Building a translational cancer dependency map for The Cancer Genome Atlas.

Cancer dependency maps have accelerated the discovery of tumor vulnerabilities that can be exploited as drug targets when translatable to patients. The Cancer Genome Atlas (TCGA) is a compendium of 'maps' detailing the genetic, epigenetic and molecular changes that occur during the pathogenesis of cancer, yet it lacks a dependency map to translate gene essentiality in patient tumors. Here, we used machine learning to build translational dependency maps for patient tumors, which identified tumor vulnerabilities that predict drug responses and disease outcomes. A similar approach was used to map gene tolerability in healthy tissues to prioritize tumor vulnerabilities with the best therapeutic windows. A subset of patient-translatable synthetic lethalities were experimentally tested, including PAPSS1/PAPSS12 and CNOT7/CNOT78, which were validated in vitro and in vivo. Notably, PAPSS1 synthetic lethality was driven by collateral deletion of PAPSS2 with PTEN and was correlated with patient survival. Finally, the translational dependency map is provided as a web-based application for exploring tumor vulnerabilities.

ß-Catenin activity induces an RNA biosynthesis program promoting therapy resistance in T-cell acute lymphoblastic leukemia.

Understanding the molecular mechanisms that contribute to the appearance of chemotherapy resistant cell populations is necessary to improve cancer treatment. We have now investigated the role of ß-catenin/CTNNB1 in the evolution of T-cell Acute Lymphoblastic Leukemia (T-ALL) patients and its involvement in therapy resistance. We have identified a specific gene signature that is directly regulated by ß-catenin, TCF/LEF factors and ZBTB33/Kaiso in T-ALL cell lines, which is highly and significantly represented in five out of six refractory patients from a cohort of 40 children with T-ALL. By subsequent refinement of this gene signature, we found that a subset of ß-catenin target genes involved with RNA-processing function are sufficient to segregate T-ALL refractory patients in three independent cohorts. We demonstrate the implication of ß-catenin in RNA and protein synthesis in T-ALL and provide in vitro and in vivo experimental evidence that ß-catenin is crucial for the cellular response to chemotherapy, mainly in the cellular recovery phase after treatment. We propose that combination treatments involving chemotherapy plus ß-catenin inhibitors will enhance chemotherapy response and prevent disease relapse in T-ALL patients.

Cuboid fracture: An extremely rare cause of limping in a 2-year-old child.

The purpose of this case report is to raise awareness of an extremely rare cause of limping, in young children. Clinicians should have a high index of suspicion of a possible cuboid fracture when evaluating a young child who presents with limping.

Pediatric Leukemia From an Orthopedic Perspective: A Case of Acute Lymphoblastic Leukemia Initially Managed as Septic Hip With Osteomyelitis.

Acute lymphoblastic leukemia (ALL) represents the most common pediatric cancer accounting for about one-third of all malignancies in childhood. The differential diagnosis for a pediatric patient manifesting with joint pain and refusal to bear weight is wide and includes trauma, transient synovitis, septic arthritis, rheumatologic disorders, and malignancy. Overt complaints from the musculoskeletal system as the initial manifestation of ALL may present in up to 30% of cases with normal laboratory tests and without hepatosplenomegaly or lymphadenopathy, perplexing the establishment of a definite diagnosis. Herein, we report the case of a three-year-old male who presented with recurrent hip pain and fever masquerading as septic arthritis recalcitrant to intravenous (IV) antibiotics, irrigation, and debridement of the hip joint with a final diagnosis of acute lymphoblastic leukemia confirmed by bone marrow biopsy.

Effect of N-acetyl-L-cysteine on inflammation after intraperitoneal mesh placement in a potentially contaminated environment: An experimental study in the rat.

