Acute Myeloid Leukemia Therapeutics Research Articles

Computational Evaluation of Bioactive Compounds in Vernonia Amygdalina as a Novel Therapeutics for Acute Myeloid Leukemia: Targeting the Hedgehog Pathway

AbstractThe drug resistance challenges in acute myeloid leukemia have been linked the hedgehog (HH) signaling pathway activation by many studies. Furthermore, studies have shown that the components of the HH pathway, including 12‐pass transmembrane receptor Patched (PTCH), the 7‐pass transmembrane signal transduction protein Smoothened (SMO), and all three GLI transcription factors, are expressed in AML. Thus, pharmacological inhibition of SMO in the HH signaling in these cells has anti‐leukemic effects. However, the first SMO inhibitor specific to AML (Glasdegib) has been approved based on improved overall survival rates, but alternative approaches are still required. Thus we employed druglikeness screening, molecular docking, molecular dynamics simulation, non‐equilibrium free energy and in silico pharmacokinetics profiling on bioactive compounds from Vernonia amygdalina against SMO protein target. From 30 bioactive compounds in V. amygdalina, 13 had drug‐like potential, 5 had binding energy between −9.2 to −9.9 kcal/mol. Cryptolepine, neocryptolepine and isocryptolepine with the least BE showed good stability, dynamics over 300 ns of MD run and higher binding affinities via the Jarzynski's identity. Hence, these bioactive compounds may serve as potential drugs for the treatment of acute myeloid leukemia by targeting the SMO protein after wet lab experiments.

Abstract LB026: Exploring MCT4 inhibitor Ful-23 and cytarabine synergy in acute myeloid leukemia therapeutics

Abstract In the ongoing pursuit of addressing the formidable challenge presented by Acute Myeloid Leukemia (AML) treatment, this comprehensive study delves into the intricate antileukemic effects of Ful-23, our newly developed Monocarboxylate Transporter 4 (MCT4) inhibitor. Positioned as a promising candidate, Ful-23's unique mechanisms and potential synergies with established chemotherapeutic agents, such as Cytarabine, are elucidated in this research. Intracellular alkalinization stands as a crucial factor contributing to the evasion of chemotherapy by cancer cells. It serves as the primary determinant for the heightened resistance exhibited by cancer cells against chemotherapy interventions. The investigation reveals the multifaceted impact of Ful-23 on AML cells, showcasing its capacity to induce intracellular lactate accumulation and acidification. These metabolic alterations correlate with a substantial inhibition of proliferation, as evidenced by an IC50 of approximately 0.55 μM, and a marked induction of apoptosis, particularly notable in the MV411 AML cell line. These findings underscore Ful-23's potential as a potent antileukemic agent with distinct cellular effects. A collaborative approach involves the combination of low concentrations of Ful-23 (0.312 μM) with Cytarabine (0.312 μM), revealing a significant enhancement in cytarabine-induced proliferation inhibition and apoptosis when Ful-23 is introduced concurrently. Importantly, the translational impact of these findings extends to a zebrafish model, utilizing both MV411 cells and transplanted primary AML patient-derived bone marrow cells. In the zebrafish AML model, the combination treatment of Ful-23 and Cytarabine demonstrates a significant enhancement in their respective antitumor effects compared to monotherapy, indicating a pronounced synergistic effect. This additional validation strengthens the clinical relevance of the observed synergistic antileukemic interactions between Ful-23 and Cytarabine. Essentially, this investigation advances our understanding of AML treatment dynamics, emphasizing the crucial role of MCT4 inhibition and elucidating the mechanism by which it leads to intracellular acidification in synergistic therapeutic approaches for this intricate and formidable disease. Citation Format: Yong Wu, Ke Wu, Jay Vadgama. Exploring MCT4 inhibitor Ful-23 and cytarabine synergy in acute myeloid leukemia therapeutics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB026.

Identifying Stress Granules As Determinants of Leukemia Stem Cell Maintenance and Stress Adaptation

The development of novel leukemic stem cell (LSC)-targeted therapeutics, which spare hematopoietic stem cells (HSC), is an urgent goal towards reducing the high, LSC-driven, relapse rates in acute myeloid leukemia (AML) patients. Towards elucidating key post-transcriptional networks underlying LSC maintenance we previously performed an in vivo pooled CRISPR dropout screen targeting 128 RNA binding proteins (RBPs) highly expressed in AML LSCs while having low expression in HSCs. This screen highlighted 12 RBPs crucial for murine AML LSC-mediated leukemic growth, of which there was a significant enrichment of stress granule (SG) RBPs. Further, gene set enrichment analysis shows the SG proteome is enriched in human AML LSCs vs non-LSCs while normal HSCs have reduced SG RBP expression relative to multipotent progenitor cells. SGs are dynamic biomolecular condensates, composed of ribonucleoprotein complexes, which may form as a protective mechanism against a variety of stress conditions. SGs are gaining increasing attention as contributors to cancer metastasis and chemoresistance, but their role in LSCs is virtually unexplored. The intriguing enrichment of SG RBPs in our screen hints at a larger role for SGs in LSC maintenance and/or stress resistance. To address this, we focused on the core SG nucleation RBP, G3BP1, a molecular switch that guides SG assembly. Fluorescence microscopy was first performed to describe SG dynamics in leukemia vs primitive normal cells where interestingly, a subset of patient AML cells contained G3BP1 SGs in the absence of added stress. In contrast, primitive healthy murine stem cells had reduced heat shock-induced SG formation in comparison to downstream normal progenitors. A series of shG3BP1 knockdown experiments performed in human leukemia cell lines and patient AML specimens demonstrated that shG3BP1-transduced cells showed increased apoptosis and differentiation and reduced colony forming capacity and competition in vitro compared to control cells. The same set of assays showed minimal impact of SG impairment to healthy umbilical cord blood cells. Importantly, in vivo xenotransplantation demonstrated that LSCs with G3BP1 knocked down were significantly compromised in their capacity to generate leukemic grafts whereas normal HSCs assessed in a similar manner display much less G3BP1 dependency. Towards determination of the SG mechanisms essential for bulk AML and LSC regulation, we performed matched RNA-sequencing and proteomics in G3BP1 knocked down AML cells as well as BioID in AML cell lines, primary patient AML cell subsets, and healthy hematopoietic cells to allow for the uncovering of an AML specific SG proteome. These analyses highlighted numerous potential SG mechanisms of cancer cell stress resistance and/or stem cell regulatory processes including sequestration of apoptotic factors and oncogenic signalling pathways. Of note, RNA-seq and proteomics in SG impaired shG3BP1 leukemia cells showed both a negative enrichment of the SG proteome and LSC signature as well as a positive enrichment of the innate immune response. Interestingly, our BioID uncovered that the AML specific SG proteome is also enriched for positive regulation of RIG-I signaling pathway (GO: Biological Process). Together this data suggests SGs play a protective role through the downregulation of overactive innate immune signalling in AML. Work using optimized small input approaches to map SG RNA and protein interactomes in LSCs vs HSCs to confirm similar regulatory relationships as well as identify LSC-context specific ones is ongoing. Overall, these results elucidate SGs as key AML LSC stress adaptive regulatory networks and highlights them as novel targets for consideration in AML therapeutics.

