Double Hit Research Articles

First Report on a Successful Screening Program for MYC Rearrangements and a Prospective Clinical Trial Based on MYC Rearrangement in Newly Diagnosed DLBCL Patients in the Netherlands

▪IntroductionPatients with aggressive B-cell lymphoma that harbor a MYC rearrangement (WHO 2008: Diffuse Large B-cell Lymphoma (DLBCL) or B-cell lymphoma with features intermediate between DLBCL and Burkitt lymphoma (BCL-U)) have a dismal prognosis following standard first-line treatment. Intensifying chemotherapy prolongs disease free survival when compared to R-CHOP. Despite this, relapses often occur and overall survival has not been improved, hence justifying trials with MYC targeting drugs.Lenalidomide has demonstrated to down-regulate MYC and its target genes in MYC rearranged B-cells and therefor potentially is an effective MYC targeting compound. Although lenalidomide seems mainly effective in ABC subtype DLBCL, the ability of MYC downregulation provides the rationale of lenalidomide administration in MYC rearranged lymphoma, that are usually of GCB subtype. The addition of lenalidomide to R-CHOP has already been demonstrated to be safe and well tolerable. Until now, no prospective studies for MYC rearranged DLBCL patients have been conducted. This is at least partially due to lack of adequate screening facilities for MYC rearrangements.To this end we initiated a registration and screening program for all newly diagnosed DLBCL patients in which performing MYC FISH was advocated and financially supported (HOVON 900). Patients with a proven MYC rearrangement were subsequently included in a clinical trial (HOVON 130: A phase II study evaluating the effect of the addition of lenalidomide to R-CHOP for patients with newly diagnosed MYC positive DLBCL and BCL-U (EudraCT 2014-002654-39)). Here we present data from the screening program and demographic data of the first 64 eligible patients included in the prospective clinical trial.Material, methods and preliminary resultsDuring the screening period for MYC rearrangements, administration of one cycle of R-CHOP21 is allowed. After being diagnosed with a MYC rearrangement, lenalidomide 15 mg on day 1-15 of R-CHOP21 is added. In total patients receive 6 cycles of R-CHOP21 plus 6 cycles of lenalidomide plus 2 cycles of rituximab monotherapy. Moreover, as these patients have a high risk of CNS relapse, intrathecal prophylaxis is given to all patients . Primary endpoint of the study is centrally reviewed end-of-treatment complete metabolic remission rate on PET-CT scan.Screening database (HOVON 900): Between August 1st 2015 and July 17th 2017, 581 newly diagnosed DLBCL patients from 62 centers covering all regions of the Netherlands have been registered. Median age at diagnosis was 65 years (range 19 to 82 years). Male: female ratio 62%: 38%. MYC FISH analysis was successful in 73% of patients. Clinical trial (HOVON 130): In total 67 patients were diagnosed with a MYC rearrangement and were included. After central pathology review, 3 patients were declared ineligible (2 transformed follicular lymphoma, 1 MYC rearrangement not confirmed). Additional FISH analysis on BCL2 and BCL6 rearrangements revealed that 27% of the cases were MYC single hit, 59% were double hit (MYC and BCL2 or BCL6) and 14% were triple hit (MYC and BCL2 and BCL6) DLBCL patients (analysis of 51 patients, to be completed).Clinical baseline characteristics of the 64 eligible patients are shown in table 1. A planned interim analysis, after 27 eligible patients finished treatment, has been performed. Safety results showed toxicities grade 2 in 41%, grade 3 in 33% and grade 4 in 15% of patients. Most common toxicities were polyneuropathy grade 2 or 3 (44%), infections grade 2 or 3 (33%) and gastrointestinal disorders grade 2 or 3 (30%). 16 severe adverse events were reported (all hospitalizations, including 5 infections). Based on these results, the DSMB concluded that no safety issues prohibit continuation of the trial. Efficacy results are blinded until the inclusion will be completed (expected Q4 2017). Updated demographic results will be available in December 2017.ConclusionA successful screening program for MYC rearrangements for all newly diagnosed DLBCL patients is initiated in the Netherlands. This program, which allows start of treatment during the screening period, enables newly diagnosed DLBCL patients with a proven MYC rearrangement to switch to a prospective biomarker-based clinical trial that evaluates the addition of lenalidomide to standard R-CHOP. [Display omitted] DisclosuresChamuleau:Genmab: Research Funding; Gilead: Research Funding. Nijland:Novartis Pharmaeuticals Corporation: Honoraria; Kite Pharma: Honoraria; Roche: Honoraria, Research Funding; Celgene: Honoraria, Research Funding; MIllennium/Takeda: Honoraria, Research Funding; Mundipharma: Honoraria; Gilead Sciences: Honoraria; BMS: Honoraria; MSD: Honoraria; Amgen: Honoraria. Kersten:MSD: Honoraria; BMS: Honoraria; Gilead Sciences: Honoraria; Mundipharma: Honoraria; Milennium/Takeda: Honoraria, Research Funding; Celgene: Honoraria, Research Funding; Roche: Honoraria, Research Funding; Kite Pharma: Honoraria; Amgen: Honoraria; Novartis Pharmaceuticals Corporation: Honoraria. Jong:BMS: Consultancy, Honoraria; Takeda: Consultancy, Honoraria; Roche: Consultancy; Celgene: Consultancy, Research Funding; Genmab: Research Funding.

Gemcitabine/Clofarabine/Busulfan: A New Myeloablative Reduced-Toxicity Conditioning Regimen with High Activity for Allogeneic Stem-Cell Transplant (alloSCT) in Aggressive Lymphomas

