Navigating the Landscape of Antibody Drug Conjugates: Current Trends and Future Research Prospects in Acute Myeloid Leukemia.

Acute myelogenous leukemia

A CD123-Targeting Antibody-Drug Conjugate (ADC), IMGN632, Designed to Eradicate Acute Myeloid Leukemia (AML) Cells While Sparing Normal Bone Marrow Cells

Gemtuzumab-ozogamicin (GO) is an antibody-drug conjugate (ADC) in which a monoclonal antibody targeting CD33 is covalently linked to the toxin calicheamicin. GO was initially approved by the United States Food and Drug Administration (FDA) for the treatment of adult patients with CD33+ acute myeloid leukemia (AML) in 2000. However, GO was recalled from the US market due to the lack of efficacy, and higher incidence of hepatotoxicities, including hepatic veno-occlusive disease (VOD), observed in phase 3 SWOG-0106 study. Since then, several other phase 3 studies have evaluated the efficacy of GO in the frontline treatment of adult patients with AML using different GO doses and schedules. The pivotal study that led to the reconsideration of GO was the French ALFA-0701 study, which used a lower and fractionated dose of GO in combination with standard chemotherapy (SC). Patients treated with the GO combination had a significantly longer survival outcome. The modified schedule also improved the toxicity profile. A systematic review and meta-analysis of over 3000 patients treated in five phase 3 studies showed that adding GO to SC improved relapse-free and overall survival. Most importantly, 6 mg/m2 dose of GO was associated with higher grade ⩾3 hepatoxicities and VOD than 3 mg/m2. The survival benefit was significant in the favorable and intermediate cytogenetic risk groups. This led to the reapproval of GO in 2017 for the treatment of patients with CD33+ AML. Currently, several clinical trials are exploring the role of GO with various combinations and in eliminating the measurable residual disease in patients with CD33+ AML.

Acute myeloid leukemia (AML) is the most common and aggressive blood cancer in adults. In particular, significant unmet medical needs exist for effective treatment strategies for acute myelomonocytic leukemia (M4) and acute monocytic leukemia (M5) AML subtypes. Antibody-drug conjugates (ADC) are a promising drug class for AML therapy, as demonstrated by the FDA-approved anti-CD33 ADC, gemtuzumab ozogamicin (Mylotarg). However, CD33 is expressed in normal hematopoietic stem cells, highlighting the critical need to identify AML-specific targets to minimize the risk of potential adverse effects. We have demonstrated that the leukocyte immunoglobulin-like receptor subfamily B4 (LILRB4) is expressed at significantly higher levels on monocytic M4 and M5 AML cells than on normal counterparts. Here, we test whether LILRB4 is a promising ADC target to kill monocytic AML cells while sparing healthy counterparts. To this end, we generated ADCs from a humanized anti-LILRB4 mAb and the antimitotic payload, monomethyl auristatin F. The conjugates constructed were characterized and evaluated for LILRB4-specific cell killing potency, toxicity to progenitor cells, pharmacokinetics, and therapeutic efficacy. Our ADC linker technology platform efficiently generated homogeneous anti-LILRB4 ADCs with defined drug-to-antibody ratios. The homogeneous anti-LILRB4 ADCs demonstrated the capacity for LILRB4-mediated internalization, suitable physicochemical properties, and high cell killing potency against LILRB4-positive AML cells. Importantly, our data indicate that these ADCs spare normal progenitor cells. One of our homogeneous conjugates exerted a remarkable therapeutic effect and no significant toxicity in a xenograft mouse model of disseminated human AML. Our findings highlight the clinical potential of anti-LILRB4 ADCs in monocytic AML therapy.