Differentiation Syndrome in Acute Leukemia: APL and Beyond.

Differentiation syndrome (DS) is a frequent and potentially life-threatening clinical syndrome first recognized with the advent of targeted therapeutics for acute promyelocytic leukemia (APL). DS was subsequently observed more broadly with targeted therapeutics for acute myeloid leukemia (AML). DS is typically characterized by fever, dyspnea, hypotension, weight gain, pleural or pericardial effusions, and acute renal failure. The incidence in patients with APL ranges from 2 to 37%, with the wide variation likely attributed to different diagnostic criteria, use of prophylactic treatment, and different treatment regimens. Treatment with corticosteroids +/- cytoreductive therapy should commence as soon as DS is suspected to reduce DS-related morbidity and mortality. The targeted anti-leukemic therapy should be discontinued in patients with severe DS. Here, we discuss the pathogenesis of DS, clinical presentations, diagnostic criteria, management strategies, and implementation of prospective tracking on clinical trials.

Fluorescence In Situ Hybridization (FISH) for Detection of c-MYC Amplifications in AML Patients: A Potential Marker for Risk Stratification

Abstract Background c-MYC is a transcription factor (located on 8q24) which regulates genes involved in proliferation, cell growth, differentiation, and apoptosis. The c-MYC amplification is related to a rapid proliferation of the affected cell clone in patients with myeloid neoplasms. Gene amplification (amp) is one of the basic mechanisms connected with over-expression of oncogenes. c-MYC is routinely analyzed in mature lymphoid neoplasms especially in Burkitt’s Lymphoma while not assessed for acute myeloid leukemia (AML).Despite the fact the c- MYC oncogene is commonly over-expressed in AML it has remained an elusive target. Routine assessment for c- MYC in AML may pave the way for targeting the c-MYC in treatment of AML patients. Several experiments are ongoing investigating both direct and indirect c-MYC targeting therapies and further establishing cMYC as an oncogene of central importance to AML pathogenesis and treatment. Aim of the Work In this study we aimed to define the prognostic value of c-MYC amplification in adult AML patients, and to relate it to other prognostic factors in AML. Patients and Methods The study was a randomized control study, conducted at the Clinical Pathology Department, Ain-Shams University Hospitals during the period from January 2019 till June 2020. It was carried on 50 adult AML patients (30 newly diagnosed and 20 on Therapy) and 10 healthy age-matched volunteers as control. All subjects underwent full history taking, thorough clinical examination, laboratory investigations including; CBC, Examination of Leishman’s stained peripheral blood films, Bone marrow aspiration and examination of Leishman’s stained bone marrow smears, Cytochemical studies using Myeloperoxidase stain, Immunophenotyping of bone marrow or peripheral blood samples and Fluorescence in situ hybridization analysis (FISH). Results we had 9 patients with c-MYC amplification (6 in the newly diagnosed group and 3 in the on therapy group). So the percentage of c-MYC amplification in our patients was 9/50 (18%). To further elucidate the effect of c-MYC amplification on our patients’ clinical, laboratory and prognostic factors, an analytical comparison was done between negative and positive patients for cMYC amplification as regard-studied parameters in each group of patients. Both group of patients showed a statistical significant association between c-MYC amplification and outcome where the majority of patients with positive c-MYC amplification achieved incomplete remission. In addition, c-MYC amplification showed a statically significant association with Trisomy 8 in both patients’ groups and with 17p deletion and hepatosplenomegaly in on therapy group. Univariate logistic regression analysis for the association between c-MYC amplification and outcome showed statistically significant association between c-MYC amplification and incomplete remission in newly diagnosed AML patients with P-value = 0.013 and odds ratio (95% CI) of 14.0. Conclusion Our study has shown that c-MYC amplification is associated with adverse prognosis, poor treatment outcome and it might be involved in the process of AML progression. Targeting cMYC may be of value in the development of new-targeted AML therapeutics.

The Immuno-Oncology and Genomic Aspects of DNA-Hypomethylating Therapeutics in Acute Myeloid Leukemia.

Hypomethylating agents (HMAs) have been used for decades in the treatment of hematologic neoplasms, and now, have gathered attention again in terms of their combination with potent molecular-targeted agents such as a BCL-6 inhibitor venetoclax and an IDH1 inhibitor ivosidenib, as well as a novel immune-checkpoint inhibitor (anit-CD47 antibody) megrolimab. Several studies have shown that leukemic cells have a distinct immunological microenvironment, which is at least partially due to genetic alterations such as the TP53 mutation and epigenetic dysregulation. HMAs possibly improve intrinsic anti-leukemic immunity and sensitivity to immune therapies such as PD-1/PD-L1 inhibitors and anti-CD47 agents. This review describes the immuno-oncological backgrounds of the leukemic microenvironment and the therapeutic mechanisms of HMAs, as well as current clinical trials of HMAs and/or venetoclax-based combination therapies.

