ALBERT

Description: Morphological interpretation is the standard in diagnosing myelodysplastic syndrome (MDS), but it has limitations, such as varying reliability in pathologic evaluation and lack of integration with genetic data. Somatic events shape morphologic features, but the complexity of morphologic and genetic changes make clear associations challenging. This article interrogates novel clinical subtypes of MDS using a machine learning technique devised to identify patterns of co-occurrence among morphologic features and genomic events. We sequenced 1,079 MDS patients and analyzed bone marrow morphological alterations and other clinical features. A total of 1,929 somatic mutations were identified. Five distinct morphologic profiles with unique clinical characteristics were defined. 77% of higher-risk patients clustered in profile-1. All lower-risk patients clustered into the remaining 4 profiles: profile-2 was characterized by pancytopenia, profile-3 by monocytosis, profile-4 by elevated megakaryocytes, and profile-5 by erythroid dysplasia. These profiles could also separate patients with different prognosis. Lower-risk MDS patients were classified into eight genetic signatures (e.g. signature-A had TET2 mutations, signature-B had both TET2 and SRSF2 mutations and signature-G had SF3B1 mutations) demonstrating association with specific morphologic profiles. Six morphologic profiles/genetic signatures' associations were confirmed in a separate analysis of an independent cohort. Our study demonstrates that non-random or even pathognomonic relationships between morphology and genotype to define clinical features can be identified. This is the first comprehensive implementation of machine learning algorithms to elucidate potential intrinsic interdependencies among genetic lesions, morphologies, and clinical prognostic in attributes of MDS.

Description: Morphology has been the pillars of the MDS diagnosis. In the genetic era, the subjectivity of pathologic evaluation in the absence of integration with mechanistic correlations is a clear limitation of current diagnostic schemes. Well-known genotype/phenotype associations, including e.g., linking of SF3B1 mutations to RARS or JAK2/SF3B1 mutations to RARS-T, suggest that somatic events can shape morphologic features and vice versa. However, the complexity of morphologic and genetic changes precludes identification of many consequential genotype/phenotype associations. As objective image analytic tools are being developed, this project aims at determining the most mutually predictive relationships between combinations of morphologic features and genomic changes. We have analyzed 1,079 MDS patients for somatic CNVs and mutations in a targeted panel of 33 genes frequently mutated in myeloid cancers. Our stringent bioanalytic pipeline removed artifacts, SNPs and errors. We applied this pipeline to discovery (2/3) and validation (1/3) cohorts. Bone marrow morphological alterations were mapped to binary features called by an independent pathologist in a blinded fashion based on uniformly defined criteria occurring in 〉10% of cells. A total of 10 such features were investigated. For instance, myeloid, erythroid and megakaryocytic dysplasia occurred in 54%, 70% and 72% of patients, respectively. 89% had at least one cytopenia, 57% multiple cytopenias, and 50% had at least one myeloproliferative feature (e.g., monocytosis). In addition, all cases were partitioned in accordance with two risk groups defined by IPSS-R (lower risk 3.5). NGS analysis identified 1,929 somatic mutations, but for proper correlation with morphologic definitions only mutations with a clonal burden 〉10% were used. Our initial univariate analysis yielded 52 significant associations (q