BACKGROUND: The use of prosthetic meshes in abdominal wall reconstruction is a well-established approach; however, in certain cases where a bowel resection coexists its application is disputed. Any underlying inflammatory process may augment adhesion formation which is a major postoperative complication. In this animal study, our aim was to investigate the effect of N-acetyl-l-cysteine (NAC) on adhesion formation and the expression of inflammatory markers when a mesh was used in a clean or a potentially contaminated environment. METHODS: Sixty male Wistar rats were randomly and equally allocated in 3 groups: A, B and C. Animals in all groups underwent laparotomy, a prosthetic mesh was placed and chemoprophylaxis with ciprofloxacin was administered. In groups B and C an enterectomy was also performed. NAC was injected intraperitoneally in group C. Adhesion formation, IL-1a, IL-6, TNF-a and histological data including fibrosis, neutrophils' infiltration and neovascularization were assessed. Mesh samples were sent for cultivation. RESULTS: Adhesion formation was significantly less and inflammation markers were also lower in group C compared to group B (p<0.05). Histological findings were significant for greater fibrosis, neutrophils' infiltration and neovascularization in group B compared to both group A and C. Regarding mesh cultures, more specimens were tested positive in group B (p <0.05). Outcomes between group A and C did not differ. CONCLUSION: NAC effectively ameliorated adhesion formation and inflammation in a potentially septic environment where a prosthetic mesh was placed.

Ex vivo drug screening defines novel drug sensitivity patterns for informing personalized therapy in myeloid neoplasms.

Precision medicine approaches such as ex vivo drug sensitivity screening (DSS) are appealing to inform rational drug selection in myelodysplastic syndromes (MDSs) and acute myeloid leukemia, given their marked biologic heterogeneity. We evaluated a novel, fully automated ex vivo DSS platform that uses high-throughput flow cytometry in 54 patients with newly diagnosed or treatment-refractory myeloid neoplasms to evaluate sensitivity (blast cytotoxicity and differentiation) to 74 US Food and Drug Administration-approved or investigational drugs and 36 drug combinations. After piloting the platform in 33 patients, we conducted a prospective feasibility study enrolling 21 patients refractory to hypomethylating agents (HMAs) to determine whether this assay could be performed within a clinically actionable time frame and could accurately predict clinical responses in vivo. When assayed for cytotoxicity, ex vivo drug sensitivity patterns were heterogeneous, but they defined distinct patient clusters with differential sensitivity to HMAs, anthracyclines, histone deacetylase inhibitors, and kinase inhibitors (P < .001 among clusters) and demonstrated synergy between HMAs and venetoclax (P < .01 for combinations vs single agents). In our feasibility study, ex vivo DSS results were available at a median of 15 days after bone marrow biopsy, and they informed personalized therapy, which frequently included venetoclax combinations, kinase inhibitors, differentiative agents, and androgens. In 21 patients with available ex vivo and in vivo clinical response data, the DSS platform had a positive predictive value of 0.92, negative predictive value of 0.82, and overall accuracy of 0.85. These data demonstrate the utility of this approach for identifying potentially useful and often novel therapeutic drugs for patients with myeloid neoplasms refractory to standard therapies.

Impending hand compartment syndrome in an infant: is clinical evaluation enough to perform a fasciotomy?

The purpose of this case report is to raise awareness of the early diagnosis and treatment of compartment syndrome in children. Late diagnosis can lead to irreversible outcomes, including myonecrosis, neurologic injury, functional problems, and even amputation. In this age group, clinical judgment may be enough to proceed with fasciotomy.

Flexion-Type Supracondylar Humeral Fracture with Ulnar Nerve Injury in Children: Two Case Reports and Review of the Literature.

