ALBERT

Description: Primary mediastinal large B-cell lymphoma (PMBL) is a type of aggressive B-cell lymphoma that typically affects young adults, characterized by presence of a bulky anterior mediastinal mass. Lymphomas with gene expression features of PMBL have been described in non-mediastinal sites, raising questions about how these tumors should be classified. Here, we investigated whether these "non-mediastinal PMBLs" are indeed PMBLs or instead represent a distinct group within DLBCL. From a cohort of 325 de novo DLBCL cases, we identified tumors from patients without evidence of anterior mediastinal involvement that expressed a PMBL expression signature (nm-PMBLsig-pos, n=16, 5%). The majority of these tumors expressed MAL and CD23 - proteins typically observed in bona fide PMBL (bf-PMBL). Evaluation of clinical features of nm-PMBLsig-pos cases revealed close associations with DLBCL, and the majority displayed a germinal center B-cell-like cell-of-origin (GCB). In contrast to bf-PMBL, nm-PMBLsig-pos patients presented at an older age, did not show pleural disease, and bone/bone marrow involvement was observed in three cases. However, while clinically distinct from bf-PMBL, nm-PMBL-sig-pos tumors resembled bf-PMBL at the molecular level with upregulation of immune response, JAK-STAT, and NF-kB signatures. Mutational analysis revealed frequent somatic gene mutations in SOCS1, IL4R, ITPKB and STAT6, as well as CD83 and BIRC3, with the latter genes being significantly more frequently affected than in GCB-DLBCL and bf-PMBL. Our data establish nm-PMBLsig-pos lymphomas as a group of DLBCL with distinct phenotypic and genetic features, and potential implications for gene expression- and mutation-based subtyping of aggressive B-cell lymphoma and related targeted therapies.

Description: Allogeneic hematopoietic cell transplant (HCT) improves survival in patients with relapsed or high risk acute myeloid leukemia (AML). Complete remission (CR) is typically a pre-requisite for transplantation, though many do not achieve a formal CR. The traditional AML treatment starts with induction chemotherapy, followed by assessment of response to guide next steps. Response criteria definitions differ between that of the National Comprehensive Cancer Network (NCCN), utilized by the majority of clinicians, and the Center for International Blood and Marrow Research (CIBMTR) data registry utilized by transplant centers, making interpretation of the impact of HCT difficult. Definitions for morphologic complete remission (CR) are the same, however complete remission with incomplete hematologic recovery (CRi) differs and the CIBMTR does not recognize the morphologic leukemia-free state (MLFS), thus mis-identifying such patients and preventing clear treatment guidelines for this population. We conducted a retrospective study, identifying a cohort of 35 AML patients at our center who underwent allogeneic HCT while in MLFS, to evaluate characteristics in patient demographics, disease status, treatment(s), and outcomes. From our cohort, the median overall survival (OS) was 14 months, however 37% were alive and in remission with median follow-up of survivors of five years. Twenty three percent had progression of disease following transplant. Non-relapse mortality (NRM) was 35% with leading cause of death being infection. Our study reveals that transplant can induce long-term survival in patients with acute leukemia who are in MLFS at the start of induction, similar to data for patients with high risk disease in early relapse or in later remissions. Early transplantation while in MLFS and not waiting for full count recovery may protect patients from toxicities of further chemotherapeutic agents or prevent unnecessary delays that may allow for infections or other barriers to arise, and requires further study. Disclosures Leblanc: American Cancer Society, BMS, Duke University, NINR/NIH, Jazz Pharmaceuticals, Seattle Genetics: Research Funding; UpToDate: Patents & Royalties: Royalties; Agios, AbbVie, and Bristol Myers Squibb/Celgene: Speakers Bureau; AstraZeneca: Research Funding; AbbVie, Agios, Amgen, AstraZeneca, CareVive, BMS/Celgene, Daiichi-Sankyo, Flatiron, Helsinn, Heron, Otsuka, Medtronic, Pfizer, Seattle Genetics, Welvie: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Rizzieri:Karyopharm: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Kite: Honoraria, Speakers Bureau; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Stemline: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Sanofi: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Mustang: Membership on an entity's Board of Directors or advisory committees; Celltrion: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Jazz: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Seattle Genetics: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; AROG: Membership on an entity's Board of Directors or advisory committees; abbvie: Membership on an entity's Board of Directors or advisory committees; Teva: Membership on an entity's Board of Directors or advisory committees; Acrobiotech: Membership on an entity's Board of Directors or advisory committees; Bayer: Membership on an entity's Board of Directors or advisory committees.