The value of the combination of gemtuzumab ozogamicin(GO) and chemotherapy for the treatment of acutemyeloid leukemia (AML) is currently analyzed within clinicaltrials. GO (6 mg/m2) and standard-dose cytarabine(100 mg/m2) is evaluated for the treatment of newly diagnosedAML in elderly patients in the SAL phase II trial.Preliminary results of the MRC AML15 trial support theapplication of GO 3 mg/m2 with standard- and high-dosecytarabine and anthracyclines for the treatment of denovo AML. Within this trial the addition of GO seems especiallyof value for favorable and intermediate cytogeneticrisk groups. The combination of GO (3 mg/m2) andhigh-dose cytarabine (3 g/m2) is safe and more effectivefor the treatment of refractory AML than previous combinationsfrom the AMLSG study group. First results provethe possibility of allogeneic stem cell transplantationafter GO therapy. Initial data of a phase II trial documentthe safety and efficacy profile of GO within a reduced-intensityconditioning protocol applying fludarabine andtotal body irradiation.

immune response (1). The dysfunction and reprogramming of the immune system play critical roles in leukemia initiation and progression (2). Immunotherapy aims to reactivate antitumor immune cells and overcome the immune escape mechanisms of leukemia in the bone marrow microenvironment. Represented by hematopoietic stem cell transplantation (3), chimeric antigen receptor (CAR) T-cell therapy (4), and T cell receptor (TCR) T-cell therapy (5), immunotherapy for leukemia has achieved tremendous success in clinical practice. However, the overall prognosis of leukemia immunotherapy still needs improvement, which warrants the discovery of biomarkers as well as the development of novel therapeutic designs.In this topic， Hino et al. summarized the role of the thymus in generating a diverse T lymphocyte repertoire and how age-related thymic involution and diseases such as acute myeloid leukemia (AML) affect T cell-mediated anti-tumor immunity. Patients with AML have impaired immune surveillance including reduced output of naïve T cells (6), restricted TCR repertoire (7), and increased frequency of regulatory T cells (8). Therefore, there is a potential correlation between thymus dysfunction and T-lymphocyte impairment with the ontogeny of AML. Many strategies for In recent years, tremendous approaches targeting the adaptive immune system have achieved great success in the treatment of hematological malignancies (9). Due to the innate immune system being the first line of defense against the external environment, its targeted therapies may offer additional hope for the treatment of hematological malignancies. CD47 as a cell surface ligand is overexpressed in a variety of malignant tumors and binds to signal-regulating protein alpha (SIRPa) on macrophages to promote tumor cells to escape phagocytosis. Blocking CD47-SIRPa axis can increase phagocytosis of macrophages and exert anti-tumor effect. Inhibitors targeting the CD47-SIRPa axis are being developed worldwide and undergoing preclinical and clinical studies. Xu et al. focused on the opportunities and challenges of anti-CD47 antibodies in hematologic malignancies. The combinations of anti-CD47 antibodies with other drugs have shown encouraging treatment response rate in patients with blood tumors, but side effects such as anemia also occur. Furthermore, bispecific antibodies and SIRPa/Fc fusion proteins appear to solve these difficulties and efficiently balance the efficacy and safety of the treatment.patients with central nervous system (CNS) involvement who received blinatumomab, a CD3/CD19 bispecific antibody. Case 1 was diagnosed with chronic myeloid leukemia and developed B-ALL with CNS involvement during treatment with dasatinib, and Case 2 was diagnosed with B-ALL and experienced early hematologic relapse and cerebral parenchyma involvement. Both patients achieved complete remission in the bone marrow and CNS after treatment with blinatumomab, suggesting that blinatumomab may be a potential promising treatment option for B-ALL patients with CNS leukemia. Xu et al. comprehensively explored the clinical benefits and safety of gemtuzumab ozogamicin (GO), an anti-CD33 humanized antibody in the treatment of AML. They summarized that GO tended to improve complete remission rates, followed by significantly improved survival outcomes including overall survival, event-free survival, relapse-free survival, and cumulative incidence of relapse. Subgroup analysis indicated that GO benefits were evident in patients with favorable-and intermediate-risk karyotypes, NPM1 mutations, and wild-type FLT3-ITD. Furthermore, administration of GO was also associated with reduced relapse rates and improved survival in certain patient subgroups, such as those aged <70 years, with de novo AML, and with CD33 + . Incorporating GO into established induction treatment strategies and a lower (<6 mg/m 2 ) dose of GO prolonged survival, while administration of GO increased risk of earth death at a higher dose ( ≥6 mg/m 2 ) and enhanced hepatic-related adverse effects. Further head-to-head randomized controlled trials need be performed to validate the therapeutic benefits and safety of GO in AML. Wu et al. systematically summarized the role of CD44 in leukemia development, which can promote the proliferation, migration, metastasis, chemoresistance and immune evasion of leukemic cells. CD44 and its variants as biomarkers are associated with poor prognosis, relapse or drug resistance in leukemia patients. Some CD44 subtypes, especially CD44v6 and CD44v3 tightly interact with the extracellular matrix, such as hyaluronic acid (HA) and osteopontin, to activate Nanog to enhance the expression of drug-resistance genes.Given the powerful tumor promoting ability of CD44 and its high affinity with HA, a number of CD44-related interventions, including anti-CD44 monoclonal antibodies, specific small molecular drugs, and HA-directed targeting of cancer cells, have been applied in the clinical treatment of leukemia. These strategies have shown encouraging results in preclinical and clinical trials. Certainly, targeting CD44 inevitably causes some side effects due to its expression on certain immune cells.

