Chemotherapy Resistance in B-ALL with Cryptic NUP214-ABL1 Is Amenable to Kinase Inhibition and Immunotherapy.

BCR-ABL1 kinase inhibitors have improved the prognosis of Philadelphia-chromosome-positive (Ph+)-acute lymphoblastic leukemia (ALL). Ph-like (or BCR-ABL1-like) ALL does not express BCR-ABL1 but commonly harbors other genomic alterations of signaling molecules that may be amenable to therapy. Here, we report a case with a NUP214-ABL1 fusion detected at relapse by multiplexed, targeted RNA sequencing. It had escaped conventional molecular work-up at diagnosis, including cytogenetic analysis and fluorescence in situ hybridization for ABL1 rearrangements. The patient had responded poorly to initial multi-agent chemotherapy and inotuzumab immunotherapy at relapse before the fusion was revealed. The addition of dasatinib targeting NUP214-ABL1 to inotuzumab resulted in complete molecular remission, but recurrence occurred rapidly with dasatinib alone. However, deep molecular remission was recaptured with a combination of blinatumomab and ponatinib, so he could proceed to allotransplantation. This case illustrates that next-generation sequencing approaches designed to discover cryptic gene fusions can benefit patients with Ph-like ALL.

A phase 1 study of the antibody-drug conjugate brentuximab vedotin with re-induction chemotherapy in patients with CD30-expressing relapsed/refractory acute myeloid leukemia.

BACKGROUND: Outcomes for patients with relapsed/refractory acute myeloid leukemia (R/R AML) remain poor. Novel therapies specifically targeting AML are of high interest. Brentuximab vedotin (BV) is an antibody-drug conjugate that is specific for human CD30. In this phase 1 dose escalation study, the authors evaluated the safety of BV combined with mitoxantrone, etoposide, and cytarabine (MEC) re-induction chemotherapy for patients with CD30-expressing R/R AML. METHODS: Using a standard dose escalation design, the authors evaluated 3 dose levels of BV (0.9 mg/kg, 1.2 mg/kg, and 1.8 mg/kg) administered once on day 1 followed by MEC on days 3 through 7. RESULTS: There were no dose-limiting toxicities noted and the maximum tolerated dose was not reached. The recommended phase 2 dose of BV was determined to be 1.8 mg/kg when combined with MEC. The side effect profile was similar to that expected from MEC chemotherapy alone, with the most common grade ≥3 toxicities being febrile neutropenia, thrombocytopenia, and anemia (toxicities were graded using the National Cancer Institute Common Terminology Criteria for Adverse Events [version 4.0]). Among the 22 patients enrolled on the trial, the composite response rate was 36%, with a composite response rate of 42% noted among those who received the highest dose of BV. The median overall survival was 9.5 months, with a median disease-free survival of 6.8 months observed among responders. Approximately 55% of patients were able to proceed with either allogeneic hematopoietic stem cell transplantation or donor lymphocyte infusion. CONCLUSIONS: The combination of BV with MEC was found to be safe in patients with CD30-expressing R/R AML and warrants further study comparing this combination with the use of MEC alone in this population (ClinicalTrials.gov identifier NCT01830777). LAY SUMMARY: The outcomes for patients with relapsed/refractory acute myeloid leukemia (R/R AML) are exceptionally poor. New and emerging treatment combinations are actively being studied in an effort to improve outcomes. The authors examined the combination of brentuximab vedotin, an antibody product that recognizes a marker called CD30, with mitoxantrone, etoposide, and cytarabine (MEC), a common chemotherapy regimen, in patients with R/R AML that expressed the CD30 marker. The authors found that the combination was safe and well tolerated. Future studies comparing this new combination with the use of MEC alone can help to inform its effectiveness for this patient population.

A complex Polycomb issue: the two faces of EZH2 in cancer.