INTRODUCTION:Stem-cell transplantation (SCT) is standard treatment of relapsed lymphomas. Patients in refractory relapse or relapsed after autoSCT are often considered for an alloSCT, aiming at a graft-vs.-lymphoma (GVL) effect. While reduced intensity conditioning regimens have substantially contributed safety to alloSCT, aggressive lymphomas often progress rapidly afterwards, partly due to tumor cell growth outpacing GVL. Outcomes are particularly poor for patients with active disease at the time of alloSCT. Thus, development of more efficacious conditioning regimens for aggressive lymphomas, avoiding mucotoxic drugs (such as melphalan), is a major need. We previously showed that the combination of 2 nucleoside analogues [gemcitabine (GEM) + clofarabine (CLO)] followed by busulfan (BU) is highly synergistic against B- and T-cell lymphoma cell lines, through inhibition of DNA damage repair. Those observations led us to clinically study a novel high-dose regimen of GEM/CLO/BU with alloSCT for aggressive lymphomas.METHODS:Eligibility included age 12-65; refractory DLBCL, T-cell and Hodgkin lymphomas; acceptable end-organ function, and an 8/8 HLA-matched (A, B, C and DRB1) sibling (MSD) or unrelated donor (MUD). GEM was escalated from dose level 1 to 3; it was given on days -6 (d-6) and d-4 as a loading dose of 75 mg/m2 followed by infusion at 10 mg/m2/min over 40-90 min (total daily dose 475, 675 or 975 mg/m2), to be followed by CLO (40 mg/m2/day x 4, d-6 to d-3) and BU (pharmacokinetically targeted to an average daily AUC of 4,000 μMol•min x 4, d-6 to d-3). AlloSCT was on d0. Patients receiving a MUD transplant received rabbit ATG (0.5, 1.5 and 2 mg/Kg on d-3, d-2, and d-1, respectively). Patients with CD20+ tumors received rituximab (375 mg/m2) on d-14, d-7, d+1 and d+8. The GVHD prophylaxis included tacrolimus (tapered after d+100) and mycophenolate mofetil (from d0 to d+60). Dose limiting toxicities (DLT) were defined as any G4-5 nonhematological organ toxicity, or as G3 skin or G3 mucositis lasting >3 days at peak severity. Dose escalation of gemcitabine followed a Bayesian design targeting a maximal DLT probability of 25%. Non-relapse mortality (NRM), relapse incidence, G II-IV or III-IV acute GVHD or chronic GVHD were calculated with the cumulative incidence method with consideration of competing risks.RESULTS:Between 11/12 and 06/17, 48 patients were enrolled: 25 DLBCL (5 double hit), 13 T-NHL and 10 HL (Table); 27 MSD, 20 MUD.All patients received PBPC. The MTD of gemcitabine was level 3. DLT was observed at the following frequencies: level 1: 12.5% (1/8), level 2: 9% (2/9), and level 3: 25% (4/16). Regimen-related toxicities were manageable, including mucositis (47% G2, 42% G3), self-limited transaminitis (21% G2, 49% G3), hyperbilirubinemia (no cases of VOD) (30% G2, 19% G3), and dermatitis (13% G2). There were no cardiac, pulmonary, renal or CNS toxicities. Neutrophils dropped to <0.1 in all patients on median d0 (d-3 to +2), and engrafted on median d+10 (d+9 to +20). Platelets engrafted on median d+12 (d+8 to +38).All patients engrafted; full donor chimerism was established early: a median of 100% (50-100%) on d+28, and 100% (98%-100%) on d+100. The rates of G II-IV and III-IV acute GVHD were 29.8% (35% MSD, 25% MUD) and 10.6% (19% MSD, 0% MUD), respectively. The rates of moderate-severe / severe chronic GVHD were 8.5% / 6.4%. 1/3 HL pts with prior exposure to anti-PD1 developed steroid-responsive hepatic chronic GVHD.The γ-H2AX levels in PBMNC (N=19) increased throughout treatment, to a median of 8.5 fold from baseline (range, 0.4-46.3) (P=0.0001) and PARP-1 decreased to 0.45 fold (0.2-6.3) (P=0.4), indicative of activation of DNA damage response and apoptosis, respectively. The overall response rates on d+30 (in 34 patients with measurable disease) are 77% (DLBCL), 100% (T-NHL), and 75% (HL). Their respective CR rates are 69%, 100% and 75%. The d+100 / 1-year NRM rates were 6.25% / 14.5%. NRM causes were infection (6) and GVHD (1). At median follow-up of 18 months (1-53) the EFS / OS rates of the entire population are 44% / 56%. Outcomes appear especially promising in the DLBCL (EFS / OS, 54% / 54%) and the T-NHL (62% / 69%) subgroups.CONCLUSIONS:GEM/CLO/BU is a new, myeloablative, reduced-toxicity alloSCT regimen with high activity in this population of heavily pretreated and refractory aggressive lymphomas, which warrants further study in disease-specific trials [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Pre-Clinical Development of Targeted Therapies for Double Hit (DH) Diffuse Large B-Cell Lymphoma (DLBCL)

Background: Gene expression profile classifies DLBCL into three subtypes with distinct pathogenesis and clinical outcomes: activated B-cell (ABC), germinal center B-cell (GCB) and primary mediastinal B-cell lymphoma (PMBL). DH-DLBCL is a distinct subgroup, with a poor clinical outcome, harboring concurrent gene translocations involving the C-MYC and either/both BCL2 or BCL6 proto-oncogenes. DH-DLBCL has inferior response rates to rituximab-based chemotherapy, and a shorter progression-free survival (PFS)/overall survival (OS). In addition, pts with DH-DLBCL who are refractory to frontline chemoimmunotherapy fare dismally; a retrospective review at our institute yielded survival rates of zero and median OS of 6 months in these patients, highlighting the need for novel therapies. DH-DLBCL is characterized by de-regulation of apoptosis and cell cycle progression, leading to rapid proliferation of neoplastic cells, and resistance to apoptotic stimuli. Upregulation of the anti-apoptotic protein Mcl-1 is implicated in chemotherapy resistance and poor clinical outcomes in ABC-DLBCL. Transcriptional regulation of MCL1 by c-MYC might further potentiate Mcl-1 induced resistance to chemotherapy in DH-DLBCL. Specific inhibitors targeting cell pro-survival proteins might provide a novel and alternative therapeutic approach in rituximab-chemotherapy resistant DH-DLBCL. We hypothesize that dual inhibition of anti-apoptotic proteins Bcl-2 and Mcl-1 using readily available agents (Venetoclax and Proteasome inhibitors) is an effective strategy in inducing lymphoma cell death. Materials and Methods: We conducted pre-clinical in vitro and in vivo models targeting Bcl-2 with Venetoclax and Mcl-1 with either a c-Myc inhibitor (JQ-1) or Proteasome inhibitor (Carfilzomib) in DHL. Val, DOHH-2, and ROS-50 cells were exposed to Venetoclax (0-10 uM), JQ-1 (0-100 uM) and Carfilzomib (CFZ) (0-50 nM) for 24, 48 and 72 hours. IC50 drug concentrations were calculated for each agent. Subsequently, DHL cells were exposed to Venetoclax in combination with JQ-1 or CFZ for 48 hours. Cell viability was determined by Cell Titerglo. Coefficient of synergy was calculated using CalcuSyn. In addition, induction of apoptosis was evaluated by flow cytometry (Annexin V/PI staining) and changes in Bcl-2 family members and Caspase activation were determined by Western blotting. Primary tumor cells isolated from B-cell lymphoma patients (N= 15) including DHL were exposed ex vivo to Venetoclax +/- Carfilzomib. For in vivo experiments 6-8 weeks old severe combined immunodeficiency (SCID) mice were inoculated with 10x106 ROS-50 cells via tail vein injection (IV). Subsequently, animals were treated with Venetoclax (100mg/kg/dose via gastric lavage on days 3-7 and 10-14); Carfilzomib (2mg/kg/dose IV days 3, 4, 10, 11), or both agents. A group of untreated animals was used as a control. Differences in survival were evaluated between treatment groups. Results: In vitro, Venetoclax, JQ-1, and CFZ induced cell death in a dose- and time-dependent manner. Significant synergistic activity was observed by combining both Venetoclax and CFZ and to a lesser degree with JQ-1. Similar findings were observed in primary tumor cells isolated from B-cell lymphoma patients. Annexin/PI staining with the combination of Venetoclax and CFZ resulted in caspase dependent apoptosis. Western blotting confirmed changes in Bcl-2 family members favoring an apoptotic state as well as PARP cleavage following in vitro exposure to Venetoclax and CFZ. In vivo the combination of Venetoclax with Carfilzomib resulted in significant increased survival increase (P<0.05) when compared to single agent Venetoclax or CFZ. Conclusion: Our data suggests that Venetoclax exhibits strong synergistic activity with Carfilzomib in vitro and in vivo . Perhaps related to the synergy observed between Venetoclax and CFZ, is the effects of proteasome inhibition on Mcl-1 levels. Combination therapy targeting the proteasome inhibition and BCL-2 may provide an alternative therapeutic approach in DH-DLBCL. DisclosuresNo relevant conflicts of interest to declare.

