ALBERT

Description: Key Points Checkpoint blockade via anti–PD-1 mAbs was associated with a high overall response rate in relapsed Hodgkin lymphoma allo-HCT patients. Checkpoint blockade via anti–PD-1 mAbs after allo-HCT can be complicated by rapid onset of severe and treatment-refractory GVHD.

Description: Previous reports suggest that peripheral blood counts are strongly influenced by environmental and genetic determinants; however few of the genetic factors that regulate these quantitative traits have been discovered. We analyzed CBC data from 395 samples collected from an 854-member Amish pedigree with von Willebrand disease. 71 individuals of the pedigree are heterozygous for a missense mutation at position 4120, represented by a single base substitution (C〉T) that predicts an arginine to cysteine change at position 1374 (R1374C) in the A1 domain of the mature von Willebrand factor molecule. The detection of genetic signals is likely to be enhanced in groups that live in a more homogeneous environment like the Amish. Linear and quadratic age effect accounted for varying proportions of the gender-specific variation in the CBC measures (from 1% to 70%). The variance component associated with additive polygenic effects was estimated for each CBC phenotype using MENDEL to obtain estimates of heritability. Significant heritability was found for platelet (h2= 0.518, p

Description: Von Willebrand Disease (VWD) is a common and highly variable bleeding disorder with incomplete penetrance and variable expressivity. Due to the complexity of VWD it is highly likely that several modifier genes influence its phenotype. We have identified and characterized an extensive Amish pedigree which is composed of 854 individuals of whom 395 have provided blood samples. 71 pedigree members are heterozygous for a missense mutation at position 4120, represented by a single base substitution (C〉T) that predicts an arginine to cysteine change at position 1374 (R1374C) in the A1 domain of the mature VWF molecule. Phenotypic characterization of the pedigree included several laboratory measures and a clinical bleeding phenotype determined by a validated bleeding score. The bleeding score was established by a questionnaire that consists of 10 questions that focus on seven areas related to spontaneous and trauma-related mucocutaneous bleeding. The lowest possible score is 0 and the highest is 7. A bleeding score of 3 or higher exhibited a specificity of 0.99 and sensitivity of 0.90 for VWD when given to 97 healthy controls and 63 unrelated individuals diagnosed with VWD. We performed association studies with candidate genes in the pedigree and correlated genotype and allele frequencies with severity of bleeding. The list of candidate variants studied included a total of 27 genetic polymorphisms in 20 genes that encode for the platelet receptors ITGA2, GP1BA, ITGB3 and GP6, for the coagulation factors F2, F5, F7, and F13 and for critical proteins involved in normal hemostasis, thrombosis, fibrinolysis and inflammation such as THBD, SELP, SELE, FGA, FGB, MTHFR, PLAT, PECAM1, CPB2, NOS3, SERPINE and CD14. These gene variants were selected based on their biological significance as well as their minor allele frequency. The differences in genotype distributions and allele frequencies between affected and unaffected pedigree members were evaluated using a proportional odds model for the association analysis. This model, which is a generalization of the usual logistic regression model for a binary outcome, can accommodate a multi-level ordered response variable such as the bleeding score. Five alleles were associated with increased severity of bleeding: Allele C at position 2361 of the SELP gene that encodes for P selectin and has been associated with decreased risk of thrombosis (p= 0.09 and p=0.003 and 0.001 when corrected for mutation status and sex, respectively). Allele C, at position 807 of ITGA2 that has been associated with increased bleeding (p= 0.01, and p= 0.002 and 0.01 when adjusted for mutation status and sex, respectively). Allele G, at position - 260 of CD14 that has been associated with decreased risk for myocardial infarction (p= 0.003, and p = 0.02 and 0.003 when adjusted for mutation status and sex, respectively). Allele T at position 13254 of the GP6 gene that has been associated with bleeding and thrombosis (p= 0.009, and p= 0.05 and 0.01 when adjusted for mutation status and sex, respectively). Finally, allele C at position 677 that has been described as protective for thrombosis (p= 0.02, and p= 0.01 and 0.018 when adjusted for mutation status and sex, respectively). In summary, our results demonstrate that specific genetic variants modify the bleeding phenotype of this pedigree and may contribute to the clinical variability observed in unrelated patients with VWD.