Outcomes for Acute Myeloid Leukemia Relapse after Allogeneic Hematopoietic Cell Transplantation Remain Poor in the Modern Era

Background: Relapse of acute myeloid leukemia (AML) following allogeneic hematopoietic cell transplantation (allo-HCT) has historically been associated with dismal prognosis with most patients failing to achieve subsequent remission and estimated 2-year overall survival (OS) of <15 %. Rapid progress in the understanding of AML biology and genomics resulted in recent FDA approval of several novel small molecules and immunotherapies for the treatment of front-line and relapsed/refractory AML, as early as 2017. We hypothesized that the availability of novel therapies for AML relapsing after allo-HCT has resulted in an improved OS in recent years. Methods: Patients with AML ≥18 years old who underwent allo-HCT at Stanford University between the years 2010 and 2021 and subsequently relapsed were included in this study. Patients were divided into two eras based on year of relapse: 2010-2016 (early era) and 2017- 2022 (later era). Descriptive statistics were used to characterize patients across the two cohorts; Kaplan Meier method was used to evaluate OS. Results: A total of 258 adults with AML relapsed following allo-HCT; 142 (55%) transplanted in the early era and 116 (45%) in the later era. Relative to the early era, patients who relapsed in the later era were more likely to be Hispanic (19.8% vs. 8.5%, P=0.01), have adverse risk leukemia (64.7% vs 52.1%, P=0.039), poorer performance status as defined by KPS <90 (46.6% vs. 16.9%, p<0.001), an HCT-comorbidity index of ≥ 3 (43.1% vs. 28.2%, P=0.012), and receive alternative donor transplant (haploidentical donors 11.2% vs 3.5%, P=0.002; umbilical cord blood 9.5% vs. 4.2%, P=0.002). Post-HCT maintenance therapy was used more frequently in the later era (16.4% vs. 2.1%, P=0.002). Median age, sex, and conditioning intensity did not significantly differ between the two cohorts. The median time to AML relapse post-HCT was similar in the early vs late era (149 days vs. 172 days, P=0.003). The management of relapse differed significantly across eras (Figure 1), as expected, with patients in the later era more commonly receiving targeted therapy with FLT3 inhibitors and IDH1/2 inhibitors (25% vs. 7.7%), hypomethylating agents (HMA) (51.7% vs. 35.2%) and BCL2 inhibitors (32.8% vs 3.5%) and less likely receiving cytotoxic chemotherapy (21.6% vs 34.5%). The use of donor lymphocyte infusion increased over time (16.9% vs. 12.1%) as did second allo-HCT (10.3% vs. 7%). Despite increased use of novel therapies in the later era, there was no improvement in median OS compared to the early era (5.1 months versus 4.5 months, p=0.43) or 2-year OS following post-HCT relapse (19.1% vs. 12.4%, P=0.26), (Figure 2). Across both eras, relapsed AML was the most common cause of death in this cohort, attributed to 83.7% of all deaths. Conclusion: Despite the greater use of novel therapies to treat post-HCT relapse of AML in recent years, OS remains poor and has not improved. Relapsed AML following allo-HCT remains a significant clinical challenge despite advances in modern AML therapeutics. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Acute Myeloid Leukemia: New Multiomics Molecular Signatures and Implications for Systems Medicine Diagnostics and Therapeutics Innovation.

Acute myeloid leukemia (AML) is a common, complex, and multifactorial malignancy of the hematopoietic system. AML diagnosis and treatment outcomes display marked heterogeneity and patient-to-patient variations. To date, AML-related biomarker discovery research has employed single omics inquiries. Multiomics analyses that reconcile and integrate the data streams from multiple levels of the cellular hierarchy, from genes to proteins to metabolites, offer much promise for innovation in AML diagnostics and therapeutics. We report, in this study, a systems medicine and multiomics approach to integrate the AML transcriptome data and reporter biomolecules at the RNA, protein, and metabolite levels using genome-scale biological networks. We utilized two independent transcriptome datasets (GSE5122, GSE8970) in the Gene Expression Omnibus database. We identified new multiomics molecular signatures of relevance to AML: miRNAs (e.g., mir-484 and miR-519d-3p), receptors (ACVR1 and PTPRG), transcription factors (PRDM14 and GATA3), and metabolites (in particular, amino acid derivatives). The differential expression profiles of all reporter biomolecules were crossvalidated in independent RNA-Seq and miRNA-Seq datasets. Notably, we found that PTPRG holds important prognostication potential as evaluated by Kaplan-Meier survival analyses. The multiomics relationships unraveled in this analysis point toward the genomic pathogenesis of AML. These multiomics molecular leads warrant further research and development as potential diagnostic and therapeutic targets.

Engineered Nanoparticles as Potential Therapeutics for Acute Myeloid Leukemia

Acute myeloid leukemia (AML) is a malignancy associated with poor prognosis. Particularly, older patients suffer greatly from the standard chemotherapy and have a 5‐year survival of only 4%. Hence, new therapeutic agents with higher specificity and lower general cytotoxicity are urgently required.We have discovered that positively charged amino‐functionalized polystyrene nanoparticles (NP‐PS+) induce accumulation of acidic vesicular organelles with elevated pH and impaired processing of procathepsin B leading to mTOR inhibition, activation of autophagy, and induction of caspase‐dependent apoptosis in leukemia cells, but not in normal human macrophages. The antileukemic effect of NP‐PS+ was also preserved in vivo, where NP‐PS+ inhibited proliferation and induced apoptosis in leukemia xenografts grown on chick chorioallantoic membranes.Similar to polystyrene particles, amino‐functionalized gold nanoparticles (NP‐Au+) exhibited selective cytotoxicity towards AML cell lines as well as primary patient‐derived AML cells. Thus, NP‐Au+ particles induced cell death in primary human leukemia cells and reduced their colony‐forming potential, whereas normal hematopoietic cells remained unaffected by the treatment with NP‐Au+. NP‐Au+ targeted specifically the oxidative mitochondrial respiration, which is, different to normal hematopoietic cells, the main source of energy production in AML blasts and leukemic stem cells. Different to NP‐Au+, conventional chemotherapeutics such ascytarabine act in a cell cycle‐dependent manner and target only on proliferating AML blasts but not on quiescent leukemic stem cells. In agreement with the in vitro data, NP‐Au+ exhibited antileukemic efficacy against primary human AML xenografted into mice applied either as monotherapy or as a cytarabine combination regimen in the absence of detectable adverse events.Thus, this engineered nanomaterial that targets particularly resistant quiescent leukemic stem cells holds great promise as a novel nanotherapeutic for the treatment of acute myeloid leukemia independent of its cytogenetic profile.

