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Supplementary Fig. 5. Kaplan-Meier analysis of the survival of patients with different mutations.

Legends for Supplementary Figures

Current standard of care for patients with pediatric acute lymphoblastic leukemia (ALL) is mainly effective, with high remission rates after treatment. However, the genetic perturbations that give rise to this disease remain largely undefined, limiting the ability to address resistant tumors or develop less toxic targeted therapies. Here, we report the use of next-generation sequencing to interrogate the genetic and pathogenic mechanisms of 240 pediatric ALL cases with their matched remission samples. Commonly mutated genes fell into several categories, including RAS/receptor tyrosine kinases, epigenetic regulators, transcription factors involved in lineage commitment, and the p53/cell-cycle pathway. Unique recurrent mutational hotspots were observed in epigenetic regulators CREBBP (R1446C/H), WHSC1 (E1099K), and the tyrosine kinase FLT3 (K663R, N676K). The mutant WHSC1 was established as a gain-of-function oncogene, while the epigenetic regulator ARID1A and transcription factor CTCF were functionally identified as potential tumor suppressors. Analysis of 28 diagnosis/relapse trio patients plus 10 relapse cases revealed four evolutionary paths and uncovered the ordering of acquisition of mutations in these patients. This study provides a detailed mutational portrait of pediatric ALL and gives insights into the molecular pathogenesis of this disease. Cancer Res; 77(2); 390–400. ©2016 AACR.

Supplementary Fig. 4. p53 pathway is dysregulated in ALL.

Supplementary Fig. 2. Mutations found in the ASXL family of genes.

Supplementary Fig. 1. Mutational spectrum of pediatric ALL.

Supplementary Table 1. Samples used for whole exome and targeted sequencing. Supplementary Table 2. Clinical information of patients. Supplementary Table 3. 560 Genes screened in the targeted sequencing. Supplementary Table 4. Sequences of shRNA or CRISPR-Cas9 sgRNA used in this study. Supplementary Table 5. Sequences of real-time PCR primers used in this study. Supplementary Table 6. Sequences of primers used for Sanger validation of CRISPR-Cas9 indel. Supplementary Table 7. Mutations of KRAS identified in this study have been found in a variety of cancers. Supplementary Table 8. Mutations of PTPN11 in other cancers occurring in the same location as in our ALL cohort. Supplementary Table 9. Recurrent mutation sites of FLT3 in different cancers. Supplementary Table 10. Cell lines or cancer samples harboring either E1099K or T1150A hotspot mutation of WHSC1. Supplementary Table 11. R1446 mutations of CREBBP recurrently occurred in a variety of cancers. Supplementary Table 12. Mutational hotspot (D1399) of EP300 occurring in other cancers occurring in the same location as in our ALL cohort.Supplementary Table 13. Rare mutations occurring in our ALL cohort (mutated in one individual each), but these mutations have been recurrently documented in the COSMIC cancer mutations database. Supplementary Table 14. Mutations of genes involved in DNA repair pathway in relapse samples.

Due to the curative potential and improvement in progression-free survival (PFS), high-dose chemotherapy followed by autologous stem cell transplantation (ASCT) is considered the standard of care for several hematologic malignancies, such as multiple myeloma, and lymphomas. ASCT typically involves support with blood product transfusion. Thus, difficulties arise when Jehovah’s Witness patients refuse blood transfusions. In order to demonstrate the safety of performing “bloodless” ASCT (BL-ASCT), we performed a retrospective analysis of 66 Jehovah's Witnesses patients who underwent BL-ASCT and 1114 non-Jehovah’s Witness patients who underwent transfusion-supported ASCT (TF-ASCT) at Cedars-Sinai Medical Center between January 2000 and September 2018. Survival was compared between the two groups. Transplant-related complications, mortality, engraftment time, length of hospital stay, and number of ICU transfers were characterized for the BL-ASCT group. One year survival was found to be 87.9% for both groups (P = 0.92). In the BL-ASCT group, there was one death prior to the 30 days post transplant due to CNS hemorrhage, and one death prior to 100 days due to sepsis. Based on our data, BL-ASCT can be safely performed with appropriate supportive measures, and we encourage community oncologists to promptly refer JW patients for transplant evaluation when ASCT is indicated.

The rapid evolution of blood and marrow transplantation (BMT), coupled with diverse outcomes associated with heterogeneous groups of patients, led to the formation of 2 important organizations early in the development of the field: the Center for International Blood and Marrow Transplant Research (CIBMTR) and the Foundation for the Accreditation of Cellular Therapy (FACT). These organizations have addressed 2 of the 9 elements identified by the National Quality Strategy (NQS) for achieving better health care, more affordable care, and healthy people and communities: a registry that promotes improvement of care and accreditation based on quality standards. More recently, a federally mandated database in the United States addresses the third element of the NQS: public reporting of treatment results. Here we describe the current process by which FACT incorporates patient outcomes reported by the CIBMTR into standards for accreditation, the requirements for accredited programs with performance below expected outcomes to maintain accreditation, and preliminary findings of an assessment of corrective action plans intended to improve outcomes.

Patient-centered medical home models are fundamental to the advanced alternative payment models defined in the Medicare Access and Children's Health Insurance Plan Reauthorization Act (MACRA). The patient-centered medical home is a model of healthcare delivery supported by alternative payment mechanisms and designed to promote coordinated medical care that is simultaneously patient-centric and population-oriented. This transformative care model requires shifting reimbursement to include a per-patient payment intended to cover services not previously reimbursed such as disease management over time. Payment is linked to quality measures, including proportion of care delivered according to predefined pathways and demonstrated impact on outcomes. Some medical homes also include opportunities for shared savings by reducing overall costs of care. Recent proposals have suggested expanding the medical home model to specialized populations with complex needs because primary care teams may not have the facilities or the requisite expertise for their unique needs. An example of a successful care model that may provide valuable lessons for those creating specialty medical home models already exists in many hematopoietic cell transplantation (HCT) centers that deliver multidisciplinary, coordinated, and highly specialized care. The integration of care delivery in HCT centers has been driven by the specialty care their patients require and by the payment methodology preferred by the commercial payers, which has included bundling of both inpatient and outpatient care in the peritransplant interval. Commercial payers identify qualified HCT centers based on accreditation status and comparative performance, enabled in part by center-level comparative performance data available within a national outcomes database mandated by the Stem Cell Therapeutic and Research Act of 2005. Standardization across centers has been facilitated via voluntary accreditation implemented by Foundation for the Accreditation of Cell Therapy. Payers have built on these community-established programs and use public outcomes and program accreditation as standards necessary for inclusion in specialty care networks and contracts. Although HCT centers have not been described as medical homes, most HCT providers have already developed the structures that address critical requirements of MACRA for medical homes.

Although substantial progress has been made in the treatment of B-cell acute lymphoblastic leukemia (B-ALL), the prognosis of patients with either refractory or relapsed B-ALL remains dismal. Novel therapeutic strategies are needed to improve the outcome of these patients. KPT-9274 is a novel dual inhibitor of p21-activated kinase 4 (PAK4) and nicotinamide phosphoribosyltransferase (NAMPT). PAK4 is a serine/threonine kinase that regulates a variety of fundamental cellular processes. NAMPT is a rate-limiting enzyme in the salvage biosynthesis pathway of nicotinamide adenine dinucleotide (NAD) that plays a vital role in energy metabolism. Here, we show that KPT-9274 strongly inhibits B-ALL cell growth regardless of cytogenetic abnormalities. We also demonstrate the potent in vivo efficacy and tolerability of KPT-9274 in a patient-derived xenograft murine model of B-ALL. Interestingly, although KPT-9274 is a dual PAK4/NAMPT inhibitor, B-ALL cell growth inhibition by KPT-9274 was largely abolished with nicotinic acid supplementation, indicating that the inhibitory effects on B-ALL cells are mainly exerted by NAD+ depletion through NAMPT inhibition. Moreover, we have found that the extreme susceptibility of B-ALL cells to NAMPT inhibition is related to the reduced cellular NAD+ reserve. NAD+ depletion may be a promising alternative approach to treating patients with B-ALL.