Comparison of the Efficacy and Feasibility of R-CODOX-M/IVAC-R Versus Conventional R-CHOP As First Line Regimen for High Risk Diffuse Large B-Cell Lymphoma

BACKGROUND AND AIMSDiffuse Large B Cell Lymphoma (DLBCL) is an heterogeneous group of diseases whose behavior can be predicted by clinical risk scores, immuno-histochemistry and cytogenetic. Among DLBCL, patient with high risk features such as high IPI score, high mitotic index, double hit lymphomas display a worse outcome and the standard frontline treatment with R-CHOP usually results in poor outcome. Consequently, many groups have explored more intensified first line regimens for those patients, such as R-DA-EPOCH and R-CODOX-M/IVAC-R.However, no study directly comparing conventional R-CHOP with more aggressive regimens are currently available, so that a clear indication of superior efficacy from an intensified approach has not been provided yet.The aim of the present was to retrospectively compare through a match-paired analysis two cohorts of patients affected by aggressive DLBCL who were treated with either conventional R-CHOP (center 1) or intensified R-CODOX-M/IVAC-R (center 2) as frontline regimen, in order to disclose to identify subgroups of patients who are more likely to benefit from intensive first line approach.PATIENTS AND METHODSHigh Risk DLBCL was defined by at least one among these features: high tumour burden, DPLs, IPI score >3 or the presence of at least 1 extranodal site.In center 1, 6 x R-CHOP is standard therapy for every patient with newly diagnosed DLBCL, whereas in center 2 patients with aggressive DLBCL are treated with R-CODOX/IVAC-R repeated for two courses.Fifty-one patients affected by aggressive DLBCL and treated with R-CHOP or R-CODOX-M/IVAC-R between January 2011 and January 2015 in the two centers were retrospectively included in a match-paired analysis. Median follow up was 36 months.Matching variables were age, IPI score, disease staging according to Ann Arbor, cell of origin according to Hans algorithm. Matching ratio was 2:1 (2 patients treated with conventional R-CHOP were matched with each patient treated with R-CODOX-M/IVAC-R).The two groups of patients had no significant differences in baseline characteristics such as age, stage, IPI and cell of origin.To disclose differences in terms of efficacy, overall response rate, as defined by international standard, overall survival and cumulative incidence of relapse were compared between the two cohorts.RESULTSOverall response rate (ORR) was 44/51 (86.3%). Patient treated with R-CODOX-M/IVAC-R had an higher probability to achieve at least a partial response (17/17, 100%, vs 27/34, 79.4%, p<0.05, respectively for R-CODOX-M/IVAC-R and R-CHOP).Specifically, after a subgroup analysis, patients with age <50 yrs (p<0.05) and stage III/IV (p<0.05) were found to have a significant improvement in ORR if treated with R-CODOX-M/IVAC-R. A trend towards improved chance of response with R-CODOX-M/IVAC-R, however without reaching statistical significance was observed among patients with germinal center DLBCL, according to Hans algorithm.Overall survival (OS) at 3 years was 88.2% (median of reached) and was not significantly different among patients treated with R-CHOP (3 year OS=86.9%) or R-CODOX-M/IVAC-R (3-year OS=93.2%). None of the other analyzed variables significantly impacted OS probability.After subgroup analysis, a trend for improved OS if treated with R-CODOX-M/IVAC-R was found among patients with IPI score >3, age <50 and germinal center DLBLC (Figure 1), again without reaching statistical significance.Patient treated with R-CODOX-M/IVAC-R had a significantly higher probability of experiencing grade III/IV hematological toxicities (p<0.05), grade III/IV mucositis (p<0.05) or grade III/IV infections (p<0.05), whereas cardiovascular and hepatic toxicities were unlikely and non significantly different among the two cohorts.CONCLUSIONSR-CODOX-M/IVAC is an highly effective regimen for patients affected by DLBCL, with an ORR significantly higher than conventional R-CHOP, however at the price of significantly higher therapy-related toxicities. Our results seem to suggest a potential benefit for younger patients with very aggressive DLBCL, where higher risk of toxicities is overcome by the improved probability of cure.Of note, patient with germinal center DLBCL seem to take a greater benefit from the intensified approach, this may be explained by the higher chemo-sensitivity that is usually observed in this setting, however our result must be confirmed in larger trials. [Display omitted] DisclosuresBocchia:Jansen: Other: Travel grant; Novartis: Other: Travel grant; Celgene: Other: Travel grant; Roche: Other: Travel grant.

Lenalidomide and Obinutuzumab with CHOP for Newly Diagnosed Diffuse Large B-Cell Lymphoma: Final Phase I/II Results

▪Introduction:Diffuse large B-cell Lymphoma (DLBCL) is the most common lymphoid cancer in the United States with 30K diagnoses each year. The standard therapy for all newly diagnosed patients is rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (RCHOP) which cures ~60% of patients. Lenalidomide (L) and obinutuzumab (O) have both shown promising efficacy and acceptable toxicity in DLBCL patients. Obinutuzumab, a humanized CD20 monoclonal antibody with increased pre-clinical affinity to the FcGRIIIa receptor in comparison to rituximab, may have increased antibody dependent cell-mediated cytotoxicity (ADCC). Lenalidomide, a potent immunomodulatory, is toxic to DLBCL via interferon signaling stimulation. As single agents, O and L achieve overall response rates (ORR) of 32% and 28%, respectively, in patients with relapsed DLBCL (Morschhauser, 2013, Hernandez-Ilizaliturri, 2011). The addition of L to RCHOP may overcome adverse outcomes of the non-germinal center (non-GCB) subtype of DLBCL. Randomized phase III clinical trials evaluating RCHOP +/- Len in non-GCB DLBCL are highly anticipated. The GOYA study found RCHOP was as effective as OCHOP in patients with unselected newly diagnosed DLBCL. We hypothesize the combination of OL-CHOP will result in enhanced NK cell-mediated cytotoxicity, down-regulation of interferon regulatory factor 4, and a complete response (CR) rate ≥ to OCHOP (57% in GOYA trial, Vitolo et al, ASH 2016).Methods: Newly diagnosed, CD20+ DLBCL patients were eligible if they have measurable disease, age ≥18 years, and adequate organ function. In the Phase Ib trial, Obinutuzumab was dosed at 1000mg IV days (d) 1, 8, and 15 during cycle 1, and d 1 on cycles 2-6. Lenalidomide dose was 15mg d 1 - 14 cycles 1 - 6 in the first cohort, with lower dose cohorts if toxicities occurred, using a 3+3 design. CHOP was dosed in standard fashion. Once the maximum tolerated dose (MTD) was identified, phase II was initiated with a planned total of at least 50 evaluable patients at the MTD. The primary objectives of this phase Ib/II trial were to determine the MTD and efficacy of LOCHOP. Correlative studies, including minimal disease assessment, cell of origin determination, and immune profiling are planned on blood and tumor samples.Results: Fifty three patients were enrolled, 6 in Phase Ib trial, 47 in Phase II with all evaluable for safety and 51 for efficacy. Two patients withdrew consent after 1 cycle of therapy. The median age was 62 years (26-83), with 42% males and a median IPI of 2 and NCCN IPI of 3. Five patients had double hit DLBCL, 5 patients had Double Expressor DLBCL, and 21 patients were non-GCB DLBCL. No dose limiting toxicities were identified in in Phase Ib. Toxicities encountered in PhIb include neutropenia (Grade 3: 50%, Grade 4 33%), thrombocytopenia (grade 3: 17%), and rash (grade 2: 17%), with similar findings in PhII. All six patients in PhIb achieved a complete response (CR). In the Phase II trial, 50 patients have end of therapy response assessment as of 8/2/17, with 45 with CR, 4 with partial response, 1 with progression. In the combined evaluable patients, the overall response rate is 98%, with CR of 90%. The CR rate was 90% in non-GCB and 92% in GCB. The median follow up 11 months and the median progression free and overall survival are not reached. There have been no deaths. Additional follow up and analyses will be presented at the meeting.Conclusions: The combination of Obinutuzumab, Lenalidomide, and CHOP is well tolerated, with no dose limiting toxicity encountered in the phase Ib trial, and efficacy is impressive in this single center single arm Phase Ib/II trial. Adverse events do not appear to be different than expected with standard RCHOP. Final results and correlative analyses will be presented at the ASH meeting [Display omitted] DisclosuresWestin:Apotex: Membership on an entity's Board of Directors or advisory committees; Novartis Pharmaceuticals Corporation: Membership on an entity's Board of Directors or advisory committees; Kite Pharma: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees. Nastoupil:TG Therapeutics: Honoraria, Research Funding; Gilead: Honoraria; Abbvie: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Genentech: Honoraria, Research Funding; Karus Therapeutics: Research Funding; Celgene: Honoraria, Research Funding. Neelapu:Merck: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Poseida Therapeutics, Inc: Research Funding; Cellectis Inc.: Research Funding; Kite Pharma: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol-Myers Squibb: Research Funding; Karus: Research Funding; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Lee:KITE PHARMA: Consultancy.

