Treatment Of Juvenile Myelomonocytic Leukemia Research Articles

Clinical Outcome of Unrelated Cord Blood Engraftment for Children with Juvenile Myelomonocytic Leukemia

Objective: To evaluate the clinical efficacy and safety of unrelated cord blood engraftment in the treatment of Juvenile Myelomonocytic Leukemia. Method: Seventy-three JMML children received cord blood transplantation since 2018. 47 children with cord blood engrafment were analyzed retrospectively. The median at diagnosis age was 35 months (12 to 99). Of them, 8 patients with NF-1 mutation, 24 with PTPN11 mutation, 2 with Nras somatic mutation, 4 with Kras somatic mutation, 8 with multiple mutation. The transplantation regimen was divided into two groups: Complementary transplantation group (CT-group) i.e. 55 patients underwent unrelated cord blood(UCB) following haploidentical stem cells transplantation(see blood 2016 128:1235 and blood 2019 134: 4181).UCB-cells engrafted finally in 31 patients,(others with haplo-engrafted).Conditioning regimen composed of Cyclophosphamide (Cy), Busulfan (Bu), Thiotepa (TT) and Fludarabine (Flu). Haploid PBSC (MNC >15×108/kg) was infused on day 0. GVHD prophylaxis consisted of PTCy on day+3 to +4,and FK506 and Mycophenolate Mofetil (MMF). Unrelated cord blood was infused on day+6. The next group was haploid DLI Bridging unrelated cord blood transplantation group (LCB-group) . Conditioning regimen composed of Cla+Ara-C for 4 days, and the haploid donor DLI was infused on day-11 (the number of infusion donor Lymphocyte was 4-8 ×108/kg) ,Cy, FLU, BU and TT started on day-8~-3. Unrelated cord blood was infused on day 0.GVHD prophylaxis consisted of FK506 and Mycophenolate Mofetil (MMF). Unrelated cord blood matching requires ≥7/10 HLA locus , and Kir assay was carried out at the same time. Cord blood with Haplotype Cen-Bx was preferred. Results: The median follow-up time was 32 months(2 to 52ms).47 patients were engrafted with cord blood in the two groups totally. In the CT group,Haplo-cells and UCB-cells engrafted finally in 24 and 31 patients, respectively.In the LCB group, only 1 of 17 cases developed primary graft failure,who achieved disease-free survival after the second haploid transplantation. Of the 47 patients with cord blood engraftment in the two groups, the median number of infusion TNC and CD34+ from cord blood were 8.1(3.3~17)×107/kg and 0.25(0.08~4.0)×106/kg, respectively. The median times to neutrophil more than 0.5×109/L and platelet more than 20×109/L were 38 (19-67) days and 36(12-67) days post-HSCT, respectively. The Overall survival (OS) and leukemia free survival (LFS) were 97.9% and 95.8%, respectively. There were no significant difference in the OS and LFS between CT group and LCB group (p=0.18 and p=0.50, respectively). The cumulative incidences of grades Ⅱ-Ⅳ° aGVHD was 29.8% (14pts).There were 4 patients with grade III° and Ⅳ° aGVHD ，all of whom were effectively improved. Chronic GVHD occurred in 13 patients who mainly had mild skin involvement. Only one of them had mild BO. The transplantation-related mortality was 2.1% (1 case died of severe pre-engraftment syndrome in LCB group). The transient reactivation of virus was mainly caused by HHV-6. One case developed HHV-6 virus encephalitis.And no VOD occurred. Summary: UnrelatedCord blood engraftment has significant clinical efficacy in the treatment of JMML,and it is relatively safe and reliable.UnrelatedCord blood could be used as a priority donor for such children. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Molecular characteristics and advances in diagnosis and treatment of juvenile myelomonocytic leukemia

幼年型粒单核细胞白血病（JMML）是一种罕见的儿童早期恶性克隆性疾病，2022年世界卫生组织公布的第5版血淋巴肿瘤分类已将其归入骨髓增殖性肿瘤（MPN）。近年来，JMML的多组学研究不断取得进展，使JMML的临床分型得以补充、分子特征被逐步认识，这有助于疾病的诊断和危险度分层，进而根据不同危险度制定更加合理的临床决策，现对JMML的分子特征及诊疗进展进行综述。.

