -57.2). The median hemoglobin level at baseline was 102g/L. Durable improvement in hemoglobin levels was found in 83% of patients with a median maximal improvement of 40g/L (IQR: 24.0, 62.0).Safety data were summarized by the cutoff date of June 1, 2021. The most commonly reported AEs were thrombocytopenia (27.7%)，neutropenia (14.9%), leukopenia (10.6%), upper respiratory infection (14.9%)，weight increased (14.9%), influenza-like disease (12.8%) and rash (10.6%). Most reported AEs (89.5%) were grade 1-2. 16 patients (34.0%) reported grade ≥ 3 TEAE while 9 patients (19.1%) reported grade ≥ 3 TRAE. There was no reported grade ≥3 atrial fibrillation and/or atrial flutter, or grade ≥3 diarrhea. Only One TRAE (2.1%) resulted in drug discontinuation.ConclusionOrelabrutinib has demonstrated substantial efficacy in treating r/r WM patients under short-term follow-up. It has shown favorable safety and tolerability profile with limited off-target adverse effects. It has the potential to be a promising treatment option for r/r WM patients. DisclosuresHu: Astellas Pharma, Inc.: Research Funding. Tian: Innocare pharma: Current Employment. Zhu: Innocare pharma: Current Employment. Zhang: Innocare pharma: Current Employment. Zhao: Innocare pharma: Current Employment. Zhang: Innocare pharma: Current Employment.

REFRACTORY CHYLOTHORAX SECONDARY TO WALDENSTROM MACROGLOBULINEMIA

TOPIC: Pulmonary Manifestations of Systemic Disease TYPE: Medical Student/Resident Case Reports INTRODUCTION: A chylothorax is defined by the presence of chyle within the pleural cavity. Chylothoraces can be observed as a complication of malignancy; however, Waldenstrom Macroglobulinemia (WM) has rarely been described as a potential cause. We present a case of a refractory chylothorax secondary to WM presenting as a periaortic mass, which eventually required thoracic duct embolization (TDE). CASE PRESENTATION: A 76-year-old male former heavy smoker underwent a low-dose chest CT that showed a large right-sided pleural effusion with suggestion of subcarinal lymphadenopathy. The patient underwent a right-sided thoracentesis, and pleural fluid appeared milky-orange (Figure 1) with triglyceride concentration of 410 mg/dL, diagnostic of a chylothorax. Contrast chest CT revealed a soft tissue mass encasing the descending aorta (Figure 2). Tissue sampling via endobronchial ultrasound (EBUS) demonstrated a monomorphic population of small lymphocytes expressing CD19, CD20, and CD22, while being negative for CD5 and CD10. Additional oncological evaluation including the detection of a MYD88 L265P mutation led to a diagnosis of WM. The patient was started on combination chemoimmunotherapy and had notable tumor response; however, he continued to require high-volume therapeutic thoracentesis (2.5 liters per week). The patient was referred to interventional radiology and underwent successful TDE without complications. Within weeks of thoracic duct embolization, the patient's milky pleural effusion decreased significantly in amount and transitioned to a serosanguinous appearance with a triglyceride concentration of 60 mg/dL. The patient had significant dyspnea relief with decreased frequency of thoracentesis leading to an improved quality-of-life. DISCUSSION: Lymphoma is the most common malignancy-associated cause of a chylothorax. WM is rarely described as a cause in the literature, especially in the setting of a periaortic mass. Malignancy-associated chylothoraces commonly improve with chemotherapy; however, this patient continued to require therapeutic thoracentesis after months of effective chemotherapy. Pedal lymphangiography identified a lymphatic leak within the chest, and TDE was successfully performed without complications. CONCLUSIONS: In patients with a non-traumatic chylothorax, malignancy should be high on the differential diagnosis. Refractory chylothorax can cause serious decrement in quality-of-life due to chronic dyspnea, malnutrition, and recurrent pleural procedures. Early referral for TDE should be considered by chest physicians in patients with refractory chylothorax. REFERENCE #1: Rudrappa M, Paul M. Chylothorax. In: StatPearls. Treasure Island: StatPearls Publishing; 2021 Jan. REFERENCE #2: Otoupalova E, Meka SG, Dogra S, Dalal B. Recurrent chylothorax: a clinical mystery. BMJ Case Rep. 2017;2017:bcr2017220750. Published 2017 Oct 6. doi:10.1136/bcr-2017-220750 DISCLOSURES: No relevant relationships by Ali Bukhari, source=Web Response No relevant relationships by Matthew Koroscil, source=Web Response No relevant relationships by Nathan Mullen, source=Web Response