Description: The functional or genetic inactivation of p53 hampers human tumor treatment. Therefore, novel therapeutic strategies are needed. ONC201 is a p53-independent inducer of apoptosis that is the founding member of the imipridone class of novel anti-cancer compounds, which possess a unique pharmacophore. We discovered that ONC201 exerts anti-tumor effects via ATF4 induction through activation of an atypical integrated stress response (ISR) (Ishizawa et al. and Kline et al, Sci Signal, 2016). Several clinical trials of ONC201 are ongoing in advanced cancers, showing a promising safety profiling and signs of clinical activity in both solid tumors and hematopoietic malignancies. In this study, we investigated the effects of ONC212, which has emerged as a highly potent member of the imipridone family, in preclinical models of hematological malignancies. ONC212 exerted potent and prominent apoptogenic effects on acute myeloid leukemia (AML) and mantle cell lymphoma (MCL) cell lines (e.g., ED50s of 141.0 nM in p53 wild-type OCI-AML3 cells, 105.7 nM in MOLM13 cells, and 265.2 nM in p53-null JeKo-1 cell lines). Time course analysis of apoptosis in OCI-AML3 cells showed that ONC212 takes more than 36 hours to start to induce apoptosis, which is similar to observations with ONC201. Next, we further examined similarities between ONC212 and ONC201 by evaluating the in vitro efficacy of ONC212 in ONC201-resistant (ONC201-R) cell lines that we have generated by chronic exposure of MCL and AML cell lines to ONC201, of which ED50s for ONC201 treatment at 72 hrs were all 〉 5 μM. Interestingly, the ONC201-R cell lines were more resistant to ONC212 than the isogenic ONC201-naïve cells (Figure 1), indicating that these cell lines are cross-resistant to ONC212. We previously proved that increased protein translation of the transcription factor ATF4 is one of the major molecular events involved in ONC201-induced apoptosis (Ishizawa et al., Sci Signal, 2016). Similarly, ATF4 protein abundance was increased by 24-hour treatment with ONC212. DDIT3 (CHOP) gene, a target of ATF4, was transcriptionally upregulated in parallel with its target genes GADD34, DR5 and TRIB3 in ONC212-treated JeKo-1 and OCI-AML3 cells by 24 hrs after treatment (Figure 2). Of note, ONC201 was reported to transcriptionally induce TRAIL in a p53-independent manner in solid tumors (Allen et al., Sci Transl Med, 2013), but it was not operational in hematological cell lines (Ishizawa et al., Sci Sig 2016). Consistently, we also confirmed that ONC212 does not increase TRAIL mRNA in MCL (JeKo-1) and AML (OCI-AML3) cells. BCL-2 is a protective factor for cells under endoplasmic reticulum stress, which is one way to activate ISR. Therefore, we investigated whether the BCL-2 inhibitor ABT-199 sensitizes hematopoietic malignant cells to ONC212. Apoptosis was significantly higher in the combination than either drug alone in MCL and AML cell lines even in THP-1 and OCI-AML3 cells that are relatively resistant to ONC201/212 and/or ABT-199 (Figure 3), suggesting that this combination could overcome the resistance to either of agents. Indeed, the combination was also synergistic in the OCI-AML3 ONC201-R cell line (Figure 3). Taken together, our preclinical studies suggest that ONC212 is a promising and potent new member of the impridone class of anti-cancer compounds that warrants further development in hematological malignancies. The combination of ONC212 with ABT-199 is attractive, considering that acquired resistance after a short-term response remains a clinical challenge with ABT-199. Disclosures Konopleva: AbbVie: Research Funding; Genentech: Research Funding. Allen:Oncoceutics Inc.: Employment. Stogniew:Oncoceutics Inc.: Employment, Equity Ownership. Andreeff:Oncoceutics Inc.: Membership on an entity's Board of Directors or advisory committees.

Description: Background: Younger patients (pts) with acute myeloid leukemia (AML) who enter a remission after intensive induction chemotherapy routinely receive at least one cycle of consolidation therapy with high dose cytarabine (HiDAC). This is commonly administered over a five-day inpatient stay, after which pts are discharged home as their blood counts nadir. It is thus a natural consequence of therapy that readmission for febrile neutropenia (FN) occurs, which can impact measures of quality and value in this population. Precise descriptions of incidence, type, and severity of infection, if identified, are lacking, and thus it is unknown to what standard cancer centers should be held for anticipated readmission. We measured these rates, and attempted to identify predictive factors for readmission. Methods: Adult AML pts ≥ 18 years of age who received at least one cycle of HiDAC consolidation (1000-3000 mg/m2 for six doses) in 2009-2019 