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Supplementary Materials and Methods, Supplementary Figures S1-S12

Supplementary Table 3 HD and MPN samples.

Supplementary Table 1 VEGFAC Top Differentially Expressed Genes by Cluster

MM ALL Donor Details

Supplementary Table 2 Gene sets for GSEA

Supplementary Table 6 qRT PCR Probes

Supplementary Table 5 Antibodies

Supplementary Table 7 NGS Panel

A lack of models that recapitulate the complexity of human bone marrow has hampered mechanistic studies of normal and malignant hematopoiesis and the validation of novel therapies. Here, we describe a step-wise, directed-differentiation protocol in which organoids are generated from induced pluripotent stem cells committed to mesenchymal, endothelial, and hematopoietic lineages. These 3D structures capture key features of human bone marrow—stroma, lumen-forming sinusoids, and myeloid cells including proplatelet-forming megakaryocytes. The organoids supported the engraftment and survival of cells from patients with blood malignancies, including cancer types notoriously difficult to maintain ex vivo. Fibrosis of the organoid occurred following TGFβ stimulation and engraftment with myelofibrosis but not healthy donor–derived cells, validating this platform as a powerful tool for studies of malignant cells and their interactions within a human bone marrow–like milieu. This enabling technology is likely to accelerate the discovery and prioritization of novel targets for bone marrow disorders and blood cancers.Significance:We present a human bone marrow organoid that supports the growth of primary cells from patients with myeloid and lymphoid blood cancers. This model allows for mechanistic studies of blood cancers in the context of their microenvironment and provides a much-needed ex vivo tool for the prioritization of new therapeutics.See related commentary by Derecka and Crispino, p. 263.This article is highlighted in the In This Issue feature, p. 247

Phylogenetic relationships in Mentha were inferred from DNA sequences of the chloroplast (cp) rpl16 intron and trnL-trnF region. The objectives were to test monophyly of Mentha and each of its sections and assess relationships among Mentha species. Based on cpDNA data, Mentha is strongly supported as monophyletic. The suggestions that M. cervina and M. cunninghamii should be placed in other genera gain no support from this analysis. Mentha cervina is sister to M. gattefossei, and the New Zealand endemic, M. cunninghamii, is allied with species from Australia (M. australis, M. diemenica, M. satureioides). No recognized section with more than one species sampled forms a monophyletic group. Section Pulegium may be monophyletic, but the third member of this section, M. grandiflora, was not sampled. Chloroplast DNA sequences of the putative allopolyploids M. canadensis and M. spicata suggest that M. arvensis and M. longifolia, respectively, may be their maternal parents.

Hematopoietically expressed homeobox (Hhex) is a T-cell oncogene. It is frequently deregulated in murine retroviral insertional mutagenesis screens and its enforced expression induces T-cell leukemia in bone marrow transduction and transplantation experiments. We discovered that HHEX is a direct transcriptional target of an LIM domain Only-2 (LMO2)-associated protein complex. HHEX clusters with LMO2-overexpressing T-ALLs and is especially overexpressed in Early T-cell Precursor (ETP) – ALL where it is a direct transcriptional target of LMO2. To further understand Hhex's function, we induced a conditional knockout in floxed Hhex mice with the Vav-iCre transgene. Mice were viable and showed normal blood cell counts with highly efficient deletion of Hhex in all hematopoietic tissues. Thymocytes from conditional knockouts showed a normal pattern of development. Most impressively, Hhex conditional knockout markedly prolonged the latency of T-ALL onset in CD2-Lmo2 transgenic mice (figure 1). Hhex conditional knockouts (Hhex cKOs) also had a significant decrease in mature B cells in the spleen and bone marrow. Interestingly, hematopoietic stem and progenitor cells plated on OP9-GFP or OP9-DL1 stromal cells showed proliferative defects and incomplete differentiation towards both B and T lineage. Also under stress conditions such as sublethal irradiation and competitive bone marrow transplants, Hhex conditional knockouts show a marked defect in both B and T lineages but an increase in early progenitor populations. Our experiments show that Hhex is a critical transcription factor in lymphoid development and in LMO2-induced T-ALL.Figure 1Hhex conditional knockout markedly prolonged the latency of T-ALL onset in CD2-Lmo2 transgenic miceFigure 1. Hhex conditional knockout markedly prolonged the latency of T-ALL onset in CD2-Lmo2 transgenic mice Disclosures: No relevant conflicts of interest to declare.