Clonal Expansion of Tet2 +/- hematopoiesis Is Driven By Inflamm-Ageing Associated IL-1 Increase in Mice

Abstract Clonal Hematopoiesis of Indeterminate Potential (CHIP) is defined as the presence of an expanded somatic blood cell clone carrying a mutation in genes that are drivers of hematologic malignancy including DNMT3A, TET2, ASXL1, TP53, JAK2, and SF3B1, at a variant allele frequency (VAF) of at least 2% in the absence of other hematological abnormalities. CHIP has a prevalence of about 10% in the 70-80 year old population, further increases with ageing and associates with an increased risk of hematological malignancies, cardiovascular disease and all-cause mortality (Genovese et al. NEJM 2014; Jaiswal, S. et al. NEJM 2014). Recent studies indicate that higher pre-malignant clonal size and mutational burden increase the chances of malignant transformation in individuals carrying CHIP (Desai, P. Nat. Med. et al., 2018; Abelson, S. et al. Nature, 2018). While age is the best predictor of CHIP development and correlates directly with pre-malignant clonal size, the specific cellular-extrinsic factors promoting CHIP clonal expansion in the context of physiological aging are still unclear. We hypothesized that ageing associated low-grade inflammation (termed "inflamm-ageing") is a driver of CHIP clonal expansion. We used standard bone marrow (BM) chimera models and developed a novel, irradiation independent, hematopoietic specific and tamoxifen inducible genetic mosaicism mouse model of Tet2 +/- driven CHIP (HSC-Scl-Cre-ERT; Tet2+/flox; R26 +/floxstop-EGFP triple transgenic mice) to determine the contribution of inflamm-aging factors to Tet2 +/- hematopoieticclonal expansion. Using these complementary models, we observe that peripheral Tet2 +/- clonal expansion rates increase with age (evident in erythroid, myeloid, lymphoid B and T lineages), which is paralleled by a significant expansion of Tet2 +/- hematopoietic stem and progenitor cell (HSPCs) populations in aged mice (12-14 months old). Importantly, Tet2 +/- clonal expansion associates with increased levels of inflammatory cytokine IL-1 in aged mice, which derives partially from Tet2 +/- mutant mature hematopoietic cells. To test the contribution of IL-1 to Tet2 +/- clonal expansion, we administered IL-1 (0.5ug/day for 14 days) to young CHIP carrying mice (2-4 months of age) and observed an IL-1R1-dependent expansion of Tet2 +/- hematopoietic mature lineages and HSPCs. Dissection of the cellular mechanisms operating downstream of IL-1/IL-1R1 revealed that Tet2 +/- clonal expansion results from increased multilineage differentiation and associates with increased HSPC cell-cycle progression (while not depending on IL-1-mediated effects on HSPC viability). Moreover, Tet2 +/- HSPCs show a higher in vitro and in vivo repopulation capacity in response to prolonged IL-1 exposure compared to their WT counterparts. Finally, to directly test the contribution of IL-1 to drive Tet2 +/- clonal expansion in the context of physiological aging, we set up genetic (BM chimeras using donor BM from Tet2 +/-; Il-1r1-/- compound mutants) or pharmacological inhibited IL-1 signaling (Anakinra, hIL-1ra) during mouse ageing. Strikingly, both approaches prevented ageing-dependent Tet2 +/- clonal expansion, thus confirming IL-1 as key "inflamm-ageing" driver of Tet2 +/- clonal expansion. Overall, our data provide proof-of-concept that IL-1 production derived from aged BM cells is a relevant and targetable driver of Tet2 +/- clonal expansion in aged mice. Disclosures Manz: CDR-Life Inc: Consultancy, Current holder of stock options in a privately-held company; University of Zurich: Patents & Royalties: CD117xCD3 TEA.