Description: Effective targeting of the acute myeloid leukemia (AML) leukemia stem cell (LSC) population may allow for deep, durable remissions and curative potential. In older, newly diagnosed AML patients who are not candidates for induction, venetoclax + azacitidine (aza) targets specific metabolic vulnerabilities of LSCs, resulting in very promising clinical outcomes. In our single institution experience treating 45 previously untreated AML patients with venetoclax + aza both in the context of the multi-institutional study NCT02203773 (N=33) and with off-label use (N=12), 36/45 (80%) achieved a complete remission (CR) or CR with incomplete count recovery (CRi). In the relapsed/refractory (R/R) setting, the efficacy of venetoclax + aza has been reported to be significantly worse. In our single-institution off-label experience (N=7), only 1/7 (14%) R/R patients had a CR/CRi (p=0.005 compared to the untreated group). R/R and untreated patients had similar baseline characteristics, although more R/R patients had an antecedent hematological disorder (Table 1). Multivariate analysis showed cytogenetic risk and R/R disease as the sole predictors of response to venetoclax + aza (Table 2). In light of existing data regarding biological changes that occur in LSCs after treatment and subsequent relapse, we aimed to determine whether laboratory analysis of LSCs from patients treated with venetoclax + aza would show differential sensitivity to this therapy in the up-front vs R/R setting that could help to explain the different clinical activity. We have previously shown that LSCs from untreated patients are uniquely reliant on oxidative phosphorylation (OXPHOS), and that venetoclax + aza targets LSCs by decreasing OXPHOS. Therefore, we tested the hypothesis that inferior responses of R/R patients to venetoclax + aza are due to changes in OXPHOS regulation in relapsed LSCs. LSCs were defined as cells bearing relatively low levels of reactive oxygen species (ROS-low), an effective means of enriching primary human LSCs. We found that in contrast with untreated patients, venetoclax + aza does not decrease viability or OXPHOS in LSCs from R/R patients (Fig1). Furthermore, R/R LSCs had altered fatty acid metabolism that contributed to these OXPHOS differences, with increased flux of fatty acids into the TCA cycle (Fig 2). In addition, R/R samples compensated for the metabolic perturbations that occurred upon exposure to venetoclax + aza through upregulation of fatty acid uptake and metabolism into the TCA cycle (Fig 3). Fatty acid metabolism is controlled by multiple genes and pathways. Integral to its activity is the gene Carnitine Palmitolytransferase 1 (CPT1), due to its pivotal role in the beta-oxidation of long chain fatty acids. Investigation of the Cancer Genome Atlas AML dataset reveals higher expression of CPT1 leads to significantly worse overall survival, suggesting increased fatty acid metabolism may drive a more resistant LSC population in R/R AML patients. We also found elevated baseline levels of CPT1 in patients who progressed on venetoclax + aza compared to those that had long term remissions (not shown). Therefore we utilized the CPT1 inhibitor etomoxir to block fatty acid metabolism. We found addition of etomoxir to cultures of R/R LSCs rescued the ability of venetoclax + aza to decrease OXPHOS and re-sensitized R/R LSCs to venetoclax + aza (Fig 4). To prove that this novel regimen targets functionally-defined R/R LSCs we performed ex vivo treatment followed by xenotransplantation of R/R patient specimens, which showed that upon etomoxir addition, engraftment potential is significantly decreased over venetoclax + aza alone (not shown). Therefore we propose a novel mechanism for the increased resistance of R/R AML patients to venetoclax + aza involving altered energy metabolism. We find increased fatty acid metabolism in R/R patient specimens, and targeting this pathway using the CPT1 inhibitor etomoxir leads to sensitization to venetoclax + aza and rescued targeting of OXPHOS, allowing for LSC eradication. Disclosures Pollyea: Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Argenx: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Research Funding; Agios: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead: Consultancy; Celyad: Consultancy, Membership on an entity's Board of Directors or advisory committees; Curis: Membership on an entity's Board of Directors or advisory committees; Karyopharm: Membership on an entity's Board of Directors or advisory committees.

