Alicia Perzolli

Although immunotherapy has revolutionized cancer treatment, its benefits in pediatric acute myeloid leukemia (AML) remain limited, partly due to an incomplete understanding of the AML microenvironment. While most studies focus on T cells, innate immune populations such as macrophages are less well explored. In pediatric AML patients, we demonstrate that immunosuppressive macrophages dominate the AML microenvironment and support growth of RUNX1::RUNX1T1 and KMT2A -rearranged leukemic blasts. In co-culture experiments, AML cells reprogrammed naïve and pro-inflammatory macrophages toward an anti-inflammatory state, characterized by suppression of NF-κB and other inflammatory pathways. This phenotypic switch requires direct cell contact and involves tunneling nanotube–mediated transfer of AML mitochondria to macrophages associated with metabolic rewiring toward oxidative phosphorylation. Functionally, these reprogrammed macrophages exhibited impaired phagocytosis, convey chemoresistance and reduced T cell– and bispecific antibody–mediated killing, facilitating immune escape. Our findings reveal a novel mechanism of immune suppression and suggest that targeting AML-macrophage interactions could enhance the efficacy of T cell–based immunotherapies.

SUMMARY: A better understanding of lymphocyte dynamics during current treatment regimens in pediatric AML is urgently needed to understand whether the application of bispecific T-cell-engagers (BiTEs) during periods of low tumor burden could be a viable treatment strategy. In this study, we found that induction 1, comprising mitoxantrone-etoposide-cytarabine in nearly all patients (as part of the NOPHO-DBH AML-2012 protocol), led to preserved or increased relative lymphocyte abundances alongside marked blast reduction in most cases. This was accompanied by a shift towards higher T-cell fractions, potentially creating a favorable window for BiTE therapy. The absence of a correlation between blast reduction and lymphocyte changes suggests that chemotherapy exerts differential effects on the lymphocyte compartment. Despite the heterogeneity of agents used in induction 2, more than half of patients showed a decline in lymphocyte levels. Nonetheless, the increase in T- and B-cells observed in most patients from the NOPHO-AML 2004 cohort after induction 2 suggests that lymphocyte recovery at this treatment stage is not uniformly impaired. Our transcriptomic and ex vivo functional data align with preclinical findings in adult AML and provide a basis for further investigations in in vivo models and early clinical trials. Such efforts should prioritize novel BiTE constructs targeting multiple tumor-associated (e.g., NCT05673057 ) or tumor-specific antigens.

Background: Proteolysis targeting chimeras (PROTACs) are heterobifunctional small molecules that utilize the ubiquitin–proteasome system to selectively degrade target proteins. This innovative technology has shown remarkable efficacy and specificity in degrading oncogenic proteins and has progressed through various stages of preclinical and clinical development for hematologic malignancies, including adult acute myeloid leukemia (AML). However, the application of PROTACs in pediatric AML remains largely unexplored. Methods: In this study, we show the potent effect of GSPT1 degradation against AML cells induced by either a GSPT1-selective cereblon modulator CC-90009 or by an off-target effect caused by a CDK6-PROTAC named GU3341. Results: Both in vitro and ex vivo experiments revealed that GSPT1 degradation significantly inhibited tumor growth, induced cell cycle arrest, and triggered apoptosis in two pediatric AML subtypes characterized by RUNX1::RUNX1T1 and FUS::ERG fusion genes. Furthermore, the degradation of GSPT1 impaired the expression of RUNX1::RUNX1T1 and its cooperating transcription factors RUNX1 and ERG. Similarly, GSPT1 degradation also reduced FUS::ERG fusion transcript levels in AML cells harboring the translocation t(16;24)(p11:q22). Conclusions: These findings suggest a new role of GSPT1 in regulating leukemic transcriptional networks and open a new therapeutic strategy to target leukemic fusion genes in pediatric AML patients.

Because of the low mutational burden and consequently, fewer potential neoantigens, children with acute myeloid leukemia (AML) are thought to have a T cell-depleted or ‘cold’ tumor microenvironment and may have a low likelihood of response to T cell-directed immunotherapies. Understanding the composition, phenotype, and spatial organization of T cells and other microenvironmental populations in the pediatric AML bone marrow (BM) is essential for informing future immunotherapeutic trials about targetable immune-evasion mechanisms specific to pediatric AML. Here, we conducted a multidimensional analysis of the tumor immune microenvironment in pediatric AML and non-leukemic controls. We demonstrated that nearly one-third of pediatric AML cases has an immune-infiltrated BM, which is characterized by a decreased ratio of M2- to M1-like macrophages. Furthermore, we detected the presence of large T cell networks, both with and without colocalizing B cells, in the BM and dissected the cellular composition of T- and B cell-rich aggregates using spatial transcriptomics. These analyses revealed that these aggregates are hotspots of CD8⁺ T cells, memory B cells, plasma cells and/or plasmablasts, and M1-like macrophages. Collectively, our study provides a multidimensional characterization of the BM immune microenvironment in pediatric AML and indicates starting points for further investigations into immunomodulatory mechanisms in this devastating disease.

While the introduction of T cell-based immunotherapies has improved outcomes in many cancer types, the development of immunotherapies for both adult and pediatric AML has been relatively slow and limited. In addition to the need to identify suitable target antigens, a better understanding of the immunosuppressive tumor microenvironment is necessary for the design of novel immunotherapy approaches. To date, most immune characterization studies in AML have focused on T cells, while innate immune lineages such as monocytes, granulocytes and natural killer (NK) cells, received less attention. In solid cancers, studies have shown that innate immune cells, such as macrophages, myeloid-derived suppressor cells and neutrophils are highly plastic and may differentiate into immunosuppressive cells depending on signals received in their microenvironment, while NK cells appear to be functionally impaired. Hence, an in-depth characterization of the innate immune compartment in the TME is urgently needed to guide the development of immunotherapeutic interventions for AML. In this review, we summarize the current knowledge on the innate immune compartment in AML, and we discuss how targeting its components may enhance T cell-based- and other immunotherapeutic approaches.

DHX30 was recently implicated in the translation control of mRNAs involved in p53-dependent apoptosis. Here, we show that DHX30 exhibits a more general function by integrating the activities of its cytoplasmic isoform and of the more abundant mitochondrial one. The depletion of both DHX30 isoforms in HCT116 cells leads to constitutive changes in polysome-associated mRNAs, enhancing the translation of mRNAs coding for cytoplasmic ribosomal proteins while reducing the translational efficiency of the nuclear-encoded mitoribosome mRNAs. Furthermore, the depletion of both DHX30 isoforms leads to higher global translation but slower proliferation and lower mitochondrial energy metabolism. Isoform-specific silencing supports a role for cytoplasmic DHX30 in modulating global translation. The impact on translation and proliferation was confirmed in U2OS and MCF7 cells. Exploiting RIP, eCLIP, and gene expression data, we identified fourteen mitoribosome transcripts we propose as direct DHX30 targets that can be used to explore the prognostic value of this mechanism in cancer. We propose that DHX30 contributes to cell homeostasis by coordinating ribosome biogenesis, global translation, and mitochondrial metabolism. Targeting DHX30 could, thus, expose a vulnerability in cancer cells.