Colon Cast in a Child With Graft Versus Host Disease.

A 4-year-old girl with juvenile myelomonocytic leukemia underwent a cord blood transplant 2 months post-diagnosis following a poor response to chemotherapy. One month after the transplant, she developed a skin rash, abdominal pain, and bloody diarrhea despite receiving oral tacrolimus and methotrexate to prevent graft versus host disease (GVHD). Administration of intravenous prednisolone relieved the skin rash but not the intestinal symptoms. Colonoscopy revealed marked redness and bleeding in the colonic mucosa. Neither corticosteroid enemas nor intravenous antibiotics improved her symptoms. Two months after the transplant, she excreted an approximately 6-cm colon cast, a full-thickness, infarcted colonic segment (Fig. 1). Colonoscopy and pathology showed a demucosed colon (Fig. 2). Hyperbaric oxygen (HBO) therapy was initiated with the suspicion of intestinal ischemia secondary to transplant-related thrombotic microangiography. Her symptoms improved within a week after therapy initiation.FIGURE 1.: The excreted colon cast.FIGURE 2.: Colonoscopic and histologic findings of the colon. Left) Colonoscopic examination revealed diffuse mucosal inflammation and peeling. Endoscopic examination could only be performed up to 15 cm due to colonic stenosis. Right) Histology of the sigmoid colon showed infiltrating lymphocytes and loss of mucous membrane.Follow-up colonoscopy performed 3 months after the HBO therapy revealed healed colonic mucosa. Giuseppe et al reported that 37% of patients experience gastrointestinal GVHD after allogeneic hematopoietic stem cell transplant (1). A colon cast is believed to be associated with mucosal ischemia because most cases are reported in patients with ischemic colitis (2,3). Although several cases in adults have been reported, the presumed first case of colon cast associated with ischemia in a child was reported in 2017 (4). Colon cast associated with GVHD is rare, with only one previously reported adult case (5). In our case, the patient had demucosed colon on colonoscopy, similar to previous cases, and showed immediate improvement after the HBO therapy. The HBO therapy may be a safe and effective treatment option for intestinal GVHD accompanied by a colon cast. Patient anonymity: this patient’s anonymity is carefully protected. There are no identifiable photographs of the patient included in this manuscript. ACKNOWLEDGMENTS T.M. wrote the initial draft of the manuscript. T.I. performed colonoscopy and hyperbaric oxygen therapy and assisted in preparing the manuscript. All other authors were involved in the treatment of juvenile myelomonocytic leukemia and critically reviewed the manuscript. All authors approved and agreed to be accountable for the manuscript.

Current Treatment of Juvenile Myelomonocytic Leukemia.

Juvenile myelomonocytic leukemia (JMML) is a rare pediatric leukemia characterized by mutations in five canonical RAS pathway genes. The diagnosis is made by typical clinical and hematological findings associated with a compatible mutation. Although this is sufficient for clinical decision-making in most JMML cases, more in-depth analysis can include DNA methylation class and panel sequencing analysis for secondary mutations. NRAS-initiated JMML is heterogeneous and adequate management ranges from watchful waiting to allogeneic hematopoietic stem cell transplantation (HSCT). Upfront azacitidine in KRAS patients can achieve long-term remissions without HSCT; if HSCT is required, a less toxic preparative regimen is recommended. Germline CBL patients often experience spontaneous resolution of the leukemia or exhibit stable mixed chimerism after HSCT. JMML driven by PTPN11 or NF1 is often rapidly progressive, requires swift HSCT and may benefit from pretransplant therapy with azacitidine. Because graft-versus-leukemia alloimmunity is central to cure high risk patients, the immunosuppressive regimen should be discontinued early after HSCT.

