[Frequency and age distribution of combined sternovertebral injuries : Analysis of routine data from German hospitals 2005-2012]. / Häufigkeit und Altersverteilung sternovertebraler Kombinationsverletzungen : Analyse aus Routinedaten deutscher Krankenhäuser 2005­2012.

BACKGROUND: Combinations of sternal and spinal fractures often occur due to high velocity accidents and are associated with a high incidence of concomitant injuries. The anterior thoracic wall is described as the fourth column of torso stability, which is why sternovertebral injuries (SVI) present a high risk of sagittal deformation of the trunk, in particular injuries of the thoracic spine. To date, no studies have been published on the frequency distribution of the involved vertebral bodies in large patient groups. OBJECTIVES: This study was intended to elaborate a frequency distribution of vertebral fractures accompanying sternal fractures (SF) and examine the risk of a vertebral fracture accompanying a SF. MATERIAL AND METHODS: A total of 48,193 cases with the main or secondary diagnosis of a SF and 897,963 cases with vertebral fractures based on routine data of German hospitals from the years 2005-2012 were evaluated. A concomitant injury to the spinal column was examined for each vertebral body and then evaluated statistically. RESULTS AND CONCLUSIONS: Of all patients with a SF 30.96% also suffered from a vertebral fracture. Of these 3.11% were SF as the main diagnosis and 60.89% the secondary diagnosis. While vertebral fractures generally occurred most frequently in the region of the thoracolumbar transition and the second cervical vertebral body, the SVI showed a further frequency peak in the range from the lower cervical spine to the middle thoracic spine. The present study was able to show a frequency distribution of accompanying vertebral body injuries in a large and representative collective in the case of SF for the first time.

[Bony injuries of the thoracic cage in multiple trauma : Incidence, concomitant injuries, course and outcome]. / Knöcherne Verletzung der Brustwand beim Polytrauma : Inzidenz, Begleitverletzungen, Verlauf und Outcome.

BACKGROUND: Thoracic trauma is considered to be responsible for 25 % of fatalities in multiple trauma and is a frequent injury with an incidence of 50 %. In addition to organ injuries, severe injuries to the bony parts of the thorax also occur and these injuries are described very differently mostly based on single center data. OBJECTIVES: The focus of this study was on a holistic presentation of the prevalence and the incidence of thoracic trauma in patients with multiple trauma from the data of the large collective of the TraumaRegister DGU® (TR-DGU) with the objective of an analysis of concomitant injuries, therapy options and outcome parameters. MATERIAL AND METHODS: A retrospective analysis was carried out based on the data set of the TR-DGU from the years 2009-2013. Inclusion criteria were an injury severity scale (ISS) score ≥ 16 and primary admission to a trauma center but isolated craniocerebral injury was an exclusion criterium. Patients were separated into two groups: those with rib fractures (RF) and those with flail chest (FC). RESULTS: A total of 21,741 patients met the inclusion criteria including 10,474 (48.2 %) suffering from either RF or FC. The mean age was 49.8 ± 19.9 years in the RF group and 54.1 ± 18.2 years in the FC group. Approximately 25 % were female in both groups, 98.1 % were blunt force injuries and the median ISS was 28.0 ± 11.2 in RF and 35.1 ± 14.2 in FC. Shock, insertion of a chest tube, (multi) organ failure and fatality rates were significantly higher in the FC group as were concomitant thoracic injuries, such as pneumothorax and hemothorax. Sternal fractures without rib fractures were less common (3.8 %) than concomitant in the RF (10.1 %) and FC (14 %) groups, as were concomitant fractures of the clavicle and the scapula. Out of all patients 32.6 % showed fractures of the thoracolumbar spine, 26.5 % without rib fractures, 36.6-38.6 % with rib fractures or monolateral FC and 48.6 % concomitant to bilateral FC. Thoracotomy was carried out only in isolated cases in RF and in 10.2 % of the FC group. Operative stabilization of the thoracic cage was carried out in 3.9-9.1 % of patients in the RF group and in 17.9-23.9 % in the FC group.

Dye efflux studies suggest that hematopoietic stem cells expressing low or undetectable levels of CD34 antigen exist in multiple species.

We previously described a method for isolating murine hematopoietic stem cells capable of reconstituting lethally irradiated recipients, which depends solely on dual-wavelength flow cytometric analysis of murine bone marrow cells stained with the fluorescent DNA-binding dye Hoechst 33342. This method, which appears to rely on the differential ability of stem cells to efflux the Hoechst dye, defines an extremely small and homogeneous population of cells (termed SP cells). We show here that dual-wavelength analysis of Hoechst dye-stained human, rhesus and miniature swine bone marrow cells reveals a small, distinct population of cells that efflux the dye in a manner identical to murine SP cells. Like the murine SP cells, both human and rhesus SP cells are primarily CD34-negative and lineage marker-negative. In vitro culture studies demonstrated that rhesus SP cells are highly enriched for long-term culture-initiating cells (LTC-ICs), an indicator of primitive hematopoietic cells, and have the capacity for differentiation into T cells. Although rhesus SP cells do not initially possess any hematopoietic colony-forming capability, they acquire the ability to form colonies after long-term culture on bone marrow stroma, coincident with their conversion to a CD34-positive phenotype. These studies suggest the existence of a hitherto unrecognized population of hematopoietic stem cells that lack the CD34 surface marker classically associated with primitive hematopoietic cells.