Interim PET-CT Result Is Not Predictive of Survival in Patients with MYC-Rearranged Non-Burkitt Aggressive B Cell Lymphoma

Abstract Background Patients with a diagnosis of MYC-rearranged non–Burkitt aggressive B-cell lymphoma (MYC-R), including those with double hit lymphoma, are at high risk of developing relapsed/refractory disease, even if treated with intensive front-line immunochemotherapy. It is common in clinical practice and clinical trials to perform an interim positron emission tomography (PET)/computed tomography (CT) scan (iPET) during front-line therapy for diffuse large B-cell lymphoma. However, the utility of the iPET result for MYC-R patients for predicting outcomes is unclear. Patients and Methods We performed a single-center retrospective study with centralized pathologic review and PET/CT image acquisition and interpretation for 28 MYC-R patients. The patients received front-line therapy with R-CHOP (rituximab, cyclophosphamide, hydroxydaunorubicin [doxorubicin], Oncovin [vincristine], prednisone) or intensive immunochemotherapy. Results Eight patients had iPET-positive (iPET+) and 20 patients had iPET-negative (iPET−) results using the Deauville visual assessment criteria. At a median follow-up length of 30.4 months, progression-free survival was 65% and overall survival was 76%, neither of which differed significantly between the iPET− and iPET+ patients. The positive predictive value of iPET for progression at 30 months was 25%, and the negative predictive value was 65%. Conclusion Although patients with MYC-R lymphoma have been reported to be at high risk of primary treatment failure, this was not predicted by iPET+ results. Thus, the iPET result should not be used to guide changes in front-line or consolidative therapy for these patients.

Umbralisib/TGR-1202 As a Novel Dual PI3K/CK1 Inhibitor Has a Unique Therapeutic Role in Silencing Oncogenes in Aggressive Lymphomas

Introduction: A number of oncogenes have been shown to promote the pathogenesis and correlate with poor survival in aggressive lymphoma. For example, deregulation of C-MYC is prevalent in diffuse large B cell lymphoma (DLBCL), chromosome translocations involving C-MYC and BCL2 define “double hit lymphoma” (DHL), and chromosome translocation involving CCND1 is pathognomonic for mantle cell lymphoma (MCL). Despite impressive progress being made in the treatment of these aggressive lymphomas, the treatment options for patients with relapsed aggressive lymphoma, especially DHL, remain inadequate. Interestingly, translation of C-MYC, BCL2, CCND1, and a number of other oncogenes is highly sensitive to inhibition of the eukaryotic translation initiation factor 4F (eIF4F) (Manier et al., 2017; Wolfe et al., 2014), which is comprised of the eIF4E, eIF4A, and eIF4G subunits. However, inhibitors of eIF4F are currently not available for the clinic. eIF4F is negatively regulated by the eIF4E binding protein 1 (4E-BP1), which in the hypo-phosphorylated form sequesters eIF4E (Haghighat et al., 1995). To allow for robust initiation of translation in cancer cells 4E-BP1 is hyper-phosphorylated by upstream signals, such as mechanistic target of rapamycin (mTOR). Our recent results indicate that casein kinase 1 epsilon (CK1ε) positively regulate translation via phosphorylating4E-BP1 (Deng et al., 2016). There may be additional poorly understood signals regulating 4E-BP1. Although no CK1ε inhibitor has been developed for the clinic, we recently discovered that the PI3Kδ inhibitor umbralisib/TGR-1202 acts additionally to inhibit CK1ε. Given that TGR-1202 has demonstrated a favorable safety profile in clinical trials and is capable of targeting both PI3Kδ and CK1ε, the drug may be particularly valuable for the silencing of oncogenes in aggressive lymphoma. Our hypothesis is the following: If hyper-phosphorylation of 4E-BP1 serves as a nexus for upregulating oncogene translation, then co-targeting signals phosphorylating 4E-BP1, such as CK1ε and mTOR, may potently and synergistically silence oncogenes in aggressive lymphomas.Materials and Methods: Cytotoxicity was studied in lymphoma cell lines and using Cell TiterGlo (Promega®). Screen of drug synergy was performed between pairs of TGR-1202, mTOR inhibitors, proteasome inhibitors, and CK1 inhibitors. The Bliss additivism model was used to determine the expected inhibition of cell growth and the excess over Bliss (EOB) values. EOB values above 0 indicate synergy, with higher values indicating higher levels of synergy. Expression of oncogenes was studied by Western blot and qPCR. 4E-BP1 was studied by Western blot and by 4E-BP1 cap-binding assay using m7GTP Sepharose beads. Translation was determined using a bicistronic reporter. Drug studies were validated by gene knockout of expected molecular targets such as CK1ε and PI3Kδ using CRISPR/Cas9. To further investigate the global effects of targeting translation initiation, we are conducting in parallel proteomic and RNA-Seq studies in 3 cell lines representing DLBCL, DHL, and MCL. Furthermore, the mechanism will be validated by rescuing experiments through over-expression of eIF4E, MYC, or CCND1. The efficacy of TGR-1202-containing regimens is being investigated in xenograft mouse models of MCL.Results: We investigated the combinatorial effect of TGR-1202 and the proteasome inhibitor carfilzomib. The combination of TGR-1202 and carfilzomib (TC) was highly synergistic in mantle cell lymphoma and double hit lymphoma cell lines (Figure 1A). TC synergistically inhibited phosphorylation of 4E-BP1 at the Ser65 in both MCL and DHL (Figure1B). The TC combination synergistically produced cleavage of poly-ADP ribose polymerase (PARP), indicating apoptosis as its mechanism (Figure 1C). TC reduced the protein expression levels of several oncogenes responsible for malignancies such as c-Myc, cyclin D1, and Bcl-2. In contrast, this combination had no significant effect on the mRNA level of c-Myc and cyclin D1 (Figure 1D).Conclusion: A novel therapeutic strategy can be developed by combining umbralisib/TGR-1202 and the proteasome inhibitor carfilzomib for the treatment of DLBCL, DHL, and MCL. The combination induces apoptosis by inhibiting the phosphorylation of 4E-BP1, leading to translational silencing and suppressed protein levels of c-Myc, cyclin D1, and Bcl-2. [Display omitted] DisclosuresAli:Onconova Therapeutics: Consultancy; Kura Oncology: Consultancy. O'Connor:Celgene: Honoraria, Research Funding; Trillium Therapeutics: Research Funding.

Functional Implication of p53-Mediated DNA Damage Response on the B-Cell Lymphoma Microenvironment Interaction in Chemoimmunotherapy