Pharmacologic Inhibition of PI3Kδ Prolongs Survival of Mutant Shp2E76K-Expressing Mice

Juvenile myelomonocytic leukemia (JMML) is an aggressive overproduction of cells in the myeloid lineage, characterized by hypersensitivity to granulocyte macrophage colony-stimulating factor (GM-CSF). There is a clear need for improved chemotherapies, as the current treatments are ineffective. The most commonly mutated gene in JMML patients is PTPN11, which encodes the protein tyrosine phosphatase Shp2. One of the downstream targets of Shp2 is phosphoinositide 3-kinase (PI3K), and we have previously focused on the hematopoietic-specific catalytic subunit p110δ (PI3Kδ). The PI3Kδ inhibitor idelalisib is FDA-approved for patients with B cell malignancies. However, the effectiveness of PI3Kδ inhibition in JMML and other myeloid malignancies has not been studied. Therefore, following our previous work in vitro which demonstrated reduced proliferation of GOF Shp2-expressing murine cells and primary JMML cells (Goodwin et. al, Blood 2014), we assessed the effect of PI3Kδ inhibition on GOF Shp2-expressing mice in vivo as the next step in exploring PI3Kδ inhibition as a potential treatment in JMML.We treated Shp2E76K/+;LysMcre+ mice between 12 and 20 weeks of age with 30mg/kg of the PI3Kδ inhibitor GS-9820 (hereafter PI3Kδ inhibitor) or vehicle BID via oral gavage for 21 days. First, we evaluated mice directly after completing 21 days of treatment (16 hours following final vehicle or PI3Kδ inhibitor dose). The PI3Kδ inhibitor-treated mice had significantly reduced spleen-to-body weight ratio in comparison to the vehicle-treated mice. We measured the colony-forming ability of bone marrow low-density mononuclear cells (LDMNCs) in response to GM-CSF and found only a slight reduction in drug-treated mice, indicating that pharmacologic inhibition of PI3Kδ does not permanently correct GM-CSF hypersensitivity. To investigate how PI3Kδ inhibitor treatment induced the functional effect of reduced spleen size, we phenotypically analyzed the bone marrow and peripheral blood for hematopoietic progenitor cells (lineage-Sca1+cKit+, LSK) and for mature myeloid cells (Gr1+Mac1+). The frequency of LSK cells was significantly reduced in the bone marrow of PI3Kδ inhibitor-treated mice. There was no difference in the frequency of Gr1+Mac1+ in the bone marrow compartment, but there was a significant increase in the peripheral blood of PI3Kδ inhibitor-treated mice. Furthermore, the Mac1+ mean fluorescence intensity in the peripheral blood was significantly higher in PI3Kδ inhibitor-treated mice. Collectively, these findings suggest that PI3Kδ inhibition induced stem/progenitor terminal differentiation and reduced the self-renewal and hyperproliferation of immature myeloid cells.A second cohort of treated mice was followed for overall survival. Survival of the PI3Kδ inhibitor-treated mice was significantly prolonged compared to their vehicle-treated counterparts. Serial peripheral blood WBC counts were similar between the two groups until approximately 20 weeks and 34 weeks after the start of treatment, when WBC counts trended upward in the vehicle-treated and drug-treated animals, respectively. These time points coincided with a sharp decrease in the number of surviving mice in the corresponding group. When moribund, mice were euthanized to assess the cause of death. The PI3Kδ inhibitor-treated animals had no difference in splenomegaly and leukocytosis compared to the vehicle-treated animals at the time of euthanasia, and composition of the bone marrow and spleen in moribund animals was not significantly different, suggesting that mice of the two groups ultimately succumbed to the same disease. Taken together, these findings suggest that time-limited PI3Kδ inhibition delays onset of mutant Shp2-induced leukemia, but does not alter the ultimate course of disease. These findings imply that continued PI3Kδ inhibition may be needed for optimal treatment.Our results demonstrate that in vivo administration of a hematopoietic-specific PI3Kδ inhibitor increases the survival of GOF Shp2-expressing mice. The more limited side effect profile of PI3Kδ inhibition compared to pan-PI3K inhibition may provide a therapeutic window to simultaneously target both the Ras-Erk and PI3K-Akt pathways in the treatment of JMML. This work supports the rationale of using PI3Kδ inhibitors in the treatment of patients with JMML and possibly other myeloid cell malignancies. DisclosuresNo relevant conflicts of interest to declare.