In the April 1, 2012, issue of Genes & Development, Simon and colleagues (pp. 651-656) demonstrated that the disruption of Ezh2 in mice is sufficient to cause T-acute lymphoblastic leukemia (T-ALL). Moreover, in concert with concurrent studies, the authors revealed that similar mechanisms are involved in human T-ALL. These data contrast with previous findings showing that increased EZH2 activity promotes cancer.

Inhibitory role of the transcription repressor Gfi1 in the generation of thymus-derived regulatory T cells.

Foxp3(+) regulatory T (T(reg)) cells are essential for the maintenance of self-tolerance and immune homeostasis. The majority of T(reg) cells is generated in the thymus as a specific subset of CD4(+) T cells, known as thymus-derived or natural T(reg) (nT(reg)) cells, in response to signals from T-cell receptors, costimulatory molecules, and cytokines. Recent studies have identified intracellular signaling and transcriptional pathways that link these signals to Foxp3 induction, but how the production of these extrinsic factors is controlled remains poorly understood. Here, we report that the transcription repressor growth factor independent 1 (Gfi1) has a key inhibitory role in the generation of nT(reg) cells by a noncell-autonomous mechanism. T cell-specific deletion of Gfi1 results in aberrant expansion of thymic nT(reg) cells and increased production of cytokines. In particular, IL-2 overproduction plays an important role in driving the expansion of nT(reg) cells. In contrast, although Gfi1 deficiency elevated thymocyte apoptosis, Gfi1 repressed nT(reg) generation independently of its prosurvival effect. Consistent with an inhibitory role of Gfi1 in this process, loss of Gfi1 dampens antitumor immunity. These data point to a previously unrecognized extrinsic control mechanism that negatively shapes thymic generation of nT(reg) cells.

ETV6-NTRK3 fusion oncogene initiates breast cancer from committed mammary progenitors via activation of AP1 complex.

To better understand the cellular origin of breast cancer, we developed a mouse model that recapitulates expression of the ETV6-NTRK3 (EN) fusion oncoprotein, the product of the t(12;15)(p13;q25) translocation characteristic of human secretory breast carcinoma. Activation of EN expression in mammary tissues by Wap-Cre leads to fully penetrant, multifocal malignant breast cancer with short latency. We provide genetic evidence that, in nulliparous Wap-Cre;EN females, committed alveolar bipotent or CD61(+) luminal progenitors are targets of tumorigenesis. Furthermore, EN transforms these otherwise transient progenitors through activation of the AP1 complex. Given the increasing relevance of chromosomal translocations in epithelial cancers, such mice serve as a paradigm for the study of their genetic pathogenesis and cellular origins, and generation of preclinical models.

Heterogeneity in histone 2B-green fluorescent protein-retaining putative small intestinal stem cells at cell position 4 and their absence in the colon.

Stem cells have been identified in two locations in small intestinal crypts; those intercalated between Paneth cells and another population (which retains DNA label) are located above the Paneth cell zone, at cell position 4. Because of disadvantages associated with the use of DNA label, doxycycline-induced transient transgenic expression of histone 2B (H2B)-green fluorescent protein (GFP) was investigated. H2B-GFP-retaining putative stem cells were consistently seen, with a peak at cell position 4, over chase periods of up to 112 days. After a 28-day chase, a subpopulation of the H2B-GFP-retaining cells was cycling, but the slow cycling status of the majority was illustrated by lack of expression of pHistone H3 and Ki67. Although some H2B-GFP-retaining cells were sensitive to low-dose radiation, the majority was resistant to low- and high-dose radiation-induced cell death, and a proportion of the surviving cells proliferated during subsequent epithelial regeneration. Long-term retention of H2B-GFP in a subpopulation of small intestinal Paneth cells was also seen, implying that they are long lived. In contrast to the small intestine, H2B-GFP-retaining epithelial cells were not seen in the colon from 28-day chase onward. This implies important differences in stem cell function between these two regions of the gastrointestinal tract, which may have implications for region-specific susceptibility to diseases (such as cancer and ulcerative colitis), in which epithelial stem cells and their progeny are involved.

