Cyrus Khandanpour

A Dominant-Negative GFI1B Mutation in the Gray Platelet Syndrome

The gray platelet syndrome is a hereditary, usually autosomal recessive bleeding disorder caused by a deficiency of alpha granules in platelets. We detected a nonsense mutation in the gene encoding the transcription factor GFI1B (growth factor independent 1B) that causes autosomal dominant gray platelet syndrome. Both gray platelets and megakaryocytes had abnormal marker expression. In addition, the megakaryocytes had dysplastic features, and they were abnormally distributed in the bone marrow. The GFI1B mutant protein inhibited nonmutant GFI1B transcriptional activity in a dominant-negative manner. Our studies show that GFI1B, in addition to being causally related to the gray platelet syndrome, is key to megakaryocyte and platelet development.

Functional inhibition of mesenchymal stromal cells in acute myeloid leukemia

Hematopoietic insufficiency is the hallmark of acute myeloid leukemia (AML) and predisposes patients to life-threatening complications such as bleeding and infections. Addressing the contribution of mesenchymal stromal cells (MSC) to AML-induced hematopoietic failure we show that MSC from AML patients (n=64) exhibit significant growth deficiency and impaired osteogenic differentiation capacity. This was molecularly reflected by a specific methylation signature affecting pathways involved in cell differentiation, proliferation and skeletal development. In addition, we found distinct alterations of hematopoiesis-regulating factors such as Kit-ligand and Jagged1 accompanied by a significantly diminished ability to support CD34+ hematopoietic stem and progenitor cells in long-term culture-initiating cells (LTC-ICs) assays. This deficient osteogenic differentiation and insufficient stromal support was reversible and correlated with disease status as indicated by Osteocalcin serum levels and LTC-IC frequencies returning to normal values at remission. In line with this, cultivation of healthy MSC in conditioned medium from four AML cell lines resulted in decreased proliferation and osteogenic differentiation. Taken together, AML-derived MSC are molecularly and functionally altered and contribute to hematopoietic insufficiency. Inverse correlation with disease status and adoption of an AML-like phenotype after exposure to leukemic conditions suggests an instructive role of leukemic cells on bone marrow microenvironment.

The human GFI136N variant induces epigenetic changes at the Hoxa9 locus and accelerates K-RAS driven myeloproliferative disorder in mice

The coding single nucleotide polymorphism GFI136N in the human gene growth factor independence 1 (GFI1) is present in 3%-7% of whites and increases the risk for acute myeloid leukemia (AML) by 60%. We show here that GFI136N, in contrast to GFI136S, lacks the ability to bind to the Gfi1 target gene that encodes the leukemia-associated transcription factor Hoxa9 and fails to initiate histone modifications that regulate HoxA9 expression. Consistent with this, AML patients heterozygous for the GFI136N variant show increased HOXA9 expression compared with normal controls. Using ChipSeq, we demonstrate that GFI136N specific epigenetic changes are also present in other genes involved in the development of AML. Moreover, granulomonocytic progenitors, a bone marrow subset from which AML can arise in humans and mice, show a proliferative expansion in the presence of the GFI136N variant. In addition, granulomonocytic progenitors carrying the GFI136N variant allele have altered gene expression patterns and differ in their ability to grow after transplantation. Finally, GFI136N can accelerate a K-RAS driven fatal myeloproliferative disease in mice. Our data suggest that the presence of a GFI136N variant allele induces a preleukemic state in myeloid precursors by deregulating the expression of Hoxa9 and other AML-related genes.

Hand-foot syndrome: common presentation in an uncommon situation.