Uncoupling the BMP receptor antagonist function from the WNT agonist function of R-spondin 2 using the inhibitory peptide dendrimer RWd

Signaling by bone morphogenetic proteins (BMPs) plays pivotal roles in embryogenesis, adult tissue homeostasis, and disease. Recent studies revealed that the well-established WNT agonist R-spondin 2 (RSPO2) is also a BMP receptor (BMP receptor type 1A) antagonist, with roles in early Xenopus embryogenesis and human acute myeloid leukemia (AML). To uncouple the BMP antagonist function from the WNT agonist function and to promote development of AML therapeutics, here we identified a 10-mer peptide (RW) derived from the thrombospondin 1 domain of RSPO2, which specifically prevents binding between RSPO2 and BMP receptor type 1A without altering WNT signaling. We also show that a corresponding RW dendrimer (RWd) exhibiting improved half-life relieves inhibition of BMP receptor signaling by RSPO2 in human AML cells, reduces cell growth, and induces differentiation. Moreover, microinjection of RWd in Xenopus embryos ventralizes the dorsoventral embryonic patterning by upregulating BMP signaling without affecting WNT signaling. Our study corroborates the function of RSPO2 as a BMP receptor antagonist and provides a proof of concept for pharmacologically uncoupling BMP antagonist from WNT agonist functions of RSPO2 using the inhibitor peptide RWd with enhanced target selectivity and limited side effects.

Isoform Selective HDAC8 Inhibitor (OCH3) Shows Potency in Selectively Targeting Primary AML Cells

The treatment outcomes for patients diagnosed with acute myeloid leukemia (AML) are still dismal. Recent advances in understanding AML indicate that the lack of efficacy is primarily due to non-specificity of currently used chemotherapeutics targeting both leukemic stem/progenitor cells (LSC) and normal hematopoietic stem cells (HSC). Thus, a critical barrier is the identification of innovative therapies that selectively target LSC. Histone deacetylase 8 (HDAC8) has been shown to enhance p53 protein deacetylation, which results in inactivation of p53, promoting LSC survival. We hypothesize that enzymatic/non-enzymatic role of HDAC8 is critical for LSC survival but not for HSCs. Then, we characterized our two tetrahydroisoquinoline (TIQ)-based selective HDAC8 inhibitors (HDAC8i) BIP and OCH3 for growth inhibition, apoptosis, activation of caspase 3, integrity of mitochondrial membrane potential (MMP), and acetylation of histone H4 in human leukemia cell lines. The growth inhibitory effects observed in cell lines were validated using bone marrow (BM) or peripheral blood (PB) cells from AML patients. Colony forming cell (CFC) assays were performed using AML BM/PB cells treated with OCH3 or BIP. OCH3 and BIP were also tested for hematotoxicity using normal CB CD34+ cells. Furthermore, we compared class I HDAC isoform engagement in human normal cord blood (CB) CD34+ cells and in SET-2 leukemia cells using our novel photoreactive probe TH1143. In CD34+ cells, TH1143 had higher level of engagement for HDAC1 and 2, whereas engagement of HDAC3 and 8 was minimal. In SET-2 cells, HDAC3 and HDAC8 displayed relatively higher engagement with TH1143 indicating HDAC engagement is likely cell type specific.The biological efficacies of OCH3 at 50uM and BIP at 25uM were noted to exert >50% growth inhibition in KG1 and in K562 leukemia cells. Both OCH3 and BIP significantly increased the number of apoptotic cells and there was an enhanced active caspase-3 activity. Furthermore, OCH3 and BIP treated cells displayed lower red/green ratio in comparison to control, indicative of poor MMP and depolarization to induce apoptosis (Table 1.a). OCH3 and BIP were further validated by using BM/PB cells from AML patients showing growth inhibition. This was also accompanied by increase in apoptotic cells by OCH3 and BIP. In contrast to BIP, OCH3 spared CB CD34+ cells as demonstrated by notably lower growth inhibition, apoptotic cells vs control when compared with primary AML cells from patients. Both OCH3 and BIP displayed minimal inhibition of CFU growth in CD34+ cells. However, HDAC8i induced significant CFU growth inhibition in primary AML samples suggesting that HDAC8i spares normal CFU progenitors but not leukemia progenitors (Table 1.b). Notably, both BIP and OCH3 lack ability to exert acetylation of histone H4, unlike broad spectrum HDAC inhibitor TSA (MFI with OCH3=0.96±0.03, BIP=0.77±0, TSA =1.63±0.15) which is consistent with isoform selectivity of OCH3 and BIP.The leukemia growth inhibitory effects at LSC level was demonstrated using ex vivo OCH3 treated AML patient derived BM/PB cells transplantation in humanized immunodeficient NSGS mice. After 10 to 12 weeks of transplantation mice receiving untreated AML cells had 7.73±2.18% while with OCH3 treatment mice had 4.84±1.37% human CD34+ leukemia cells, a 38% reduction in CD34+ leukemia cells, despite only a single ex vivo exposure to OCH3. Furthermore, in a second model, NSGS humanized mice were transplanted (IV) with primary leukemia cells from AML patients and after 4 weeks injected (IP) with OCH3 or vehicle control. After 12 weeks of transplantation in this second model human primary AML cell burden was 5.74±1.31% (OCH3) and 18.13±12.76% (vehicle control), while mice transplanted with normal CD34+ cells treated similarly with OCH3 or vehicle control displayed no detectable inhibition of human myeloid cell chimerism (OCH3:12.28 ± 3.31% vs vehicle control: 17.92±11.96%).Taken together, our data indicate that HDAC8 isoform inhibitor, OCH3 displayed significant inhibition of primary AML patient derived leukemia cells growth in vitro and in vivo in contrast to normal CD34+ cells. Selective inhibition of HDAC8 is sufficient to cause growth inhibition in primary AML progenitors including LSCs in vivo while sparing normal HSCs thus offer opportunities for further development of HDAC8i as new experimental therapeutics in AML. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Modulation of RNA Splicing Enhances Response to BCL2 Inhibition in Acute Myeloid Leukemia