BACKGROUND Supracondylar humeral fracture is a common fracture in the pediatric population. Although extension-type is the most common fracture pattern (97% to 98%), flexion-type supracondylar fractures are rarely encountered (2% to 3%). The combination of a flexion-type supracondylar humeral fracture with an ulnar nerve injury represents a real challenge for an orthopaedic surgeon. CASE REPORT We report 2 cases of flexion-type supracondylar humeral fracture with ulnar nerve injury that open reduction and fixation was necessary because closed reduction could not achieve an acceptable result. An anterior approach to the elbow joint was chosen to explore whether any neurovascular structures were entrapped between the fragments. The ulnar nerve was not found to be compressed in the fracture site. After anatomic reduction, cross K-wire fixation of the fracture was performed. At 6-month follow-up, ulnar nerve injuries (in both patients) were resolved. CONCLUSIONS These case reports enhance the existing literature that flexion-type supracondylar fractures with ulnar nerve injury are associated with higher rates of open reduction. Orthopaedic surgeons should be aware, and family members of those patients should be informed, that the likelihood of an open reduction in these types of injuries is extremely high. Open reduction is needed not only to achieve an anatomic reduction of the fracture but to make sure that the ulnar nerve is not entrapped between the proximal and distal fragment.

The transcription factor NFAT5 limits infection-induced type I interferon responses.

Type I interferon (IFN-I) provides effective antiviral immunity but can exacerbate harmful inflammatory reactions and cause hematopoietic stem cell (HSC) exhaustion; therefore, IFN-I expression must be tightly controlled. While signaling mechanisms that limit IFN-I induction and function have been extensively studied, less is known about transcriptional repressors acting directly on IFN-I regulatory regions. We show that NFAT5, an activator of macrophage pro-inflammatory responses, represses Toll-like receptor 3 and virus-induced expression of IFN-I in macrophages and dendritic cells. Mice lacking NFAT5 exhibit increased IFN-I production and better control of viral burden upon LCMV infection but show exacerbated HSC activation under systemic poly(I:C)-induced inflammation. We identify IFNß as a primary target repressed by NFAT5, which opposes the master IFN-I inducer IRF3 by binding to an evolutionarily conserved sequence in the IFNB1 enhanceosome that overlaps a key IRF site. These findings illustrate how IFN-I responses are balanced by simultaneously opposing transcription factors.

Correlation of the Hallux Sesamoids' Orientation with Various Anatomical Parameters in Patients with Hallux Valgus Deformity.

Introduction An awareness of the anatomical parameters of the foot such as the position and orientation of the sesamoid bones can be of great value for the etiology and diagnostic approach to patients with hallux valgus (HV). The purpose of this study was to evaluate the basic anatomical features and measurements related to the characteristics of HV in cadaveric material. Materials and methods The study sample included 12 cadaveric limbs with HV and 10 cadaveric limbs without HV as a control group. We measured the HV angle (HVA), the first to second intermetatarsal angle (IMA), and the first to fifth IMAs of all samples. We also recorded the shape of the first metatarsal head and the position/orientation of the sesamoid bones. Results The mean values of the HVA (p = 0.000), the first to second IMA (p = 0.000), and the first to fifth IMA (p = 0.000) differed between the HV and non-HV group. The position of the sesamoid ossicles between the HV and the non-HV group was statistically significant (p = 0.001). While we noted the round shape of the first metatarsal head was predominant in HV samples, we found no statistically significant difference in the first metatarsal head shape between the test and control groups. Conclusion The HVA, the first to second IMA, the first to fifth IMA, and the orientation of the sesamoid ossicles differed significantly between cadaveric samples with HV and those without HV.

Selective Spleen Embolization of Splenomegaly to Improve Thrombocytopenia Facilitating Open Aortic Aneurysm Repair: A Staged Approach.

We present an 82-year-old man with a history of hairy cell leukemia, having an 11-cm abdominal aortic aneurysm, who also had severe thrombocytopenia (about 20 000 platelets/µL) and splenomegaly at presentation. The patient had unfavorable anatomy for endovascular aneurysm repair, and therefore, an open procedure was planned. To reduce risk for perioperative bleeding and optimize patient preoperative status, a staged approach was employed. Initially, several sessions of embolization of 2 splenic artery branches were performed with the intent to decrease spleen size and to increase platelet count thus decreasing the perioperative bleeding risk. Then, after successfully increasing platelet count (280 000 PLT/µL), open repair of the aneurysm was conducted. This case demonstrates that selective splenic embolization in patients with hypersplenism and subsequent thrombocytopenia who are in need for major surgery may achieve a significant rise in platelet count and optimize patient's preoperative status in order to avoid bleeding complications.