Introduction: Inflammatory breast cancer (IBC) is a rare aggressive form of breast cancer. It is well known that the long-term survival and progression-free survival of IBC are worse than that of non-IBC. We report the long term outcomes of patients with IBC and non-IBC who had undergone high-dose chemotherapy (HDC) with autologous hematopoietic cell transplantation (AHCT). Methods: All 3387 patients with IBC or non-IBC who underwent HDC with AHCT between1990-2002 and registered with CIBMTR were included in this analysis. Transplant-related mortality (TRM), disease relapse/progression, progression-free survival (PFS) and overall survival (OS) were compared between the two cohorts. Multivariate Cox regression model was used to determine the independent impact of stage on outcomes. Results: 527 patients with IBC and 2,860 patients with non-IBC were included; the median age at transplantation (47 vs 46 years old) and median follow-up period in the 2 groups (167 vs 168 months) were similar. The most common conditioning regimen was cyclophosphamide and carboplatin based in both groups (54% in IBC and 50% in non-IBC). AHCT was well tolerated in both groups. TRM was similar in both groups (one year TRM was 2% for IBC and 3% for non-IBC, p=0.16). The most common cause of death was disease progression or relapse (81% in IBC and 75% in non-IBC). The median survival for both IBC and non-IBC was the same at 40 months. The PFS at 10 years was 27% (95% CI: 23-31%) for IBC and 24% (95% CI: 22-26%) for non-IBC (p=0.21), and the OS at 10 years was 31% (95% CI: 27-35%) for IBC and 28% (95% CI: 26-30%) for non-IBC (p=0.16). In univariate analysis, patients with stage III IBC and no active diseases at transplantation had lower PFS and OS than that in non-IBC. In multivariate analysis, controlling for age, disease status at AHCT, hormonal receptor status, time from diagnosis to AHCT, and performance status at AHCT, patients with stage III IBC had higher mortality (HR 1.16, 95% CI: 1-1.34, p= 0.0459), worse PFS (HR: 1.17, 95% CI: 1.01-1.36, p= 0.0339) and higher risk of disease relapse/progression (HR: 1.24, 95% CI: 1.06-1.45, p= 0.0082) as compared to stage III non-IBC. Amongst all patients a higher stage disease was associated with worse PFS, OS and disease relapse/progression. Conclusions: Long-term outcomes of stage III IBC patients who underwent AHCT were poorer than that in non-IBC patients confirming that the poor prognosis of IBC even in the setting of HDC with AHCT.

Diagnosis and relapse: cytogenetically normal acute myelogenous leukemia without FLT3 -ITD or MLL -PTD

Background Clonal VDJ rearrangement of B/T cell receptors (B/TCRs) occurring during B/T lymphocyte development has been used as a marker to track the clonality of B/T cell populations. Methods We systematically profiled the B/T cell receptor repertoire of 936 cancer cell lines across a variety of cancer types as well as 462 Epstein-Barr Virus (EBV) transformed normal B lymphocyte lines using RNA sequencing data. Results Rearranged B/TCRs were readily detected in cell lines derived from lymphocytes, and subclonality or potential biclonality were found in a number of blood cancer cell lines. Clonal BCR/TCR rearrangements were detected in several blast phase CML lines and unexpectedly, one gastric cancer cell line (KE-97), reflecting a lymphoid origin of these cells. Notably, clonality was highly prevalent in EBV transformed B lymphocytes, suggesting either transformation only occurred in a few B cells or those with a growth advantage dominated the transformed population through clonal evolution. Conclusions Our analysis reveals the complexity and heterogeneity of the BCR/TCR rearrangement repertoire and provides a unique insight into the clonality of lymphocyte derived cell lines. Electronic supplementary material The online version of this article (10.1186/s12885-018-4840-5) contains supplementary material, which is available to authorized users.

Aim: Patients treated with allogeneic hematopoietic stem cell transplantation (HSCT) may experience oral complications associated with chronic graft-versus-host disease (cGVHD). These complications may significantly affect quality of life, even many years post-HSCT. Current treatment options for oral cGVHD are limited and often include steroid or other immunomodulatory medications, which may not adequately control the oral condition. A non-immunosuppressive intervention for symptomatic relief in oral cGVHD would thus be a welcome addition to the treatment paradigm. Materials and methods: We report seven cases of oral cGVHD that were treated with photobiomodulation therapy (PBM), previously known as low-level laser therapy (LLLT). Patients underwent at least two PBM treatments per week in addition to local treatment with steroids, and if on systemic therapies, these were either unchanged or dosage was reduced during the period of PBM therapy. Follow-up data is presented for 4 weeks of treatment. Results: Oral pain, sensitivity, and dry mouth improved in most patients. These findings suggest PBM therapy may represent an additional approach for management of oral cGVHD, and suggest that controlled studies should be conducted to confirm the efficacy and safety of PBM therapy in oral cGVHD and to determine optimal PBM therapy protocols.

Partial tandem duplication of MLL (MLL-PTD) characterizes acute myeloid leukemia (AML) patients often with a poor prognosis. To understand the order of occurrence of MLL-PTD in relation to other major AML mutations and to identify novel mutations that may be present in this unique AML molecular subtype, exome and targeted sequencing was performed on 85 MLL-PTD AML samples using HiSeq-2000. Genes involved in the cohesin complex (STAG2), a splicing factor (U2AF1) and a poorly studied gene, MGA were recurrently mutated, whereas NPM1, one of the most frequently mutated AML gene, was not mutated in MLL-PTD patients. Interestingly, clonality analysis suggests that IDH2/1, DNMT3A, U2AF1 and TET2 mutations are clonal and occur early, and MLL-PTD likely arises after these initial mutations. Conversely, proliferative mutations (FLT3, RAS), typically appear later, are largely subclonal and tend to be unstable. This study provides important insights for understanding the relative importance of different mutations for defining a targeted therapeutic strategy for MLL-PTD AML patients.

Acute promyelocytic leukemia (APL) is a subtype of myeloid leukemia characterized by differentiation block at the promyelocyte stage. Besides the presence of chromosomal rearrangement t(15;17), leading to the formation of PML-RARA (promyelocytic leukemia-retinoic acid receptor alpha) fusion, other genetic alterations have also been implicated in APL. Here, we performed comprehensive mutational analysis of primary and relapse APL to identify somatic alterations, which cooperate with PML-RARA in the pathogenesis of APL. We explored the mutational landscape using whole-exome (n=12) and subsequent targeted sequencing of 398 genes in 153 primary and 69 relapse APL. Both primary and relapse APL harbored an average of eight non-silent somatic mutations per exome. We observed recurrent alterations of FLT3, WT1, NRAS and KRAS in the newly diagnosed APL, whereas mutations in other genes commonly mutated in myeloid leukemia were rarely detected. The molecular signature of APL relapse was characterized by emergence of frequent mutations in PML and RARA genes. Our sequencing data also demonstrates incidence of loss-of-function mutations in previously unidentified genes, ARID1B and ARID1A, both of which encode for key components of the SWI/SNF complex. We show that knockdown of ARID1B in APL cell line, NB4, results in large-scale activation of gene expression and reduced in vitro differentiation potential.

Oral mucormycosis is a rare and high risk of infection in patients following hematopoietic cell transplantation few cases in the literature. We review the literature and present an additional case to emphasize the subtle changes that resulted in positive outcome when diagnosed and managed in a comprehensive transplant team.A patient was diagnosed with gingival mucormycosis on day +25 following a hematopoietic stem cell transplant for lymphoblastic transformation of chronic myeloid leukemia. The patient was diagnosed with minor and nonspecific symptoms and was successfully treated with local dental extraction, a short course of liposomal amphotericin B and 4 months of oral posaconazole.The good outcome of this case highlights the subtle clinical changes that present early in mucormycosis and the importance of early detection and treatment of post-transplant oral infections by an experienced multidisciplinary team.