Endogenous oncogenic K-ras(G12D) stimulates proliferation and widespread neoplastic and developmental defects.

Activating mutations in the ras oncogene are not considered sufficient to induce abnormal cellular proliferation in the absence of cooperating oncogenes. We demonstrate that the conditional expression of an endogenous K-ras(G12D) allele in murine embryonic fibroblasts causes enhanced proliferation and partial transformation in the absence of further genetic abnormalities. Interestingly, K-ras(G12D)-expressing fibroblasts demonstrate attenuation and altered regulation of canonical Ras effector signaling pathways. Widespread expression of endogenous K-ras(G12D) is not tolerated during embryonic development, and directed expression in the lung and GI tract induces preneoplastic epithelial hyperplasias. Our results suggest that endogenous oncogenic ras is sufficient to initiate transformation by stimulating proliferation, while further genetic lesions may be necessary for progression to frank malignancy.

Ixazomib in addition to chemotherapy for the treatment of acute lymphoblastic leukemia in older adults.

We sought to assess the safety of adding ixazomib, an oral proteasome inhibitor, to a multi-agent treatment regimen for older adults with acute lymphoblastic leukemia (ALL). Patients 51 to 75 years of age with newly diagnosed ALL were screened. Induction consisted of prednisone (P), vincristine (V), and doxorubicin (D). For BCR-ABL1+ patients, dasatinib was added. On Days 1, 8, 15 of induction, ixazomib was given orally. After induction patients received 1 cycle of consolidation in which ixazomib was given on Days 1, 8, 15. After consolidation, patients in remission (CR) were offered stem cell transplantation. Among the 19 patients treated, 15 (79%) [90% CI, 58-92%] achieved CR or CRi. At 2 years, the overall survival was 47% [95%CI, 29-72%]. In this study the dose of 2.3 mg of ixazomib in combination was the MTD for older patients with ALL and is the recommended dose for future phase 2 studies.

A nonrandomized phase I and biomarker trial of regorafenib in advanced myeloid malignancies.

We conducted a single-center, open-label, dose escalation, and expansion phase I trial of the antiangiogenic multikinase inhibitor regorafenib in patients with advanced myeloid neoplasms. We enrolled 16 patients with relapsed/refractory acute myeloid leukemia (AML), myeloproliferative neoplasms (MPN), chronic myelomonocytic leukemia (CMML), or myelodysplastic syndrome (MDS). A 3 + 3 dose escalation design was used with two planned dose levels (120 or 160 mg daily) and one de-escalation level (80 mg daily). An additional 10 patients were treated on an expansion cohort. The recommended phase two dose of regorafenib was 160 mg daily, with no dose-limiting toxicities. The best overall disease response by International Working Group criteria included one partial and stable disease in 11 patients. Tissue studies indicated no change in Ras/mitogen-activated protein kinase (MAPK) pathway activation in responders. Pharmacodynamic changes in plasma VEGF, PlGF, and sVEGFR2 were detected during treatment. Baseline proinflammatory and angiogenic cytokine levels were not associated with clinical response. Single-agent regorafenib demonstrated an acceptable safety profile in relapsed/refractory myeloid malignancy patients. Most patients achieved stable disease, with modest improvements in cell counts in some MDS patients. Biomarker studies were consistent with on-target effects of regorafenib on angiogenesis. Future studies should investigate the role of regorafenib in combination therapy approaches.

Chromatin protein L3MBTL1 is dispensable for development and tumor suppression in mice.