Resistance to therapy is one of the most significant challenges in the treatment of acute myeloid leukemia (AML). While great efforts have uncovered genetic mechanisms of resistance to certain AML-directed therapies, to date, treatment resistance in AML has only partially explained by acquired genetic alterations. Here, we performed genome-wide CRISPR/Cas9 screens to identify drug-gene interactions that modulate therapeutic response to treatments commonly used in AML. Interestingly, our findings uncovered several genes that regulate pre-mRNA splicing whose loss strongly synergized with venetoclax, a BH3 mimetic that blocks the antiapoptotic protein BCL-2. To further delineate the role of RNA processing in response to AML treatments, we performed secondary CRISPR screens with a domain-focused gRNA library targeting 490 RNA processing factors in the presence of various AML drugs. Overall, these genetic screens identified a number of RNA splicing factors whose loss-of-function sensitized AML cells to BCL2 inhibition (Fig. A).Among the top gene candidates whose loss promoted venetoclax efficacy was the splicing factor RBM10 (Fig.B). Strikingly, loss of RBM10 exclusively synergized with venetoclax-based treatments across AML therapeutics, including in TP53 mutant lines (Fig.C-D). Moreover, RBM10 loss restored venetoclax sensitivity to AML cell line variants with acquired venetoclax resistance. Interestingly, while many RNA splicing factors are pan-essential, generation of an Rbm10 conditional knockout mouse revealed that Rbm10 is completely dispensable for steady-state normal hematopoiesis (Fig.E). Since RBM10 has not been studied previously in hematopoiesis, we mapped the impact of RBM10 on mRNA expression and splicing using RNA-seq and direct RNA binding partners genome-wide by eCLIP-Seq (Fig. F). RBM10 loss was strongly associated with downregulation of BCL2A1, an anti-apoptotic factor whose expression is correlated with venetoclax resistance in AML (Fig.G-H). This was dependent on RBM10's ability to bind RNA and expression of BCL2A1 cDNA fully rescued the growth-inhibitory effect of RBM10 KO-venetoclax treated AML cells. Overall, the above data support RBM10 as a synthetic lethal vulnerability in venetoclax therapy.Beyond RBM10, our genetic screens also identified several splicing factors belonging to the family of serine and arginine-rich (SR) proteins whose loss synergized with venetoclax treatment (Fig. I). SR proteins are essential for pre-mRNA splicing and are substrates for phosphorylation by conserved family of kinases, such as Cdc2-like kinases (CLKs) and (dual-specificity tyrosine-regulated kinases) DYRKs. We therefore utilized a series of selective pan-CLK/DYRK1A inhibitors, including SM09419 and SM08502, that potently suppress SR protein phosphorylation. Interestingly, BCL2 is one of the top genetic dependencies upon DYRK1A genetic suppression in prior work from the DepMap (Fig. J). Pharmacologic inhibition of CLK/DYRK1A exhibited high in vitro efficacy at nanomolar range across a diverse range of AML subtypes including cell lines with acquired venetoclax resistance (Fig.K). Consistent with this, combined SM09419 and venetoclax displayed synergistic anti-leukemic effects and venetoclax-sensitive AML cell lines (Fig.L). Taken together, these data support the notion of targeting CLK/DYRK1A in the context of BCL2 inhibition.In this study, we systematically defined gene interactions that mediate the response to a wide range of AML drugs. Recent studies have begun to show that dysfunctional RNA processing promotes AML development. However, the role of RNA processing in modulating drug responsiveness in AML is not well understood. Here, we have uncovered that synthetic lethal targeting of splicing factors, such as RBM10, increases sensitivity of AML cells to BCL2 inhibition. Therapeutically, pharmacologic inhibition of SR protein function via inhibiting CLK/DYRK1A-mediated phosphorylation of splicing factors is an effective strategy used in combination with venetoclax or to overcome venetoclax resistance. Overall, our findings underscore the central importance of RNA splicing in drug response and provides a therapeutic rationale for modulating RNA splicing to enhance current AML therapies. [Display omitted] DisclosuresMcMillan: Prizer: Ended employment in the past 24 months. Bossard: Biosplice Therapeutics: Current Employment. Aifantis: AstraZeneca: Research Funding; Foresite (FL2020-010) LLC: Consultancy. Abdel-Wahab: H3B Biomedicine: Consultancy, Research Funding; Foundation Medicine Inc: Consultancy; Merck: Consultancy; Prelude Therapeutics: Consultancy; LOXO Oncology: Consultancy, Research Funding; Lilly: Consultancy; AIChemy: Current holder of stock options in a privately-held company, Membership on an entity's Board of Directors or advisory committees; Envisagenics Inc.: Current holder of stock options in a privately-held company, Membership on an entity's Board of Directors or advisory committees.

Abstract 1430: In vivo PoC study and design for Phase I clinical study of SJP1604, a nucleolin-targeted therapeutic agent to treat entire population of relapsed/refractory acute myeloid leukemia patients

Abstract SJP1604 (APTA-16) is an aptamer-drug conjugate (APTA-DC) specifically targeting nucleolin, which is highly expressed only on the cell membrane of acute myeloid leukemia (AML) cells including relapsed/refractory aute myeloid leukemia (R/R AML). We previously presented the outstanding anti-leukemic activity of SJP1604 on AML cells including drug-resistant AML cells while standard chemotherapies showed no efficacy in vitro and in vivo. Therefore, we were able to conclude that only SJP1604 exhibits excellent resistance overcoming efficacy regardless of genetic defects of AML cells. Additionally, we recently identified that SJP1604 markedly eradicated AML cell engraftment with decreasing expression level of nucleolin protein in the bone marrow cells of MOLM-13 xenograft mouse model. These findings demonstrate that SJP1604 contributes to anti-leukemic effects through the regulation of nucleolin protein. We also figured out that nucleolin as a clinical biomarker was significantly highly expressed on AML and R/R AML patient-derived bone marrow cells. In particular, nucleolin expression is highly overexpressed in the bone marrow cells of entire population of R/R AML patients. Upto recent, considered as one of the major targeted therapeutics for AML, FLT3 inhibitors can target 25-30% of AML patients only. Therefore, our results mentioned above are very noteworthy finding showing the high potential of SJP1604 to be further developed as the first innovative targeted therapy to treat entire population of R/R AML patients. Herein, we show a phase I clinical trial design of SJP1604 to perform an open-label, first-in-human, dose escalation study to evaluate the safety, tolerability, pharmacokinetics and anti-tumor activity in patients with R/R AML. Citation Format: Jihyun Um, Dohyeong Lee, So Hee Kim, Eun-Hye Huh, Yongbin Park, Sung Hwan Moon, Soo Jin Lee, Je-Hwan Lee, Min-Hyo Ki. In vivo PoC study and design for Phase I clinical study of SJP1604, a nucleolin-targeted therapeutic agent to treat entire population of relapsed/refractory acute myeloid leukemia patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1430.