Description: Current chemotherapeutic regimens for acute myeloid leukemia (AML) often fail to eliminate leukemic stem cells (LSCs) which contribute to disease relapse. A key step towards the development of more effective therapies is the identification of vulnerabilities that are unique to LSCs. Here, we sought to identify LSC-specific metabolic dependencies by performing a flow cytometry-based screen of 110 metabolically-focused drugs against a primary human AML sample. This sample harbored distinct subsets defined by CD34 and CD38 expression, and LSC activity assayed by xenotransplantation was restricted to the CD34+CD38- fraction. Through this screen, we found that inhibitors of nicotinamide phosphoribosyltransferase (NAMPT), which catalyzes the rate-limiting step in the NAD+ salvage pathway, preferentially depleted CD34+CD38- cells, implicating NAMPT inhibitors as potential anti-LSC agents. To evaluate the therapeutic potential of NAMPT inhibitors, we focused on KPT-9274, a small-molecule NAMPT inhibitor currently under clinical development for other cancer types. Treatment with KPT-9274 depleted the CD34+CD38- fraction across multiple primary human AML samples through induction of apoptosis. The preferential sensitivity of CD34+CD38- cells to NAMPT inhibition correlated with a lower basal level of intracellular NAD+ and greater dependency on NAMPT activity for NAD+ generation relative to the other fractions. In contrast, normal CD34+ HSPCs were largely resistant to the cytotoxic effects of KPT-9274 due to their capacity to utilize the Preiss-Handler pathway for NAD+ generation. Consistent with the in vitro findings, KPT-9274 treatment significantly reduced LSC activity as determined by secondary engraftment potential in 2 of 3 patient-derived xenograft (PDX) models of human AML and had minimal impact on normal HSC activity in mice engrafted cord blood cells. To gain mechanistic insights into how NAMPT inhibition induces cell death, we performed transcriptomic analysis of sorted CD34+CD38- cells treated with KPT-9274. This analysis revealed a striking upregulation of genes involved in cholesterol and lipid synthesis including the stearoyl-CoA desaturase (SCD) gene. The upregulated genes were highly enriched for known targets of the sterol regulatory element binding protein (SREBP) transcription factors. Functional studies demonstrated that this transcriptional response was protective against the cytotoxic effect of NAMPT inhibition in AML cells. To uncover the metabolic basis of this protective effect, we performed global metabolomic profiling of AML cells treated with KPT-9274 and observed a decrease in the ratio of monounsaturated fatty acids (MUFAs) to saturated fatty acids (SFAs) upon drug treatment. This drop in MUFA:SFA ratio reflected a reduction in SCD activity which catalyzes the desaturation of SFAs to MUFAs in a NADPH-dependent reaction. Since depletion of intracellular MUFAs could trigger apoptosis, we hypothesized that the SREBP response might protect against cell death through upregulation of SCD activity and consequent increase in MUFA synthesis. In line with this hypothesis, we found that exogenous oleic acid, a MUFA, completely rescued cell death induced by KPT-9274, while treatment with SCD inhibitors sensitized AML cells to the cytotoxic effects of NAMPT inhibition. To explore the translational application of our findings, we tested whether dipyridamole (DP), a clinically approved anti-platelet agent with inhibitory activity against SREBP signaling, can be repurposed to enhance the anti-leukemic effects of KPT-9274. We showed that treatment with DP, at non-toxic concentrations, potentiated the cytotoxicity of KPT-9274 against AML cells in vitro. Importantly, in vivo combination treatment with KPT-9274 and DP effectively targeted LSC activity in a PDX model that was refractory to KPT-9274 as single agent. In summary, our findings demonstrate that LSCs are preferentially dependent on NAMPT activity for survival over non-LSCs and normal HSCs. We further uncovered that NAMPT inhibition results in dysregulation of lipid homeostasis and induces a lipogenic response coordinated by SREBPs that protects AML cells against NAD+ depletion. These findings offer insights into drug combination strategies to enhance the efficacy of NAMPT inhibitors and provide the rationale for testing NAMPT inhibitors in the treatment of AML in clinical trials. Disclosures Dick: Bristol-Myers Squibb/Celgene: Research Funding. Wang:Trilium therapeutics: Patents & Royalties: There is an existing license agreement between TTI and University Health Network and J.C.Y.W. may be entitled to receive financial benefits further to this license and in accordance with UHN's intellectual property policies. .