Description: Previous studies have demonstrated the importance of energy metabolism as it relates to numerous aspects of leukemia stem cell (LSC) biology. Specifically, in acute myelogenous leukemia (AML), it has been shown that LSCs have a unique reliance on oxidative phosphorylation (OXPHOS), and that inhibition of B-cell lymphoma 2 (BCL-2) acts to down-regulate OXPHOS and eradicate LSCs in pre-clinical models. In the clinical setting, when BCL-2 inhibitor venetoclax is combined with azacitidine, high response rates (~80%) in elderly de novo AML patients have been observed (PMID: 29339097). Nonetheless, a significant portion of these patients ultimately experience disease progression. The mechanism of resistance and characteristics of these patients is poorly understood. Our preliminary data shows that the venetoclax + azacitidine (ven/aza) regimen targets LSCs through alteration of energy metabolism. Specifically, the regimen disrupts the TCA cycle leading to decreases in ATP production and inhibition of OXPHOS. This metabolic targeting is central to ven/aza efficacy and we hypothesize that resistance and progression of AML patients is due to compensatory mechanisms that restore sufficient levels of OXPHOS (PMID:3333149). Investigation of such mechanisms led us to explore the potential activity of MCL1. Previous studies have shown MCL1 can influence venetoclax resistance, however little is known about MCL1's role in metabolism, although a report in breast cancer cells suggests MCL1 modulates OXPHOS (PMID: 28978427). In leukemia, MCL1 expression has been shown to be partially upregulated through mutations in protein tyrosine phosphatase non- receptor type 11 (PTPN11), and PTPN11 mutations have been shown to increase LSC frequency. Thus, we hypothesized that PTPN11 mutations may confer resistance to venetoclax-based regimens at least partially by up-regulation of MCL1. To test this hypothesis, we investigated the relationship between PTPN11 mutations, MCL1, and the metabolic phenotype. In comparison to specimens with a wild type allele, LSCs isolated from PTPN11 mutant patient specimens showed increased levels of OXPHOS as well as glycolysis, amino acids, and fatty acids, suggesting an ability to utilize multiple energy sources for survival. PTPN11 mutant specimens also show decreased sensitivity to venetoclax, suggesting OXPHOS is not affected by venetoclax to the same degree as PTPN11 wild type specimens (fig. 1). Furthermore, when we introduced a mutated allele of PTPN11 into a primary AML specimen we observed increased oxidative phosphorylation and glycolysis, which correlated with decreased in vitro sensitivity to venetoclax (fig. 2). To test the potential role of MCL1 in PTPN11 mutant specimens, we employed a small molecule MCL-1 inhibitor. Metabolic analysis of specimens treated with the MCL-1 inhibitor showed decreased OXPHOS in PTPN11 mutant specimens (fig 3). Further, PTPN11 mutant specimens exhibit increased sensitivity to the MCL-1 inhibitor (fig. 4). To investigate a potential mechanistic link to clinical observations, we next examined 45 older, previously untreated AML patients from our institution who received ven/aza, both in the context of the multi-institutional study NCT02203773 (N=33) and with off-label use (N=12). Of 12 variables examined, only the presence of PTPN11 predicted shorter response duration (table 1). In addition, of the 9 patients who progressed ven/aza, 2 (22%) acquired PTPN11 mutations upon progression, further suggesting PTPN11 may represent a resistance mechanism to this regimen. Notably, PTPN11 is not preferentially detected in patients who progress after regimens other than ven/aza. In conclusion, AML containing PTPN11 mutations exhibit a unique energy metabolism profile. These specimens also appear to have increased sensitivity to MCL-1 inhibitors. The presence of PTPN11 mutations represents both a novel method for predicting response to ven/aza and a potential strategy for targeting patients who progress. We propose that addition of an MCL-1 inhibitor for treatment of AML patients bearing PTPN11 or related mutations may increase therapeutic responses. Disclosures Savona: Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte: Membership on an entity's Board of Directors or advisory committees, Research Funding; Boehringer Ingelheim: Consultancy. Fesik:Boehringer Ingelheim: Consultancy. Pollyea:Celyad: Consultancy, Membership on an entity's Board of Directors or advisory committees; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Curis: Membership on an entity's Board of Directors or advisory committees; Agios: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Membership on an entity's Board of Directors or advisory committees; AbbVie: Consultancy, Research Funding; Karyopharm: Membership on an entity's Board of Directors or advisory committees; Argenx: Consultancy, Membership on an entity's Board of Directors or advisory committees; Gilead: Consultancy.