Analysis of the Short-Term Efficacy of Decitabine Combined with Low-Dose Cytarabine in the Treatment of JMML

Objective: Juvenile myelomonocytic leukemia (JMML) has both the disease characteristics of myelodysplastic syndrome and myeloproliferative neoplasm. Currently, hematopoietic stem cell transplantation (HSCT) is the only possible cure. Most of the newly diagnosed pediatric patients with JMML have high tumor burden, rapid disease progression, and may not tolerate HSCT. This study explored the short-term efficacy of decitabine combined with low-dose chemotherapy in the treatment of JMML before transplantation. Methods: A retrospective analysis of the patient files of 9 cases of JMML was performed from January 2019 to May 2020. All patients were given decitabine 20mg/m2× 5 days, supplemented with a small dose of cytarabine (50-100mg/m2×3~5 days), and/or etoposide (50mg/m2×3~5 days) chemotherapy. Each treatment interval is 3~4 weeks, bridging with HSCT after 3~4 treatment courses. Results: The median age of onset of 9 cases of JMML was 2 years old (0.5~4 y), male to female ratio was 8:1, the median size of spleen was 6.4cm (2.9~9.8cm) under the costal arch, and WBC was 28.03×109/L (7.3~127.69×109/L), monocytes were 9.25×109/L (1.66~15.79×109/L) at diagnosis. There were 8 cases in the high-risk group and 1 case in the low-risk group. Second-generation sequencing results show that 7 cases carried PTPN11 somatic mutations. Five in 9 cases had two kinds of classic JMML mutations, and 1 case had only NRAS mutation. Seven patients had normal chromosomal karyotypes, and 3 patients had abnormal 8, 11, and 18 chromosomes, respectively. Median treatment courses with decitabine are 3 courses (1~5 courses), the response rate of one course is 77.8% (7/9), the response rate of three courses is 80% (4/5), one case from high-risk group achieved complete remission after treated with 4 courses of decitabine and low-dose chemotherapy. The five-month progression-free survival rate was 77.8% (7/9). Conclusion: Treating JMML with decitabine combined with low-dose chemotherapy, can reduce patients' tumor burden, improve the general condition, and obtain approximately 80% clinical response rate. Decitabine combined with low-dose chemotherapy can be used as a treatment option for JMML before HSCT. Disclosures No relevant conflicts of interest to declare. OffLabel Disclosure: Safety and effecacy of decitabine applicated in JMML patients have not been confirmed.

Efficacy Analysis of Unrelated Cord Blood Transplantation in the Treatment of Juvenile Myelomonocytic Leukemia

To explore the clinical efficacy and safety of unrelated umbilical cord blood transplantation (UCBT) in the treatment of Juvenile myelomonocytic leukemia (JMML). The clinical data of 5 children with JMML who were treated with unrelated UCBT from October 2011 to July 2019 were retrospectively analyzed. The age of onset for the five children (male) ranged from 0.4 to 5.0 years old, with a median age of 1.5 years old. All the patients received myeloablative conditioning regimen without ATG to whom cyclosporine A (CsA) with short-term mycophenolate mofetil (MMF) was given for GVHD prophylaxis. Four children acquired engraftment. One patient received secondary haploidentical hematopoietic stem cell transplantation because of the failure in the first unrelated UCBT. Grade Ⅲ to Ⅳ aGVHD occurred in 2 cases and was controlled, and none of the patients developed cGVHD. Three cases achieved long-time disease free survival，and no patient relapsed. UCBT is an effective treatment for children with JMML.

Molecular assessment of pretransplant chemotherapy in the treatment of juvenile myelomonocytic leukemia.