Correlation between Manchester Grading Scale and American Orthopaedic Foot and Ankle Society Score in Patients with Hallux Valgus.

OBJECTIVE: To evaluate the correlation between the Manchester Grading Scale and the American Orthopaedic Foot and Ankle Society (AOFAS) score in patients with a hallux valgus deformity. SUBJECTS AND METHODS: The study sample included 181 feet of 122 patients with hallux valgus and 424 feet of 212 individuals without hallux valgus deformity as the control group. The severity of hallux valgus, utilizing a relative nonmetric scale, the Manchester Grading Scale, and the metric AOFAS score, was determined for all individuals in the hallux valgus and control groups. SPSS version 18 (Chicago, Ill., USA) was used for data analysis. RESULTS: According to the Manchester Grading Scale, the 424 feet of the normal group were classified as 'no deformity'. In the hallux valgus group, 85 feet were classified as 'mild deformity', 67 as 'moderate deformity' and 29 as 'severe deformity'. The AOFAS total score in the control group was 99.14. In the hallux valgus group, patients with mild or moderate deformity had total scores of 86.20 and 68.19, respectively. For those with severe hallux valgus, the total score was 44.69 and the differences were statistically significant (p = 0.000). Using the Pearson correlation, strong negative correlations were found between the AOFAS score and the hallux valgus angle (HVA; r = -0.899, p = 0.000). Strong negative correlations were demonstrated between the AOFAS score and the first intermetatarsal angle (IMA) as well (r = -0.748, p = 0.000). CONCLUSIONS: The AOFAS score was negatively associated with the Manchester Grading Scale, HVA and first IMA. As the severity of hallux valgus increased, the AOFAS score seemed to decrease.

Relationship between pedographic analysis and the Manchester scale in hallux valgus.

OBJECTIVE: The aim of this study was to evaluate the correlation between the Manchester scale and foot pressure distribution in patients with hallux valgus deformity. METHODS: The study included 152 feet of 87 patients with hallux valgus and a control group of 391 feet of 241 individuals without hallux valgus deformity. The severity of hallux valgus was determined using the Manchester scale grading system. Plantar loading patterns in 10 foot areas were determined for all participants. RESULTS: According to the Manchester scale, 72% of the participants had no, 12.9% mild, 10.7% moderate and 4.4% severe deformity. The Manchester scale grade was highly correlated with both hallux valgus angle and first intermetatarsal angle (p=0.00). Significant differences between the four grades were present for mean pressure under the hallux and the first and second metatarsal heads only (p=0.00). The load distribution under these areas was higher as the hallux valgus progressed from mild to more severe. In all groups, the highest pressure was observed under the second metatarsal head. CONCLUSION: The Manchester scale was strongly associated with both the hallux valgus angle and the first intermetatarsal angle. The progression from mild to moderate and severe deformation is associated with peak pressure raise at the hallux, first and second metatarsal heads. The Manchester scale appears to be a useful tool to provide information for the degree of deformity and the pressure under painful foot areas.

Plerixafor added to G-CSF-supported paclitaxel-ifosfamide-cisplatin salvage chemotherapy enhances mobilization of adequate numbers of hematopoietic stem cells for subsequent autografting in hard-to-mobilize patients with relapsed/refractory germ-cell tumors: a single-center experience.