Introduction Treatment of acute myeloid leukemia (AML) has changed dramatically in the past ten years with the approval of targeted agents, the first of which was the anti-CD33 antibody-drug conjugate gemtuzumab ozogamicin (GO). Despite withdrawal from the market after accelerated approval, GO was reapproved and now has a well-established role in treating select AML patients. CD33 has proven to be an important target for drug development in AML as evidenced by the improvement in survival with GO treatment. Areas covered The review summarizes the development of GO, its mechanism of action, initial studies and approval, withdrawal from the market, and subsequent reapproval after the results of several large randomized studies became available. We also provide an overview of its current role in the treatment landscape of AML. Expert opinion Multiple phase 3 trials with GO have established a significant benefit with GO in induction therapy for favorable risk AML. Additional studies support the use of GO in relapsed/refractory AML and APL. Despite the withdrawal of GO from the market after initial approval, GO has proven to improve survival of select AML patients when added to induction chemotherapy and in relapsed disease.

Introduction Acute Myeloid Leukemia (AML) is a challenging blood cancer characterized by a high rate of relapse and often unfavorable outcomes. Immunotherapies can pave the way for a changing paradigm in AML treatment and improve therapeutic outcomes, ultimately leading to a possible cure for this challenging disease. This narrative review aims to summarize the progress of immunotherapy and highlight the future landscape of these measures in the context of AML. Area covered By searching English-language literature and querying the PubMed database using pertinent Medical Subject Headings, this review traces the development of AML immunotherapy, from the first antibody-drug conjugate, gemtuzumab ozogamicin, to newer approaches, including other monoclonal antibody formats, immune checkpoint inhibitors, chimeric antigen receptor T-cell therapy, and vaccinations. Expert review Although there have been significant advances, the outcomes of both traditional and novel therapies are still unsatisfactory. Immunotherapies could eliminate leukemia stem cells, which contribute to treatment resistance and disease relapse in AML. The positioning of these new therapeutic measures in development within the management algorithm for AML and their precise place in each patient’s therapeutic plan are future challenges for enhancing targeted, personalized clinical programs.