Despite the intensity of hematopoietic stem cell transplantation (HCT), relapse remains the most common cause of death in juvenile myelomonocytic leukemia (JMML). In contrast to other leukemias where therapy is used to reduce leukemic burden prior to transplant, many patients with JMML proceed directly to HCT with active disease. The objective of this study was to elucidate whether pre-HCT therapy has an effect on the molecular burden of disease and how this affects outcome post-HCT. Twenty-one patients with JMML who received pre-HCT therapy and were transplanted at UCSF were analyzed in this study. The mutant allele frequency of the driver mutation was assessed before and after pre-HCT therapy, using custom amplicon next-generation sequencing. Of the 21 patients, seven patients (33%) responded to therapy with a significant reduction in their mutant allele frequency and were classified as molecular responders. Six of these patients received moderate-intensity chemotherapy, one patient received only azacitidine. The 5-year progression-free survival after HCT of molecular responders was 100%versus 61% for nonresponders (P=.12). Survival of molecular nonresponders was not improved by use of high-intensity conditioning, but patients were salvaged if they experienced severe graft versus host disease. There were no baseline clinical characteristics that were associated with response to pre-HCT therapy. Despite the myelodysplastic nature of JMML, patients treated with pre-HCT therapy can achieve molecular remissions. These patients experienced a trend toward improved outcomes post-HCT. Importantly, molecular testing can be helpful to distinguish between responders and nonresponders and should become an integral part of clinical care.

Diagnosis and treatment of juvenile myelomonocytic leukemia

ABSTRACTObjective: To investigate clinical features, diagnosis, treatment strategies and prognosis of juvenile myelomonocytic leukemia (JMML).Methods: The clinical data of 21 patients with JMML who were diagnosed in our hospital from January 2013 to May 2018 were retrospectively analyzed.Results: Among the 21 children with JMML, 16 were male and 5 were female. Out of the 21 children who were diagnosed with JMML, 7 were lost after treatment while the remaining 14 received A-3V chemotherapy regimen of South Korea. The effective response rate was 78.5%. The three-year overall survival (OS) rate and three-year disease-free survival (DFS) rate were (76.2 ± 14.8)% and (66.2 ± 14)%, respectively. Single factor analysis showed that PLT count ≤33×109/L, LDH level >500 U/L and HbF level >10% and chemotherapy only were the significant factors that lead to poor prognosis in children. Cox multivariate analysis showed that the choice of treatment options affected the prognosis of JMML children. By taking prognostic factors for long-term efficacy into account, patients with treatment strategy of chemotherapy alongside hematopoietic stem cell transplantation (HSCT) have a better prognosis.Conclusion: The PLT count, LDH level, HbF level and choice of treatment plan are important for the evaluation of prognosis for children with JMML. Although there is a lack of consistency in terms of donors but the A-3V scheme is relatively stable, so HSCT should be preferred for children with poor prognostic factors.

Clinical Study on Treatment of Juvenile Myelomonocytic Leukemia with Haploidentical-Hematopoietic Stem Cell Transplantation

To analyze the therapeutic efficacy of haploidentical-hematopoietic stem cell transplantation (hi-HSCT) for patients with juvenile myelomonocytic leukemia (JMML). The engraftment of hematopoietic stem cells, incidence of graft versus host disease (GVHD), infection, relapse, and survival of 6 JMML patients received hi-HSCT were retrospectively analyzed. Six (6 males) JMML patients received hi-HSCT from haplo-HLA-matched related donors. The results showed that the hematopoictic stem cells in all 6 patients were grafted successfully. Two cases of JMML died of pulmenary infections, other 4 cases survive without disease. Acute GVHD occurred in 3 patients and chronic GVHD occurred in 1 patients. The overall survival, disease free survival and relapse rates were 66.7%, 66.7%, 0%, respectively. The hi-HSCT is an effective method for treatment of patients with JMML, but there also is a serial problems to be resolved.

Use of Rapamycin in a Patient With Juvenile Myelomonocytic Leukemia: A Case Report.

The relapse rate for children with juvenile myelomonocytic leukemia (JMML) status post hematopoietic stem cell transplantation (HSCT) approaches 50% within 5 years. Graft-versus-leukemia (GVL) is thought to play important role in the treatment of JMML. For this reason, careful management of immunosuppressive drugs after HSCT is crucial. This case report demonstrates that rapamycin and GVL represent a viable medical strategy for the management of pediatric patients with JMML who relapse following status post-HSCT.

Diagnosis and treatment of juvenile myelomonocytic leukemia.