An appreciable percentage of patients with relapsed/refractory germ-cell tumors (GCTs), candidates for high-dose chemotherapy (HDC) and autologous hematopoietic cell transplantation (HCT), fail to mobilize adequate hematopoietic stem cells (HSCs) numbers with granulocyte colony-stimulating factor (G-CSF)±salvage chemotherapy. Plerixafor has shown a potential to mobilize adequate CD34+HSCs numbers in this context. Here, we applied plerixafor in combination with G-CSF after salvage chemotherapy in 'poor' mobilizers with relapsed/refractory GCTs for HDC+HCT. Patients with relapsed/refractory GCTs (n=10) received salvage paclitaxel-ifosfamide-cisplatin (TIP) chemotherapy+G-CSF to mobilize adequate HSCs to support HDC, mainly with two courses of high-dose thiotepa-etoposide-carboplatin (TEC). Patients failing to achieve the minimum collection threshold of 2.0×10/kg CD34+ cells, to support at least one cycle of HDC, were administered plerixafor before the anticipated HSC collection during subsequent cycle(s). Overall, seven patients mobilized adequate CD34+ cells (>5.0×10/kg) aiming to support two cycles of HDC. Three patients did not mobilize adequate numbers of CD34+ cells after previous G-CSF plus salvage TIP, and plerixafor was added in subsequent cycle(s). This led to a collection of adequate CD34+ cells, able to support HDC with TEC (1-2 cycles). Hematopoietic engraftment for neutrophils (absolute neutrophil count>500/µl) and platelets (platelet count>20 000/µl) with plerixafor-mobilized HSCs occurred after a median of 9 and 14 days, respectively. Salvage TIP+G-CSF leads to successful HSC mobilization in patients with less heavily pretreated GCTs, whereas the addition of plerixafor to G-CSF+TIP led to mobilization of adequate HSCs that supported autografting after one to two TEC cycles.

CD41 expression marks myeloid-biased adult hematopoietic stem cells and increases with age.

The hematopoietic stem cell (HSC) compartment is heterogeneous, yet our understanding of the identities of different HSC subtypes is limited. Here we show that platelet integrin CD41 (αIIb), currently thought to only transiently mark fetal HSCs, is expressed on an adult HSC subtype that accumulates with age. CD41+ HSCs were largely quiescent and exhibited myeloerythroid and megakaryocyte gene priming, governed by Gata1, whereas CD41- HSCs were more proliferative and exhibited lymphoid gene priming. When isolated without the use of blocking antibodies, CD41+ HSCs possessed long-term repopulation capacity on serial transplantations and showed a marked myeloid bias compared with CD41- HSCs, which yielded a more lymphoid-biased progeny. CD41-knockout (KO) mice displayed multilineage hematopoietic defects coupled with decreased quiescence and survival of HSCs, suggesting that CD41 is functionally relevant for HSC maintenance and hematopoietic homeostasis. Transplantation experiments indicated that CD41-KO-associated defects are long-term transplantable, HSC-derived and, in part, mediated through the loss of platelet mass leading to decreases in HSC exposure to important platelet released cytokines, such as transforming growth factor ß1. In summary, our data provide a novel marker to identify a myeloid-biased HSC subtype that becomes prevalent with age and highlights the dogma of HSC regulation by their progeny.

Induced pluripotent stem cell-derived human platelets: one step closer to the clinic.

The era of induced pluripotent stem (iPS) cells carries with it the promise of virtually unlimited sources of autologous cells for regenerative medicine. However, efficiently differentiating iPS cells into fully functional mature cell types remains challenging. A new study reporting the formation of fully functional platelets from human iPS (hiPS) cells improves upon recent efforts to generate this enucleated cell type, which remains in high demand for therapeutic transfusions. Notably, their lack of nucleus renders platelets unable to retain the pluripotent or tumorigenic properties of iPS cells.

Hematopoietic stem cell development in the placenta.

The placenta is a highly vascularized organ that mediates fetal-maternal exchange during pregnancy and is thereby vital for the survival and growth of the developing embryo. In addition to having this well-established role in supporting pregnancy, the placenta was recently shown to function as a hematopoietic organ. The placenta is unique among other fetal hematopoietic organs, as it is capable of both generating multipotential hematopoietic cells de novo and establishing a major hematopoietic stem cell (HSC) pool in the conceptus, while protecting HSCs from premature differentiation. The mouse placenta contains two distinct vascular regions that support hematopoiesis: the large vessels in the chorionic plate where HSCs/progenitors are thought to emerge and the labyrinth vasculature where nascent HSCs/progenitors may colonize for expansion and possible functional maturation. Defining how this cytokine- and growth factor rich organ supports HSC generation, maturation and expansion may ultimately help to establish culture protocols for HSC expansion or de novo generation from pluripotent cells.