L3MBTL1, a paralogue of Drosophila tumor suppressor lethal(3)malignant brain tumor (l(3)mbt), binds histones in a methylation state-dependent manner and contributes to higher order chromatin structure and transcriptional repression. It is the founding member of a family of MBT domain-containing proteins that has three members in Drosophila and nine in mice and humans. Knockdown experiments in cell lines suggested that L3MBTL1 has non-redundant roles in the suppression of oncogene expression. We generated a mutant mouse strain that lacks exons 13-20 of L3mbtl1. Markedly reduced levels of a mutant mRNA with an out-of-frame fusion of exons 12 and 21 were expressed, but a mutant protein was undetectable by Western blot analysis. L3MBTL1(-/-) mice developed and reproduced normally. The highest expression of L3MBTL1 was detected in the brain, but its disruption did not affect brain development, spontaneous movement, and motor coordination. Despite previous implications of L3mbtl1 in the biology of hematopoietic transcriptional regulators, lack of L3MBTL1 did not result in deficiencies in lymphopoiesis or hematopoiesis. In contrast with its demonstrated biochemical activities, embryonic stem (ES) cells lacking L3MBTL1 displayed no abnormalities in H4 lysine 20 (H4K20) mono-, di-, or trimethylation; had normal global chromatin density as assessed by micrococcal nuclease digests; and expressed normal levels of c-myc. Embryonic fibroblasts lacking L3MBTL1 displayed unaltered cell cycle arrest and down-regulation of cyclin E expression after irradiation. In cohorts of mice followed for more than 2 years, lack of L3MBTL1 did not alter normal lifespan or survival with or without sublethal irradiation.

Neutrophils in a mouse model of autoantibody-mediated arthritis: critical producers of Fc receptor gamma, the receptor for C5a, and lymphocyte function-associated antigen 1.

OBJECTIVE: Neutrophils represent a prominent component of inflammatory joint effusions and are required for synovial inflammation in mouse models, but the mechanisms are poorly understood. In this study, we developed a system with which to test the importance of the production of specific factors by neutrophils in a mouse model of arthritis. METHODS: Neutrophil-deficient Gfi-1(-/-) mice were administered sublethal doses of radiation and were then engrafted with donor bone marrow cells (BMCs), which resulted in the production of mature neutrophils within 2 weeks. By reconstituting with BMCs from mice lacking selected proinflammatory factors, we generated mice that specifically lacked these factors on their neutrophils. Arthritis was initiated by transfer of K/BxN serum to identify the role of defined neutrophil factors on the incidence and severity of arthritis. RESULTS: Neutrophils lacking the signaling chain of stimulatory Fc receptors (FcRgamma(-/-)) were unable to elicit arthritis, but neutrophils lacking FcgammaRIII still did so. Neutrophils lacking the chemotactic or adhesion receptor C5a receptor (C5aR) or CD11a/lymphocyte function-associated antigen 1 (LFA-1) also failed to initiate arthritis but could enter joints in which inflammation had been initiated by wild-type neutrophils. Neutrophils unable to produce interleukin-1alpha (IL-1alpha) and IL-1beta (IL-1alpha/beta(-/-)) or leukotrienes (5-lipoxygenase [5-LOX(-/-)]) produced arthritis of intermediate severity. The inability of neutrophils to make tumor necrosis factor or to express receptors for tumor necrosis factor or IL-1 had no effect on arthritis. CONCLUSION: A novel transfer system was developed to identify neutrophil production of FcRgamma, C5aR, and CD11a/LFA-1 as critical components of autoantibody-mediated arthritis. Neutrophil production of IL-1 and leukotriene B(4) likely contributes to inflammation but is not essential. Molecular requirements for neutrophil influx into joints become more permissive after inflammation is initiated.

The Second Oncogenic Hit Determines the Cell Fate of ETV6-RUNX1 Positive Leukemia.