Description: Background: Ibrutinib (I) and venetoclax (V) have each demonstrated modest single-agent activity in relapsed/refractory follicular lymphoma (FL) (Gopal A, JCO 2018; Davids M, JCO 2017). Preclinical data have shown synergy with these agents in B-cell cell lines (Kuo H, Mol Cancer Ther 2017). Based on these observations, we proposed the first trial to combine I and V in FL. Results from the phase Ib portion of this multi-institutional investigator-initiated trial are presented here (NCT02956382). Methods: This phase Ib/II trial is open at Georgetown/Lombardi CCC, Hackensack/John Theurer CC, and University of Washington/Fred Hutchinson/Seattle Cancer Care Alliance. Eligibility criteria include WHO grade 1-3a FL, 〉1 prior systemic therapy, measurable disease warranting therapy by standard criteria or physician discretion, ECOG performance status 〈 2, adequate marrow, hepatic, renal function. Patients (pts) were enrolled in a standard phase I 3+3 design at a starting dose level (DL) of I 420 mg daily, V 400 mg daily (DL0). The highest initially planned dose level was DL3: I 560 mg daily, V 800 mg daily. There was no dose ramp up of V based on monotherapy experience in FL. Pts at high risk for tumor lysis syndrome (TLS), defined as node ≥ 8 cm and/or significant lymphocytosis, were hospitalized for initial dose. Pts received study drugs until progression or unacceptable toxicity. Response was assessed by PET-CT and bone marrow biopsy (if marrow involvement present at time of enrollment). Results: Sixteen pts were enrolled between November 2017 - May 2020. Median age was 66 years (range 50-87); 75% were male; 75% were Stage III/IV, 94% had WHO grade 1/2 FL (Table 1). FLIPI score at enrollment was 25% low risk, 44% intermediate risk, 31% high risk. Two pts were considered high risk for TLS. Pts received a median of 2 prior therapies (range 1-8); 19% were refractory to last line of therapy. Cohort enrollment was: DL0 (n=3), DL1 (n=6), DL2 (n=6), DL3 (n=1). The protocol was amended to close DL3 based on pharmacokinetic data from DL2 indicating a 1.8-fold higher mean steady-state ibrutinib plasma exposure compared to ibrutinib 560 mg monotherapy and concern for potential toxicity. Grade 3 adverse events (AE) included neutropenia (25%), thrombocytopenia (13%), lung infection (13%), upper respiratory infection (6%), neutropenic fever (6%), atrial fibrillation (6%), ALT/AST elevations (6%), mucositis (6%), failure to thrive in setting of progression (6%), abdominal pain (6%). There were no grade 4/5 AE. Grade 1/2 AE occurring in 〉 20% of pts included diarrhea (75%), nausea (63%), bruising (38%), rash (31%), headache (31%), constipation (25%), fatigue (25%). There was no evidence of clinical TLS; 19% had grade 1 hyperuricemia. The pt enrolled at DL3 had grade 1 diarrhea, grade 1 neutropenia. One dose limiting toxicity (DLT) occurred at DL1 (I 560 mg, V 400 mg): grade 3 neutropenia with fever and infection. There were no other DLTs. Therefore, DL2 (I 560 mg, V 600 mg) was determined to be the recommended phase 2 dose (RP2D). The ORR was 69% (0.413, 0.890); CR 25% (0.073, 0.524). The ORR at the RP2D was 83% (CR 33%). Responses by dose level are listed in Table 2. The regimen demonstrated activity in the bone marrow; 2 pts had eradication of involvement and 1 had a decrease from 60% to 0.5% by flow cytometry. Response by lines of prior therapy: 1 (86%, 6/7), 〉 2 (56%, 5/9). Most pts (91%) had a response by time of first assessment (12 weeks). The median progression-free survival (PFS) was 8.3 months (5.6 months, NA) (Figure 1). Of note, 2 responding pts chose to withdraw from study due to travel and were censored in the PFS analysis at time of discontinuation. One remained in a CR at least 9 months after study withdrawal as documented by PET-CT performed off protocol. No pts discontinued due to toxicity. Conclusion: In the first clinical trial to combine a BTK inhibitor and a BCL-2 inhibitor in relapsed/refractory FL, we found the I-V doublet to demonstrate a toxicity profile similar to that seen in mantle cell lymphoma and CLL. While our sample size is small, there was no evidence of clinical TLS, despite omission of the V ramp up. Preliminary results of anti-tumor activity are encouraging and further evaluation at the RP2D (I 560 mg, V 600 mg) is ongoing in the phase II trial. The combination of ibrutinib and venetoclax may provide an effective option for FL, utilizing a targeted approach distinct from other novel agents currently approved for this malignancy. Disclosures Ujjani: Verastem