Combined endovascular/surgical management of a ruptured para-anastomotic aneurysm of the left common iliac artery.

A 75-year old man presented with signs and symptoms of acute abdomen and a clinical picture of hypovolemic shock. An emergency CT scan revealed a ruptured para-anastomotic left common iliac artery aneurysm. The patient had undergone an elective abdominal aortic aneurysm repair operation and placement of an aortoiliac bifurcated graft 10 years before. Para-anastomotic aneurysms had developed in all 3 (aortic and the 2 iliac) anastomosis. As the patient was highrisk, a combined endovascular/surgical approach was undertaken. The patient was discharged 4 days later.This article discusses the applicability of endovascular procedures in emergency settings to high-risk patients.

Mef2C is a lineage-restricted target of Scl/Tal1 and regulates megakaryopoiesis and B-cell homeostasis.

The basic helix-loop-helix transcription factor stem cell leukemia gene (Scl) is a master regulator for hematopoiesis essential for hematopoietic specification and proper differentiation of the erythroid and megakaryocyte lineages. However, the critical downstream targets of Scl remain undefined. Here, we identified a novel Scl target gene, transcription factor myocyte enhancer factor 2 C (Mef2C) from Scl(fl/fl) fetal liver progenitor cell lines. Analysis of Mef2C(-/-) embryos showed that Mef2C, in contrast to Scl, is not essential for specification into primitive or definitive hematopoietic lineages. However, adult VavCre(+)Mef2C(fl/fl) mice exhibited platelet defects similar to those observed in Scl-deficient mice. The platelet counts were reduced, whereas platelet size was increased and the platelet shape and granularity were altered. Furthermore, megakaryopoiesis was severely impaired in vitro. Chromatin immunoprecipitation microarray hybridization analysis revealed that Mef2C is directly regulated by Scl in megakaryocytic cells, but not in erythroid cells. In addition, an Scl-independent requirement for Mef2C in B-lymphoid homeostasis was observed in Mef2C-deficient mice, characterized as severe age-dependent reduction of specific B-cell progenitor populations reminiscent of premature aging. In summary, this work identifies Mef2C as an integral member of hematopoietic transcription factors with distinct upstream regulatory mechanisms and functional requirements in megakaryocyte and B-lymphoid lineages.

Isolation and analysis of hematopoietic stem cells from the placenta.

Hematopoietic stem cells (HSCs) have the ability to self-renew and generate all cell types of the blood lineages throughout the lifetime of an individual. All HSCs emerge during embryonic development, after which their pool size is maintained by self-renewing cell divisions. Identifying the anatomical origin of HSCs and the critical developmental events regulating the process of HSC development has been complicated as many anatomical sites participate during fetal hematopoiesis. Recently, we identified the placenta as a major hematopoietic organ where HSCs are generated and expanded in unique microenvironmental niches (Gekas, et al 2005, Rhodes, et al 2008). Consequently, the placenta is an important source of HSCs during their emergence and initial expansion. In this article, we show dissection techniques for the isolation of murine placenta from E10.5 and E12.5 embryos, corresponding to the developmental stages of initiation of HSCs and the peak in the size of the HSC pool in the placenta, respectively. In addition, we present an optimized protocol for enzymatic and mechanical dissociation of placental tissue into single-cell suspension for use in flow cytometry or functional assays. We have found that use of collagenase for single-cell suspension of placenta gives sufficient yields of HSCs. An important factor affecting HSC yield from the placenta is the degree of mechanical dissociation prior to, and duration of, enzymatic treatment. We also provide a protocol for the preparation of fixed-frozen placental tissue sections for the visualization of developing HSCs by immunohistochemistry in their precise cellular niches. As hematopoietic specific antigens are not preserved during preparation of paraffin embedded sections, we routinely use fixed frozen sections for localizing placental HSCs and progenitors.