were included. Our primary aim was to identify predictive factors for readmission after the first cycle of consolidation chemotherapy. The following pt characteristics and co-morbid conditions were analyzed: age, gender, body mass index (BMI), smoking status, AML cytogenetic risk status, history of diabetes, peripheral vascular disease, cardiovascular disease, chronic pulmonary disease, hepatic impairment, and other cancers. Secondary aims included: estimating rates of all-cause readmissions among all HiDAC cycles, defining the rate of FN readmissions, estimating rates of intensive care unit (ICU) admissions, clinical (e.g., probable pneumonia per imaging) and microbiologically-documented infections, prophylactic (ppx) medications used, and mortality. Statistical analyses interrogated potential risk factors for evidence of association with hospital readmission after the first cycle of consolidation chemotherapy. Results: We identified 182 AML pts who fit inclusion criteria. The median age was 50 years (range 19-73); 55% were female and 45% were male. Statistical analyses revealed no association with readmission after cycle 1 for cytogenetic risk (p=0.85), history of heart failure (p= 0.67), chronic pulmonary disease (p=1), connective tissue disease (p=0.53), cerebrovascular accident (p=0.63), diabetes (p=0.63), gender (p=0.07), history of lymphoma (p=0.53), other solid tumors (p=0.53), liver disease (p=1), myocardial infarction (p=0.71), peripheral vascular disease (p=1), or smoking status (p= 0.52). For 480 HiDAC cycles analyzed (88% at 3000 mg/m2), the overall readmission rate was 50% (242/480), of which 85% (205/242) were for FN. Those readmissions which were not FN were for cardiac complications (chest pain, EKG changes), non-neutropenic fevers or infections, neurotoxicity, bleeding or clotting events, or other symptoms associated with chemotherapy (nausea/vomiting, pain, etc.). Median time to FN hospital admission was 18 days (range 6-27) from the start of HiDAC. Of the 205 FN readmissions, 57% had documented infections. Of these infections, 41% were bacteremia, 23% fungal, 16% sepsis, 12% other bacterial, and 8% viral. Of 480 HiDAC cycles, ppx medications prescribed included: 92% fluoroquinolone (442/480), 81% anti-viral (389/480), 30 % anti-fungal (142/480), and 3% colony stimulating factor (14/480). Only 7% (14/205) of FN readmissions resulted in an ICU admission, and 1% (3/205) resulted in death. Conclusions: Approximately half of patients treated with consolidation therapy following intensive induction therapy can be expected to be readmitted to the hospital. The majority of FN readmissions were associated with clinical or microbiologically documented infections and are not avoidable, however ICU admission and death associated with these complications are rare. Readmission of AML pts following HiDAC is expected, and therefore, should be excluded from measures of value and quality. Disclosures Waldron: Amgen: Consultancy. Hobbs:Amgen: Research Funding; SimulStat Inc.: Consultancy. Advani:Macrogenics: Research Funding; Abbvie: Research Funding; Kite Pharmaceuticals: Consultancy; Pfizer: Honoraria, Research Funding; Amgen: Research Funding; Glycomimetics: Consultancy, Research Funding. Nazha:Incyte: Speakers Bureau; Abbvie: Consultancy; Daiichi Sankyo: Consultancy; Jazz Pharmacutical: Research Funding; Novartis: Speakers Bureau; MEI: Other: Data monitoring Committee; Tolero, Karyopharma: Honoraria. Gerds:Imago Biosciences: Research Funding; Roche: Research Funding; Celgene Corporation: Consultancy, Research Funding; Pfizer: Consultancy; CTI Biopharma: Consultancy, Research Funding; Incyte: Consultancy, Research Funding; Sierra Oncology: Research Funding. Sekeres:Syros: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Millenium: Membership on an entity's Board of Directors or advisory committees. Mukherjee:Partnership for Health Analytic Research, LLC (PHAR, LLC): Consultancy; McGraw Hill Hematology Oncology Board Review: Other: Editor; Projects in Knowledge: Honoraria; Celgene Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Honoraria; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol-Myers Squibb: Speakers Bureau; Takeda: Membership on an entity's Board of Directors or advisory committees.