Resistance against chemotherapy remains a challenge in the current treatment of B cell malignancies. In previous work we could show that macrophages are an essential element in the synergistic response to the administration of the DNA damaging agent Cyclophosphamide and monoclonal antibodies. The combination treatment strongly increases tumor clearance overall survival in the treatment refractory humanized BCL2-MYC-dependent double hit lymphoma hMB mouse model.Here we observed that the alkylating agent Cyclophosphamide is inducing the secretion of distinct cytokines like VEGF-A, CCL4, IL8, IL10 and TNFα by the leukemic cells. This in turn leads to a repolarization of tumor associated macrophages from a suppressive to an activated phenotypic state, which is designated as acute secretory activating phenotype (ASAP). We analyzed the functional implications of the DNA damage response pathway for the generation of the ASAP and synergy in chemoimmunotherapy. When disrupting DNA damage response pathway by knock-down of key elements like ATM, DNA-PK or p53 in the leukemic cells we could prevent the formation of the stimulatory ASAP effect on macrophage phagocytic capacity. Particularly p53 status seems to possess a central regulating role on macrophage mediated malignant cell depletion. TP53 activation via Nutlin-3A treatment of lymphoma cell enhances ADCP in in p53 wildtype cells, while not displaying enhancement in p53-deficient lymphoma cells. Utilizing the hMB model for modeling in vivo treatment of lymphoma bearing mice we could demonstrate diminished ASAP and ADCP for p53-deficient lymphoma treated with cyclophosphamide in vivo . Using primary human CLL patient cells comparing both wildtype and p53-deficient status, the p53-deficient CLL cells failed to induce the stimulatory, cytokine-mediated effect on macrophage phagocytosis in response to combination treatment as seen with the p53 proficient CLL cells. This observation is underlined by in vivo modeling of CLL therapy by treating Eµ-TCL1/p53wt/wt as well as Eµ-TCL1p53-/- mice. Here, low-dose cyclophosphamide treated Eµ-TCL1p53-/- mice failed to induce an antibody mediated stimulatory effect on macrophage phagocytosis capacity as seen with Eµ-TCL1/p53wt/wt mice.In this work, we are able to recapitulate the effect of ASAP mediated increase of macrophage mediated antibody dependent cellular phagocytosis in the Eµ-Tcl1 mouse model and therefore demonstrate that the ASAP is not only restricted to double hit lymphoma, but also shows functional relevance in low-grade lymphoma such as CLL. Moreover, we indicate that p53 functional status influences the generation of the ASAP in independent models in vitro and in vivo as in primary CLL patient cells. Mediated by this process p53 regulates the interraction with tumor asociated macrophages and the tumor microenvironment.Finally our ongoing research offers possibility to reveal and tailor new treatment approaches for chemorefractory patients. DisclosuresHallek:AbbVie: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Celgene: Consultancy, Honoraria, Research Funding; F. Hoffmann-LaRoche: Consultancy, Honoraria, Research Funding; Gilead: Consultancy, Honoraria, Research Funding; Janssen-Cilag: Consultancy, Honoraria, Research Funding; Mundipharma: Consultancy, Honoraria, Research Funding.

Stratification By MYC Protein Expression Level Improves the Prognostic Impact of MYC Translocation in Patients with Large B-Cell Lymphoma

Introduction: In patients with Large B-cell Lymphoma (LBCL) the prognostic significance of MYC translocations is not clearly established. We aimed to investigate if additional identification of MYC translocation partner gene and/or MYC and BCL2 protein expression levels could improve patient prognostication associated with MYC translocations. We also wanted to evaluate the prognostic effect of double hit MYC + BCL2/BCL6 translocation (DH) compared to MYC single hit (SH).Methods: A consecutively collected training cohort of 64 patients and an independent validation cohort of 28 patients all with MYC translocation positive LBCL were investigated. Translocations were identified with fluorescent in situ hybridization (FISH) and MYC and BCL2 protein expression with immunohistochemistry (IHC).Results: MYC> 75% was seen in 50% of patients and was related to frequent bone marrow involvement, high IPI and elevated LDH. MYC>75% was also significantly associated with reduced progression free survival (PFS) and overall survival (OS) in univariate analysis (PFS: HR 6.8 (95%CI 1.5-31), p=0.004. OS: HR 4.3 (95%CI 0.9-21), p=0.05) (Fig. 1). In multivariable analysis adjusting for IPI MYC>75% was independently prognostic for PFS but not OS (PFS: HR 5.4 (95%CI 1.1-25), p=0.04. OS: HR 3.1 (95%CI 0.6-16), p=0.2). DH was seen in 71% of patients and did not confer a worse outcome compared to MYC SH. Immunoglobulin (IG) MYC translocation partner gene was found in 55% and was related to MYC protein expression levels (p=0.047). Translocation partner gene was not prognostic for PFS (p=0.8) or OS (p=0.6). High BCL2 expression was related to high MYC expression (p=0.003) and did not add to the prognostic effect observed for MYC. The findings were confirmed in a comparable, independent validation cohort of 28 MYC translocation positive LBCL. The patients included in the survival analyses were treated with R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone) or R-CHOEP (R-CHOP + etoposide).Conclusions: These findings suggest that stratification by MYC protein expression level significantly improves the prognostic impact of MYC translocation in patients with large B-cell lymphoma. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

A Simulation Analysis to Evaluate the Effect of Prospective Biomarker Testing on Progression-Free Survival (PFS) in DLBCL

Introduction. Novel treatment regimens that combine chemotherapy with targeted agents are being developed for DLBCL. Evaluation of these targeted agents in clinical trials may require prospective biomarker testing, such as immunohistochemistry (IHC), to select patients based on relevant molecular features. However, prospective biomarker testing takes days and there is concern that the delay in treatment may prevent enrollment of patients with the most aggressive disease, biasing the trial population. Here, using data from the Phase 3 GOYA trial (NCT01287741) in first-line DLBCL, we evaluate the impact on PFS of a treatment delay that simulates the additional screening time required for prospective IHC testing.Methods. In GOYA, previously untreated DLBCL patients were randomized 1:1 to obinutuzumab or rituximab plus 6 or 8 cycles of CHOP. Median times from clinical diagnosis to initiation of screening (“DTIS interval”), and from initiation of screening to randomization (“screening time”) were determined in patients with IPI 2-5 (n=1124 patients with timeline information available, no prospective testing). A two-step analysis of PFS was conducted by dividing the evaluable patient population into three groups of varying DTIS intervals (Figure 1, Step 1), and then into additional groups of varying screening times (Figure 1, Step 2). Groups for screening times were 1) <6 days (based on median screening time of 6 days in GOYA; 2) between 6-9 days (based on median screening time of 9 days in Roche clinical trial with prospective IHC testing - NCT02366143); and 3) above 9 days. We asked whether the clinical outcome (PFS), assessed by Kaplan-Meier survival analysis, correlated with either the DTIS interval (Step1) or the screening times (Step 2).Results. Median time from diagnosis to randomization was 24 days (range 2 to 1106 days, 95th percentile of 66 days). Patients with times from diagnosis to randomization below 15 days (n=250) had significantly worse outcome than patients that were randomized in 15 or more days (n=413 for 15-28 days and n=466 for >28 days; p<0.0001; Table 1; Figure 2 A). 3-year PFS was 56% for patients with <15 days, 70% for 15-28 and 73% for >28 days from diagnosis to randomization. In Step 1 of the two-step analysis, DTIS times of <8 days, 8-14 days, and >14 days were observed for 190, 228 and 706 patients, respectively. Patients with a DTIS interval of <8 days were enriched with BCL2-positive and Double Hit status (Table 2). Patients with a DTIS interval of <8 days had shorter PFS than patients with longer DTIS intervals (p=0.0011 Table 1; Figure 2 B); 3-year PFS was 55% for patients with <8 days, 66% for patients with 8-14 days and 72% for patients with >14 days from diagnosis to screening; no additional difference in 3-year PFS was observed in subgroups beyond 14 days. In Step 2, we asked if a longer screening interval (screening to randomization in Figure 1), simulating additional time for prospective testing, affected patient outcome. Of the 190 patients with the shortest DTIS interval (<8 days from diagnosis to screening) and the shortest PFS from Step 1, 70 patients had screening times of <6 days, 75 between 6-9 days and 45 above 9 days (max 26 days). All cohorts showed indistinguishable PFS curves (p=0.7652; Table 1; Figure 2C). Similarly, no differences in PFS were observed for groups of increasing screening times in the DTIS cohorts of 8-14 days and >14 days (not shown).Conclusion. In GOYA, short PFS was associated with <15 days from diagnosis to randomization and <8 days from diagnosis to screening, possibly attributable to high-risk biology, such as high expression of BCL2 and DH. Despite seemingly expedited work-up, these high-risk patients do poorly, highlighting the need for targeted therapies and innovative trial designs. In this example, additional screening time similar to the time required for prospective testing did not adversely affect PFS. Our results may have implications for designing precision medicine trials in DLBCL. [Display omitted] DisclosuresSzafer-Glusman:Genentech: Employment. Liu:F. Hoffmann-La Roche: Employment. Peale:Genentech / Roche: Employment, Equity Ownership. Farazi:Genentech: Employment. Ray:Genentech/Roche: Employment. Horn:F. Hoffmann-La Roche Ltd: Employment. Oestergaard:F. Hoffmann-La Roche Ltd: Employment. Kornacker:F. Hoffmann-La Roche: Employment; Roche: Equity Ownership. Sehn:Janssen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; Seattle Genetics: Consultancy, Honoraria; Roche/Genentech: Consultancy, Honoraria. Fingerle-Rowson:F. Hoffmann-La Roche Ltd: Employment, Equity Ownership. Venstrom:Genentech, Inc.: Employment. Byrtek:Genentech: Employment, Equity Ownership. Punnoose:Genentech: Employment.