Abstract 2933: Inhibition of FLT3 and AXL by gilteritinib controls systemic tumor growth in luciferase-transduced MV-4-11 acute myeloid leukemia

Abstract Mutations and amplification of the fms-like receptor tyrosine kinase FLT3 gene, such as those found in MV-4-11 cells, are common in acute myeloid leukemia (AML). Such mutations result in aberrant activation of FLT3, increased growth and survival of tumor cells, and the development of refractory forms of AML with poor patient outcomes. In addition, AXL receptor tyrosine kinase imparts chemotherapeutic resistance in AML. MV-4-11 is a human biphenotypic B myelomonocytic leukemia with t(4;11)(q21;q23) chromosomal translocation commonly found in AML patients; it possesses a homozygous in-frame tandem repeat insertion in the FLT3 allele resulting in constitutively phosphorylated FLT3. Commonly employed preclinical orthotopic models of AML are primarily evaluated based on morbidity and mortality, which provide limited real time information on tumor burden and dissemination. The purpose of this study is to develop a MV-4-11 mouse model that allows the quantification and anatomical localization of the AML cells by monitoring bioluminescent signal in live animals and to utilize this model to investigate the efficacy of a FLT3/AXL inhibitor, gilteritinib. This poster describes a) the production of luciferase-expressing human MV-4-11 (MV-4-11-luc) cells via lentiviral transduction, b) the characterization of MV-4-11-luc progression in two models of immunocompromised mice, c) the efficacy of the FLT3/AXL kinase inhibitor gilteritinib against systemic MV-4-11-luc compared with two standard AML therapies, cytarabine and cyclophosphamide and d) the development of a humanized MV-4-11 tumor bearing model in NCG mice. Bioluminescence imaging revealed dose dependent tumor growth localized to the hind limbs and peritoneal cavity, which disseminated and intensified over time. Characteristic moribundity associated with disease progression was observed in this model. By comparison, robust growth of both parental and luc-transduced MV-4-11 was also observed in the highly immunodeficient NCG mice with or without prior myeloablation. Efficacy studies in NCG mice bearing parental or luciferase-tagged MV-4-11 cells demonstrated potent anti-tumor activity using the FLT3/AXL inhibitor. Bioluminescent imaging of orthotopic MV-4-11-luc xenografts in immunocompromised mice is a quantitative and sensitive method that provides anatomical context for assessing the efficacy of AML therapeutics. Ongoing and future work is focused on a) genetic profiling of MV-4-11 tumors after treatment with gilteritinib, b) further characterization of a humanized MV-4-11 model to evaluate immune-modulating therapeutics, and c) utilizing the model to evaluate activity of cell therapies that target AML. Citation Format: Na Li, Vivek Mahajan, Lindsay Butler, Elizabeth Rainbolt, Shasta Kidder, Kay Meshaw, Diana Gietl, Susan Yeyeodu, Anya Avrutskaya, Jing Wan, Thi Bui, Chassidy Hall, David P. Harris. Inhibition of FLT3 and AXL by gilteritinib controls systemic tumor growth in luciferase-transduced MV-4-11 acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2933.

Apoptosis Induction and Cell Cycle Profiling after Treatment with Low-Dose Decitabine and/or Bortezomib in the AML Cell Line Kasumi-1

Introduction : Acute myeloid leukemia (AML) is a heterogeneous clonal hematopoietic neoplasm characterized by maturation arrest in the myeloid lineage. The standard of care regimen of induction for AML patients is an anthracycline and cytarabine combination, followed by consolidation therapy. Nevertheless, relapse still remains a major hurdle for successful AML chemotherapy and alternative treatment strategies are much needed. Epigenome-modifying drugs, such as decitabine (D), a DNA methyltransferase inhibitor (DMTi), have shown promise as therapeutics for AML in low doses, while higher doses are associated with cellular differentiation and cytotoxicity. On the other hand, oxidative stress inducers have been strongly implicated in the targeting of LSCs, since increased oxidation is associated with reduced self-renewal, which in turn leads to either differentiation or death of haemopoietic cells. Nevertheless, the induction of oxidative stress alone is not sufficient for AML treatment.Aims: Since only approximately 50% of patients treated with DMTis show a hematological improvement (HI) or better, and few alternative treatments exist for patients who fail to respond, the aim of this study is to investigate the possible reversal of decitabine resistance phenotype in AML, in terms of apoptosis and cell cycle profiling, by the combination of decitabine with oxidative stress inducing agents, such as the proteasome inhibitor bortezomib (BZ).Methods : The AML Kasumi-1 cells, carrying the t(8;21) and the KIT mutation N822, were cultured in RPMI 1640, supplemented with 20% FBS, and incubated in 5% CO2 at 37°C. Cells were treated with low-dose decitabine (10nΜ, 50nΜ, 100nM, 200nM and 400nΜ), with or without bortezomib (10nM) for 24h. Flow cytometry was used for apoptosis (Annexin V/PI staining) and cell cycle profiling (DAPI staining). One-way Anova and LSD/ Bonferroni methods were applied for the statistical analysis of the results.Results : Our data indicate significant alterations in cell death and cell cycle stages in Kasumi-1 cells following D and BZ combination treatment compared to the control (untreated cells) and single treatments. Apoptosis was statistically significantly increased compared to control (15,15%) after only 100nM and 400nM of single D treatment (27,2% and 28,25%, p=0.037 and p=0.026, respectively) and after D/BZ treatment for all D concentrations tested [31,5%, 33,4%, 54,65%, 47,1% and 55,55%, from lower to higher concentration, with p=0.008 and p=0.004 for 10nM and 50nM respectively, and p<0.000 for 100nM, 200nM and 400nM). Compared to single treatments, the D/BZ combination significantly increased apoptosis by 101,92%, 72%, 100,9%, 113,12% and 96,63% (p=0.009, p=0.019, p<0.000) and reduced the live cell population by 23,51%, 22,21%, 54,9%, 41% and 49,81% (p=0.006, p=0.011, p<0.000) for 10nΜ, 50nΜ, 100nM, 200nM and 400nΜ of D/BZ treatments, respectively. Cell cycle profiling also highlighted a much greater sensitivity of Kasumi-1 cells in D/BZ combinations compared to single treatments, with a significant increase in the G1 population by 11,84% (p<0.000), 12,55% (p<0.000), 6,62% (p=0.021), 8,38 (p=0.006) for all D/BZ combinations (from lower to higher) tested, except 400nM D/BZ. A significant decrease in S phase was observed after all combination treatments tested, compared to single ones, by 36,26%, 39,25%, 26%, 30,99% and 30,8% (lower to higher D concentration), (p<0.000), while finally, G2/M population was increased by 56,35% (p<0.000), 85,34% (p<0.000), 34,29% (p=0.02), 81,92% (p<0.000) and 138,45% (p<0.000), respectively.Conclusions : Our data indicate that the addition of bortezomib -a proteasome inhibitor which, among others, is capable of inducing oxidative stress- to low-dose decitabine significantly enhances apoptosis and decreases live cell population of Kasumi-1, with the combinations of 100nM and 200nM of D with BZ appearing as the most successful ones. Moreover, cell cycle profiling revealed that D/BZ treatment synergistically leads to G1 and G2 arrest, hence prohibiting cells to either synthesize DNA (S phase) or proceed to mitosis. Although these observations need to be further investigated at a molecular level, they appear very encouraging for better understanding the mechanisms underlying primary resistance to decitabine and offer new directions for a much needed, more successful epigenetic therapy in AML. DisclosuresNo relevant conflicts of interest to declare.