Introduction: Intensive chemotherapy remains the only therapeutic option that provides a reasonable chance of long-term survival for adults with acute myeloid leukemia (AML), particularly for those younger than 60 years of age. However, the vast majority of patients with AML are older than 60 years and the risks associated with intensive approaches are not only substantially greater but are also associated with significantly lower response rates. While a subset of older adults who are medically fit and have favorable biological features may be suitable candidates for intensive chemotherapy, many older adults are not candidates due to comorbidities, impaired performance status, and/or poor prognosis due to cytogenetic and molecular features. Outside of supportive care, there are limited treatment options that provide durable responses for older patients with AML, thereby warranting the development of novel therapeutic approaches. Decitabine (Dacogen, Astex/Eisai), a nucleoside analog, is FDA approved for the treatment of myelodysplastic syndrome (MDS) and has shown encouraging antileukemic efficacy for the management of older adults with AML. Combined with its favorable side-effect profile, decitabine has been at the center of several trials in the treatment of older adults with AML. Furthermore, decitabine is approved for the treatment of elderly AML in Europe.Areas covered: This paper will discuss the biological and clinical rationale for using decitabine as frontline therapy for AML patients unable to tolerate and/or benefit from intensive chemotherapy. A review of selected published manuscripts and abstracts on the subject of elderly AML and a search of PubMed for manuscripts in English using a combination of index words including AML, decitabine, elderly, hypomethylation, and 2′-deoxy-5-azacytidine was conducted and articles of interest selected.Expert opinion: Conventional treatment strategies for AML have not changed appreciably over the last 40 years. The intensive strategies necessary for attaining long-term survival are best tolerated by individuals younger than 60 – 65 years of age and a subset of medically fit older adults with favorable biological features. Unfortunately, many older adults with AML are medically unfit for intensive therapy due to significant comorbidities or poor functional status. In this setting, epigenetic therapy with hypomethylating agents represents a reasonable approach for the frontline treatment of older patients with AML unwilling or unsuitable for intensive chemotherapy or clinical trial. Decitabine has proven clinical benefits for patients with MDS and more recently in patients with AML. Response rates gained from low-dose decitabine (20 mg/m2 i.v. daily for 5 or 10 days every 28 days) are comparable to more intensive strategies. Compared to other widely accepted low-intensity programs such as low-dose cytarabine (20 mg/m2 subcutaneously once daily for 10 days) and supportive care, the 5-day decitabine regimen provides a modest survival benefit, as well.

Induction Therapy by Ara-C Plus Daunorubicin Versus Ara-C Plus Gemtuzumab Ozogamicin: Interim Analysis of a Randomized Phase II Trial of the SAL In Elderly Patients with Acute Myeloid Leukemia

BrUOG 345: Fractionated Gemtuzumab Ozogamicin Followed By Non-Engraftment Donor Leukocyte Infusions for Relapsed/Refractory Acute Myeloid Leukemia

Gemtuzumab Ozogamicin (GO) Inclusion to Induction Chemotherapy Eliminates Leukemic Initiating Cells and Significantly Improves Survival in Mouse Models of Acute Myeloid Leukemia

Objectives: Acute myeloid leukemia (AML) is a heterogeneous disease with diverse genetic abnormalities present in all ages, but mainly prevalent in an elderly population with an average age of above 60 years. The treatment of elderly AML remains a formidable challenge as the long-term outcomes of elderly AML patients have not improved in the last three decades, with ≤5% overall survival recorded at 5 years, calling for novel treatment options. Epigenetic therapy has a significant impact on the management of hematologic malignancies. Recent findings show that using very low dosages of decitabine depletes DNA methyl transferase 1 (DNMT1) without cytotoxicity. Decitabine-mediated epigenetic therapy does not induce FLT3 (fms-like tyrosine kinase 3) ligand, which hinders the effectiveness of FLT3 inhibitors. The inhibition of FLT3 and DNMT1 is associated with terminal myeloid differentiation of AML cells. Therefore, combining the inhibitors of FLT3 and DNMT1 may be an effective therapeutic approach for the treatment of poor-risk AML. Ponatinib is a third-generation receptor tyrosine kinase inhibitor. Various in vitro and in vivo preclinical studies demonstrated antileukemic activities of ponatinib against AML cells bearing FLT3-ITD mutations. Methods: Apoptosis and CD11b were measured by flow cytometry. FLT3 signaling and DNMT1 levels were analyzed by immunoblotting. Results: FLT3-ITD expressing AML cell lines MV4-11, MOLM-13, and MOLM-14 were used in the study. Co-treatment with decitabine (10-100 nM) and ponatinib (2 nM) in AML cells induced apoptosis in association with PARP cleavage. Increased levels of pro-apoptotic protein BAD and decreased levels of anti-apoptotic protein MCL1 were observed in these treated cells. The combination of decitabine and ponatinib also showed similar effects in primary AML cells expressing FLT3-ITD. The Western blot analysis demonstrated that treatment of decitabine decreased levels of DNMT1 and ponatinib inhibited FLT3 signaling and activation of downstream effectors STAT5, AKT, and ERK1/2 with induction of PU.1 a key regulator of myeloid differentiation. Co-treatment with ponatinib and decitabine using low concentrations in MV4-11 and MOLM14 cells induced myeloid differentiation. The percentage of differentiated cells was measured by increased surface expression of CD11b analyzed by flow cytometry and granulocytic/monocytic morphology examined by Wright-Giemsa staining. Conclusions: Mechanistically, the hypomethylating agent initiates the differentiation process and FLT3 inhibition augments differentiation leading to apoptosis of AML cells. Altogether, these preclinical findings of downregulation of DNMT1 and induction of PU.1 are a novel differentiation approach to induce apoptosis in AML cells and warrant future therapeutic potential for the treatment of AML patients expressing FLT3 mutations. Citation Format: Sudhakiranmayi Kuravi, Myles Taylor, Tara L. Lin, Jensen Roy, Joseph McGuirk, Ramesh Balusu. A targeted differentiation therapy for the treatment of acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4821.