ETV6-RUNX1 is almost exclusively associated with childhood B-cell acute lymphoblastic leukemia (B-ALL), but the consequences of ETV6-RUNX1 expression on cell lineage decisions during B-cell leukemogenesis are completely unknown. Clinically silent ETV6-RUNX1 preleukemic clones are frequently found in neonatal cord blood, but few carriers develop B-ALL as a result of secondary genetic alterations. The understanding of the mechanisms underlying the first transforming steps could greatly advance the development of non-toxic prophylactic interventions. Using genetic lineage tracing, we examined the capacity of ETV6-RUNX1 to instruct a malignant phenotype in the hematopoietic lineage by cell-specific Cre-mediated activation of ETV6-RUNX1 from the endogenous Etv6 gene locus. Here we show that, while ETV6-RUNX1 has the propensity to trigger both T- and B-lymphoid malignancies, it is the second hit that determines tumor cell identity. To instigate leukemia, both oncogenic hits must place early in the development of hematopoietic/precursor cells, not in already committed B-cells. Depending on the nature of the second hit, the resulting B-ALLs presented distinct entities that were clearly separable based on their gene expression profiles. Our findings give a novel mechanistic insight into the early steps of ETV6-RUNX1+ B-ALL development and might have major implications for the potential development of ETV6-RUNX1+ B-ALL prevention strategies.

Gfi-1 restricts proliferation and preserves functional integrity of haematopoietic stem cells.

Haematopoietic stem cells (HSCs) sustain blood production throughout life. HSCs are capable of extensive proliferative expansion, as a single HSC may reconstitute lethally irradiated hosts. In steady-state, HSCs remain largely quiescent and self-renew at a constant low rate, forestalling their exhaustion during adult life. Whereas nuclear regulatory factors promoting proliferative programmes of HSCs in vivo and ex vivo have been identified, transcription factors restricting their cycling have remained elusive. Here we report that the zinc-finger repressor Gfi-1 (growth factor independent 1), a cooperating oncogene in lymphoid cells, unexpectedly restricts proliferation of HSCs. After loss of Gfi-1, HSCs display elevated proliferation rates as assessed by 5-bromodeoxyuridine incorporation and cell-cycle analysis. Gfi-1-/- HSCs are functionally compromised in competitive repopulation and serial transplantation assays, and are rapidly out-competed in the bone marrow of mouse chimaeras generated with Gfi-1-/- embryonic stem cells. Thus, Gfi-1 is essential to restrict HSC proliferation and to preserve HSC functional integrity.

Incidence of Invasive Fungal Infections in Acute Myeloid Leukemia Without Antifungal Prophylaxis.

BACKGROUND: Antifungal prophylaxis during induction for acute myeloid leukemia (AML) varies according to local rates of invasive fungal infections (IFIs). We evaluated fluconazole prophylaxis and no antifungal prophylaxis, as a natural interrupted time-series study to assess survival and infection complications. PATIENTS AND METHODS: We identified patients with AML ≥ 18 years old undergoing induction chemotherapy during 2 time periods: period 1, fluconazole prophylaxis from August 1, 2013 to September 30, 2015, and period 2, no prophylaxis from October 1, 2015 to December 31, 2017. The primary outcome was incidence of proven or probable IFI. Secondary outcomes included types of IFIs and 60-day overall survival (OS). IFI was defined by the 2002 European Organization for Research and Treatment of Cancer/Mycoses Study Group Consensus criteria. RESULTS: One hundred forty-four patients received induction chemotherapy over the 2 time periods. In the prophylaxis versus no-prophylaxis groups, the rate of proven or probable IFIs was 4 (5%) of 87 versus 12 (21%) of 57 (P = .01). The total number of proven IFIs was 3 (3%) of 87 versus 4 (7%) of 57 (P = .44), whereas probable IFIs were 1 (1%) of 87 versus 8 (14%) of 57 (P < .01). No difference was observed in fungemia. Incidence of IFIs was too low to detect resistance patterns. OS at 60 days was improved in with fluconazole prophylaxis compared with no prophylaxis (hazard ratio, 0.329; 95% confidence interval, 0.12-0.89; P = .028). CONCLUSION: Observed rates of proven or probable IFI were lower in the fluconazole prophylaxis group versus the no-prophylaxis group. Sixty-day OS was higher with fluconazole prophylaxis. Further study is required to evaluate how fluconazole may impart the differences in survival seen in this analysis.

Clinical response to larotrectinib in adult Philadelphia chromosome-like ALL with cryptic ETV6-NTRK3 rearrangement.