Oncology: Consultancy, Honoraria; Genentech: Consultancy, Honoraria; MorphoSys: Consultancy; Atara: Consultancy, Honoraria; AstraZeneca: Consultancy, Honoraria, Research Funding; Abbvie: Consultancy, Honoraria, Research Funding; Gilead/Kite: Consultancy, Research Funding. Lai:Agios: Consultancy; Macrogenics: Consultancy; Astellas: Speakers Bureau; Jazz: Speakers Bureau; Abbvie: Consultancy. Leslie:Seattle Genetics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; KitePharma: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; BMS: Speakers Bureau; Celgene: Speakers Bureau; BeiGene: Honoraria, Speakers Bureau; Pharmacyclics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; AstraZeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Epizyme: Honoraria, Speakers Bureau; Karyopharm: Honoraria, Speakers Bureau; Bayer: Consultancy, Membership on an entity's Board of Directors or advisory committees; ADC therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees; TG Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees. Broome:sanofi: Honoraria; argenx: Honoraria; apellis: Honoraria; Alexion: Honoraria. Gopal:IgM bio, BMS, merck: Research Funding; imab bio, takeda,astrazeneca,gilead: Research Funding; Seattle Genetics; Janssen; IMab Bio; TG Therapeutics; Astra Zeneca; Merck; Gilead; ADC Therapeutics; Nurix; TG therapeutics, Cellectar; Actinium: Consultancy; Seattle Genetics; Janssen; Takeda; IgM Bio; IMab Bio; BMS; Astra Zeneca; Merck; Gilead: Research Funding. Smith:Beigene: Consultancy; Millenium/Takeda: Consultancy; AstraZeneca: Consultancy; Portola: Research Funding; Seattle Genetics: Research Funding; Pharmacyclics: Research Funding; Merck: Research Funding; Incyte: Research Funding; Ignyta: Research Funding; Genentech: Research Funding; De Novo Biopharma: Research Funding; Bristol Meyers Squibb: Research Funding; Ayala: Research Funding; Acerta Pharma BV: Research Funding; AstraZeneca: Research Funding; Bayer: Research Funding; Karyopharm: Consultancy. Till:Mustang: Patents & Royalties, Research Funding. Lynch:Morphosys: Consultancy; Takeda: Research Funding; Bayer: Research Funding; TG therapeutics: Research Funding; Incyte: Research Funding; Juno: Research Funding; Cyteir: Research Funding; Genentech: Research Funding; Rhizen: Research Funding. Shadman:Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; TG therapeutics: Research Funding; Celgene: Research Funding; Sunesis: Research Funding; Gilead: Research Funding; AstraZeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol Meyers Squibb: Consultancy, Membership on an entity's Board of Directors or advisory committees; Verastem: Consultancy, Membership on an entity's Board of Directors or advisory committees; ADC Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees; Atara Biotherapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Cellectar: Consultancy, Membership on an entity's Board of Directors or advisory committees; Pharmacyclics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Mustang Bio: Research Funding; BeiGene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Acerta Pharma: Ended employment in the past 24 months; MophoSys: Consultancy, Membership on an entity's Board of Directors or advisory committees; Sound Biologics: Consultancy, Membership on an entity's Board of Directors or advisory committees. Maloney:Novartis: Consultancy, Honoraria; Bioline Rx: Consultancy, Honoraria; Juno Therapeutics: Consultancy, Honoraria, Patents & Royalties: Patents are pending, but not issued, licensed, no royalties, no licensees., Research Funding; A2 Biotherapeutics: Consultancy, Current equity holder in publicly-traded company, Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Kite, a Gilead Company: Consultancy, Honoraria, Research Funding; Gilead Sciences: Consultancy, Honoraria; Amgen: Consultancy, Honoraria; MorphoSys: Consultancy, Honoraria; Genentech: Consultancy, Honoraria; Pharmacyclics: Consultancy, Honoraria. Cheson:TG Therapeutics: Speakers Bureau; Symbio: Membership on an entity's Board of Directors or advisory committees; Kite: Consultancy; Karyopharm: Consultancy, Membership on an entity's Board of Directors or advisory committees; Morphosys: Consultancy; Trillium: Research Funding; Abbvie: Consultancy, Research Funding; Jannsen: Consultancy; Pharmacyclics: Consultancy, Research Funding; Parexel: Consultancy; GSK: Membership on an entity's Board of Directors or advisory committees. OffLabel Disclosure: We are presenting data regarding the use of venetoclax and ibrutinib in follicular lymphoma.