Juvenile myelomonocytic leukemia (JMML) is a rare myelodysplastic/myeloproliferative disorder that occurs during infancy and early childhood; this disorder is characterized by hypersensitivity of the myeloid progenitor cells to granulocyte-macrophage colony-stimulating factor invitro. JMML usually involves somatic and/or germline mutations in the genes of the RAS pathway, including PTPN11, NRAS, KRAS, NF1, and CBL, in the leukemic cells. Almost all patients with JMML experience an aggressive clinical course, and hematopoietic stem cell transplantation (HSCT) is the only curative treatment. A certain proportion of patients with somatic NRAS and germline mutations in CBL, however, have spontaneous resolution. A suitable treatment after diagnosis and conditioning regimen prior to HSCT are yet to be determined, but several clinical trials have been initiated throughout the world to develop suitable pre- or post-allogeneic HSCT treatments and new targeted therapies that are less toxic, to improve patient outcome.

Molecular mechanism and progress in treatment of juvenile myelomonocytic leukemia (JMML)

Molecular mechanism and progress in treatment of juvenile myelomonocytic leukemia (JMML)

若年性骨髄単球性白血病(JMML)の分子機構と治療

若年性骨髄単球性白血病(JMML)の分子機構と治療

Molecular-Targeted Therapies for Hematologic Malignancies

Major advances in the disciplines of hematology, genetics, biochemistry, and chemistry over the past decades have empowered investigators with the background and methods required for development of customized molecular-targeted therapies. The ability to identify signaling pathways that are dysregulated, to determine the associated mutations, and to develop chemical drugs toward a desired correction is now a realistic work flow. The landmark demonstration that BCR-ABL could be molecularly targeted and could have a major impact upon disease progression really ignited the field of targeted therapies for hematologic malignancies. Through much hard work, we now know the key drivers of some hematologic malignancies, and depending on the particular disease, we have an arsenal of agents available to act at multiple nodal points. Some enabling technologies that have been a key for these advances include small molecule screening, high throughput whole genome sequencing, mouse models for cancer, and gene and microRNA expression array analyses. In this special issue, we present a collection of seven articles that contribute to our understanding of molecular targets and the development of approaches for their inhibition or rationale use of existing agents or their derivatives. These targets range from very early initiators of malignancy to molecules that are already advanced into clinical studies. The first paper in this issue by A. Fathi and T. Abdel-Wahab “Mutations in epigenetic modifiers in myeloid malignancies and the prospect of novel epigenetic-targeted therapy” addresses the series of mutations identified that alter DNA and/or histone lysine methylation. These early epigenetic changes predispose to leukemogenesis and are relevant in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) patients. The second paper by J. Okabe-Kado et al. “Extracellular NM23 protein as a therapeutic target for hematologic malignancies” describes extracellular NM23-H1 protein and its relationship with altered signaling pathways and growth/survival in AML. The third paper by S. Verma et al. “Gab adapter proteins as therapeutic targets for hematologic disease” describes the Grb2-associated adapter proteins (Gabs) as potential therapeutic targets playing major roles in regulation of multiple signaling pathways. These first three papers describe potential new targets that require further validation and greater specificity, but could have significant impact on the initiation and progression of hematologic malignancy. The fourth paper by X. Liu et al. “Molecular targets for the treatment of juvenile myelomonocytic leukemia” describes the outstanding progress made in identifying the mutations associated with activation of the Ras pathway in juvenile myelomonocytic leukemia (JMML), several of which are already targets of drugs that are being tested such as Ras and SHP-2. This paper additionally points toward protein:protein interactions as potential therapeutic targets such as SHP-2/Gab2 in hematopoiesis. The fifth paper by P. Argyriou et al. “The role of mTOR inhibitors for the treatment of B-cell lymphomas” focuses on the downstream activation of the mTOR pathway and the development and testing of new rapalogues and ATP-competitive inhibitors for clinical use. The mTOR pathway is central to cell survival and metabolism and represents a common target for many types of cancers. The sixth paper by F. Tzifi et al. “The role of Bcl2 family of apoptosis regulator proteins in acute and chronic leukemias” describes the exciting new advances in understanding and targeting the Bcl-2 family of proteins and gives a comprehensive update on new agents that are in clinical studies targeting survival in acute and chronic leukemias. All of these three papers describe bona fide targets that are already subject to significant validation and commercial drug development. The seventh paper by P. Koehler et al. “Engineered T cells for the adoptive therapy of B cell-chronic lymphocytic leukemia (B-CLL)” in the issue addresses immunotherapy using T-cell therapies against CD19, which has recently been very successful and received widespread attention for treatment of chronic lymphocytic leukemia. This approach promises to provide sustained targeted therapy based on cell surface phenotype and although it has to deal with issues such as B-cell deficiency and intravenous immunoglobuin infusions, such approaches when combined with chemotherapy are very promising as a form of targeted gene-based therapy. In summary, the articles in this special issue address the spectrum of new targeted therapy development, from basic understandings of structure-function to mature rationale drugs already being tested in patients. We sought to cover the full spectrum of therapeutic development and are pleased to present a series of papers that do just that. As editors of this issue, we appreciate the important contributions of the authors of these review articles and hope that this issue will encourage expanded translational research toward developing novel therapies for hematologic malignancies. Kevin D. Bunting Cheng-Kui Qu Michael H. Tomasson