Description: Cellular stress response has dual aspects; cell-protective or lethal. Mitochondria have their unique organellar response termed "mitochondrial unfolded protein response (UPRmt)" induced by damaged mitochondrial (mt) matrix proteins. While recent discoveries have successfully targeted BCL2, a regulator of mt integrity in acute myeloid leukemia (AML), the significance of UPRmt is unknown. We hypothesized that priming UPRmt towards cell death would be a novel therapeutic strategy for AML. UPRmt is generally induced by dysregulation of mt protein pools. Therefore, to test if UPRmt signaling is also operational in AML cells, we selected classical or putative UPRmt inducers; the mt translation inhibitors tetracycline and tigecycline, the mt protein transport inhibitor MitoBlock6, and the mtDNA damaging agent ethidium bromide. In OCI-AML3 and HL60 cells, these agents indeed induced the transcription factor ATF5, which was reported as a central inducer of UPRmt, and its targets (e.g., LonP, HSPA9), triggering apoptosis in AML cells. In addition, we here report imipridones (ONC201 and ONC212), the activators of mt protein degradation, as novel UPRmt inducers. We recently reported that imipridones non-covalently bind the mt protease ClpP and allosterically activate it. They induced prominent apoptosis in primary AML progenitor and leukemia initiating cells (LICs) in vitro and in vivo, but not in normal bone marrow cells, following "mitochondrial proteolysis" with reduction of selective mt matrix proteins (e.g., SDHB, NDUFA12) and resultant inhibition of oxidative phosphorylation (Oxphos) (Ishizawa, Zarabi et al, Cancer Cell 2019). We then postulated that dysregulation of mt protein pools by mitochondrial proteolysis can also induce UPRmt. Indeed, our gene expression profiles of ONC201-treated Z138 and Jeko-1 cells were highly enriched for previously published UPRmt gene signatures, and UPRmt effectors were induced also in AML cells. Of potentially high clinical significance is the finding of synergistic anti-leukemia effects of imipridones when combined with the selective BCL2 inhibitor venetoclax, in vitro and in vivo (Ishizawa et al. Science Signaling 2016, and Nii et al. Blood 2019). However, its underlying molecular mechanism is unclear. Since BCL2 is reported to be induced by UPRmt, we hypothesized that BCL2 is critical for the ClpP-mediated UPRmt to have the cell protective effects, contrary to lethal effects, as dual aspects of stress response. We utilized the tetracycline-inducible system of an activated mutant (Y118A) form of ClpP in OCI-AML3 cells, and demonstrated that venetoclax treatment sensitizes OCI-AML3 cells to genetic activation of ClpP towards apoptosis. Furthermore, other UPRmt inducers (tetracycline, tigecycline, and MitoBlock6) in combination with venetoclax also synergistically induced apoptosis in AML cells, suggesting that BCL2 inhibition and UPRmt induction generally exerts synergistic anti-leukemia effects. We next focused on the enhanced effect observed for the combination of imipridones with venetoclax as compared to other UPRmt inducers, searching for other targets that could further enhance the synergy. We then hypothesized that the synergism between ClpP activation and BCL2 inhibition involves SDHB, a respiratory chain complex II subunit degraded by activated ClpP but not targeted by any of other UPRmt inducers. Consistently, SDHB knockdown sensitized OCI-AML3 cells to venetoclax-induced apoptosis, indicating that SDHB reduction and UPRmt by ClpP activation concomitantly enhance the cell lethality by BCL2 inhibition. Collectively, UPRmt is a new potential therapeutic target for AML, which significantly enhances the cell death effects of BCL2 inhibition on AML cells. In particular, ClpP activation induces UPRmt and, concomitantly, downregulates SDHB, thus targeting the respiratory chain complex II, which results in improved synergistic leukemia cell apoptosis when combined with BCL2 inhibition. Oxphos is also a hallmark of drug resistant AML stem cells, which supports the notion that Oxphos inhibition by this combination targets LICs. Based on promising preclinical anti-tumor efficacy, ONC201 as a single agent is being evaluated in early phase clinical trials, showing clinical responses in AML and midline gliomas. A clinical trial testing the combinatorial strategy of targeting ClpP and Bcl-2 is under development. Disclosures Borthakur: Novartis: Research Funding; NKarta: Consultancy; Eisai: Research Funding; Oncoceutics: Research Funding; BioLine Rx: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Cyclacel: Research Funding; Strategia Therapeutics: Research Funding; Eli Lilly and Co.: Research Funding; Arvinas: Research Funding; Merck: Research Funding; AstraZeneca: Research Funding; PTC Therapeutics: Consultancy; Agensys: Research Funding; Argenx: Membership on an entity's Board of Directors or advisory committees; FTC Therapeutics: Membership on an entity's Board of Directors or advisory committees; GSK: Research Funding; Incyte: Research Funding; Janssen: Research Funding; AbbVie: Research Funding; BMS: Research Funding; Oncoceutics, Inc.: Research Funding; Bayer Healthcare AG: Research Funding; BioTheryX: Membership on an entity's Board of Directors or advisory committees; Tetralogic Pharmaceuticals: Research Funding; Cantargia AB: Research Funding; Polaris: Research Funding; Xbiotech USA: Research Funding. Stogniew:Oncoceutics, Inc.: Employment. Oster:Oncoceutics, Inc.: Employment. Kantarjian:BMS: Research Funding; AbbVie: Honoraria, Research Funding; Ariad: Research Funding; Amgen: Honoraria, Research Funding; Jazz Pharma: Research Funding; Pfizer: Honoraria, Research Funding; Cyclacel: Research Funding; Immunogen: Research Funding; Agios: Honoraria, Research Funding; Actinium: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Research Funding; Takeda: Honoraria; Astex: Research Funding; Daiichi-Sankyo: Research Funding. Schimmer:Novartis Pharmaceuticals: Consultancy; Medivir Pharmaceuticals: Research Funding; Jazz Pharmaceuticals: Consultancy; Otsuka Pharmaceuticals: Consultancy. Andreeff:Eutropics: Equity Ownership; Daiichi Sankyo, Inc.: Consultancy, Patents & Royalties: Patents licensed, royalty bearing, Research Funding; Jazz Pharmaceuticals: Consultancy; Celgene: Consultancy; Aptose: Equity Ownership; Reata: Equity Ownership; 6 Dimensions Capital: Consultancy; AstaZeneca: Consultancy; Amgen: Consultancy; Breast Cancer Research Foundation: Research Funding; CPRIT: Research Funding; NIH/NCI: Research Funding; Center for Drug Research & Development: Membership on an entity's Board of Directors or advisory committees; Cancer UK: Membership on an entity's Board of Directors or advisory committees; NCI-CTEP: Membership on an entity's Board of Directors or advisory committees; German Research Council: Membership on an entity's Board of Directors or advisory committees; Leukemia Lymphoma Society: Membership on an entity's Board of Directors or advisory committees; NCI-RDCRN (Rare Disease Cliln Network): Membership on an entity's Board of Directors or advisory committees; CLL Foundation: Membership on an entity's Board of Directors or advisory committees; BiolineRx: Membership on an entity's Board of Directors or advisory committees; Oncolyze: Equity Ownership; Oncoceutics: Equity Ownership; Senti Bio: Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Ishizawa:Daiichi Sankyo: Patents & Royalties: Joint submission with Daiichi Sankyo for a PTC patent titled "Predictive Gene Signature in Acute Myeloid Leukemia for Therapy with the MDM2 Inhibitor DS-3032b," United States, 62/245667, 10/23/2015, Filed.

Description: Key Points FZR1 loss causes increased sensitivity of B-ALL cells to oncogene- or chemotherapy-induced DNA damage. Prolonged loss of FZR1 contributes to the development of treatment-resistant clones in mouse and human B-ALL.

Description: Background: CD123 is frequently expressed in hematologic malignancies including AML. CD123 has been a potential immunotherapeutic target in AML due to its association with leukemic stem cells that play an essential role in disease progression and relapse. Our previous study using T-cells secreting CD123/CD3-bispecific T-cell engagers (BiTEs) (CD123-ENG T-cells) has shown activity in preclinical studies, recognizing and killing acute myeloid leukemia (AML) blasts in vitro and in vivo. CD123-ENG T-cells secrete bispecific molecules that recognize CD3 (T-cells) and CD123 (AML blasts), and are able to direct transduced T-cells and recruit bystander T-cells to kill CD123-positive blasts. Venetoclax is a BCL-2 inhibitor that can restore functional apoptosis signaling in AML cells, and has been FDA approved for the treatment of AML patients in combination with hypomethylating agents. To improve the efficacy of CD123-ENG T-cells we explored efficacy in AML by combining targeted immunotherapy (CD123-ENG T cells) with targeted inhibition of anti-apoptotic BCL-2 (venetoclax) in vitro and in vivo models of AML. Methods : CD123-ENG T-cells were generated by retroviral transduction and in vitro expansion. Non-transduced (NT) T-cells served as control. In vitro, GFP+ MOLM-13 AML cells were pretreated with venetoclax (0, 10µM, and 20µM) for 24 hours prior to co-culture with CD123-ENG or NT T-cells at an effector/target ratio of 1:10. After 16 hours, MOLM-13 AML cells were analyzed by flow cytometry and quantitated using counting beads; cytotoxicity was calculated relative to untreated MOLM-13 control. The anti-AML activity of the combination was further evaluated in a MOLM-13-luciferase xenograft AML mouse model. Leukemia progression was assessed by bioluminescence imaging. The frequency of MOLM13 AML and human T cells in periphera blod (PB) was determined by flow cytometry. Results: In vitro, we demonstrated that pretreatment of Molm13 AML cells with venetoclax enhanced the cytolytic activity of CD123-ENG T-cells compared to NT- or no T-cell controls. Interestingly, venetoclax sensitized Molm13 to CD123-ENG T-cell killing in a dose-dependent manner (Fig.1; 50%/31% killing by CD123-ENG T-cells versus 27%/14% of killing by NT T cells post pretreatment with 10µM or 20µM ventoclax, p