Acute myeloid leukemia (AML) with an FLT3 internal tandem duplication (FLT3-ITD) mutation is an aggressive hematologic malignancy with a grave prognosis. To identify the mutational spectrum associated with relapse, whole-exome sequencing was performed on 13 matched diagnosis, relapse, and remission trios followed by targeted sequencing of 299 genes in 67 FLT3-ITD patients. The FLT3-ITD genome has an average of 13 mutations per sample, similar to other AML subtypes, which is a low mutation rate compared with that in solid tumors. Recurrent mutations occur in genes related to DNA methylation, chromatin, histone methylation, myeloid transcription factors, signaling, adhesion, cohesin complex, and the spliceosome. Their pattern of mutual exclusivity and cooperation among mutated genes suggests that these genes have a strong biological relationship. In addition, we identified mutations in previously unappreciated genes such as MLL3, NSD1, FAT1, FAT4, and IDH3B. Mutations in 9 genes were observed in the relapse-specific phase. DNMT3A mutations are the most stable mutations, and this DNMT3A-transformed clone can be present even in morphologic complete remissions. Of note, all AML matched trio samples shared at least 1 genomic alteration at diagnosis and relapse, suggesting common ancestral clones. Two types of clonal evolution occur at relapse: either the founder clone recurs or a subclone of the founder clone escapes from induction chemotherapy and expands at relapse by acquiring new mutations. Relapse-specific mutations displayed an increase in transversions. Functional assays demonstrated that both MLL3 and FAT1 exert tumor-suppressor activity in the FLT3-ITD subtype. An inhibitor of XPO1 synergized with standard AML induction chemotherapy to inhibit FLT3-ITD growth. This study clearly shows that FLT3-ITD AML requires additional driver genetic alterations in addition to FLT3-ITD alone.

Purpose The Bruton's tyrosine kinase inhibitor ibrutinib has demonstrated clinical activity in B-cell malignancies. The DAWN study assessed the efficacy and safety of single-agent ibrutinib in chemoimmunotherapy relapsed/refractory follicular lymphoma (FL) patients. Methods DAWN was an open-label, single-arm, phase II study of ibrutinib in patients with FL with two or more prior lines of therapy. Patients received ibrutinib 560 mg daily until progressive disease/unacceptable toxicity. The primary objective was independent review committee–assessed overall response rate (ORR; complete response plus partial response). Exploratory analyses of T-cell subsets in peripheral blood (baseline/cycle 3) and cytokines/chemokines (baseline/cycle 2) were performed for available samples. Results Between March 2013 and May 2016, 110 patients with a median of three prior lines of therapy were enrolled. At median follow-up of 27.7 months, ORR was 20.9% (95% CI, 13.7% to 29.7%, which did not meet the 18% lower-bound threshold for the primary end point). Twelve patients achieved a complete response (11%; 95% CI, 5.8% to 18.3%). Median duration of response was 19.4 months (range, 1 to ≥ 33 months), with a median progression-free survival of 4.6 months and a 30-month overall survival of 61% (95% CI, 0.51% to 0.70%). Lymphoma symptoms resolved in 67%. Seven of 32 patients who experienced initial radiologic progression responded upon continuing therapy (pseudoprogression). The most common adverse events were diarrhea, fatigue, cough, and muscle spasms; 48.2% of patients reported serious adverse events. In patients who experienced a response, regulatory T cells were downregulated at C3D1 ( P = .02), and Th1-promoting (antitumor) cytokines interferon-γ and interleukin-12 increased ( P ≤ .035). Conclusion With an ORR of 20.9%, ibrutinib failed to meet its primary efficacy end point in chemoimmunotherapy in patients with relapsed/refractory FL, although responses were durable and associated with a reduction in regulatory T cells and increases in proinflammatory cytokines.

Topic Significance & Study Purpose/Background/Rationale Patients being considered for allogeneic Blood and Marrow Transplant (Allo-BMT) undergo rigorous evaluations of which can influence Allo-BMT plan and outcomes. The goal of this project was to improve the patient selection process prior to admission for Allo-BMT. The process was modeled after the Universal Protocol (UP) used prior to surgery. UP is a National Patient Safety Goal, supported by Joint Commission, Institute of Medicine, and World Health Organization. Our center aimed to standardize the approach to determine patient suitability and treatment plan with the goal of improving 1-year survival. A UP approach was employed prior to BMT to secure consensus on patient suitability and treatment plan. Methods, Intervention, & Analysis A 1-page comprehensive BMT-Time Out (BMT-TO) checklist was created with critical domains selected to provide programmatic workflow (Figure 1). All Allo-BMT recipients underwent systematic BMT-TO reviews through progressive handoffs beginning July 2016. Checklists were audited monthly for completeness and adherence to standard operating procedures. Blood Marrow Transplant Information Patient System ("bmTIPS™") generated Allogeneic Transplant Intake Dashboard (ATID) reports, providing real-time numerical data on intake distribution patterns (Figure 2). The 2017 Allo-BMT 1-year survival data reported by Center for Blood and Marrow Transplant Research(CIBMTR) were used to set confidence levels and forecast CY18,19 projected 1-year survivals. Findings & Interpretation ATID for calendar years (CY)13,14, and 15, showed acceptance rates at presentation of 92%(n=98), 94%(n=105), and 77%(n=89), respectively. ATID for CY16, 17, 18 after implementation of BMT-TO showed lower acceptance rates: 83%(n=89), 86%(n=92), 85%(n=72), respectively. CIBMTR results showed improvement in our center's Allo-BMT 1-year survival rates with use of BMT-TO checklist (Figure 3): CY14(64%/reported); CY15=64%(n=45/reported); CY16=78%(n=28/reported); CY17=89%(n=37/predicted). Discussion & Implications Implementation of systematic review of patient selection-suitability data and BMT treatment plans using structured BMT-TO checklist before Allo-BMT, correlated with improved allogeneic 1-year survivals. Further evaluation and validation of the BMT-TO form is warranted.

Relapse acute lymphoblastic leukemia (ALL) is the leading cause of childhood cancer deaths. Although relapse usually occurs in the bone marrow (medullary), extramedullary relapse occasionally occurs. Currently, the clonal origin and evolution of extramedullary relapse remain elusive. We selected two pediatric B-ALL patients who experienced testicular ALL relapse and interrogated their leukemic cells (diagnosis, remission, bone marrow relapse and testicular relapse) with whole exome sequencing. Case D483 (5.6 years old at diagnosis of ALL) developed bone marrow and testicular relapse 5 years after diagnosis of B-ALL. At diagnosis he was treated as an intermediate risk with hyperdiploid-ALL with the absence of any well-known ALL fusion-oncogene. Mutations of KRAS (G12D) and CREBBP (S1436C) were found in the founding leukemic clone at diagnosis and persisted in the bone marrow and testis at relapse). Mutation of CREBBP has been frequently found in ALL (particularly in hyperdiploid subtype) and is correlated with increased incidence of relapsed ALL. A MEF2B mutation (R17Q) was found in the bone marrow and testicular relapse sample. Missense mutation of this gene is frequently found in diffuse large B cell lymphoma (DLBCL); this protein regulates the expression of the proto-oncogene BCL6 and contributes to malignant transformation. Second child, case D727 (1.3 years old at diagnosis) harbored a MLL-AF9 fusion and was assigned as a high risk-ALL at diagnosis. Two NT5C2 mutations occurred at relapse, being present at different VAF in bone marrow and testicle: missense mutation R367Q was present with a VAF of 33.5% in bone marrow and 4.5% in testicle; while D407V was present with a VAF of 6.5% in bone marrow and 35.5% in the testicular relapse. NT5C2 encodes a 5'-nucleotidase involved in purine metabolism. The missense mutations (R367Q and D407V) identified here, have been reported as recurrent mutational hotspots of NT5C2 in relapse ALL and have been functionally validated. These mutations increase the 5'-IMP nucleotidase activity of NT5C2 protein leading to resistance to 6-mercaptopurine, a drug that was a component of the treatment regime of this patient. To understand the evolutionary trajectories of these two ALL cases, we analyzed clonal evolution based on their sequencing data. In patient D483, the relapse leukemia was directly evolved from the diagnosis leukemia clone: all of the mutations at diagnosis were persisted at relapse, and four mutated genes (MEF2B, KCNG1, AIM1, OTUD5) were acquired at both bone marrow and testicular relapse with different variant allele frequency (VAF). In patient D727, however, a faction of mutations present at diagnosis were subsequently lost at relapse, suggesting that relapsed leukemia arose from an ancestral subclone that developed before the overt leukemia at diagnosis. The mutational pattern and VAF cluster analysis results suggest that relapse in the patients' testicle represents an independently subclones from the relapse in their bone marrows. Taken together, our sequencing results suggest that relapse of patient D483 was directly evolved from the diagnosis leukemic clone; while the relapse leukemia cells (both bone marrow and testicle) of patient D727 was likely derived from a common ancestral clone, and the testicular relapse arose independently from the bone marrow relapse leukemia. Disclosures Lill: Sanofi: Speakers Bureau; California Cord Blood Services: Consultancy; Kite: Research Funding.