Description: Introduction: Life expectancy for long-term survivors of allogeneic hematopoietic stem cell transplant (allo-HCT) is significantly lower compared to that of the age-matched general population, despite a relatively low primary disease relapse rate 〉2 years post-transplant. Long-term transplant-related complications instead account for most mortality in this patient population. These include chronic graft-versus-host disease (cGVHD), infection, organ failure, and secondary cancers. In addition, gender is emerging as a critical determinant of outcome in the immediate post-transplant setting. However, less is known regarding gender effects on outcomes of long-term survivors. We retrospectively investigated the impact of recipient gender and donor-recipient gender mismatch on outcomes of long-term survivors of allo-HCT. Methods: We performed a retrospective analysis using Duke University's Adult Blood and Marrow Transplant database, supplemented by individual patient chart review. Inclusion criteria consisted of long-term survivors of first allo-HCT, excluding syngeneic, between 1995 - 2015 for a hematologic malignancy. A long-term survivor is defined as having been alive with documented follow-up to at least five years following allo-HCT. Patient characteristics were summarized as count (%) for categorical variables and median (interquartile range) for continuous variables. Fisher's exact tests or t-tests were used to compare difference between groups. Overall survival was estimated using the Kaplan-Meier method and multivariable Cox proportional hazard model. Patients who had received cord blood allograft were excluded from donor-recipient gender survival analyses. SAS version 9.4 (SAS Institute, Cary, NC) and R 3.5.0 were used to perform statistical analyses. Results: Over this 20-year period, 1103 patients underwent allo-HCT, with 247 (22%) meeting inclusion criteria. Of these 247, males and females had similar demographic and treatment characteristics (Table 1). However, significantly more deaths after the 5-year landmark occurred in male recipients (Figure 1, p value=0.003). To estimate whether this was due to the general population-wide shorter life expectancy for males, we performed Kaplan-Meier estimates of survival for patients aged 〈 50 years at transplant and aged ≥ 50 years at transplant, with similar results for both groups (Figure 2, p value=0.006). Interestingly, donor gender did not have a significant impact on overall survival in multivariate analysis (Table 2), and differences in overall survival of donor-recipient gender pairs was driven by patient gender (Figure 3, p value =0.007). A separate multivariate analysis of interaction between donor-recipient gender pairs further demonstrated that significant differences in overall survival were due to patient gender (Table 3). In addition to patient gender, only cGVHD retained significance as a covariate with impact on overall survival in multivariate analysis (Table 2, Table 3). Men experienced slightly higher rates and increased severity of cGVHD, and a greater percentage of cGVHD-related mortality as compared to females, though these findings were not statistically significant (Table 1). Conclusion: In this long-term survival analysis of allo-HCT adult patients, one of the only to include follow-up to 15 years, our results show that women survive significantly longer than men irrespective of their age at transplant. This outcome is independent of other common pre-transplant prognostic indicators such as donor gender or performance status at transplant. Inferior survival for males is consistent with survival outcomes described in another large and fully risk annotated HCT cohort, and in solid organ transplants such as lung and kidney. Gathering evidence suggests a biologic basis for long-term gender-determined outcomes, possibly due to differing rates or severity of cGVHD or sustained alloimmune tolerance in females. Male patients should be counseled on their possible increased long-term risk. Prospective and larger studies are warranted to validate these retrospective clinical results. Disclosures Rizzieri: Gilead: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Jazz: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Seattle Genetics: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Stemline: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Kite: Honoraria, Speakers Bureau; Karyopharm: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; abbvie: Membership on an entity's Board of Directors or advisory committees; AROG: Membership on an entity's Board of Directors or advisory committees; Bayer: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Celltrion: Membership on an entity's Board of Directors or advisory committees; Mustang: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees; Teva: Membership on an entity's Board of Directors or advisory committees; Acrobiotech: Membership on an entity's Board of Directors or advisory committees.