Following several relapses with follicular lymphoma, a 61-year-old female patient was diagnosed in July 2011 with diffuse large cell B-cell lymphoma. According to the German ALL GMALL B-ALL/NHL 2002 study protocol, the patient was treated with six cycles of chemotherapy containing high doses of methotrexate. The treatment was well tolerated until the application of the final course. Following treatment with vincristine, methotrexate, cyclophosphamide and adriamycin, the patient developed a pronounced hand– foot syndrome (HFS) (Fig. 1 Panel A). Also known as palmoplantar erythrodysesthesia, HFS is the result of toxic damage to the skin. Moreover, HFS has become a frequent side effect of targeted cancer therapies (e.g. sorafenib, sunitinib), newer drugs (e.g. capecitabine, pegylated liposomal doxorubicin) and following exposure to common substances (e.g. anthracyclines, taxanes, pyrimidine analogues). Clinically, HFS is characterised by swelling, reddening and desquamation of the palms and soles, which is accompanied by numbness and severe pain. Treatment recommendations include the discontinuation or reduction in exposure to the reactive substance, cooling of the skin, topical treatment with steroids, urea, uridine or DMSO, as well as systemic therapy (e.g. pyridoxine, steroids). In the present case, the affected skin of the patient was cooled, treated topically with steroids and uridine, and the patient received an oral prescription of pyridoxine and appropriate pain medication. After 2 weeks, the skin lesions had begun to regress (Fig. 1 Panel B, C) and eventually the patient recovered completely.

Gfi1b: a key player in the genesis and maintenance of acute myeloid leukemia and myelodysplastic syndrome

Differentiation of hematopoietic stem cells is regulated by a concert of different transcription factors. Disturbed transcription factor function can be the basis of (pre)malignancies such as myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML). Growth factor independence 1b (Gfi1b) is a repressing transcription factor regulating quiescence of hematopoietic stem cells and differentiation of erythrocytes and platelets. Here, we show that low expression of Gfi1b in blast cells is associated with an inferior prognosis of MDS and AML patients. Using different models of human MDS or AML, we demonstrate that AML development was accelerated with heterozygous loss of Gfi1b, and latency was further decreased when Gfi1b was conditionally deleted. Loss of Gfi1b significantly increased the number of leukemic stem cells with upregulation of genes involved in leukemia development. On a molecular level, we found that loss of Gfi1b led to epigenetic changes, increased levels of reactive oxygen species, as well as alteration in the p38/Akt/FoXO pathways. These results demonstrate that Gfi1b functions as an oncosuppressor in MDS and AML development.

GFI1 is required for RUNX1/ETO positive acute myeloid leukemia

Author(s): Marneth, Anna E; Botezatu, Lacramioara; Hones, Judith M; Israel, Jimmy CL; Schutte, Judith; Vassen, Lothar; Lams, Robert F; Bergevoet, Saskia M; Groothuis, Laura; Mandoli, Amit; Martens, Joost HA; Huls, Gerwin; Jansen, Joop H; Duhrsen, Ulrich; Berg, Tobias; Moroy, Tarik; Wichmann, Christian; Lo, Mia-Chia; Zhang, Dong-Er; van der Reijden, Bert A; Khandanpour, Cyrus

High Frequencies of Anti-Host Reactive CD8+ T Cells Ignore Non-Hematopoietic Antigen after Bone Marrow Transplantation in a Murine Model.

Background: Graft versus host disease (GvHD) occurs in 20% of cases with patients having an MHC I matched bone marrow transplantation (BMT). Mechanisms causing this disease remain to be studied. Methods: Here we used a CD8+ T cell transgenic mouse line (P14/CD45.1+) and transgenic DEE mice bearing ubiquitously the glycoprotein 33-41 (GP33) antigen derived from the major lymphocytic choriomeningitis virus (LCMV) epitope to study mechanisms of tolerance in anti-host reactive CD8+ T cells after BMT. Results: We found that anti-host reactive CD8+ T cells (P14 T cells) were not negatively selected in the thymus and that they were present in wild type (WT) recipient mice as well as in DEE recipient mice. Anti-host reactive CD8+ T cells ignored the GP33 antigen expressed ubiquitously by host cells but they could be activated ex vivo via LCMV-infection. Lipopolysaccharides (LPS) induced transient cell damage in DEE mice bearing anti-host reactive CD8+ T cells after BMT, suggesting that induction of host inflammatory response could break antigen ignorance. Introducing the GP33 antigen into BM cells led to deletion of anti-host reactive CD8+ T cells. Conclusion: We found that after BMT anti-host reactive CD8+ T cells ignored host antigen in recipients and that they were only deleted when host antigen was present in hematopoietic cells. Moreover, LPS-induced immune activation contributed to induction of alloreactivity of anti-host reactive CD8+ T cells after BMT.