Current standard of care for patients with pediatric acute lymphoblastic leukemia (ALL) is mainly effective, with high remission rates after treatment. However, the genetic perturbations that give rise to this disease remain largely undefined, limiting the ability to address resistant tumors or develop less toxic targeted therapies. Here, we report the use of next-generation sequencing to interrogate the genetic and pathogenic mechanisms of 240 pediatric ALL cases with their matched remission samples. Commonly mutated genes fell into several categories, including RAS/receptor tyrosine kinases, epigenetic regulators, transcription factors involved in lineage commitment, and the p53/cell-cycle pathway. Unique recurrent mutational hotspots were observed in epigenetic regulators CREBBP (R1446C/H), WHSC1 (E1099K), and the tyrosine kinase FLT3 (K663R, N676K). The mutant WHSC1 was established as a gain-of-function oncogene, while the epigenetic regulator ARID1A and transcription factor CTCF were functionally identified as potential tumor suppressors. Analysis of 28 diagnosis/relapse trio patients plus 10 relapse cases revealed four evolutionary paths and uncovered the ordering of acquisition of mutations in these patients. This study provides a detailed mutational portrait of pediatric ALL and gives insights into the molecular pathogenesis of this disease. Cancer Res; 77(2); 390–400. ©2016 AACR.

Reduced-intensity conditioning/nonmyeloablative conditioning regimens are increasingly used in allogeneic hematopoietic cell transplantation (HCT). Reports have shown CD34(+) dose to be important for transplantation outcome using myeloablative conditioning. The role of CD34(+) dose of peripheral blood progenitor cells (PBPC) has not been previously analyzed in a large population undergoing reduced-intensity conditioning/nonmyeloablative HCT. We studied 1054 patients, ages 45 to 75 years, with acute myeloid leukemia or myelodysplastic syndrome who underwent transplantation between 2002 and 2011. Results of multivariate analysis showed that PBPC from HLA-matched siblings containing4 × 10(6) CD34(+)/kg was associated with higher nonrelapse mortality (hazard ratio [HR], 2.03; P = .001), overall mortality (HR, 1.48; P = .008), and lower neutrophil (odds ratio [OR], .76; P = .03) and platelet (OR, .76; P = .03) recovery. PBPC from unrelated donors with CD34(+) dose6 × 10(6) CD34(+)/kg was also associated with higher nonrelapse (HR, 1.38; P = .02) and overall mortality (HR, 1.20; P = .05). In contrast to reports after myeloablative HCT, CD34(+) dose did not affect relapse or graft-versus-host disease with either donor type. An upper cell dose limit was not associated with adverse outcomes. These data suggest that PBPC CD34(+) doses4 × 10(6) CD34(+)/kg and6 × 10(6) CD34(+)/kg are optimal for HLA-matched sibling and unrelated donor HCT, respectively.

Objectives We sought to investigate the magnitude and clinical importance of thrombocytopenia post transcatheter aortic valve replacement (TAVR). Background Thrombocytopenia has been observed after TAVR but has not been well studied. Methods Major thrombocytopenia (platelet count

Introduction: Adult T-cell Leukemia-Lymphoma (ATL) is a rare malignancy of T-cells infected with HTLV-1 and has the worst prognosis of the T-cell lymphomas. CC chemokine receptor 4 (CCR4) is overexpressed on ATL cells (>90% of ATL patients [pts]). Mogamulizumab (Moga) is a monoclonal antibody directed against CCR4 that has been approved in Japan for the treatment of CCR4+ ATL. There is no standard or effective treatment for relapsed/refractory (R/R) ATL pts outside of Japan. Methods: Pts from the USA, EU, and Latin America with R/R ATL (acute, lymphoma, and chronic subtypes) were randomized 2:1 to treatment with Moga, 1.0 mg/kg, given weekly for the first 4-week cycle and then biweekly, or to 1 of 3 investigator choice (IC) regimens (gemcitabine/oxaliplatin, DHAP, or pralatrexate). Pts randomized to the IC arm were permitted to cross over to Moga upon progression. The primary endpoint was confirmed objective response rate (ORR), defined as a response that is maintained for approximately 8 weeks, in those randomized to the Moga arm (based on modified Tsukasaki criteria). ORR was assessed by the treating investigator (IA) and in blinded fashion by independent review (IR). Key secondary endpoints were ORR for IC and after cross over to Moga, duration of response (DoR), progression-free survival (PFS), and overall survival (OS). Results: 71 pts were randomized (47 to Moga, 24 to IC). Tissue /blood from 65/71 pts (91.5%) expressed CCR4. Although randomized, there were baseline imbalances in known ATL prognostic factors. Pts with adverse prognostic features were more often randomized to Moga: these features included older age, poor ECOG performance status of 2, bone marrow involvement, and refractory to (vs having relapsed from) their most recent ATL therapy. Based on data through 31 March 2016, ORR (all responses, confirmed and unconfirmed) by IA was 34.0% (16/47) in the Moga group and 0% (0/24) in the IC group; these rates were largely concordant with ORR findings by IR of 27.7% (13/47) for the Moga group and 8.3% (2/24) for the IC group. Confirmed ORR for Moga was 14.9%; there were no confirmed responses in the IC group. 18 IC pts crossed over to Moga and 3 responded, including one confirmed response. The median (m) DoR for Moga was 5.65 months [mo] (95% CI 3.63, not reached). Two pts had responses lasting >9 mo. The 16 responders in the Moga arm (as assessed by IA) were evenly distributed over ATL subtypes: chronic 5/16 (31%), acute 5/16 (31%), and lymphoma 6/16 (38%). Of the 65 pts with any exposure to Moga (randomized plus crossover), response according to subtype was: chronic 5/10 (50%), acute 7/31 (23%), and lymphoma 7/24 (29%). While the study was not powered to demonstrate a difference between the treatment arms, the Kaplan-Meier plots of PFS and OS for the ITT population are presented below. The PFS curves deviate modestly in favor of the Moga group. The mPFS for Moga was 0.93 mo (min-max: 0.03-26.20 mo) with 21.5% of subjects progression free at 3 mo, while the mPFS for IC was 0.88 mo (0-4.23 mo) with 12.5 % of subjects progression free at 3 mo. The OS curves overlap. The mOS for the Moga arm was 4.90 mo (0.27-37.33 mo) with 38.9% alive at 12 mo, and the mOS for the IC arm was 6.87 mo ( 0.57-33.87 mo) with 37.5% alive at 12 mo. The OS curves may have been affected by the imbalance in prognostic factors and are confounded by the crossover of 75% of IC pts to Moga. Treatment emergent adverse events (TEAEs) occurring more often in the Moga group than the IC group were infections (51.1% vs 20.8%); respiratory disorders (48.9% vs 29.2%); infusion related reactions (46.8% vs 0%); and skin disorders (42.6% vs 8.3%). TEAEs ≥Grade 3 were 29/47 (61.7%) for Moga and 13/24 (54.2%) for IC. With the exception of known Moga-related events (infusion related reactions and drug eruptions), TEAE rates are similar when adjusted for exposure time to Moga or IC; for example, the AE rate per pt-mo of exposure for infections is 0.296 for the Moga group vs. 0.234 for the IC group. Conclusions: In the largest randomized clinical trial of pts with R/R ATL thus far conducted, commonly used cytotoxic regimens provided limited to no therapeutic benefit whereas treatment with Moga resulted in objective responses, a trend for favorable PFS, no clear survival advantage or disadvantage, and adverse events that are predictable, thereby supporting Moga's therapeutic potential in this setting. Figure Figure. Disclosures Phillips: Takeda: Speakers Bureau; Kyowa Kirin: Research Funding; Celgene: Speakers Bureau. Hermine:AB science: Consultancy, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding, Speakers Bureau; Celgene: Research Funding; Novartis: Research Funding; Alexion: Research Funding. Lill:Kite: Research Funding; California Cord Blood Services: Consultancy; Sanofi: Speakers Bureau. Feldman:Seattle Genetics: Consultancy, Research Funding, Speakers Bureau; Abbvie/Pharmacyclics/Janssen: Speakers Bureau; Celgene: Consultancy, Speakers Bureau. Sawas:Gilead Sciences: Speakers Bureau; Seattle Genetics: Honoraria. Cook:Kyowa Kirin Pharmaceutical Development, Inc.: Consultancy. Kurman:Kyowa Kirin Pharmaceutical Development, Inc: Consultancy. George:Kyowa Kirin Pharmaceutical Development: Employment. Dwyer:Kyowa Kirin Pharmaceutical Development, Inc.: Employment. Leoni:Kyowa Kirin Pharmaceutical Development, Inc.: Employment. Gonsky:Bioclinica: Other: Compensation for independent review of trial data. Horwitz:Infinity Pharmaceuticals: Research Funding; Celgene: Consultancy, Honoraria, Research Funding; Spectrum Pharmaceuticals: Consultancy, Honoraria, Research Funding; Kyowa Kirin Pharmaceutical: Research Funding; Millennium Pharmaceuticals: Consultancy, Honoraria, Research Funding; Seattle Genetics: Consultancy, Honoraria, Research Funding; Huya Bioscience International: Consultancy, Honoraria; Forty Seven, Inc.: Consultancy, Honoraria; ADC Therapeutics: Other: Travel.