Real World Outcome in a Patient Cohort with Newly Diagnosed Diffuse Large B Cell Lymphoma

Intro:Recent phase 3 trials targeted to patients with adverse prognostic factors in DLBCL (non GC, double hit/expressors) have faced problems in interpretation because control groups scored clearly better than expected, (Lym2034, Pyramid, ReModl). This suggests that the application of inclusion/exclusion criteria, combined with a delay in treatment caused by their confirmation, may lead to a selection bias that excludes the patient group most in need for treatment improvement.Aims:To characterize the magnitude and nature of exclusion bias we conducted a retrospective real world study on unselected patients diagnosed with DLBCL between 2011-2015 and followed up for at least 2 years at the University Hospital Ghent, a tertiary care centre serving an area of approximately 1.5 million inhabitants.Four questions were asked:- How many patients are considered fit enough in a real world population of DLBCL to receive standard treatment (R-(mini)CHOP, -like)?- What is the PFS/OS in this group compared to the total patient population?- How is the PFS/OS influenced by time from diagnosis to initiation of treatment?- How is treatment choice and PFS/OS influenced by comorbidities?Patients and methods:A total of 162 patients with DLBCL were included. The median age was 65 years (range 16-91) with amale/female ratio 56%/44%. Four % of patients had an ECOG >2 and the distribution of the IPI-score was 0-1: 44 patients (27%), IPI 2: 27 (17%), IPI 3: 49 (30%), IPI ≥ 4: 41 patients (25%). NCCN-IPI was low (0-1) in 16 pts, low interm (2-3) in 48 pts, high-int (4-5) in 71 pts and high(>5) in 22 pts (14%). All patients had a pathological diagnosis of DLBCL (excl PMBL). In 25 patients there were signs of transformed lymphoma with DLBCL, 12 were double expressor (DEL), 3 double hit lymphoma (DHL). Cell of origin was assessed by IHC, Hans Algorythm in 131 patients (72 GC, 57 non GC). 133 patients were treated with standard R-(mini)CHOP or R-CHOP like regimens (81%) outside clinical trials (RCT). 17 patients were considered unfit for at least R-mini CHOP: 5 received R-CvP, 8 R-mono, 4 palliative care. Four young patients with aggressive presentation received more intensive regimens. 107/162 completed full treatment as planned (66%) 12 patients were entered in RCT and received at least R-CHOP like regimens. Statistical significance was tested using a log-rank based analysis on actuarial survival curvesResults:Median follow-up was 61 mo (range 19-323).Response assessment was based on PET-CT as defined by the Lugano 2014 criteria.Overall, 143/162 could be treated with R-(mini)-CHOP like regimens (88%), 12 in RCT and 131 out of RCT. 123/162 76% of patients responded, 18 died (11%) and 19/162 (12%) were refractory to 1st line treatment. 37 of 123 responders relapsed, 10 within 6 months, 28 others after a median of 44 months.In the subgroup of 131 patients receiving standard R-(mini)CHOP like treatment outside RCT, 101 reached CR (77%), 4 PR, 15 PD. 9 died and 28 relapsed following CR. Two years PFS is 67% in the standard R-(mini)CHOP group outside RCT, and 83% in 12 patients entering RCT. 2 years overall survival is 71% in the 131 patients receiving standard R-(mini)CHOP and 92% in the 12 patients in RCT.There was no difference in PFS and overall survival in patients with GC versus non GC based on IHC, (p=0.503) or by the presence of signs of transformation (p=0.131).Time from diagnosis to initiation of treatment divided the patient population in patients receiving treatment within 16 days (n=66), 16-30 days (n=55) and beyond 30 days (n=41).Two years PFS and OS in these 3 groups were 50%, 76%, 79% (p=0,004) and 59%, 72%, 83% (p=0,017), respectively.Significant comorbidities were observed in 114/162 patients. Two years OS in patients without comorbidity was 92% compared to 54% in patients with comorbidity. (p<0.05)Conclusion:Standard treatment in DLBCL can be given to up to 88% of patients, but it selects for fitter patients with less comorbidities and better PFS/OS and in RCT it is even more so (2yr OS 67, 71 and 91%). Patients requiring early treatment who cannot be subject of central review as part of study inclusion because of potential treatment delay fare worse than patients in whom treatment can be delayed. These combined factors may explain part of the better results observed in RCT control groups compared to real world data. DisclosuresNo relevant conflicts of interest to declare.

Impact of Immunohistochemistry-Defined Cell of Origin and MYC Rearrangements on Outcomes in Patients with PET-Defined Stage I/II Diffuse Large B-Cell Lymphoma Treated with R-CHOP ± Radiotherapy: An International Multi-Center Retrospective Study

Impact of Immunohistochemistry-Defined Cell of Origin and MYC Rearrangements on Outcomes in Patients with PET-Defined Stage I/II Diffuse Large B-Cell Lymphoma Treated with R-CHOP ± Radiotherapy: An International Multi-Center Retrospective Study

Primary thyroid lymphoma with double expression of MYC and B-cell lymphoma-2 Gene

Primary lymphomas of the thyroid (PTLs) are rare entities. We report a case of high-grade B-cell lymphoma double hit (DH) ofthe thyroid gland with dual expression of MYC and BCL-2 gene in a 55-year-old woman. Most PTLs have a good prognosis tocombined modality treatment but identification of DH lymphomas or double-expressor lymphomas is important as these are resistant tochemotherapy and radiotherapy and pose a challenge for the treatment of the same.

Stage 2 Registered Report: Variation in neurodevelopmental outcomes in children with sex chromosome trisomies: testing the double hit hypothesis.

Background: The presence of an extra sex chromosome is associated with an increased rate of neurodevelopmental difficulties involving language. The 'double hit' hypothesis proposes that the adverse impact of the extra sex chromosome is amplified when genes that are expressed from the sex chromosomes interact with autosomal variants that usually have only mild effects. We predicted that the impact of an additional sex chromosome on neurodevelopment would depend on common autosomal variants involved in synaptic functions. Methods: We analysed data from 130 children with sex chromosome trisomies (SCTs: 42 girls with trisomy X, 43 boys with Klinefelter syndrome, and 45 boys with XYY). Two comparison groups were formed from 370 children from a twin study. Three indicators of phenotype were: (i) Standard score on a test of nonword repetition; (ii). A language factor score derived from a test battery; (iii) A general scale of neurodevelopmental challenges based on all available information. Preselected regions of two genes, CNTNAP2 and NRXN1, were tested for association with neurodevelopmental outcomes using Generalised Structural Component Analysis. Results: There was wide phenotypic variation in the SCT group, as well as overall impairment on all three phenotypic measures. There was no association of phenotype with CNTNAP2 or NRXN1 variants in either the SCT group or the comparison groups. Supplementary analyses found no indication of any impact of trisomy type on the results, and exploratory analyses of individual SNPs confirmed the lack of association. Conclusions: We cannot rule out that a double hit may be implicated in the phenotypic variability in children with SCTs, but our analysis does not find any support for the idea that common variants in CNTNAP2 or NRXN1 are associated with the severity of language and neurodevelopmental impairments that often accompany an extra X or Y chromosome. Stage 1 report: http://dx.doi.org/10.12688/wellcomeopenres.13828.2.