We are investigating signaling molecules involved in apoptotic cell death in acute myeloid leukemia (AML) cells after treatment with the targeted drug gemtuzumab ozogamicin (GO) and the conventional drug daunorubicin, with focus on caspase-2. New effective therapeutic drugs are highly desirable in the treatment of AML. Although a majority of treated patients initially respond to conventional treatment, most patients will relapse within 1-2 years. Also, the treatment is associated with severe off-target effects. GO is a novel therapeutic approach, consisting of a monoclonal CD33 antibody coupled to calicheamicin, which cause DNA double strand breaks (DSB). We demonstrated that activation of Bak and Bax is critical for GO- and daunorubicin- induced apoptotic signaling in AML cells. The activation was followed by mitochondrial depolarization, caspase-3 activation and nuclear fragmentation. We continued to examine potential proapoptotic signaling events upstream of mitochondria in HL60 AML cells in vitro. We identified caspase-2 and the pro-apoptotic BH3-only protein Bid as two candidates. By analyzing caspase-2 by western blotting after GO treatment we found that GO caused cleavage of caspase-2 after 24h, which was prior to mitochondria-mediated signaling. Also, full-length Bid was cleaved to its truncated, active form tBid. The importance of caspase-2 activation in the signaling cascade after GO treatment was demonstrated using z-VDVAD-fmk, a caspase-2 inhibitor. Preincubation with this inhibitor reduced caspase-3 activation with around 50%, suggesting that caspase-2 is an important molecule, located upstream of caspase-3, in the signaling between DNA double stranded breaks and apoptosis signaling in AML cells. In conclusion, we demonstrate that GO and daunorubicin, at clinically applicable concentrations, induce pro-apoptotic signaling which involve activation of caspase-2, and activation of the BH3-only protein Bid. Blocking caspase-2 activity reduced caspase-3 activation with 50%. This suggests multiple signaling pathways capable of activating caspase-3. The caspase-2 pathway appears central for downstream caspase-3 activity, after GO treatment in AML. Our findings may highlight possible resistance mechanisms in AML, which might have profound therapeutic implications. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4090. doi:10.1158/1538-7445.AM2011-4090