Background Final Transplant Center-Specific Survival Reports are published annually by CIBMTR to display center's 1-year expected and actual survival. CIBMTR Data Audits are conducted every 4-years to determine a center's error rate by evaluating accuracy and completeness of data reported. Insurance contracts and accreditations are directly affect by these reports. Our center aimed to create a process improvement (PI) procedure to ensure highly accurate data was submitted to CIBMTR by incorporating BMT physician reviews with DCCs using a checklist. Method Firstly, 100% of data submitted to CIBMTR within the past 5 years (CY 12-17), were internally audited. Internal audit revealed common errors in comorbidity index, Karnofsky Performance Status, and disease stage. Secondly, our center voluntarily requested a CIBMTR SIA to provide objective validation of forms accuracy. Results CIBMTR SIA demonstrated an overall passing score, showcasing a marked improvement in comparison to previous audit (Figure 1). The common errors influencing survival outcomes were used to design Pre-Transplant Essential Data (TED) and Post-TED checklists (Figure 2). Physician 1:1 data reviews were conducted bi-monthly to corroborate data and/or to resolve discrepancies with DCCs. Following implementation of pre-TED/post-TED checklist and physician 1:1 reviews our center met CIBMTR's expected 1-year outcomes survival, fulfilling insurance contract(s), and accreditation(s) requirements (Figure 3). Conclusion Internal and external data audits led to the creation of the Pre-TED/Post-TED checklists. Implementation of physician 1:1 reviews positively impacted the data accuracy of CIBMTR data submitted. In conclusion, data accuracy improvements can potentially impact reported survival outcomes.

The National Marrow Donor Program, in partnership with the American Society for Blood and Marrow Transplantation, sponsored and organized a series of symposia to identify complex issues affecting the delivery of hematopoietic cell transplantation (HCT) and to collaboratively develop options for solutions. "Hematopoietic Cell Transplantation in 2020: A System Capacity Initiative" used a deliberative process model to engage professional organizations, experts, transplant centers, and stakeholders in a national collaborative effort. Year 2 efforts emphasized data analysis and identification of innovative ideas to increase HCT system efficiency, address future capacity requirements, and ensure adequate reimbursement for HCT programs to meet the projected need for HCT. This report highlights the deliberations and recommendations of Year 2 and the associated symposium held in September 2011.

Busulfan, cyclophosphamide, and etoposide (BuCyE) is a commonly used conditioning regimen for autologous stem cell transplantation (ASCT). This multicenter, phase II study examined the safety and efficacy of BuCyE with individually adjusted busulfan based on preconditioning pharmacokinetics. The study initially enrolled Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL) patients ages 18 to 80 years but was amended due to high early treatment-related mortality (TRM) in patients > 65 years. BuCyE outcomes were compared with contemporaneous recipients of carmustine, etoposide, cytarabine, and melphalan (BEAM) from the Center for International Blood and Marrow Transplant Research. Two hundred seven subjects with HL (n = 66) or NHL (n = 141) were enrolled from 32 centers in North America, and 203 underwent ASCT. Day 100 TRM for all subjects (n = 203), patients > 65 years (n = 17), and patients ≤ 65 years (n = 186) were 4.5%, 23.5%, and 2.7%, respectively. The estimated rates of 2-year progression-free survival (PFS) were 33% for HL and 58%, 77%, and 43% for diffuse large B cell lymphoma (DLBCL; n = 63), mantle cell lymphoma (MCL; n = 29), and follicular lymphoma (FL; n = 23), respectively. The estimated rates of 2-year overall survival (OS) were 76% for HL and 65%, 89%, and 89% for DLBCL, MCL, and FL, respectively. In the matched analysis rates of 2-year TRM were 3.3% for BuCyE and 3.9% for BEAM, and there were no differences in outcomes for NHL. Patients with HL had lower rates of 2-year PFS with BuCyE, 33% (95% CI, 21% to 46%), than with BEAM, 59% (95% CI, 52% to 66%), with no differences in TRM or OS. BuCyE provided adequate disease control and safety in B cell NHL patients ≤ 65 years but produced worse PFS in HL patients when compared with BEAM.

With increasing use of high dose chemotherapy with autologous and allogeneic transplants the need for the transplant physician workforce requires reassessment. The types of transplants and patients are also shifting toward transplants being done in patients with more comorbidities and more commonly these types of patients require more work effort per patient from the transplant physician. Additionally, HSCT survivors often require ongoing care at the transplant center due to the inability of the primary care workforce or the hematology/oncology workforce to absorb caring for post complex post transplant patients. The adult transplant workforce has had very few physicians join under age 40. Nearly 50% of adult transplant physicians are over age 50 whereas only 28% of pediatric transplant physicians are over age 50. By 2020, it is projected that we will need 1,264 new adult transplant physicians and 94 pediatric transplant physicians. Training time for a physician is approximately 15 years. The capping of both medical school slots and residency slots since the early ’80s is now having a very big impact on supply, but other factors are also affecting supplies such as generational differences, lifestyle expectations, and the change of the medical workforce from being mostly men. Workforce shortages are being reported for many specialities. Workforce problems are also present for nurses, pharmacists and medical technologists. So increasing use of general internists and mid-level providers may not exist as a solution. Transplant physicians must be actively engaged in the medical education process to show young medical students and residents who are not committed to another sub specialty career the excitement and challenges of a career in bone marrow transplantation, so that our field will have providers for the future.

Although a significant minority of patients with cyanotic congenital heart disease (CCHD) are thrombocytopenic, the pathogenesis and prevalence have not been established. This study was designed to address these 2 issues. We included 105 patients with CCHD (60 men and 45 women; aged 21 to 54 years). Systemic arterial oxygen saturations were 69% to 78%. Hematocrits were 62% to 74% with normal iron indexes. In 26 of 105 patients (25%), platelet counts were