PRELIMINARY ACTIVITY OF DEVIMISTAT (CPI 613) IN PATIENTS WITH RELAPSED OR REFRACTORY BURKITT LYMPHOMA/LEUKEMIA

Introduction: Patients (pts) with primary refractory or relapsed Burkitt lymphoma/leukemia (BL) or high-grade B-cell lymphoma with MYC and BCL2 rearrangements (double hit, DHL) and/or BCL6 (triple hit, THL) have a dismal prognosis. A dysregulated MYC oncogene has downstream effects on proliferation and highly glycolytic metabolism with tricarboxylic acid (TCA) cycle intermediates as biosynthetic precursors. Devimistat is a non-redox active analogue of lipoic acid, a required cofactor for two key TCA cycle mitochondrial enzymes: pyruvate dehydrogenase and alpha ketoglutarate dehydrogenase. Disruption results in a shutdown of ATP and biosyntheticintermediate production leading to cancer cell death. In a phase I trial (n = 26) a pt with multiply refractory BL had a partial remission sustained over one year prior to resection. She remains in remission 7 years later. As of March 2021, 20 clinical studies in over 700 pts with various cancers have been conducted (ongoing/completed) with devimistat. We initiated a phase II trial to further explore efficacy. Methods: NCT03793140 is a multicenter study enrolling 17 patients on each of two cohorts BL or DHL/THL. Pts must have had one prior therapy or are refusing standard of care, measurable disease or isolated bone marrow involvement, and must not be within 3 months of a prior stem cell transplant. Active central nervous system (CNS) parenchymal disease is excluded, but leptomeningeal disease is allowed if the CSF is negative for more than 4 weeks and the maintenance intrathecal/intraOmmaya therapy is ongoing. Devimistat is given by central line over 2 hours daily x 5 days for two 14-day cycles and then as maintenance every 21 days. Pts are evaluable for response if they receive at least 4/5 days of the first cycle. At least 1response in the first 9 pts by cycle 3 was needed to expand to 17 pts in total in that cohort. Results: 9 pts were enrolled in the DHL/THL arm. Number of prior therapies was 3(1-6). No responses and only 1 stable disease resulted in cohort closure. Thus far, 8 BL pts were enrolled. Number of prior therapies was 3 (2-4). 2 pts were inevaluable. 1/6 pts had a response. This HIV+ pt had a thigh mass having received 4 prior therapies. He had a near complete metabolic remission after 3 cycles of devimistat. PET/CT assessment: Thigh lesion baseline October 19, 2020: 10.8 x 6.5 cm SUV 24. After cycle 3: unmeasurable, SUV 2. He is currently in cycle 7, having had a 4-week treatment delay of cycle 5 due to CoVID 19 infection. ECOG improved from 3 to 0. Adverse events (AE): As of March 03, 2021, no serious AEs related to study drug. 4 pts had grade 3 events at least possibly related: 2 neutropenia, 1 thrombocytopenia and 1 elevated bilirubin. 1 patient had a dose reduction for grade 2 alanine aminotransferase increase. Conclusions: Although our results are preliminary, the near complete remission in this patient is promising in a disease where no viable treatment options exist.

Malignant plasmacytes in bone marrow detected by flow cytometry as a predictor for the risk stratification system of multiple myeloma.

Multiple myeloma (MM) is a clonal disorder characterized by the proliferation of plasma cells and their accumulation within the bone marrow (BM). The flow cytometric analysis is an essential method for the hematological diseases because of high sensitivity. This study evaluates the indication role of malignant plasmacytes (PCs) in BM detected by flow cytometry for the risk stratification of MM. Whole BM samples from 92 newly diagnosed MM patients were included in the study. We collected 106 cells each sample by flow cytometry. Then we analyzed the correlation between the malignant PCs in BM and the characteristics of patients. In this study, patients were stratified according to different baseline characteristics and the median ratio of the malignant PCs were compared. The significant statistical differences (p < 0.05) were: Hb < 100 g/L versus ≥100 g/L; β2-microglobulin <3.5 mg/dL versus 3.5-5.5 mg/dL versus >5.5 mg/dL; LDH > 250 U/L versus LDH 250 U/L; ISS I versus ISS II versus ISS III; R-ISS I versus II versus III. The detailed data are showed in Table 2. The significant correlations were observed between the malignant PCs in BM and (Figure 1): plasma cell of biopsy, hemoglobin, β2-microglobulin, lactate dehydrogenase (LDH), creatinine. "Double hit" or "triple hit" are defined as containing any two or three of the high risk cytogenetic abnormalities (t(4;14), t(14;16), t(14;20); del17q; TP53 mutation; 1q21 gain) by mSMAR. "Double or triple hit" had independently unfavorable significance for overall survival. As expected, the malignant PCs of "double or triple hit" group is significantly higher than the group B (one high risk genetic factor) and the group A (normal cytogenetic) (p < 0.0001 and p < 0.019). Multiparametric flow cytometry is a highly sensitive method to identify and quantify malignant PCs. And the ratio of malignant PCs detected by MFC showed strongly correlation with the severity of the pathology of MM. Malignant PCs in BM detected by flow cytometry could be regarded as a predictor for the risk stratification system of MM. Thus, it should be considered applying in the routine evaluation of MM at diagnosis and after therapy.

CD37 and CD44 evaluation by flow cytometry: can these markers improve B cell lymphoma characterization?

PurposeDiffuse large B cell lymphoma (DLBCL) is a heterogenous entity with many prognostic ancillary tests that are based on immunohistochemistry. These include cell-of-origin determination by the Hans criteria and “double expresser” characterization. Additionally, MYC fluorescence in situ hybridization is required to distinguish DLBCL from its morphologically similar counterpart, “double hit lymphoma” or high-grade B cell lymphoma (HGBCL), a “Burkitt-like” lymphoma with a worse prognosis than DLBCL. All such ancillary tests require judicious triage of oftentimes limited specimens. Loss of expression of CD37 has been associated with worse outcomes in DLBCL. Absence of CD44 is more commonly seen in Burkitt lymphoma than in DLBCL.MethodsVery few lymphoma studies have evaluated CD37 or CD44 expression by flow cytometric analysis, which permits a more quantitative assessment. To evaluate the utility of these two markers, we retrospectively reviewed seventy-three B cell non-Hodgkin lymphoma specimens in which expression of CD37 and CD44 was evaluated by flow cytometry. Median fluorescence intensity ratio (MFIR) was calculated by dividing the median fluorescence intensity of the lymphoma population by that of background non-B mononuclear cells.ResultsIn lymphomas with DLBCL-like morphology, loss of CD44 expression was associated with MYC rearrangement (MFIR rearranged 0.3082 vs intact 1.09, p=0.003). CD44 loss in DLBCL was also associated with germinal center B cell-like (GCB) cell-of-origin (MFIR GCB 0.4139 vs non-GCB 1.499, p=0.036) as has been seen in other studies. In DLBCL, loss of CD37 expression was not associated with cell-of-origin, double expresser status, or MYC rearrangement.ConclusionsWhile our findings require confirmation in a larger case series, decreased CD44 expression by flow cytometry in an otherwise limited biopsy with DLBCL-like features may provide the impetus for FISH testing and ultimate classification.

Carfilzomib-based induction/consolidation with or without autologous transplant (ASCT) followed by lenalidomide (R) or carfilzomib-lenalidomide (KR) maintenance: Efficacy in high-risk patients.