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Aging, hematopoiesis, and the myelodysplastic syndromes

Blood Advances ◽

10.1182/bloodadvances.2017009852 ◽

2017 ◽

Vol 1 (26) ◽

pp. 2572-2578 ◽

Cited By ~ 12

Author(s):

Stephen S. Chung ◽

Christopher Y. Park

Keyword(s):

Myelodysplastic Syndromes ◽

Bone Marrow Failure ◽

Hematologic Malignancies ◽

Relative Increase ◽

Extrinsic Factors ◽

Hematopoietic Stem ◽

Clonal Hematopoiesis ◽

Increased Risk ◽

The Many ◽

The Relationship

Abstract The aging hematopoietic system undergoes numerous changes, including reduced production of red blood cells and lymphocytes as well as a relative increase in the production of myeloid cells. Emerging evidence indicates that many of these changes are due to selection pressures from cell-intrinsic and cell-extrinsic factors that result in clonal shifts in the hematopoietic stem cell (HSC) pool, resulting in predominant HSC clones that exhibit the functional characteristics associated with HSC aging. Given the recent descriptions of clonal hematopoiesis in aged populations, the increased risk of developing hematologic malignancies in individuals with clonal hematopoiesis, and the many similarities in hematopoietic aging and acquired bone marrow failure (BMF) syndromes, such as myelodysplastic syndromes (MDS), this raises significant questions regarding the relationship between aging hematopoiesis and MDS, including the factors that regulate HSC aging, whether clonal hematopoiesis is required for the development of MDS, and even whether BMF is an inevitable consequence of aging. In this article, we will review our current understanding of these processes and the potential intersections among them.

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Single-cell multiomics reveals increased plasticity, resistant populations and stem-cell-like blasts in KMT2A-rearranged leukemia

Blood ◽

10.1182/blood.2021013442 ◽

2021 ◽

Author(s):

Changya Chen ◽

Wenbao Yu ◽

Fatemeh Alikarami ◽

Qi Qiu ◽

Chia-hui Chen ◽

...

Keyword(s):

Single Cell ◽

Young Patients ◽

Cytotoxic Lymphocytes ◽

Extrinsic Factors ◽

Hematopoietic Stem ◽

Younger Patients ◽

Factors Affecting ◽

Immune Mediated ◽

Younger Age ◽

Increased Risk

KMT2A-rearranged (KMT2A-r) infant ALL is a devastating malignancy with a dismal outcome, and younger age at diagnosis is associated with increased risk of relapse. To discover age-specific differences and critical drivers that mediate poor outcome in KMT2A-r ALL, we subjected KMT2A-r leukemias and normal hematopoietic cells from patients of different ages to single cell multi-omics analyses. We uncovered the following critical new insights: leukemia cells from patients younger than 6 months have significantly increased lineage plasticity. Steroid response pathways are downregulated in the most immature blasts from younger patients. We identify a hematopoietic stem and progenitor-like (HSPC-like) population in the blood of younger patients that contains leukemic blasts and form an immunosuppressive signaling circuit with cytotoxic lymphocytes. These observations offer a compelling explanation for the ability of leukemias in young patients to evade chemotherapy and immune mediated control. Our analysis also revealed pre-existing lymphomyeloid primed progenitors and myeloid blasts at initial diagnosis of B-ALL. Tracking of leukemic clones in two patients whose leukemia underwent a lineage switch documented the evolution of such clones into frank AML. These findings provide critical insights into KMT2A-r ALL and have clinical implications for molecularly targeted and immunotherapy approaches. Beyond infant ALL, our study demonstrates the power of single cell multi-omics to detect tumor intrinsic and extrinsic factors affecting rare but critical subpopulations within a malignant population that ultimately determines patient outcome.

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FoxM1 Haploinsufficiency Drives Clonal Hematopoiesis

Blood ◽

10.1182/blood-2019-128429 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 726-726

Author(s):

Chunjie Yu ◽

Yue Sheng ◽

Zhijian Qian

Keyword(s):