In this study, we performed whole-exome and targeted sequencing on 85 MLL-PTD AML patients. These AMLs have oncogenic tandem duplication of the MLL gene. At least one well-known oncogenic driver mutation was identified in over 90% of the MLL-PTD patients. In line with earlier sequencing studies of other AML subtypes and the TCGA-AML-sequencing project, DNMT3A was the most often mutated epigenetic regulator (25%); IDH1/2 hotspot mutations were identified in 31% of patients. TET family was the third most prominently mutated epigenetic regulator (TET1 (5%), TET2 (16.3%). Mutations of epigenetic regulators also occurred in polycomb-associated proteins (EZH2, ASXL family members), chromatin remodelers (ARID2, ARID1A), genes associated with histone acetylation (CREBBP, EP300, KAT6A, KAT6B) and histone methylation (MLL2, MLL3). Proliferation-related pathway was extensively mutated, with 54 of 80 MLL-PTD patients (67.5%) carrying at least one mutation of proliferative genes. Specifically, FLT3 mutations were found in 46% of patient samples. Notably, some FLT3-ITD patients had more than one type of internal tandem duplication (ITD) insertion, probably reflecting existence of multiple subclones in these leukemias. We found highly prevalent mutations of cohesin genes: STAG2 (16%), SMC1A (6%), SMC3 (1%), RAD21 (1%) and CTCF (6%). Cohesin pathway is more frequently mutated in MLL-PTD patients (26%) than the AML samples from either TCGA (13%) or a meta-analysis of 1000 AML (9.1%). Remarkably, an extremely high proportion of the mutations had a strong tendency to disrupt the coding sequence in STAG2, emphasizing their crucial tumor-suppressor role in this AML subtype (16% in MLL-PTD vs 3% in TCGA-AML. RNA processing pathway was also strikingly altered in MLL-PTD patients. The most prominently mutated genes within this category were the splicing factors. They included U2AF1 (13%, S34F/Y), SRSF2 (3%), SF3A1 (5%), ZRSR2 (3%), DHX15 (1%) and CWC22 (1%). Multiple mutations co-occur with MLL-PTD which are usually acquired in a sequential manner. A potential ordering for acquisition of many mutations include IDH2/DNMT3A/U2AF1/TET2→MLL-PTD→RAS-receptor tyrosine kinase based on the following reasons: #1, real-time-PCR showed that MLL-PTD was absent in remission while mutations of IDH2, DNMT3A, TET2 and U2AF1 were still retained with a high VAF. This suggests that MLL-PTD was acquired after mutations of IDH2, DNMT3A, TET2 and U2AF1; #2, MLL-PTD is highly stable during disease progression as compared with mutations of the RAS-RTK. On the other hand, RAS-RTK mutations frequently exist as subclonal mutations and tend to be unstable during disease progression. These observations support a notion that MLL-PTD was acquired prior to RAS-RTK. Taken together, MLL-PTD is acquired after those remission-persisting, initiating mutations (IDH2, DNMT3A, TET2 and U2AF1), but prior to lesions of the proliferation-related drivers. Citation Format: Lingwen Ding, Qiaoyang Sun, Kar-Tong Tan, Wenwen Chien, Anand Mayakonda, Dechen Lin, Xinyi Loh, Jinfen Xiao, Manja Meggendorfer, Tamara Alpermann, Manoj Garg, Su-Lin Lim, Vikas Madan, Norimichi Hattori, Yasunobu Nagata, Satoru Miyano, Allen Yeoh Eng Juh, Hsin-An Hou, Yan-Yi Jiang, Yan-Yi Jiang, Sumiko Takao, Li-Zhen Liu, Siew-Zhuan Tan, Siew-Zhuan Tan, Michael Lill, Mutsumi Hayashi, Akitoshi Kinoshita, Hagop M. Kantarjian, Steven M. Kornblau, Seishi Ogawa, Torsten Haferlach, Henry Yang, H. Phillip Koeffler. Mutational profiling of MLL-PTD acute myeloid leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2450. doi:10.1158/1538-7445.AM2017-2450

We sought to investigate the magnitude and clinical importance of thrombocytopenia post transcatheter aortic valve replacement (TAVR).Thrombocytopenia has been observed after TAVR but has not been well studied.Major thrombocytopenia (platelet count100 × 10(9) /L) was studied following aortic valve interventions in a single center. Changes in platelets were compared in 246 patients undergoing balloon-expandable TAVR and a similar population of 57 cases undergoing surgical aortic valve replacement (SAVR in the US PARTNER IA trial).An early drop in platelets was seen on the day of intervention. The drop day 1 post procedure was similar but slightly greater with SAVR vs. TAVR. In both platelet counts continued to drop, reaching a nadir of approximately 50-60% of the baseline platelet count at day 2-3, starting to recover after day 5. Early major thrombocytopenia occurred post TAVR in 37% of patients but was not significantly related to major bleeding (OR 0.89, 95% CI 0.51-1.60, P = 0.69) or risk of stroke (HR 0.61, 95% CI 0.16-2.20, P = 0.45); there was a trend to greater acute kidney injury (OR 1.76, 95% CI 0.95-3.26, P = 0.073) and mortality (HR 1.47, 95% CI 0.98-2.22, P = 0.065). Major thrombocytopenia was persistent in 7.7% of patients and this was independently associated with mortality (HR 3.65, 95% CI 1.63-8.16, P = 0.002).Post-TAVR thrombocytopenia is a common phenomenon and its magnitude appears similar to that seen after SAVR. It is most often transient, not associated with adverse sequelae and, unless persistent, should be managed in an expectant fashion. © 2014 Wiley Periodicals, Inc.

Pediatric ALL is the most common childhood tumor and the leading cause of childhood cancer deaths. To gain a better understanding of the landscape of somatic mutations in ALL, we performed whole exome and targeted sequencing of 240 pediatric B-ALL patients with their matched remission samples. The significantly mutated genes fall into several common categories: RAS/receptor tyrosine kinases, epigenetic regulators, transcription factors involved in lineage commitment and p53/cell cycle pathway. RAS/receptor tyrosine kinases: the most frequently mutated genes were members of RAS signaling (NRAS, KRAS, FLT3, PTPN11). Besides the well know hotspot mutations [G12D/V/C (NRAS 13 cases, KRAS 13 cases), G13D (NRAS 14 cases, KRAS 11 cases) and Q61H/L/R/K (NRAS 15 cases, KRAS 1 case)], novel mutational sites were also identified for KRAS: A146T/P (3 cases), K117N/T (4 cases) and V14I (1 case). High frequency missense mutations of PTPN11 clustered in SH2 domain (included the canonical hotspot A72T (5 cases) and E76K/V (4 cases)) and tyrosine-phosphatase catalytic domain (G503R/V). For FLT3, well-appreciated activating hotspot mutations in the kinase domain (D835Y/Y842C) and several novel recurrent mutationswere identified. Epigenetic regulators: hotspot mutations were identified in histone H3K36 methyltransferase WHSC1. Mutation E1099K located in the SET domain, was identified in 10 patients as well as two of the 5 ALL cell lines that we sequenced (RS4;11, SEM). Stable silencing of E1099K mutant WHSC1 in RS4;11 cells by either lentiviral shRNA or CRISPR guide RNA (sgRNA) markedly reduced clonogenic growth both in vitro and in vivo, underscoring the critical role of WHSC1 in lymphoid malignancies. Two highly-related histone/non-histone acetyltransferases, CREBBP and EP300, were also prominently mutated in our cohort. Mutations of CREBBP predominantly occurred in the acetyltransferase domain, particularly in the hotspot R1446C/H. Mutations of chromatin remodeling genes (ARID1A and ARID2) have been identified in a number of cases. Silencing of ARID1A in ALL cell lines by lentiviral shRNA resulted in upregulation of the pro-growth regulator c-MYC, while forced expression of ARID1A reduced c-MYC luciferase reporter activity. In addition, silencing of ARID1A by either shRNA or CRISPR-sgRNA resulted in enhanced clonogenic growth, suggesting that ARID1A may be involved in the c-MYC pathway and modulates the ALL cell proliferation. Mutations of epigenetic regulators were also found in the polycomb complex (EZH2, EED, SUZ12), chromatin/nucleosome structure modifying proteins (CHD2, CHD3, CHD4), TET family proteins [TET1 (2 cases), TET2 (5 cases)] and histone modification proteins (HDAC1, SIRT1, BCOR, BRD8, lysine demethylase PHF2/KDM6A, histone acetyltransferase KAT6B). Transcription factors and p53/cell cycle pathway: a number of alterations of transcription factors essential for hematopoietic and lymphoid differentiation were noted including the lineage regulator PAX5 (5 missense, 3 indels) and ETV6 (6 cases, 3 were frameshift indel and 1 was a splice-site mutations). In addition, mutations were also found in other lineage transcription factors (IKZF2, IKZF3, EBF1), WT1 (6 cases, including 3 indels and 1 stop-gain mutations), RUNX family member [RUNX2 (7 cases), RUNX1 (1 case)], ERG1 (3 cases), GATA1/3 (1 case each) and CTCF. Somatic mutations of genes involved in the p53 pathway occurred in 18 patients, including TP53, ATM and the kinases that regulate p53 activities (HIPK1, HIPK2). Germline TP53 pathogenic variants were found in these 2 patients. Taken together, we extensively interrogated the mutational landscape of a large cohort of pediatric ALL samples by exome and targeted resequencing. This study provides a detailed mutational portrait of pediatric ALL and gives new insights into the molecular pathogenesis of this disease. Disclosures Kantarjian: Amgen: Research Funding; ARIAD: Research Funding; Bristol-Myers Squibb: Research Funding; Pfizer Inc: Research Funding; Delta-Fly Pharma: Research Funding; Novartis: Research Funding. Ogawa:Sumitomo Dainippon Pharma: Research Funding; Kan research institute: Consultancy, Research Funding; Takeda Pharmaceuticals: Consultancy, Research Funding.

