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A goal in precision medicine is to use patient-derived material to predict disease course and intervention outcomes. Here, we use mechanistic observations in a preclinical animal model to design an ex vivo platform that recreates genetic susceptibility to T-cell-mediated damage. Intestinal graft-versus-host disease (GVHD) is a life-threatening complication of allogeneic hematopoietic cell transplantation. We found that intestinal GVHD in mice deficient in Atg16L1, an autophagy gene that is polymorphic in humans, is reversed by inhibiting necroptosis. We further show that cocultured allogeneic T cells kill Atg16L1-mutant intestinal organoids from mice, which was associated with an aberrant epithelial interferon signature. Using this information, we demonstrate that pharmacologically inhibiting necroptosis or interferon signaling protects human organoids derived from individuals harboring a common ATG16L1 variant from allogeneic T-cell attack. Our study provides a roadmap for applying findings in animal models to individualized therapy that targets affected tissues., (© 2020 by The American Society of Hematology.)

The hemophagocytic lymphohistiocytoses (HLH) comprise a heterogeneous group of disorders characterized by dysregulated activation of T cells and macrophages. Although some patients with HLH harbor perforin gene mutations, the cause of the remaining cases is not known. The phenotype of HLH bears a strong resemblance to X-linked lymphoproliferative disease (XLP), an Epstein-Barr virus (EBV)-associated immunodeficiency resulting from defects in SH2D1A, a small SH2 domain-containing protein expressed in T lymphocytes and natural killer cells. Here it is shown that 4 of 25 male patients with HLH who were examined harbored germline SH2D1A mutations. Among these 4 patients, only 2 had family histories consistent with XLP. On the basis of these findings, it is suggested that all male patients with EBV-associated hemophagocytosis be screened for mutations in SH2D1A. Patients identified as having XLP should undergo genetic counseling, and be followed long-term for development of lymphoma and hypogammaglobulinemia.

In this study we examine the effects of amino acid deprivation on the growth and differentiation of the human HL-60 myeloid leukemia cell line. The HL-60 cell line was chosen for study because of its ability to differentiate along either a granulocytic or monocytic pathway under appropriate culture conditions. Differentiation was determined by changes in cell morphology, nonspecific esterase (NSE) content, hydrogen peroxide (H2O2) production, and expression of the cell surface differentiation antigens LeuM3 (CD14) and OKM1 (CD11). Using a model system in which HL-60 cells were cultured in medium that selectively lacked one amino acid (AA), it was seen that deprivation of HL-60 cells for essential (but not nonessential) AAs results in decreased cell growth and viability and in differentiation of 30% to 60% of the surviving population of cells specifically along the monocytic pathway. This differentiation is irreversible as well as time- and dose-dependent. Culture of HL-60 cells in essential AA-deficient medium potentiated the differentiative effects of recombinant human interferon-gamma (IFN-gamma), recombinant human tumor necrosis factor (TNF), and dihydroxyvitamin D3 (D3), all of which have previously been shown to induce monocytic differentiation of HL-60 cells. Differentiated cells had decreased DNA and RNA synthesis, but protein synthesis was unchanged compared with control cells. The protein synthesis inhibitor cycloheximide prevented differentiation, indicating the necessity of protein synthesis in this process. Cell cycle analysis revealed that an increased proportion of cells cultured in AA-deficient medium was arrested in G0-G1 (80% and 50% for AA-deficient and control cells, respectively). These results suggest that alterations of AA metabolism and subsequent perturbations in DNA and RNA synthesis may be important in initiating differentiation or in augmenting cytokine-induced differentiation of HL-60 cells into more mature, nonreplicating, monocyte-like cells.

Previously we showed that starvation of HL-60 promyelocytic leukemia cells for a single essential amino acid induced irreversible differentiation into more mature monocyte-like cells. Although not an essential amino acid, glutamine is important in the growth of normal and neoplastic cells. The glutamine analogue, alpha S,5S-alpha-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid (acivicin) inhibits several glutamine-utilizing enzymes and therefore depletes cells of certain metabolic end products. The current study was designed to examine in vitro the effects of acivicin on growth and differentiation of several established human myeloid leukemia cell lines, including the HL-60 cell line, and of freshly isolated cells from patients with acute nonlymphocytic leukemia (ANLL). Four-day culture of HL-60 cells with acivicin at concentrations of 0.1 to 10.0 micrograms/mL (0.56 to 56 nmol/L) decreased cell growth by 33% to 88% as compared with untreated control cells. Viability of cells was greater than 92% for untreated cells and 93% to 41% for acivicin-treated cells. Cells treated with acivicin differentiated along a monocytic pathway as shown by increased H2O2 production and alpha-naphthyl butyrate esterase (NSE) content. Differentiation was time and dose dependent, and was irreversible. Changes in H2O2 production and NSE content were partially abrogated by co-culture with 10 mmol/L exogenous cytidine and guanosine but not by co-culture with other nucleosides or glutamine. At these concentrations of acivicin, differentiation was associated with expression of the N-formyl-methyl-leucyl-phenylalanine-receptor (FMLP-R) on 8% to 29% of cells as compared with 8% for control cells. Acivicin potentiated the differentiating effects of interferon-gamma, tumor necrosis factor, dihydroxyvitamin D3, dimethylsulfoxide, and retinoic acid. Culture of cells from the U937 (monoblastic), K562 (erythroleukemia), and KG-1 (myeloblastic) cell lines resulted in decreased growth and viability, but not consistently in differentiation. Acivicin decreased survival of freshly isolated ANLL cells and increased their H2O2 production and NSE content. These results suggest that the glutamine analogue acivicin may be useful as a differentiating agent with antileukemia activity in patients with ANLL.