Cell Cycle ◽

Progenitor Cells ◽

Low Dose ◽

Conflicts Of Interest ◽

Clonal Expansion ◽

Brdu Incorporation ◽

Quiescent State ◽

Dapi Staining ◽

Hematopoietic Stem ◽

Increased Risk

Hematopoiesis is an orchestrated process in which hematopoietic stem cells (HSCs) can self-renew and produce all lineages of blood cells. Majority of HSCs are in a quiescent state with a low growth rate. However, some genetic mutations that occur in HSCs impel HSCs to exit the quiescent state and to proliferate excessively, which enables mutant HSCs to outcompete normal HSCs and leads to clonal expansion of mutant HSCs. Myelodysplastic syndromes (MDSs) as a clonal disease, arise from the expansion of mutant HSCs and are characterized by morphologic dysplasia, ineffective hematopoiesis and an increased risk of transformation to acute myeloid leukemia. FoxM1 is one of transcription factors in the family of Fox ('Forkhead box') proteins. Analysis of public database revealed that the expression level of FOXM1 was decreased significantly in CD34 + cells from a subset of patients with MDS as compared to healthy individuals. Thus, we sought to determine whether haploinsufficiency of FOXM1 contributes to the development of MDS in mice. Our study showed that haploinsufficiency of Foxm1 led to an expansion of hematopoietic stem/progenitor cells in mice. Since FoxM1 has previously been implicated in regulation of cell cycle, we determined the cell cycle status of Foxm1 heterozygous HSCs. By BrdU incorporation assay, we showed that Foxm1 heterozygous HSCs have an increased S phase and G2/M phase as compared to control HSCs from wildtype mice. Additional analysis with Hochest33342/Pyronin-Y staining and Ki67/DAPI staining showed a significant decrease in the number of quiescent (G0) cells in Foxm1 heterozygous HSCs as compared to control HSCs. These results suggest that FoxM1 haploinsufficiency promotes HSCs to exit quiescence and to enter cell cycle, thereby leading to exhaustion of HSCs. To further assess the function of Foxm1 heterozygous HSCs in vivo, we performed competitive repopulation assay. We found that Foxm1 haploinsufficiency HSCs exhibited competitive repopulation advantage in the first and secondary recipient mice, but displayed significantly decreased capacity of repopulation in tertiary recipient mice as compared to control HSCs, suggesting that Foxm1 haploinsufficiency promoted clonal expansion of HSCs, which leads to an exhaustion of HSCs eventually. HSC proliferation can be induced by some specific immune effectors such as Toll-like receptor 4 (TLR4). Lipopolysaccharide (LPS) stimulates HSC proliferation by activating TLR4 signaling pathway. Low dose of LPS treatment over time accelerated the development of MDS in mice. Interestingly, low dose of LPS injection chronically induced defects in hematopoiesis in Foxm1 haploinsufficient mice but not the control wildtype mice. Recipient mice transplanted with Foxm1 heterozygous BM cells but not the control BM cells developed MDS-like disease with cytopenia and a decreased number of hematopoietic stem/progenitor cells after LPS stimulation. Moreover, we found that nearly half of aged Foxm1 haploinsufficient mice (20 months) developed splenomegaly. Analysis of histologic sections in Foxm1 haploinsufficient mice showed that the mice developed hematopoietic dysplasia including dysplastic megakaryocytes with bizarre-shaped nuclei in bone marrow and extramedullary hematopoiesis with accumulation of myeloid cells in spleen. RNA-seq analysis indicated that haploinsufficiency of Foxm1 perturbed multiple stem cell-maintenance mechanisms especially in metabolic processes. Taken together, our studies suggest that Foxm1 haploinsufficiency in mice causes clonal expansion of HSCs and promotes MDS-like disease, which underscores the significant role of FOXM1 downregulation in the initiation and development of human MDS. Disclosures No relevant conflicts of interest to declare.

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Dominant T-Cell Clonal Expansion Associated with Acute Graft-Versus-Host Disease after Allogeneic Hematopoietic Stem Cell Transplantation in Pediatric Patients.

Blood ◽

10.1182/blood.v104.11.1243.1243 ◽

2004 ◽

Vol 104 (11) ◽

pp. 1243-1243

Author(s):

Xiaohua Chen ◽

James Knowles ◽

Raymond Barfield ◽

Dawn Lawrence ◽

Rupert Handgretinger

Keyword(s):