Background: While first line therapy for FL with chemotherapy or chemoimmunotherapy produces durable responses, most patients (pts) with FL eventually relapse. Those who become resistant to chemoimmunotherapy have limited treatment options. Ibrutinib, an inhibitor of BTK (Bruton's tyrosine kinase) and ITK (interleukin-2-inducible T-cell kinase), has demonstrated robust clinical activity in various B-cell non-Hodgkin lymphomas. Previous Phase 1 and 2 studies have shown promising results with single-agent ibrutinib in pts with relapsed or refractory FL (Bartlett NL. Blood. 2014; 800 & Fowler N. Blood. 2015; 2706). The aim of the DAWN study was to provide additional efficacy and safety data with single-agent ibrutinib in a chemoimmunotherapy-refractory FL pt population. Methods: The DAWN study (FLR2002) was an open-label, multicenter, single-arm, Phase 2 study of ibrutinib in pts with chemoimmunotherapy-refractory FL (NCT01779791). Eligible pts had ≥ 2 prior lines of therapy with documented progressive disease (PD) within 12 months after last dose of chemotherapy in a chemoimmunotherapy regimen comprising an anti-CD20 monoclonal antibody. All pts received 560 mg oral ibrutinib once daily until disease progression or unacceptable toxicity. A protocol amendment allowed clinically stable patients with radiographic PD to remain on ibrutinib to allow for the possibility of tumor flare and delayed responses. The primary objective was Independent Review Committee (IRC)-assessed objective response rate (ORR; complete response [CR] + partial response [PR]). Secondary/ exploratory objectives included duration of response (DoR), time to next treatment (TTNT), progression-free survival (PFS), overall survival (OS), resolution of lymphoma-related symptoms, and safety. Results: The final analysis included 110 pts enrolled at 45 centers in 10 countries. The median age was 61.5 years, median number of prior therapies was 3; 40.9% of pts had refractory disease (defined as failure to achieve at least PR to the last prior regimen). Median follow-up is 27.7 months and median (range) ibrutinib exposure was 7.0 (1-37) months. The mean (SD) ibrutinib exposure was 11.2 (9.9) months. The ORR by IRC was 20.9% (23/110) and CR rate was 10.9% (12/110) with a median duration of response of 19.4 months; of the 23 pts who responded, 5 had refractory disease. ORR was 24.7% in pts with non-bulky disease (longest diameter ≤6 cm). The disease control rate (CR + PR + stable disease [SD for ≥ 6 months]) was 56.3% (62/110). 30 pts were allowed to remain on ibrutinib after initial radiographic PD (termed pseudo-progression); 7 pts had IRC-confirmed response when allowed to remain on therapy. Median TTNT on ibrutinib was 16 months compared with 10 months on the last line of treatment prior to enrollment. Median PFS was 4.6 months. The median OS has not been reached; the 24-month OS was 63% (95% CI, 0.53-0.72). Of 39 pts who had lymphoma-related symptoms at baseline, resolution of symptoms was seen in 26 pts (66.7%), including 10 whose best response was SD and 8 pts who had PD. The median duration of symptom resolution was 10.4 months. The most common adverse events (AEs) were diarrhea (50.9%; 51/56 pts had grade 1/2), fatigue (40.0%; 38/44 pts had grade 1/2), and cough (35.5%; all were grade 1/2). Serious AEs were reported in 48.2% of pts. Major hemorrhage and atrial fibrillation were consistent with previous reports at 3.6% and 9.1%, respectively. Seven pts (6.4%) discontinued ibrutinib because of an AE (none due to AF); 1 pt required a dose reduction due to AE. Biomarker analyses identifying classifiers for response or progression are ongoing. Conclusion: Treatment with single-agent ibrutinib in pts with chemoimmunotherapy-refractory FL achieved durable responses of 20.9%. The clinical benefit for these pts is also supported by disease control rate, resolution of symptoms, TTNT and OS. Safety results in this study were consistent with the current known profile of ibrutinib; the majority of events reported were grade 1/2. In pts with chemoimmunotherapy-refractory FL who have limited treatment options, treatment with ibrutinib resulted in a clinical benefit with acceptable safety and good tolerability. Figure 1. Figure 1. Figure 2. Figure 2. Disclosures Gopal: Paid Consultancy- Gilead, Janssen, Seattle Genetics, Spectrum, Research funding- Gilead, Janssen, Pfizer, BMS, Merck, Teva, Takeda, Spectrum, Seattle Genetics: Consultancy, Honoraria, Research Funding. Schuster:Celgene: Consultancy, Honoraria, Research Funding; Gilead: Research Funding; Novartis: Consultancy, Honoraria, Research Funding; Nordic Nanovector: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen Research & Development: Research Funding; Pharmacyclics: Consultancy, Research Funding; Genentech: Consultancy, Honoraria; Hoffman-LaRoche: Research Funding; Merck: Research Funding. Fowler:Roche: Consultancy, Research Funding; TG Therapeutics: Consultancy; Infinity: Consultancy, Research Funding; Jannsen: Consultancy, Research Funding; Gilead: Research Funding; Celgene: Consultancy, Research Funding. Hess:Roche: Honoraria; Celgene: Honoraria; Roche, CTI, Pfizer, Celgene: Research Funding; Janssen: Honoraria; Pfizer: Honoraria; Novartis: Honoraria. Yacoub:Alexion: Honoraria; Seattle Genetics: Consultancy, Honoraria, Speakers Bureau; Incyte: Consultancy, Honoraria, Speakers Bureau. Martin:Novartis: Consultancy; Teva: Research Funding; Celgene: Consultancy, Honoraria; Gilead: Consultancy, Other: travel, accommodations, expenses; Acerta: Consultancy; Janssen: Consultancy, Honoraria, Other: travel, accommodations, expenses. Vitolo:Celgene: Honoraria; Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Takeda: Honoraria; Gilead: Honoraria; Roche: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Jurczak:Morphosys: Consultancy; Acerta, Novartis, Pfizer, Celgene, Gillead, Janssen, Celtrion, Bayer, Morphosys, Takeda, Servier, Teva, and Roche: Research Funding; Sandoz - Novartis, Morphosys, Roche: Speakers Bureau. Osmanov:Seattle Genetics Inc.: Research Funding. Gartenberg:Janssen Research & Development: Employment. Vermeulen:Janssen Research & Development: Employment. Balasubramanian:Janssen Research & Development: Employment. Wang:Janssen Research & Development: Employment. Deshpande:Janssen Research & Development: Employment. Salles:Celgene: Consultancy, Honoraria; Mundipharma: Honoraria; Gilead: Honoraria, Research Funding; Amgen: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Janssen: Consultancy, Honoraria; Roche/Genentech: Consultancy, Honoraria, Research Funding.