8002 Background: Cytogenetic abnormalities (CA) are one of the most powerful prognostic factors in multiple myeloma (MM). In the FORTE study, carfilzomib-lenalidomide-dexamethasone induction/consolidation with ASCT (KRd_ASCT) significantly improved progression-free survival (PFS) vs KRd without ASCT (KRd12, HR 0.64) or carfilzomib-cyclophosphamide-dexamethasone (KCd) plus ASCT (KCd_ASCT, HR 0.53). KR maintenance significantly improved PFS vs R (HR 0.63). Methods: MM patients (pts) were randomized to KRd_ASCT vs KCd_ASCT vs KRd12 and, thereafter, to KR vs R maintenance. Subgroup analyses according to FISH evaluated the impact of each single high-risk (HiR) CA [del17p, t(4;14), t(14;16), del1p and 1q gain (3 copies) or amp1q (≥4 copies)] and that of combined CA, defining HiR by the presence of ≥1 HiR CA and double-hit (DH) by the presence of ≥2 HiR CA. Pts negative for all the HiR CA were considered at standard risk (SR). The primary objective was the impact of treatment on PFS according to pt risk. Results: 396 out of 474 enrolled pts were included in the analysis with complete FISH data: 243 HiR, 105 DH and 153 SR. Among HiR pts, 60 had del17p, 65 t(4;14), 20 t(14;16), 44 del1p, 126 1q gain and 49 amp1q. SR pts benefited from intensification with KRd_ASCT vs KRd12 (HR 0.47, p = 0.05) and KCd_ASCT (HR 0.38, p = 0.01), with a 4-year PFS of 80%, 67% and 57%, respectively. In HiR pts, KRd_ASCT improved PFS vs KRd12 (HR 0.6, p = 0.04) and KCd_ASCT (HR 0.57, p = 0.01), with a 4-year PFS of 62%, 45% and 45%, respectively. The advantage with KRd_ASCT vs KRd12 (HR 0.53, p = 0.07) and KCd_ASCT (HR 0.49; p = 0.03) was also observed in DH pts (4-year PFS 55%, 31% and 33%, respectively). Analyses by single CA were limited by the small number of pts in each subgroup, but a trend towards a PFS benefit from KRd_ASCT vs KRd12 was seen in pts with del17p (HR 0.61, p = 0.3), t(4;14) (HR 0.59, p = 0.2) and 1q gain (HR 0.45, p = 0.02). In pts with del1p, KRd_ASCT (HR 0.24, p = 0.06) and KRd12 (HR 0.33, p = 0.09) showed superiority over KCd_ASCT, while amp1q pts had the worst outcome regardless of treatment (KRd_ASCT vs KCd_ASCT, HR 1.16, p = 0.73; KRd12 vs KCd_ASCT, HR 1.34, p = 0.45). KR improved PFS vs R in SR (3-year PFS 90% vs 73%, HR 0.42, p = 0.06), HiR (3-year PFS 69% vs 56%, HR 0.6, p = 0.04) and DH pts (3-year PFS 67% vs 42%, HR 0.53, p = 0.1). Despite the small subgroups, a beneficial trend with KR vs R was observed in pts with del17p (HR 0.59, p = 0.37), t(4;14) (HR 0.59, p = 0.3), 1q gain (HR 0.54, p = 0.07) and del1p (HR 0.23, p = 0.08), while amp1q pts showed the worst outcome and no benefit from KR vs R (HR 0.83, p = 0.7). Conclusions: KRd_ASCT and KR maintenance are highly effective in SR and also in HiR and DH pts, with impressive 4-year PFS from diagnosis (KRd_ASCT: HiR 62%, DH 55%) and 3-year PFS from maintenance (KR: HiR 69%, DH 67%), thus supporting their use in HiR pts, who represent an unmet medical need. Clinical trial information: NCT02203643.

AB0739 MULTISYSTEM INFLAMMATORY SYNDROME ASSOCIATED WITH SARS-COV-2 INFECTION AND E.COLI SEPSIS: THE POTENTIAL ROLE OF PROCALCITONINE AS A RAPID DIAGNOSTIC BIOMARKER TO DISTINGUISH TWO DIFFERENT PHASES OF SYSTEMIC INFLAMMATORY RESPONSE SYNDROME

Background:Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is typically very mild and often asymptomatic in children. A complication is the rare multisystem inflammatory syndrome in children (MIS-C) associated with COVID-19, presenting 4-6 weeks after infection as high fever, organ dysfunction, and strongly elevated markers of inflammation. The pathogenesis is unclear but has overlapping features with Kawasaki disease suggestive of vasculitis and a likely autoimmune etiology.Objectives:We report a case of multisystem inflammatory syndrome in children (MIS-C) in patient with SARS-CoV-2 infection and Enteropathogenic Escherichia coli (EPEC) sepsis due to acute enteritis, observed at end of December 2020 to a tertiary-care center (San Carlo Hospital), in Basilicata region (Italy).Methods:This healthy 12-year- old male patient was tested positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Clinical presentations was characterized by fever, abdominal pain, gastrointestinal complaints and evanescent rash. Laboratory values were remarkable for high levels of procalcitonin, C-reactive protein (CRP), D-dimers, B-type natriuretic peptide (BNP), and troponin. He also had low albumin levels. Autoantibodies tests were negative. Chest tomography showed ground-glass opacities in less than 25% of the lungs, small bilateral pleural effusion and increased cardiac area; abdominal tomography showed enlargement of the lymphnodes and ascites. Evaluation for other infectious etiologies showed molecular test positivity on fecal samples for EPEC E. coli. He received broad spectrum intravenous antibiotics (macrolids and quinolones and then carbapenems). On the seventh day the enteritis resolved and procalcitonin normalized, however he continued to have lymphopenia, thrombocytopenia, hypoalbuminemia, elevated levels of CRP, D-dimers, ferritin, troponin, and increased BNP. On the ninth day he was feverish again and developed severe cardiac and respiratory failure requiring advanced respiratory support and admission to the intensive care unit. He received IVIG (intravenous immunoglobulin at 2 g/Kg, glucocorticoids (Methylprednisolone 1mg/kg) and enoxaparin.Results:The patient was discharged asymptomatic at home after 28 days of hospital stay.Conclusion:We observed multisystem inflammatory syndrome in children (MIS-C) in a previously healthy patient with SARS-CoV-2 infection and E.coli sepsis, who became critically ill with multisystem involvement. In this case viral and bacterial infections could be considered as a double hit for the etiopathogenesis of MIS-C. The trend of procalcitonin was better than C-reactive protein for differentiating bacterial from non-bacterial phase of systemic inflammatory response syndrome (SIRS) in this critically ill child. Although the accuracy of both tests is moderate. Diagnostic accuracy could be enhanced by combining these tests with bedside clinical judgment.

MYC single-hit large B-cell lymphoma: clinicopathologic difference from MYC-negative large B-cell lymphoma and MYC double-hit/triple-hit lymphoma

MYC-rearranged large B-cell lymphoma with BCL2 and/or BCL6 rearrangement, double-hit (DH) or triple-hit (TH) lymphoma, is associated with poor survival after standard treatment. To investigate the clinical impact of single-hit (SH) MYC rearrangement, we analyzed 241 cases of diffuse largeB-cell lymphoma (DLBCL) for MYC, BCL2, and BCL6 rearrangement by fluorescence in situ hybridization. Fifty-five of 241 (22.8%) cases showed MYC rearrangements. Twenty-three cases were diagnosed as DLBCL; 18 as high-grade B-cell lymphoma (HGBCL)-DH; 3 as HGBCL-TH; and 11 as HGBCL, not otherwise specified. Both DH and TH lymphomas showed high-grade morphology (P=0.002), higher stage (P=0.022), and more frequent germinal center B-cell-likephenotype (P=0.008). SH lymphomas displayed high-grade morphology (P=0.002) but were not different from MYC-negative lymphomas in cell of origin, clinical stage, international prognostic index (IPI), or extranodal involvement. Patients with DH/TH lymphomas had worse overall survival (OS) (P=0.016) and progression-free survival (PFS) (P<0.001), while OS and PFS of SH lymphomas were not different from those of MYC-negative lymphomas. There was no survival difference between cases of BCL2 and BCL6 rearrangements. Poorer prognostic factors included higher ECOG class, higher IPI, and DH or TH translocation for OS, and higher IPI and DH or TH translocation for PFS. Higher IPI was an independent prognostic factor for OS and PFS. In conclusion, large B-cell lymphomas with single MYC rearrangement showed high-grade morphology but were otherwise not different from MYC-negative lymphomas.