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Our current understanding of human hematopoiesis has undergone significant transformation throughout the years, challenging conventional views. The evolution of high-throughput technologies has enabled the accumulation of diverse data types, offering new avenues for investigating key regulatory processes in blood cell production and disease. In this review, we will explore the opportunities presented by these advancements for unraveling the molecular mechanisms underlying normal and abnormal hematopoiesis. Specifically, we will focus on the importance of enhancer-associated regulatory networks and highlight the crucial role of enhancer-derived transcription regulation. Additionally, we will discuss the unprecedented power of single-cell methods and the progression in using in vitro human blood differentiation system, in particular induced pluripotent stem cell models, in dissecting hematopoietic processes. Furthermore, we will explore the potential of ever more nuanced patient profiling to allow precision medicine approaches. Ultimately, we advocate for a multiparameter, regulatory network-based approach for providing a more holistic understanding of normal hematopoiesis and blood disorders.

Guilherme Castela,1– 3 Joana Providência,1,2 Madalena Monteiro,1 Bárbara Oliveiros,2 Sónia Silva,4 Manuel Brito,4 Egídio Machado,5 Joaquim Neto Murta,1,2 Miguel Castelo-Branco,2,3,* Zélia Correa6,* 1Department of Ophthalmology, Centro Hospitalar e Universitário de Coimbra, Coimbra, Portugal; 2Faculty of Medicine, University of Coimbra (FMUC), Coimbra, Portugal; 3Coimbra Institute for Biomedical Imaging and Translational Research (CIBIT), Coimbra, Portugal; 4Department of Pediatric Oncology, Centro Hospitalar e Universitário de Coimbra, Coimbra, Portugal; 5Department of Medical Imaging, Centro Hospitalar e Universitário de Coimbra, Coimbra, Portugal; 6University of Miami, Bascom Palmer Eye Institute, Miami, FL, USA*These authors contributed equally to this workCorrespondence: Guilherme Castela, Department of Ophthalmology, Centro Hospitalar e Universitário de Coimbra, Praceta Prof Mota Pinto, Coimbra, 3004-561, Portugal, Tel +351919702206, Email gcastela@hotmail.comPurpose: We aim to report about effectiveness and safety in the context of our centers’ setting in the management of retinoblastoma with intra-arterial chemotherapy (IAC) in a 5-year retrospective analysis of the Portuguese population.Patients and Methods: Retrospective analysis of consecutive cases of retinoblastoma selected to initiate IAC between 2015 and 2020, at the Portuguese National Reference Center. All included patients underwent complete ophthalmological evaluation under anesthesia with fundus photography. Diagnosis and classification of retinoblastoma was made according to the International Classification of Intraocular Retinoblastoma (ICRB). The patients were further divided into two groups: Group I for primary IAC and Group II for secondary IAC. Tumor recurrence or relapses, systemic metastasis and deaths were documented. Main efficacy outcome included ocular salvage and recurrence-free survival rates estimated using the Kaplan–Meier method.Results: Twenty-eight eyes (19 eyes included in Group I and 9 eyes included in Group II) were eligible and a total of 130 IAC procedures were performed, with a median number of sessions of 4 (range 1– 8) for each treated eye, during a median follow-up of 21 months (range 4– 64). Of the included eyes, 22 (78.6%) were preserved. An overall survival of 100% was achieved. Considering the preserved eyes, the overall median decimal visual acuity achieved at the last visit was 0.15 (range 0.02– 0.8). Three patients had permanent adverse events related to IAC (cataract, vitreous hemorrhage and choroidal ischemia). Considering the survival analysis of recurrence, the mean survival without recurrence was 84.2% for Group I and 66.7% for Group II, and the mean survival without enucleation was 78.6% (no events in Group II).Conclusion: IAC has been shown to be an effective and safe treatment for children with intraocular retinoblastoma. This study demonstrates that IAC is effective even in moderate sample sizes, when a multidisciplinary approach is available.Keywords: retinoblastoma, intra-arterial chemotherapy, ocular oncology, pediatric oncology

Background Adjuvants are key for effective vaccination against cancer and chronic infectious diseases. Saponin-based adjuvants (SBAs) are unique among adjuvants in their ability to induce robust cell-mediated immune responses in addition to antibody responses. Recent preclinical studies revealed that SBAs induced cross-presentation and lipid bodies in otherwise poorly cross-presenting CD11b+ murine dendritic cells (DCs).Method Here, we investigated the response of human DC subsets to SBAs with RNA sequencing and pathway analyses, lipid body induction visualized by laser scanning microscopy, antigen translocation to the cytosol, and antigen cross-presentation to CD8+ T cells.Results RNA sequencing of SBA-treated conventional type 1 DC (cDC1) and type 2 DC (cDC2) subsets uncovered that SBAs upregulated lipid-related pathways in CD11c+ CD1c+ cDC2s, especially in the CD5− CD163+ CD14+ cDC2 subset. Moreover, SBAs induced lipid bodies and enhanced endosomal antigen translocation into the cytosol in this particular cDC2 subset. Finally, SBAs enhanced cross-presentation only in cDC2s, which requires the CD163+ CD14+ cDC2 subset.Conclusions These data thus identify the CD163+ CD14+ cDC2 subset as the main SBA-responsive DC subset in humans and imply new strategies to optimize the application of saponin-based adjuvants in a potent cancer vaccine.