IDH1/IDH2 Inhibition in Acute Myeloid Leukemia.

Recently, the discovery of biological and clinical properties of mutated isoforms 1 and 2 mutations of isocitrate dehydrogenases (IDH) 1 and 2, affecting approximately 20% of patients with acute myeloid leukemia (AML), lead to the development of an individualized treatment strategy. Promoting differentiation and maturation of the malignant clone targeting IDH is an emerging strategy to promote clinical responses in AML. Phase I/II trials have shown evidence of safety, tolerability, and encouraging evidence of efficacy of two small molecule inhibitors targeting IDH2 and IDH1 gene mutations, respectively enasidenib and ivosidenib. In this review, the contribution of IDH1/IDH2 mutations in leukemogenesis and progress of targeted therapeutics in AML will be highlighted.

Novel Targeted Therapeutics in Acute Myeloid Leukemia: an Embarrassment of Riches.

Acute myeloid leukemia (AML) is an aggressive malignancy of the bone marrow that has a poor prognosis with traditional cytotoxic chemotherapy, especially in elderly patients. In recent years, small molecule inhibitors targeting AML-associated IDH1, IDH2, and FLT3 mutations have been FDA approved. However, the majority of AML cases do not have a targetable mutation. A variety of novel agents targeting both previously untargetable mutations and general pathways in AML are currently being investigated. Herein, we review selected new targeted therapies currently in early-phase clinical investigation in AML. The DOT1L inhibitor pinometostat in KMT2A-rearranged AML, the menin inhibitors KO-539 and SYNDX-5613 in KMT2Ar and NPM1-mutated AML, and the mutant TP53 inhibitor APR-246 are examples of novel agents targeting specific mutations in AML. In addition, BET inhibitors, polo-like kinase inhibitors, and MDM2 inhibitors are promising new drug classes for AML which do not depend on the presence of a particular mutation. AML remains in incurable disease for many patients but advances in genomics, epigenetics, and drug discovery have led to the development of many potential novel therapeutic agents, many of which are being investigated in ongoing clinical trials. Additional studies will be necessary to determine how best to incorporate these novel agents into routine clinical treatment of AML.

An Update on the Clinical Evaluation of Antibody-Based Therapeutics in Acute Myeloid Leukemia.

The advent of several targeted agents has revolutionized the treatment of acute myeloid leukemia (AML) in recent times; however, majority of patients are still not cured. In the ongoing quest for rationally targeted treatment strategies in AML, scientific endeavors have focused on identifying new antigen targets on the leukemic cells for therapeutic exploitation including strategies to directly deliver toxins into the leukemic blasts as well as strategies that harness host immunity to favorably impact clinical outcomes. Gemtuzumab ozogamicin, a CD33 directed antibody-drug conjugate, has provided the proof of concept for the potential efficacy of monoclonal antibody-based therapies in AML. This article provides an overview of immunologically relevant antigen targets expressed on the leukemic cells and synopsizes the clinical results evaluating targeted antibody-based therapeutic approach in AML. AML blasts and leukemic stem cells express several antigens, including CD33, CD47, CD70, CD123, and CLEC12A. The past several years have seen the burgeoning of cell-specific immunotherapy concepts, including checkpoint inhibitors, antibody-toxin conjugates, and bispecific antibodies in the treatment of AML. The first-in-class anti-CD47 antibody magrolimab and anti-CD70 antibody cusatuzumab in combination with hypomethylating agent (HMA) azacitidine, in newly diagnosed AML, and flotetuzumab, a bispecific DART® (dual-affinity retargeting) antibody to CD3ε and CD123 as salvage option in relapsed/refractory AML appear promising. The development of antibody-based immunotherapeutic approach in AML has been encouraging. Ongoing research will define the choice of an appropriate complementary therapeutic agent in antibody-based combination therapy, and whether one or more than one antigen should be simultaneously targeted. Further studies will likely refine the role of antibody-based therapy in post hematopoietic cell transplant setting.