Philadelphia chromosome (Ph)-like acute lymphoblastic leukemia (ALL) is a subtype of Ph-negative ALL that molecularly resembles Ph-positive ALL. It shares the adverse prognosis of Ph-positive ALL, but lacks the BCR-ABL1 fusion oncogene. Instead, Ph-like ALL is associated with alternative mutations in signaling pathways. We describe a case of Ph-like ALL that harbored 2 genomic alterations, which activated signaling, an NRASGly12Asp mutation, and an ETV6-NTRK3 rearrangement. Initially, the NRAS mutation was detected at high frequency, whereas the gene fusion was only detectable with a targeted next-generation sequencing-based fusion assay, but not by fluorescence in situ hybridization analysis. The disease failed to respond to multiagent chemotherapy but investigational CD19-directed chimeric antigen receptor T-cell therapy resulted in a complete remission. However, the leukemia relapsed after 6 weeks. Intriguingly, the NRAS mutation was extinguished during the chimeric antigen receptor T-cell therapy and did not contribute to the relapse, which was instead associated with a rise in ETV6-NTRK3. The relapsed leukemia progressed with further chemo- and immunotherapy but was controlled for 6 weeks with substantial leukemic cytoreduction using the TRK inhibitor larotrectinib. Unfortunately, recovery of normal hematopoiesis was only marginal and the patient eventually succumbed to infections. These results demonstrate that larotrectinib has clinical activity in ETV6-NTRK3-associated Ph-like ALL.

Alisertib plus induction chemotherapy in previously untreated patients with high-risk, acute myeloid leukaemia: a single-arm, phase 2 trial.

BACKGROUND: Increased aurora A kinase (AAK) expression occurs in acute myeloid leukaemia; AAK inhibition is a promising therapeutic target in this disease. We therefore aimed to assess the activity of alisertib combined with 7 + 3 induction chemotherapy in previously untreated patients with high-risk acute myeloid leukaemia. METHODS: We did a single-arm, phase 2 trial of patients recruited from the Dana-Farber/Harvard Cancer Center in the USA. Eligible patients had previously untreated acute myeloid leukaemia, an Eastern Cooperative Oncology Group performance status of 0-2, and were at high risk of disease as defined by the presence of an adverse-risk karyotype, the presence of secondary acute myeloid leukaemia arising from previous myelodysplastic syndrome or myeloproliferative neoplasm, the presence of therapy-related acute myeloid leukaemia, or being 65 years or older. Enrolled patients received 7 + 3 induction chemotherapy of continuous infusion of cytarabine (100 mg/m2 per day on days 1-7) and intravenous bolus of idarubicin (12 mg/m2 per day on days 1-3). Oral alisertib (30 mg) was given twice per day on days 8-15. Patients could receive up to four consolidation cycles with cytarabine and alisertib, and alisertib maintenance for 12 months. The primary endpoint was a composite including the proportion of patients achieving complete remission and those with a complete remission with incomplete neutrophil or platelet count recovery. Analyses were per-protocol. This study is registered with Clinicaltrials.gov, number NCT02560025, and has completed enrolment. FINDINGS: Between Dec 31, 2015, and Aug 1, 2017, we enrolled a total of 39 eligible patients. 19 (49%) of 39 patients had secondary acute myeloid leukaemia and three (8%) had therapy-related acute myeloid leukaemia. At mid-induction, 33 (85%) of 39 patients showed marrow aplasia, six (15%) received re-induction. The median follow-up was 13·7 months (IQR 12·7-14·4). Composite remission was 64% (two-stage 95% CI 48-79), with 20 (51%) of 39 patients achieving complete remission and five (13%) achieving complete remission with incomplete neutrophil or platelet count recovery. The most common grade 3 or 4 adverse events included febrile neutropenia (16 [41%] of 39), neutropenia (12 [31%]), thrombocytopenia (13 [33%]), anaemia (11 [28%]), anorexia (nine [23%]), and oral mucositis (four [10%]). No treatment-related deaths were observed. INTERPRETATION: These results suggest that alisertib combined with induction chemotherapy is active and safe in previously untreated patients with high-risk acute myeloid leukaemia. This study met criteria to move forward to a future randomised trial. FUNDING: Millennium Pharmaceuticals.