Molecular Targets for the Treatment of Juvenile Myelomonocytic Leukemia

Significant advances in our understanding of the genetic defects and the pathogenesis of juvenile myelomonocytic leukemia (JMML) have been achieved in the last several years. The information gathered tremendously helps us in designing molecular targeted therapies for this otherwise fatal disease. Various approaches are being investigated to target defective pathways/molecules in this disease. However, effective therapy is still lacking. Development of specific target-based drugs for JMML remains a big challenge and represents a promising direction in this field.

Childhood Myelodysplastic Syndrome: Focus on the Approach to Diagnosis and Treatment of Juvenile Myelomonocytic Leukemia

Expansion of myeloid blasts with suppression of normal hematopoiesis is a hallmark of acute myeloid leukemia (AML). In contrast, myeloproliferative neoplasms (MPNs) are clonal disorders characterized by overproliferation of one or more lineages that retain the ability to differentiate. Juvenile myelomonocytic leukemia (JMML) is an aggressive MPN of childhood that is clinically characterized by the overproduction of monocytic cells that can infiltrate organs, including the spleen, liver, gastrointestinal tract, and lung. Major progress in understanding the pathogenesis of JMML has been achieved by mapping out the genetic lesions that occur in patients. The spectrum of mutations described thus far in JMML occur in genes that encode proteins that signal through the Ras/mitogen-activated protein kinase (MAPK) pathways, thus providing potential new opportunities for both diagnosis and therapy. These genes include NF1, NRAS, KRAS, PTPN11, and, most recently, CBL. While the current standard of care for patients with JMML relies on allogeneic hematopoietic stem-cell transplant, relapse is the most frequent cause of treatment failure. Rarely, spontaneous resolution of this disorder can occur but is unpredictable. This review is focused on the genetic abnormalities that occur in JMML, with particular attention to germ-line predisposition syndromes associated with the disorder. Current approaches to therapy are also discussed.

A Novel Assay for Juvenile Myelomonocytic Leukemia Based on Aberrant Signaling Networks Measured Via Phospho-Specific Flow Cytometry Reduces Diagnosis Time from Weeks to Days.