Description: The promising activity of BET protein inhibitors (BETi’s) is compromised by adaptive or innate resistance in acute myeloid leukemia (AML). Here, modeling of BETi-persister/resistance (BETi-P/R) in human postmyeloproliferative neoplasm (post-MPN) secondary AML (sAML) cells demonstrated accessible and active chromatin in specific superenhancers/enhancers, which was associated with increased levels of nuclear β-catenin, TCF7L2, JMJD6, and c-Myc in BETi-P/R sAML cells. Following BETi treatment, c-Myc levels were rapidly restored in BETi-P/R sAML cells. CRISPR/Cas9-mediated knockout of TCF7L2 or JMJD6 reversed BETi-P/R, whereas ectopic overexpression conferred BETi-P/R in sAML cells, confirming the mechanistic role of the β-catenin–TCF7L2–JMJD6–c-Myc axis in BETi resistance. Patient-derived, post-MPN, CD34+ sAML blasts exhibiting relative resistance to BETi, as compared with sensitive sAML blasts, displayed higher messenger RNA and protein expression of TCF7L2, JMJD6, and c-Myc and following BETi washout exhibited rapid restoration of c-Myc and JMJD6. CRISPR/Cas9 knockout of TCF7L2 and JMJD6 depleted their levels, inducing loss of viability of the sAML blasts. Disruption of colocalization of nuclear β-catenin with TBL1 and TCF7L2 by the small-molecule inhibitor BC2059 combined with depletion of BRD4 by BET proteolysis-targeting chimera reduced c-Myc levels and exerted synergistic lethality in BETi-P/R sAML cells. This combination also reduced leukemia burden and improved survival of mice engrafted with BETi-P/R sAML cells or patient-derived AML blasts innately resistant to BETi. Therefore, multitargeted disruption of the β-catenin–TCF7L2–JMJD6–c-Myc axis overcomes adaptive and innate BETi resistance, exhibiting preclinical efficacy against human post-MPN sAML cells.

Description: Studies of the relationship between the gastrointestinal microbiota and outcomes in allogeneic hematopoietic stem cell transplantation (allo-HCT) have thus far largely focused on early complications, predominantly infection and acute graft-versus-host disease (GVHD). We examined the potential relationship of the microbiome with chronic GVHD (cGVHD) by analyzing stool and plasma samples collected late after allo-HCT using a case-control study design. We found lower circulating concentrations of the microbe-derived short-chain fatty acids (SCFAs) propionate and butyrate in day 100 plasma samples from patients who developed cGVHD, compared with those who remained free of this complication, in the initial case-control cohort of transplant patients and in a further cross-sectional cohort from an independent transplant center. An additional cross-sectional patient cohort from a third transplant center was analyzed; however, serum (rather than plasma) was available, and the differences in SCFAs observed in the plasma samples were not recapitulated. In sum, our findings from the primary case-control cohort and 1 of 2 cross-sectional cohorts explored suggest that the gastrointestinal microbiome may exert immunomodulatory effects in allo-HCT patients at least in part due to control of systemic concentrations of microbe-derived SCFAs.

Description: Gray platelet syndrome (GPS) is a rare recessive disorder caused by biallelic variants in NBEAL2 and characterized by bleeding symptoms, the absence of platelet α-granules, splenomegaly, and bone marrow (BM) fibrosis. Due to the rarity of GPS, it has been difficult to fully understand the pathogenic processes that lead to these clinical sequelae. To discern the spectrum of pathologic features, we performed a detailed clinical genotypic and phenotypic study of 47 patients with GPS and identified 32 new etiologic variants in NBEAL2. The GPS patient cohort exhibited known phenotypes, including macrothrombocytopenia, BM fibrosis, megakaryocyte emperipolesis of neutrophils, splenomegaly, and elevated serum vitamin B12 levels. Novel clinical phenotypes were also observed, including reduced leukocyte counts and increased presence of autoimmune disease and positive autoantibodies. There were widespread differences in the transcriptome and proteome of GPS platelets, neutrophils, monocytes, and CD4 lymphocytes. Proteins less abundant in these cells were enriched for constituents of granules, supporting a role for Nbeal2 in the function of these organelles across a wide range of blood cells. Proteomic analysis of GPS plasma showed increased levels of proteins associated with inflammation and immune response. One-quarter of plasma proteins increased in GPS are known to be synthesized outside of hematopoietic cells, predominantly in the liver. In summary, our data show that, in addition to the well-described platelet defects in GPS, there are immune defects. The abnormal immune cells may be the drivers of systemic abnormalities such as autoimmune disease.