T Cell ◽

Pediatric Patients ◽

Acute Gvhd ◽

Cell Clone ◽

Early Stage ◽

Clonal Expansion ◽

Hematopoietic Stem ◽

Cell Clones ◽

T Cell Clone ◽

Cell Clonal Expansion

Abstract Allogeneic hematopoietic stem cell transplantation (AHSCT) provides a valuable therapy for a variety of malignancies, solid tumors, hematological disorders and autoimmune diseases. One of the major complications in recipients after transplantation is graft-versus-host disease (GvHD), which can cause delayed T-cell reconstitution and therefore increase the risk of infection and mortality. It has been suggested that T cells play an important role in the development of GvHD. Several previous studies have identified T-cell clones associated with acute GvHD in adult patients. However, T-cell clonal expansion in the early stage of post-transplantation might also be caused by T-cells which are unrelated to acute GvHD. In this study, we used TCRβ CDR3 size spectratyping to detect T-cell clonal expansion among the polyclonal complexities of peripheral blood. We examined the TCRβ repertoire distribution at the onset of acute GvHD in 10 pediatric patients. We then compared that result with the TCRβ pattern at the same post-AHSCT stage in another 10 pediatric patients without acute GvHD. Method: An RT-PCR assay for Vβ CDR3 size distribution was performed. Each fluorescent PCR product was detected by an automated 310 DNA sequencer. The data were analyzed by GeneScan software. Results and Discussion: As expected, the TCRβ repertoire distribution was much skewed for all patients in both groups, because the immune suppression occurred at early stage of post-AHSCT. There were about 4–8 T-cell clones expanded in each patient for both groups. The TCRβ subfamilies among those clones varied significantly from patient to patient. However, Vβ2, Vβ9, Vβ13 and Vβ20 were commonly found in both groups, indicating in these cases that clonal expansion was not related to the occurrence of acute GvHD. Notably, TCR Vβ16 was found in 9 of 18 samples from 7 of 10 patients with acute GvHD. This TCR subfamily was not dominant in the patients without acute GvHD. This finding strongly suggests that the T-cell clone carrying TCR Vβ16 subfamily might be associated with acute GvHD in pediatric patients. The characterization of this T-cell clone is under study.

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Aging, hematopoiesis, and the myelodysplastic syndromes

Hematology ◽

10.1182/asheducation-2017.1.73 ◽

2017 ◽

Vol 2017 (1) ◽

pp. 73-78 ◽

Cited By ~ 6

Author(s):

Stephen S. Chung ◽

Christopher Y. Park

Keyword(s):

Myelodysplastic Syndromes ◽

Bone Marrow Failure ◽

Hematologic Malignancies ◽

Relative Increase ◽

Extrinsic Factors ◽

Hematopoietic Stem ◽

Clonal Hematopoiesis ◽

Increased Risk ◽

The Many ◽

The Relationship

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Single-cell multi-omics reveals elevated plasticity and stem-cell-like blasts relevant to the poor prognosis of KMT2A-rearranged leukemia

10.1101/2020.12.06.413930 ◽

2020 ◽

Author(s):

Changya Chen ◽

Wenbao Yu ◽

Fatemeh Alikarami ◽

Qi Qiu ◽

Chia-hui Chen ◽

...

Keyword(s):

Single Cell ◽

Single Cell Analysis ◽

Cytotoxic Lymphocytes ◽

Extrinsic Factors ◽

Hematopoietic Stem ◽

Factors Affecting ◽

Young Infants ◽

The Poor ◽

Immune Mediated ◽

Increased Risk

SummaryInfant ALL is a devastating malignancy caused by rearrangements of the KMT2A gene (KMT2A-r) in approximately 70% of patients. The outcome is dismal and younger age at diagnosis is associated with increased risk of relapse. To discover age-specific differences and critical drivers that mediate the poor outcome in KMT2A-r ALL, we subjected KMT2A-r leukemias and normal hematopoietic cells from patients of different ages to multi-omic single cell analysis using scRNA-Seq, scATAC-Seq and snmC-Seq2. We uncovered the following critical new insights: Leukemia cells from infants younger than 6 months have a greatly increased lineage plasticity and contain a hematopoietic stem and progenitor-like (HSPC-like) population compared to older infants. We identified an immunosuppressive signaling circuit between the HSPC-like blasts and cytotoxic lymphocytes in younger patients. Both observations offer a compelling explanation for the ability of leukemias in young infants to evade chemotherapy and immune mediated control. Our analysis also revealed pre-existing lymphomyeloid primed progenitor and myeloid blasts at initial diagnosis of B-ALL. Tracking of leukemic clones in two patients whose leukemia underwent a lineage switch documented the evolution of such clones into frank AML. These findings provide critical insights into KMT2A-r ALL and have potential clinical implications for targeted inhibitors or multi-target immunotherapy approaches. Beyond infant ALL, our study demonstrates the power of single cell multi-omics to detect tumor intrinsic and extrinsic factors affecting rare but critical subpopulations within a malignant population that ultimately determines patient outcome.

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Single-Cell Multiomics Reveals Increased Plasticity, Resistant Populations and Stem-Cell-like Blasts in KMT2A-Rearranged Leukemia

Blood ◽

10.1182/blood-2021-146102 ◽

2021 ◽

Vol 138 (Supplement 1) ◽

pp. 2203-2203

Author(s):

Changya Chen ◽

Wenbao Yu ◽

Fatemeh Alikarami ◽

Qi Qiu ◽

Chia-hui Chen ◽

...