Guilherme Castela,1,2 Joana Providência,1,2 Madalena Monteiro,1 Sónia Silva,3 Manuel Brito,3 Joaquim Neto Murta,1,2 Zélia Correa,4 Miguel Castelo Branco2 1Department of Ophthalmology, Centro Hospitalar e Universitário de Coimbra, Coimbra, Portugal; 2Faculty of Medicine, University of Coimbra, Coimbra, Portugal; 3Department of Pediatric Oncology, Centro Hospitalar e Universitário de Coimbra, Coimbra, Portugal; 4Bascon Palmer Eye Institute, Miami, FL, USACorrespondence: Guilherme CastelaDepartment of Ophthalmology, Centro Hospitalar e Universitário de Coimbra, Avenida Dias da Silva 164, Coimbra, 3000, PortugalEmail gcastela@hotmail.comObjective: The aim of our study was to characterize the evacuated African patients diagnosed with retinoblastoma and referred to the Portuguese national referral center (Centro Hospital e Universitário de Coimbra, University of Coimbra), identifying inequalities in the stage of diagnosis and prognostic results.Design: Retrospective observational study of evacuated African patients diagnosed with retinoblastoma and referred to the Portuguese National Referral Center (Centro Hospital e Universitário de Coimbra, University of Coimbra).Results: The study included 15 patients between October 2015 and October 2020 from Angola, Cape Verde, Guinea-Bissau and São Tomé and Príncipe. Seven (46.7%) children presented bilateral retinoblastoma. The median age at the time of diagnosis was 20.9 (interquartile range, 16– 41) months. The presenting symptoms were leukocoria (86.7%), strabism (53.3%) and buphthalmus (40%). In terms of tumor staging, five (33.3%) children presented with extraocular retinoblastoma and 10 (66.7%) children presented with intraocular retinoblastoma. At presentation, no pineal involvement was diagnosed but two (13.3%) children presented with central nervous system involvement at the time of the first observation. Children were treated with enucleation, exenteration, systemic chemotherapy, intra-arterial chemotherapy and/or supportive palliative care. During the follow-up period (mean 27.2 ± 18.2 months), the overall survival was 73.3%.Conclusion: A small proportion of African children are being referred to our center, when considering the expected incidence of retinoblastoma in these countries, and referred children arrive at advanced stages of the disease, compromising treatment outcomes. Considering retinoblastoma is now a curable disease, national and international interventions are required to attempt a better management of children born in low-income countries.Keywords: retinoblastoma, low income countries, treatment of advanced disease

A t(9;11)(p22;q23) translocation produces the MLL-AF9 fusion protein, which is found in up to 25% of de novo AML cases in children. Despite major advances, obtaining a comprehensive understanding of context-dependent MLL-AF9-mediated gene programs during early hematopoiesis is challenging. Here, we generated a human inducible pluripotent stem cell (hiPSC) model with a doxycycline dose-dependent MLL-AF9 expression. We exploited MLL-AF9 expression as an oncogenic hit to uncover epigenetic and transcriptomic effects on iPSC-derived hematopoietic development and the transformation into (pre-)leukemic states. In doing so, we observed a disruption in early myelomonocytic development. Accordingly, we identified gene profiles that were consistent with primary MLL-AF9 AML and uncovered high-confidence MLL-AF9-associated core genes that are faithfully represented in primary MLL-AF9 AML, including known and presently unknown factors. Using single-cell RNA-sequencing, we identified an increase of CD34 expressing early hematopoietic progenitor-like cell states as well as granulocyte-monocyte progenitor-like cells upon MLL-AF9 activation. Our system allows for careful chemically controlled and stepwise in vitro hiPSC-derived differentiation under serum-free and feeder-free conditions. For a disease that currently lacks effective precision medicine, our system provides a novel entry-point into exploring potential novel targets for personalized therapeutic strategies.

In this paper we give algorithms that allow to compute the set of every elementary numerical semigroups with given genus, Frobenius number and multiplicity. As a consequence we obtain formulas for the cardinality of these sets. [ABSTRACT FROM AUTHOR]