Target-Informed Repurposing of Immunotherapies in AML - a Transcriptome Based Approach for Identifying Immediately Available Therapeutics

Acute Myeloid Leukemia (AML) is an aggressive hematopoietic malignancy with limited therapeutic options and a great need for innovative treatment. However, the identification of novel molecular biomarkers remains challenging, and significant lag time from target discovery to clinical development impedes timely implementation of novel therapies. We previously demonstrated aberrant expression of non-hematopoetic (NH) antigens on leukemic cells in AML; one such antigen, mesothelin (MSLN), has recently begun AML clinical development, providing evidence for the use of NH directed therapeutics in AML. Given this observation, we questioned whether we could build a computational platform to identify misregulated genes in AML that are targeted by currently available immunotherapies in other malignancies. Early phase trial data for these targets will provide vital safety and tolerability information, thereby supporting a rapid path to AML clinical development. To this end, we curated a knowledgebase of antibody-drug conjugate (ADC) and chimeric antigen receptor T-cell (CAR T) therapies by querying all cancer immunotherapy trials registered in the U.S. National Library of Medicine clinical trials database (https://clinicaltrials.gov) and the Journal of Antibody-Drug Conjugates (https://adcreview.com) in November of 2019. In total, our knowledgebase contained 893 clinical trials assessing 141 prospective gene targets. We interrogated transcriptome expression of these targets using ribodepleted RNA-seq data from the TARGET AML and TpAML initiatives (N = 1,394, age &amp;lt;0 to 30 years) and the TCGA LAML cohort (N = 173, age 18 - 88 years). We selected for therapeutic targets expressed in AML by employing a minimum transcript expression threshold of 5 TPM in &amp;gt;50% of our AML cases. This led to the identification of 35 targets of interest for further analysis (Figure 1A). A total of 131 immunotherapies (70 ADCs and 61 CAR Ts) directed against these targets are in various stages of clinical development (Figure 1B). These included 62 AML-directed therapies targeting a total of 13 antigens; among these, CD33, CD123, FLT3, CD117, and CLEC12A have been well described in the context of AML. The remaining 69 therapies (43 ADCs and 26 CAR-Ts) target 30 NH antigens. Nine (30%) of these NH antigens, including CD74, BSG, TFRC, CXCR4, and CD44, all possesed a median of &amp;gt;90 TPM in our AML cohort and were subjects of therapies in Phase I or later. Additionally, these genes were universally expressed across major fusion groups (Figure 1C), suggesting potential as therapeutic targets regardless of age or genetic abnormality. A table of NH targets with high expression in AML is provided in Figure 1D. Conversely, we also identified targets for which expression was uniquely restricted to rare, high-risk, and therapeutically challenging AML variants. Upregulated genes in these variants were identified via differential expression analysis of CBFA2T3-GLIS2, NUP98-NSD1, and NUP98-KDM5A fusions. Genes were filtered using a log fold-change (logFC) threshold of 1 and FDR-adjusted p &amp;lt; 0.05, then cross-referenced with the full knowledgebase to identify 37 rare-variant enriched targets. The top 5 most highly enriched targets (ranked by logFC) were FOLR1, NCAM1, MMP14, TNF, and DLK1. The association of NCAM1 with CBFA2T3-GLIS2 has been previously described. However, FOLR1, a member of the folate receptor family that is overexpressed in epithelial-derived tumors, is a novel target in the context of AML. Like NCAM1, FOLR1 expression in our AML population is almost entirely limited to the CBFA2T3-GLIS2 fusion: 36/43 FOLR1-expressing patients possessed a CBFA2T3-GLIS2 fusion, comprising 94.74% of all CBFA2T3-GLIS2 fusions in our cohort. We screened transcriptome data from a large AML cohort for expression of immunotherapeutic targets with potential utility in AML. This enabled the identification of both broadly expressed and subtype-restricted putative targets. Future expansion of this methodology into other cellular therapies (CAR NK or BiTE) and the examination of new data sources could reveal additional targets with robust expression in AML. Demonstration of cell surface expression of these targets, and appropriate preclinical evidence of efficacy, could lay the groundwork for quickly moving these therapies to clinical trials in AML. Figure Disclosures No relevant conflicts of interest to declare.

FDA Analysis of Screen Failures By Race in Clinical Trials of Acute Myeloid Leukemia

Background: Ethnic and racial minorities are underrepresented in oncologic clinical trials. One potential barrier to participation may be narrow eligibility criteria which contribute to an increased rate of screen failure among minorities. Prior work in multiple myeloma demonstrated a higher rate of screen failure among black or African-American (AA) patients and lower rate of screen failure among Asian patients compared with white patients (1). Here, we examine screen failure rates by race in patients screened for participation in clinical trials of acute myeloid leukemia (AML). Methods: Trials submitted to the U.S. Food and Drug Administration (FDA) to support approval of new treatments for AML between 2016-2019 were examined. Trials which included information on screen failures and race were included. Trial, screen failure status, basic demographic information (age, gender, race, and ethnicity), country of participant, and reason for screen failure (if relevant) were abstracted from the trials. Screen failure rates were calculated based on this information. Results: Twenty-one trials of AML therapies were identified, of which screen failure and race information were available in fourteen. Of these, information on reason for screen failure was available in nine. A total of 6,471 patients were identified across fourteen trials (Table 1). Race was recorded as unknown for 2,934 (46%) of patients. A total of 3,372 (52%) screen failures occurred. Minorities were more likely to be screen failures at 201/603 (33%) compared to whites at 27%. Of minorities, screen failures occurred in 29% of black/AA patients, 36% of Asians, and 36% of patients of any other race. A total of 579 patients were included in the nine trials that included data on reasons for screen failures (Table 2). Lack of appropriate mutation (i.e. IDH1/2, FLT3) was the most common reason for screen failure overall and was more common in black/AA (41%) and Asian patients (66%) compared to white patients (29%). Other issues related to the disease course (e.g. not at the appropriate point in treatment, not treated within the time limit set in the protocol) led to screen failure in 28% black/AA patients versus 9-22% for other races (19% overall). White patients were more likely than patients of other races to be screen failures due to cardiac issues (4%) and laboratory abnormalities (6%). Conclusions: Minority patients were numerically more likely to be screen failures compared to whites in trials of AML therapeutics, which exemplifies the known issue of lower minority recruitment in clinical trials. Minority patients were more likely to be screen failures due to lack of appropriate mutation and less likely due to lab abnormalities or cardiac issues. The significance of this finding is uncertain given the limited information on reasons for screen failure and the small number of minority patients in the database. Further investigation into screen failures in minority patients with AML is indicated. These results underscore the importance of including racial and ethnic minorities in all phases of translational and clinical research to further precision oncology for all patients since oncogenic driver mutation frequencies may vary among different populations. (1) Kanapuru B, Fernandes L FDA analysis of multiple myeloma trials supporting approval; Presented at FDA-AACR workshop to examine under-representation of African Americans in multiple myeloma clinical trials 2/13/20 Disclosures No relevant conflicts of interest to declare.