Keyword(s):

Single Cell ◽

Young Patients ◽

Cytotoxic Lymphocytes ◽

Extrinsic Factors ◽

Hematopoietic Stem ◽

Younger Patients ◽

Factors Affecting ◽

Immune Mediated ◽

Younger Age ◽

Increased Risk

Abstract KMT2A-rearranged (KMT2A-r) infant ALL is a devastating malignancy with a dismal outcome, and younger age at diagnosis is associated with increased risk of relapse. To discover age-specific differences and critical drivers that mediate poor outcome in KMT2A-r ALL, we subjected KMT2A-r leukemias and normal hematopoietic cells from patients of different ages to single cell multi-omics analyses. We uncovered the following critical new insights: leukemia cells from patients younger than 6 months have significantly increased lineage plasticity. Steroid response pathways are downregulated in the most immature blasts from younger patients. We identify a hematopoietic stem and progenitor-like (HSPC-like) population in the blood of younger patients that contains leukemic blasts and form an immunosuppressive signaling circuit with cytotoxic lymphocytes. These observations offer a compelling explanation for the ability of leukemias in young patients to evade chemotherapy and immune mediated control. Our analysis also revealed pre-existing lymphomyeloid primed progenitors and myeloid blasts at initial diagnosis of B-ALL. Tracking of leukemic clones in two patients whose leukemia underwent a lineage switch documented the evolution of such clones into frank AML. These findings provide critical insights into KMT2A-r ALL and have clinical implications for molecularly targeted and immunotherapy approaches. Beyond infant ALL, our study demonstrates the power of single cell multi-omics to detect tumor intrinsic and extrinsic factors affecting rare but critical subpopulations within a malignant population that ultimately determines patient outcome. Disclosures Bernt: Merck: Other: Spouse is an employee of Merck.; Syndax: Research Funding.

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T-Cell Depleted (TCD) Hematopoietic Stem Cell Transplantation (HCT) For Adult Patients With Acute Myelogenous Leukemia (AML) In First and Second Remission: Long-Term Disease Free Survival(DFS) With a Significantly Reduced Risk Of Graft-Versus-Host Disease(GvHD)

Blood ◽

10.1182/blood.v122.21.3387.3387 ◽

2013 ◽

Vol 122 (21) ◽

pp. 3387-3387

Author(s):

Izaskun Ceberio ◽

Patrick Hilden ◽

Sean M. Devlin ◽

Molly Maloy ◽

Jenna D Goldberg ◽

...

Keyword(s):