Background: Ibrutinib, an inhibitor of Bruton's tyrosine kinase (BTK), has demonstrated robust clinical activity in various B-cell malignancies, whose mechanism of action, beside BTK signaling inhibition in B-cells may also depend partially on immune modulation as it also an inhibitor of ITK (interleukin-2-inducible T-cell kinase), a key regulator of T-cell activity. Immune-modulatory cancer therapy, such as check-point inhibitors, may result in different response patterns than traditional cytotoxic therapy, eg, resembling "tumor flare." In a recent study of single-agent ibrutinib for treatment of refractory follicular lymphoma (FL), 7 patients (pts) had documented tumor responses after initial radiological progressive disease (PD), a phenomenon termed "pseudo-progression." We describe these cases that led to a study protocol amendment. Methods: The FLR2002 (DAWN) study was an open-label, multicenter, single-arm, phase 2 study of ibrutinib in pts with chemoimmunotherapy-refractory (ie, ≥ 2 prior lines of therapy & PD within 12 months after last dose of chemotherapy in a chemoimmunotherapy regimen) FL (NCT01779791). All pts received ibrutinib (560 mg QD) until PD or unacceptable toxicity. A protocol amendment was made to address the novel situation of delayed responses that were observed after PD to allow for continuation of ibrutinib in pts with radiological evidence of PD (any new lesion or increase by ≥ 50% of previously involved sites from nadir), but who were clinically stable/improving/exhibiting signs of tumor flare without documented PD by other methods. The primary end point was Independent Review Committee (IRC)-assessed overall response rate (ORR; complete response [CR] + partial response [PR]). T-cell subsets in peripheral blood were monitored by flow cytometry at various timepoints. Results: The sentinel case was a pt diagnosed with grade 1 FL in 2008 and subsequently treated with 3 lines of therapy including autologous stem cell transplant. At time of enrollment, pt had stage III disease, an ECOG score: 0 and FLIPI (Follicular Lymphoma International Prognostic Index) score: 1. At Week 12, the pt was assessed as having PD. Ibrutinib was discontinued, and no other anti-cancer treatment was instituted. However, PET & CT scans performed 4 months later showed CR. Thereafter, 2 other investigators reported similar phenomena, prompting the aforementioned protocol amendment. Investigator assessment of the pt in the sentinel case was confirmed by IRC and ibrutinib treatment was resumed. The pt remains on treatment in CR for 30 months at the time of this report. On the basis of the protocol amendment, approximately 30 pts were approved to continue ibrutinib treatment after radiographic PD. Of these, an additional 6 pts were identified (7 in total) with IRC-confirmed "pseudo-progression" (confirmed by imaging) at a median of 22.0 (range: 11.6-59.6) weeks after starting Ibrutinib. Three of these pts (2 CR, 1 PR) maintained their response for > 1 year and 2 continue to respond. Two pts (1 CR, 1 PR) maintained responses for > 8 months, but eventually progressed. One pt maintained a PR > 1 year, but discontinued ibrutinib therapy due to an adverse event (grade 3 diarrhea). Sufficient sample was available in 2 of these 7 pts and examination of the T-cell profiles showed a strong downregulation of regulatory Tcells (Tregs) at early timepoints after initiation of therapy in both, an effect that was significant also in other responders (CR+ PR, mean decrease 17 to 12.9% CD4, p=0.02) but not in non-responders (SD+PD, 11.5 to 10.4% CD4, ns). Conclusion: Experience from the DAWN study suggests that pts with relapsed or refractory FL treated with ibrutinib may experience initial pseudo-progression followed by robust and durable responses when continued on ibrutinib. This effect may be related to the immune-modulatory effects of ibrutinib as seen by the decrease in Tregs after start of therapy. Biomarker studies are continuing to further understand this phenomenon. Nevertheless, these and other observations have prompted a revision in response criteria that recognize specific response patterns such as these in pts treated with immune modulating agents. Therefore current and future clinical trials should consider the observed phenomenon and physicians should be aware of this pattern of response when utilizing immune-related or targeted therapies to avoid premature discontinuation of therapy. Disclosures Salles: Amgen: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Celgene: Consultancy, Honoraria; Gilead: Honoraria, Research Funding; Janssen: Consultancy, Honoraria; Roche/Genentech: Consultancy, Honoraria, Research Funding; Mundipharma: Honoraria. Cheson:Pharmacyclics: Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead: Research Funding; Acerta: Membership on an entity's Board of Directors or advisory committees, Research Funding. Schuster:Pharmacyclics: Consultancy, Research Funding; Novartis: Research Funding; Gilead: Research Funding; Janssen Research & Development: Research Funding; Nordic Nanovector: Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Research Funding; Genentech: Consultancy; Hoffman-LaRoche: Research Funding. Radford:GSK: Equity Ownership; Astra-Zeneca: Equity Ownership; Seattle Genetics: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Takeda: Consultancy, Honoraria, Research Funding, Speakers Bureau. Schaffer:Janssen Research & Development: Employment. Damle:Janssen Research & Development: Employment. Gartenberg:Janssen Research & Development: Employment. Vermeulen:Janssen Research & Development: Employment. Balasubramanian:Janssen Research & Development: Employment. Deshpande:Janssen Research & Development: Employment.

Acute promyelocytic leukemia (APL) is a subtype of myeloid leukemia characterized by differentiation block at the promyelocyte stage. Besides the presence of chromosomal rearrangement t(15;17), leading to the formation of PML-RARA (promyelocytic leukemia-retinoic acid receptor alpha) fusion, other genetic alterations have also been implicated in APL. Here, we performed comprehensive mutational analysis of primary and relapse APL to identify somatic alterations, which cooperate with PML-RARA in the pathogenesis of APL. We explored the mutational landscape using whole-exome (n=12) and subsequent targeted sequencing of 398 genes in 153 primary and 69 relapse APL. Both primary and relapse APL harbored an average of eight non-silent somatic mutations per exome. We observed recurrent alterations of FLT3, WT1, NRAS and KRAS in the newly diagnosed APL, whereas mutations in other genes commonly mutated in myeloid leukemia were rarely detected. The molecular signature of APL relapse was characterized by emergence of frequent mutations in PML and RARA genes. Our sequencing data also demonstrates incidence of loss-of-function mutations in previously unidentified genes, ARID1B and ARID1A, both of which encode for key components of the SWI/SNF complex. We show that knockdown of ARID1B in APL cell line, NB4, results in large-scale activation of gene expression and reduced in vitro differentiation potential.

International audience; We evaluated outcomes and associated prognostic factors in 233 patients undergoing allogeneic hematopoietic cell transplantation (HCT) for primary myelofibrosis (MF) using reduced-intensity conditioning (RIC). The median age at RIC HCT was 55 yr. Donors were a matched sibling donor (MSD) in 34% of RIC HCTs, an HLA well-matched unrelated donor (URD) in 45%, and a partially matched/mismatched URD in 21%. Risk stratification according to the Dynamic International Prognostic Scoring System (DIPSS) was 12% low, 49% intermediate-1, 37% intermediate-2, and 1% high. The probability of survival at 5 yr was 47% (95% confidence interval [CI], 40% to 53%). In a multivariate analysis, donor type was the sole independent factor associated with survival. Adjusted probabilities of survival at 5-yr were 56% (95% CI, 44% to 67%) for MSD, 48% (95% CI, 37% to 58%) for well-matched URD, and 34% (95% CI, 21% to 47%) for partially matched/mismatched URD (P = .002). The relative risk (RR) for NRM was 3.92 (P = .006) for well-matched URD and 9.37 (P < .0001) for partially matched/mismatched URD. Trends toward increased NRM (RR, 1.7; P = .07) and inferior survival (RR, 1.37; P = .10) were observed in DIPSS intermediate-2/high-risk patients compared with DIPSS low/intermediate-1 risk patients. Our data indicate that RIC HCT is a potentially curative option for patients with MF, and that donor type is the most important factor influencing survival in these patients.

This paper summarizes a pilot, sequential dose-escalation study of PEG-rHuMGDF in patients with advanced malignancies who had delayed platelet recovery after autologous stem cell transplantation (ASCT). Patients were randomized to receive either placebo (n = 11) or PEG-rHuMGDF at 5 (n = 9), 10 (n = 6), or 25 (n = 7) microg/kg/day by subcutaneous injection for 14 days and were monitored for 5 weeks. Across all treatment groups, eight patients had platelet recovery to > or = 20 x 10(9)/l by day 21. The proportion of patients achieving platelet recovery, the median number of days and units of platelet transfusions were similar for the placebo and the PEG-rHuMGDF groups. PEG-rHuMGDF was well tolerated at all dosages. The incidence rates of adverse events in all groups were similar. No deaths on study, no drug-related serious adverse events, and no development of neutralizing antibodies to MGDF occurred.