Juvenile myelomonocytic leukemia (JMML) is an aggressive clonal malignancy characterized by over-production of myeloid lineage cells that infiltrate hematopoietic and non-hematopoietic tissues. The diagnosis and treatment of JMML can present great challenges for clinicians. Infants or young children are suspected of having this disease when presenting with hepatosplenomegaly and a peripheral blood monocytosis - criteria that are often present in more prevalent but potentially less lethal conditions such as viral infections. The diagnostic workup includes several clinical tests as well as the confirmation of abnormal numbers of granulocyte-macrophage colony forming units (CFU-GM) grown in low concentrations of granulocyte-macrophage colony stimulating factor (GM-CSF). The colony assays are laborious and can take up to 4–6 weeks for results. Using phospho-specific flow cytometry, a technique that allows simultaneous single cell measurements of cell type and signal, we have developed an assay that shows a specific cell signature in JMML patients that can reduce diagnosis time from weeks to days. The assay combines traditional surface marker phenotyping with signaling responses in low doses of GM-CSF to identify a population of CD38 intermediate cells that hyper-phosphorylate Stat 5 (pStat5). We applied this assay to primary cells obtained from 10 JMML patients and demonstrate that the cell signature is specific to JMML with respect to a wide panel of other diagnoses. For comparison purposes, we developed a statistical measure to summarize the signature and monitor its expression across varying doses of GM-CSF. The measure also ensures consistency by eliminating the need for additional manual data processing steps (e.g. gating) prior to comparison—an important step towards developing a diagnostic tool. In addition, this characteristic phospho-specific flow signature disappeared in patient samples in remission and reappeared at relapse—indicating that the signature may indeed be representative of the malignant population in JMML. Further characterization of these cells is ongoing. We conclude that phospho-specific flow cytometry is a robust methodology that can be used in primary human cells as a method to rapidly diagnosis JMML as well as to follow patients on therapy. Importantly, it is a powerful technical tool that will allow improved understanding of the complex signaling pathways that are perturbed in cancer cells.

Non‐hematopoietic stem cell transplantation treatment of juvenile myelomonocytic leukemia: A retrospective analysis and definition of response criteria

Juvenile myelomonocytic leukemia (JMML) is a rare myeloproliferative disease of infancy. Allogeneic hematopoietic stem cell transplantation (HSCT) is currently the only curative treatment modality, while the role of anti-leukemic therapy prior to HSCT is uncertain. A comparative evaluation of the efficacy of different clinical protocols and great variety of anti-neoplastic drugs applied pre-HSCT is hampered by the lack of uniform criteria of response. Classification schemas applied in other forms of leukemia are of little value, because in JMML therapy may result in divergent responses in solid organs compared to peripheral blood (PB). We therefore defined separate response criteria for white blood count (WBC), platelet count, liver size, and spleen size. We then retrospectively evaluated the efficacy of 129 treatment courses other than HSCT administered to 63 children with JMML. Treatment consisted of intensive therapy according to AML-type chemotherapy, maintenance-type combination therapy, and single agent therapy. To account for the variability observed in the natural course of disease, we also evaluated 32 episodes of "no therapy." Best responses within 3 months of initiation of therapy were highly variable for the four response criteria. In contrast to platelet count and liver size, there was a significant correlation between WBC or spleen size and therapy. Response rates for WBC and spleen size were best for purine analogs, etoposide, and cytarabine as single agents or for maintenance-type combination therapy. To rigorously test future therapeutic strategies in this rare disease an international consensus on the definition of response criteria will be helpful.

Novel regimen for the treatment of juvenile myelomonocytic leukemia (JMML)

The effective treatment for juvenile myelomonocytic leukemia (JMML) patients lacking access to stem cell transplantation remains unavailable. Here, we describe a promising result obtained with novel regimen comprised of a combination of chemotherapy and differentiation therapy. Five patients diagnosed as JMML were treated with a standard regimen (cytosine arabinoside (Ara-C) 100 mg/m 2 per day continuous infusion (days 0–6), etoposide 100 mg/m 2 per day (days 0–4), vincristine 1.5 mg/m 2 per d (day 9) and isotretinoin 75–100 mg/m 2 per day (days 10–20)). All patients responded to the standard regimen. Three of the five were later treated with salvage a regimen (Ara-C 100 mg/m 2 per day continuous infusion (days 0–4), etoposide 100 mg/m 2 per day (days 0–4) and Ara-C 15 mg/m 2 per day SC (days 6–15)) when immature myeloid cells reappeared in the peripheral blood, the spleen increased or blast crisis occurred. Immature myeloid cells disappeared again after one cycle of salvage regimen in all patients. All patients are alive now with a median follow up duration of 27 months (8–69 months). Although the number of patients enrolled was limited, the standard and salvage regimens were found to be safe and effective alternatives for JMML patients without a matched donor. These regimens also could be used safely before stem cell transplantation.