Prognostic Factors ◽

Conflicts Of Interest ◽

Conditioning Regimen ◽

Case Series ◽

Myelogenous Leukemia ◽

Low Grade ◽

Hematopoietic Stem ◽

Risk Of Death ◽

Increased Risk

Abstract Intro Allogeneic-HCT is recommended for AML patients (pts) in CR2, in CR1 with poor-risk cytogenetics, and should be considered for those in CR1 with intermediate-risk. Non-relapse mortality (NRM) and GVHD remain major causes of treatment failure. Ex vivo TCD can prevent GvHD but large case series have not been published. Methods A retrospective chart review was conducted to evaluate 178 pts with AML in CR1 and CR2 undergoing TCD-HCT between 2001 and 2011. All pts received myeloablative-conditioning. The majority received ATG for graft rejection prophylaxis. Acute (A) and chronic(c) GVHD were assessed by standard criteria. No GVHD prophylaxis was administered post-transplant. Soybean agglutination+sheepRBC rosetting (sRBCR) was used for BM TCD. CD34+ selection +/- sRBCR was used for PBSC TCD. Pt characteristics were compared using Pearson's chi-squared and Fisher's exact tests. Prognostic factors relating to overall survival (OS) and DFS, including age, gender, leukemia etiology, cytogenetic-risk group, donor-type, TCD-method, conditioning-regimen, HLA match grade, HCT-specific comorbidity index and immune reconstitution were evaluated using log-rank test statistics. Differences in cumulative incidence (CI) rates were evaluated using Gray's test. Cox proportional-hazards regression was used to further adjust for pt risk factors for OS and DFS. Results Pt characteristics and outcomes are summarized in Tables 1 and 2. Median follow-up of survivors is 52 mo (12-134). 177 pts engrafted. One died pre-engraftment, 7 developed late graft-failure (GF), and 3 are alive after a 2nd HCT. One yr incidence of aGVHD was low (grade 2-4 13%, 3-4 3%). Only 1 pt developed cGVHD by NIH consensus criteria. Univariate association between CR status (1 vs 2), OS and DFS was not statistically significant (p=0.17 and 0.16, respectively). After adjusting for HLA status, age, sex, cytogenetic risk, and regimen, CR2 pts had an increased risk of death (HR: 1.90 (1.14-3.16), p= 0.014). In CR1 pts, cytogenetics was associated with relapse incidence (p=0.003) and was highest in patients with adverse cytogenetics (31%, 95%CI 16-48) and <10% in intermediate I-II risk pts. Overall CI relapse at 1 and 2 yrs was 13% and 16%, respectively. Causes of death were: relapse (n=29), infection (n=25), GVHD (n=7), organ toxicities (n=5), GF (n=2) and other (n=7). Female gender was significantly associated with decreased OS and DFS (p<0.002 and 0.003, respectively). Two yr estimates of OS and DFS in females vs males was 51% vs. 71% and 49% vs. 69%, respectively. These differences were due to a higher NRM in females receiving the chemotherapy based regimen (p<0.001). Gender difference was not observed in pts receiving TBI-based regimens (p=0.599). This difference persisted after adjusting for other common prognostic factors in a multivariate model. For the entire group, 2yr OS and DFS was 67% and 62%, respectively. For CR1 pts, 2 yr OS and DFS was 70% and 64%, respectively (Fig.1). Conclusion These results support the use of TCD HCT in AML pts in CR. Durable DFS and long-term OS can be achieved with low rates of GVHD without compromising the graft-vs-leukemia effect. Disclosures: No relevant conflicts of interest to declare.

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Therapeutic Modulation of Autophagy in Leukaemia and Lymphoma

Cells ◽

10.3390/cells8020103 ◽

2019 ◽

Vol 8 (2) ◽

pp. 103 ◽

Cited By ~ 12

Author(s):

Mojgan Djavaheri-Mergny ◽

Sylvie Giuriato ◽

Mario P. Tschan ◽

Magali Humbert

Keyword(s):

Blood Cell ◽

Clonal Expansion ◽

Genetic Mutations ◽

Haematological Malignancies ◽

Self Renewal ◽

Hematopoietic Stem ◽

Increased Risk ◽

Cell Diversity ◽

Stem And Progenitor Cells

Haematopoiesis is a tightly orchestrated process where a pool of hematopoietic stem and progenitor cells (HSPCs) with high self-renewal potential can give rise to both lymphoid and myeloid lineages. The HSPCs pool is reduced with ageing resulting in few HSPC clones maintaining haematopoiesis thereby reducing blood cell diversity, a phenomenon called clonal haematopoiesis. Clonal expansion of HSPCs carrying specific genetic mutations leads to increased risk for haematological malignancies. Therefore, it comes as no surprise that hematopoietic tumours develop in higher frequency in elderly people. Unfortunately, elderly patients with leukaemia or lymphoma still have an unsatisfactory prognosis compared to younger ones highlighting the need to develop more efficient therapies for this group of patients. Growing evidence indicates that macroautophagy (hereafter referred to as autophagy) is essential for health and longevity. This review is focusing on the role of autophagy in normal haematopoiesis as well as in leukaemia and lymphoma development. Attenuated autophagy may support early hematopoietic neoplasia whereas activation of autophagy in later stages of tumour development and in response to a variety of therapies rather triggers a pro-tumoral response. Novel insights into the role of autophagy in haematopoiesis will be discussed in light of designing new autophagy modulating therapies in hematopoietic cancers.

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Donor Chip Causes Donor-Derived Clonal Hematopoiesis As an Early Complication of Allogeneic Stem Cell Transplantation

Blood ◽

10.1182/blood.v128.22.987.987 ◽

2016 ◽

Vol 128 (22) ◽

pp. 987-987

Author(s):

Christopher J. Gibson ◽

James A. Kennedy ◽

Sarah Nikiforow ◽

John E. Dick ◽

Jerome Ritz ◽

...