Acute myeloid leukemia (AML) is one of the most frequently occurring and fatal types of leukemia. Initiated by genetic alterations in hematopoietic stem and progenitor cells, rapidly proliferating cancer cells (leukemic blasts) infiltrate the bone marrow and damage healthy hematopoiesis. Subgroups of AML are defined by underlying molecular and cytogenetic abnormalities, which are decisive for treatment and prognosis. For AML patients that can be intensively treated, the first line treatment remains a combination of cytarabine and anthracycline, which was developed in the 1970s. While this treatment regimen clears the disease and reinstates normal hematopoiesis (complete remission, CR) in 60% to 80% of patients below the age of 60, CR rates in patients above the age of 60 are only 40% to 50%. Relapse and refractory disease are the major cause of death of AML patients, despite large efforts to improve risk-adjusted post-remission therapy with further chemotherapy cycles and, if possible, allogeneic bone marrow transplantation. Elderly patients are particularly difficult to treat because of age-related comorbidities and because their disease tends to relapse more often than the disease of younger patients. Thus, the cure rates of AML vary with age, with 5-year survival rates of about 50% in young patients, and less than 20% in patients above the age of 65 years. With the median age of AML patients being 68 years, the need for novel therapeutic options is immense. The recent approval of eight new agents (venetoclax, midostaurin, gilteritinib, glasdegib, ivosidenib, enasidenib, gemtuzumab ozogamicin and CPX-351 (liposomal cytarabine and daunorubicin)) has added considerably to the therapeutic armamentarium of AML and has increased cure rates in specific subgroups of AML. However, the high heterogeneity among patients, clonal evolution and commonly occurring drug resistance, which cause the high relapse rates, remain a substantial problem in the treatment of AML. Therefore, a better understanding of currently used therapeutics and further development of novel therapeutics is urgently needed. In recent years, attention has increasingly focused on therapeutic strategies to interfere with the metabolic requirements of cancer cells. The last three decades have provided extensive insights into the diversity and flexibility of AML metabolism. AML cells use different sources of nutrients compared to normal hematopoietic progenitor cells and reprogram their metabolic pathways to fulfill their exquisite anabolic and energetic needs. As a result, they develop high metabolic plasticity that enables them to thrive in the bone marrow microenvironment, where oxygen and nutrient availability are subject to constant change. Cancer cells, specifically AML cells, have a strong dependency for the amino acid glutamine. Glutamine serves in energy production, redox control, cell signaling as well as an important nitrogen source. The only enzyme capable of de novo glutamine synthesis is glutamine synthetase (GS). GS catalyzes glutamine production from glutamate and ammonium. In AML, the metabolic role and dependency of GS is poorly understood. Here, we investigated the effects of GS deletion on AML growth, and its functional relevance in AML metabolism. Genetic deletion of GS resulted in a significant decrease of cell growth in vitro, and impaired leukemia progression in vivo in a xenotransplantation mouse model. Interestingly, the dependency of AML cell growth on GS was shown to be independent of its functional role in glutamine synthesis. Glutamine starvation did not increase the dependency of the AML cells on GS, nor did increased glutamine availability rescue the GS-knockout-associated growth disadvantage. Instead, functional studies revealed the role of GS in the detoxification of ammonium. GS-deficient cells showed elevated ammonium secretion as well as a higher sensitivity towards the toxic metabolite. Exogenous provision of 15N-labeled ammonium was detoxified by GS-driven incorporation into glutamine. Studies on cells that had gained resistance to GS-knockout-mediated growth inhibition indicated enzymes involved in the urea cycle and the arginine biogenesis pathway to compensate for a loss of GS. Together, these findings unveiled GS as an important ammonium scavenger in AML. Clinical studies on AML patients revealed increased ammonium concentrations in the blast-infiltrated bone marrow compared to peripheral blood. In line with this finding, proteome and transcriptome analysis of AML blasts showed a significant upregulation of GS in AML compared to healthy progenitors, further indicating its importance in ammonium detoxification. Analyzing pathways that contribute to ammonium production revealed protein uptake followed by amino acid catabolism as a yet not identified mechanism supporting AML growth. Protein endocytosis and subsequent proteolytic degradation were shown to rescue AML cells from otherwise growth-inhibiting glucose or amino acid depletion. Furthermore, protein metabolization led to the reactivation of the mammalian target of rapamycin (mTOR) signaling pathway, which was deactivated upon leucine and glutamine depletion, revealing protein consumption as an important alternative source of amino acids in AML. ...