Tre-P-28 - Primary cutaneous B-cell lymphoma leg-type related to an anti-TNF-alpha therapy

<h3>Introduction:</h3> Drug-induced primary cutaneous B-cell lymphomas are rare. <h3>Observation:</h3> A 63 year old woman was referred for a lumbar cutaneous lesion which had been rapidly growing for 1 month. Her past medical history included multiple allergies and a rheumatoid arthritis which was treated with oral methotrexate 10 mg per week and adalimumab. Methotrexate had been taken for 8 years, while adalimumab had been initiated 6 months earlier. Physical examination revealed a solitary thick, violaceous plaque of 6 X 4 cm in diameters, without enlarged lymph nodes. No fatigue, fever or pruritus were present. The first clinical hypothesis was a cutaneous B-cell lymphoma. The histological examination of a skin biopsy specimen showed a monomorphous dermal proliferation of large centroblast-like cells with large round, sometimes irregular, nuclei. Only rare reactive small cells were present. The infiltrate spared the epidermis, with a Grenz zone. The tumor cells were positive for CD45, CD20, bcl2, bcl6, MUM1, highly proliferating (Ki67 95%), and negative for CD3, CD30, CD10 and EBV/EBER. No CD21 or CD23 positive follicular dendritic cell network was identified. Fluorescence in situ hybridization showed no BCL2, BCL6 or CMYC rearrangement, while high resolution melting identified the MYD88 L265P mutation. A B-cell rearrangement was detected in the lesion. Whole body FDG-PET/CT showed a single hypermetabolic activity corresponding to the cutaneous lombar lesion, without distant tumours. Although the Borrelia burgdorferi serology was negative, a 15 days course of doxycyline 200 mg daily was given. Once the diagnosis of primary cutaneous large B-cell lymphoma, leg type (PCLBCL-LT) was made, adamimumab (but not methotrexate) was discontinued. The tumor completely disappeared within 4 months. No other TNF inhibitors were later given. No recurrence was observed after a follow-up time of 4 years. <h3>Discussion:</h3> PCDLBCL-LT are aggressive B-cell lymphomas mainly (but not exclusively) located on the leg, characterized by a monomorphous proliferation of large CD20+, Bcl2+, Mum1+, CD10 - B cells, frequently exhibiting the MYD88 L265P mutation, as observed in our patient. In contrast with indolent subtypes of primary cutaneous B-cell lymphomas, spontaneous regressions in PCLBCL-LT are exceptional, and constantly followed by recurrences. Progression of undiagnosed cutaneous lymphomas after anti-TNF-alpha therapy (mainly cutaneous T-cell lymphomas misdiagnosed as psoriasis or eczema) have been reported [1]. However, the role of anti-TNF-alpha in the occurrence of new lymphoproliferative disorders remains controversial [2,3]. To our knowledge, no PCLBCL-LT related to TNF-alpha inhibitors has been reported to date. Although methotrexate may play a role as a cofactor for the development of this B-cell malignancy, the negative EBV staining and the long-term complete remission after adalimumab withdrawal while methotrexate was continued, strongly argue for a predominant role of the anti-TNF-alpha in the occurrence of this lymphoma.