Keyword(s):

Stem Cell ◽

Board Of Directors ◽

Hematologic Malignancy ◽

Clonal Evolution ◽

Donor Cell ◽

Advisory Committees ◽

Hematopoietic Stem ◽

Clonal Hematopoiesis ◽

Increased Risk ◽

Rbc Indices

Background Allogeneic stem cell transplantation (HSCT) involves the transfer of healthy donor hematopoietic cells, including hematopoietic stem cells (HSCs) and mature immune effector cells, to recipients with high-risk hematologic malignancies. The success of HSCT is fundamentally dependent on engraftment of normal donor-derived hematopoiesis. Rarely, donor-derived cells undergo transformation to myeloid leukemia as a late complication of HSCT (donor cell leukemia, DCL), a process linked in some cases to pre-existing mutations in donor hematopoietic stem cells (HSCs). In healthy older adults, HSC mutations can cause clonal hematopoiesis of indeterminate potential (CHIP), which is associated with increased risk of hematologic malignancy and an increased risk of all-cause mortality. Among donors over age 40, the frequency of donor CHIP can be estimated to be 5 to 20-fold higher than the frequency of donor cell leukemia, suggesting that non-leukemic endpoints of donor derived clonality may be more prevalent and consequential than previously recognized. Methods We performed targeted next generation sequencing in patients who developed peripheral cytopenias or myeloid neoplasms in the context of 100% donor cell chimerism after allogeneic HSCT. We also prospectively sequenced potential donors who had abnormal CBC parameters, including WBC differential and RBC indices. To determine whether the mutations detected in recipients were of donor derivation, and to assess clonal evolution over time, we performed ultra-deep genotyping at 30,000-100,000X or droplet digital PCR (ddPCR) in the donor stem cell product and serial post-SCT samples. Results We identified four allogeneic SCT recipients with 100% chimerism who met inclusion criteria, two with late-onset hematologic malignancy (latency 23 and 10 years) and two with progressive cytopenias but no evidence of hematologic malignancy on bone marrow examination (latency 18 and 13 months). In each of these patients, sequencing of bone marrow or peripheral blood specimens revealed at least one somatic mutation (SF3B1 K666Q; U2AF1 S34F; DNMT3A T868N; DNMT3A Q356X) that could be used to assess the possibility of donor derivation. In all patients, we identified at least one mutation present in both the diagnostic post-HSCT recipient sample and the pre-HSCT banked donor stem cell product. Figure 1 shows post-HSCT clonal evolution from one case. In another case, we evaluated contemporaneous samples from the donor and recipient obtained at the time of diagnosis of DCL in the recipient (23 years after HSCT). We identified two key findings: 1) a shared SF3B1 K666 mutation in donor and recipient, and 2) divergent clonal evolution, with ASXL1 C789X in the donor, and ASXL1 G642fs, SETBP1 D868N, and SUZ12 D605E in the recipient (Figure 2). In this context, the recipient had a morphologic diagnosis of MDS/MPN overlap, while the donor had abnormal RBC indices but no cytopenias. Since CHIP is associated with subtle abnormalities in hematologic parameters even in the absence of overt cytopenias, we evaluated potential donors with non-exclusionary CBC abnormalities, such as abnormal white blood cell differential or elevated MCV. In two cases, we prospectively identified DNMT3A mutations (R736Cand R882H, variant allele fractions 11.5% and 2%, respectively), consistent with donor CHIP. In one case, the donor with CHIP was used for transplantation. At 90 days after HSCT, donor cell chimerism was 99%; sequencing of recipient peripheral blood revealed the donor-derived DNMT3A mutation and the recipient remained persistently anemic, macrocytic, and thrombocytopenic. Conclusion We identified healthy stem cell donors with clonal hematopoiesis, marked by mutations in canonical genetic drivers of myeloid malignancies. In each instance, the aberrant clone engrafted in the transplant recipient, underwent selective expansion, and caused development of abnormal hematopoietic function. Our findings suggest that donor-derived clonal hematopoiesis may be common, especially in grafts from older donors, and that donor CHIP may confer an unrecognized and increased risk of leukemic and non-leukemic endpoints. We further demonstrate that donor CHIP can be identified prospectively, thus highlighting an emerging challenge in HSCT that may influence donor selection strategies as the clinical significance of donor CHIP is systematically elucidated. Disclosures Ritz: Kiadis: Membership on an entity's Board of Directors or advisory committees. Soiffer:Juno: Membership on an entity's Board of Directors or advisory committees; Kiadis: Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees. Lindsley:MedImmune: Research Funding; Takeda Pharmaceuticals: Consultancy.

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