Description: Background: Venetoclax (ven) was approved for older untreated acute myeloid leukemia (AML) patients due to high response rates and durable remissions. As a participating site in the dose escalation study, we observed deeper/more durable responses in some who received 〉400mg ven. We also noted 16/33 discontinued azacitidine (aza) after achieving a response; 9 relapsed and 7 remained in long term remission on ven only. Based on these observations, we designed a study that hypothesized: A)Higher initial doses of ven would allow deeper/more durable responses, and B)Multi modality high sensitivity measurable residual disease (MRD) testing could identify patients able to discontinue aza and remain on maintenance ven. Methods: This is an ongoing phase 2 study (NCT03466294) of 42 untreated AML patients ≥60 who decline/are ineligible for induction. Patients have adequate organ function and white blood cell counts grade 2 related AEs were leukopenia (53%), thrombocytopenia (44%) and neutropenia (35%); there were no related grade 5 AEs. The overall response rate was 70% (21/30; CR=19, MLFS=2). Median number of cycles to achieve best response was 1. Significant blast reductions were seen on day 8; of the 28 with interpretable day 8 BMBXs, 10 achieved MLFS on day 8. 4 completed ≥1 cycle and were refractory. An additional 4 did not complete cycle 1: 1 died of disease and 3 elected to come off therapy (all subsequently died of disease). Four (19%) responders relapsed, after a median 180 days (27-279). With median follow up of 214 days, median response duration has not been reached. 10 patients died, after a median 65 days (29-256); 1/30 died within 30 days. Median overall survival has not been reached. Of the 26 who completed ≥1 cycle, 19 were MRD- by MPFC, including 18/19 who achieved CR. Of these 26, 3 were not monitored by ddPCR: for 2 patients this was due to the absence of detectable baseline mutations and for 1 patient it was due to refractory disease. The remaining 23 had ddPCR monitoring; 3 became MRD- by this modality (Fig 2). All 3 were also MRD- by MPFC and per protocol discontinued aza and initiated ven maintenance (Fig 1). MRD negativity by both parameters occurred after cycles 1, 2 and 3, respectively. One MRD- patient relapsed after 216 days; two remain in remission after 301 and 124 days. An additional 4 who achieved MRD+ responses discontinued aza at their insistence (and in violation of the protocol); 1 relapsed after 279 days, and 3 remain in ongoing remission. Univariate predictors of refractory disease were FAB M0/M1 (OR 0.070, p=0.02) and RAS pathway mutations (OR 14.25, p=0.02). Conclusions: Higher initial doses of ven are tolerated in this population. Blast reduction occurs quickly in many patients (day 8), for this low intensity regimen. Response rates are consistent with lower doses of ven. Very deep responses, as measured by highly sensitive MRD methods (MPFC and ddPCR are capable of sensitivity up to 0.02%), are attainable. Longer follow up time will determine if higher ven doses and MRD-driven decisions related to continuation of aza result in more durable responses. Increased maturation of blasts and RAS pathway mutations are predictors for refractory disease. Disclosures Lyle: Pfizer: Membership on an entity's Board of Directors or advisory committees; Daiichi Sankyo Incyte: Membership on an entity's Board of Directors or advisory committees; Agios: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees. Pollyea:Agios: Consultancy, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees; Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees; Gilead: 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; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Celyad: Consultancy, Membership on an entity's Board of Directors or advisory committees; Diachii Sankyo: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Forty-Seven: Consultancy, Membership on an entity's Board of Directors or advisory committees.

Description: Background: Vincristine- or vinblastine-induced peripheral neuropathy (VIPN) is a common adverse event during lymphoma treatment that can profoundly impact quality of life and survivorship. VIPN often leads to dose-reduction or discontinuation of these agents. However, there is limited data (Annals of Hematology, Utsu 2016; Annals of Hematology, Morth 2018) evaluating the impact of dose reduction or omission of vinca alkaloids on progression-free survival (PFS) or overall survival (OS). We report on a series of lymphoma patients (pts) who had vincristine (Vn) or vinblastine (Vb) dose-reduced or omitted from their initial chemotherapy. Methods: We performed a retrospective chart review of lymphoma pts who were seen in consultation at the University of Colorado from 2015 to 2018. Data included demographics, lymphoma subtypes, Vn- or Vb-containing treatment regimens, PFS, and OS. PFS and OS were examined using Kaplan-Meier method, and stratification techniques were used to compare survival based on full dose versus dose-reduced/omitted treatments. Cox regression methods were used to assess the impact of refractory or progressive disease, disease stage, and prognostic score on PFS. Results: 373 pts who received Vn- or Vb-containing chemotherapy were identified. Of those, 228 pts had complete records available for review for treatment, response and survival data. 133 pts (58.3%) were male. Median age was 56 years (range 19 to 85 years). The most common diagnoses were diffuse large B-cell (59.2%), Hodgkin (26.3%), follicular (8.3%), PTLD (3.1%), Burkitt, (2.6%), and gray zone (0.4%) lymphomas. Chemotherapy regimens administered were REPOCH (35.1%), RCHOP (32.9%), and ABVD/AVD (21.9%). Incidence of neuropathy during treatment was 36.4%. The median follow up is 23.7 months (range 2.5 to 150.7 months). We categorized patients into 2 groups based on chemotherapy dosing: full dose (Group 1: n=155/228; 68%) and dose-reduced or omitted (Group 2: n= 73/228; 32%). There was not a significant difference in stage or prognostic score between Group 1 and Group 2. Group 2 was further sub-categorized into Subgroup 2A (patients with Vn/Vb omitted at any point: n= 46/228; 20.2%) and Subgroup 2B (patients with Vn/Vb dose-reduced: n=27/228; 11.8% ). Of the 73 patients in Group 2, 67 patients had Vn/Vb omitted or reduced due to neuropathy; 6 patients had the drugs omitted (5/73) or reduced (1/73) due to GI side effects. Complete response (CR) rates between Group 1 and Group 2 were similar (81.3% vs 87.7%, respectively). Patients in Group 1 had a higher number of patients with relapsed/refractory disease compared to Group 2 (p