Inducible histone K-to-M mutations are dynamic tools to probe the physiological role of site-specific histone methylation in vitro and in vivo.

Development and differentiation are associated with profound changes to histone modifications, yet their in vivo function remains incompletely understood. Here, we generated mouse models expressing inducible histone H3 lysine-to-methionine (K-to-M) mutants, which globally inhibit methylation at specific sites. Mice expressing H3K36M developed severe anaemia with arrested erythropoiesis, a marked haematopoietic stem cell defect, and rapid lethality. By contrast, mice expressing H3K9M survived up to a year and showed expansion of multipotent progenitors, aberrant lymphopoiesis and thrombocytosis. Additionally, some H3K9M mice succumbed to aggressive T cell leukaemia/lymphoma, while H3K36M mice exhibited differentiation defects in testis and intestine. Mechanistically, induction of either mutant reduced corresponding histone trimethylation patterns genome-wide and altered chromatin accessibility as well as gene expression landscapes. Strikingly, discontinuation of transgene expression largely restored differentiation programmes. Our work shows that individual chromatin modifications are required at several specific stages of differentiation and introduces powerful tools to interrogate their roles in vivo.

The Mst1 Kinase Is Required for Follicular B Cell Homing and B-1 B Cell Development.

The Mst1 and 2 cytosolic serine/threonine protein kinases are the mammalian orthologs of the Drosophila Hippo protein. Mst1 has been shown previously to participate in T-cell and B-cell trafficking and the migration of lymphocytes into secondary lymphoid organs in a cell intrinsic manner. We show here that the absence of Mst1 alone only modestly impacts B cell homing to lymph nodes. The absence of both Mst1 and 2 in hematopoietic cells results in relatively normal B cell development in the bone marrow and does not impact migration of immature B cells to the spleen. However, follicular B cells lacking both Mst1 and Mst2 mature in the splenic white pulp but are unable to recirculate to lymph nodes or to the bone marrow. These cells also cannot traffic efficiently to the splenic red pulp. The inability of late transitional and follicular B cells lacking Mst 1 and 2 to migrate to the red pulp explains their failure to differentiate into marginal zone B cell precursors and marginal zone B cells. Mst1 and Mst2 are therefore required for follicular B cells to acquire the ability to recirculate and also to migrate to the splenic red pulp in order to generate marginal zone B cells. In addition B-1 a B cell development is defective in the absence of Mst1.

Transcription factor Etv6 regulates functional differentiation of cross-presenting classical dendritic cells.

An IRF8-dependent subset of conventional dendritic cells (cDCs), termed cDC1, effectively cross-primes CD8+ T cells and facilitates tumor-specific T cell responses. Etv6 is an ETS family transcription factor that controls hematopoietic stem and progenitor cell (HSPC) function and thrombopoiesis. We report that like HSPCs, cDCs express Etv6, but not its antagonist, ETS1, whereas interferon-producing plasmacytoid dendritic cells (pDCs) express both factors. Deletion of Etv6 in the bone marrow impaired the generation of cDC1-like cells in vitro and abolished the expression of signature marker CD8α on cDC1 in vivo. Moreover, Etv6-deficient primary cDC1 showed a partial reduction of cDC-specific and cDC1-specific gene expression and chromatin signatures and an aberrant up-regulation of pDC-specific signatures. Accordingly, DC-specific Etv6 deletion impaired CD8+ T cell cross-priming and the generation of tumor antigen-specific CD8+ T cells. Thus, Etv6 optimizes the resolution of cDC1 and pDC expression programs and the functional fitness of cDC1, thereby facilitating T cell cross-priming and tumor-specific responses.