Molecular mechanisms that control expression of the B lymphocyte antigen receptor complex.

The B cell receptor for antigen (BCR) is a complex of membrane immunoglobulin (mIg) and at least two other proteins, Ig alpha (mb-1) and Ig beta (B29). This complex promotes surface expression of the BCR and acts to transduce an activation signal. We have used a system of mu heavy chain constructs transfected into murine B cell lines to probe structure-function relationships in the BCR complex. One mutant mu chain, in which two polar transmembrane residues (Tyr587, Ser588) are replaced with valine, fails to associate with Ig alpha and Ig beta and is incapable of transducing signals as a result of mIg cross-linking. This mutant is expressed on the surface at high levels when transfected into a plasmacytoma line that lacks Ig alpha, whereas wild-type mu is retained in this cell line in the endoplasmic reticulum. Pulse-chase and immunoprecipitation analyses indicate that the mutant is more rapidly released from calnexin than the wild-type mu. Further, transfection of Ig alpha into this Ig alpha-negative cell line allows release of the mu chain from calnexin and surface expression of the BCR. These results identify the transmembrane residues of mu heavy chain that control binding to calnexin and Ig alpha, and suggest that calnexin-dependent intracellular retention is an important control mechanism for expression of the BCR complex.

Intracellular targeting of antigens internalized by membrane immunoglobulin in B lymphocytes.

An important function of membrane immunoglobulin (mIg), the B cell antigen receptor, is to endocytose limiting quantities of antigen for efficient presentation to class II-restricted T cells. We have used a panel of mIg mutants to analyze the mechanism of mIg-mediated antigen presentation, and specifically to explore the ability of mIg to target internalized antigen to intracellular processing compartments. Transfected mIgs carrying substitutions for the transmembrane Tyr587 residue fail to efficiently present specifically bound antigen. However, these mutants internalize antigen normally, and their defect cannot be attributed to a lack of mIg-associated Ig alpha/Ig beta molecules. A novel functional assay for detecting antigenic peptides in subcellular fractions shows that wild-type mIg transfectants generate class II-peptide complexes intracellularly, whereas only free antigenic peptides are detectable in the mutant mIg transfectants. Furthermore, an antigen competition assay reveals that antigen internalized by the mutant mIgs fails to enter the intracellular processing compartment accessed by wild-type mIg. Therefore, mIg specifically targets bound and endocytosed antigen to the intracellular compartment where processed peptides associate with class II molecules, and the transmembrane Tyr587 residue plays an obligatory role in this process. Targeting of internalized antigen may be mediated by receptor-associated chaperones, and may be a general mechanism for optimizing the presentation of specifically bound and endocytosed antigens in b lymphocytes and other antigen-presenting cells.

'Off-the-shelf' allogeneic CAR T cells: development and challenges.

Autologous chimeric antigen receptor (CAR) T cells have changed the therapeutic landscape in haematological malignancies. Nevertheless, the use of allogeneic CAR T cells from donors has many potential advantages over autologous approaches, such as the immediate availability of cryopreserved batches for patient treatment, possible standardization of the CAR-T cell product, time for multiple cell modifications, redosing or combination of CAR T cells directed against different targets, and decreased cost using an industrialized process. However, allogeneic CAR T cells may cause life-threatening graft-versus-host disease and may be rapidly eliminated by the host immune system. The development of next-generation allogeneic CAR T cells to address these issues is an active area of research. In this Review, we analyse the different sources of T cells for optimal allogeneic CAR-T cell therapy and describe the different technological approaches, mainly based on gene editing, to produce allogeneic CAR T cells with limited potential for graft-versus-host disease. These improved allogeneic CAR-T cell products will pave the way for further breakthroughs in the treatment of cancer.

Inhibition of lymphocyte proliferation by detergent-solubilized mouse liver membranes.

The role of phenomena analogous to fibroblast contact inhibition in lymphocyte growth regulation is controversial, although it is clear that direct cell-cell contact is vital to immunoregulation and accessory cell function. An extract of mouse liver plasma membrane proteins, referred to as suppressive liver extract (SLE), that suppresses the growth of 3T3 fibroblasts also inhibited the mitogen-induced proliferation of murine lymphocytes. A dose of 20 micrograms/ml SLE was less than 95% suppressive of proliferation in both mouse T and mouse B cells treated with a variety of mitogens. B cell growth factor, while increasing DNA synthesis overall in mitogen-stimulated B cells, did not change the extent of SLE suppression, which suggests that the SLE does not interfere with lymphocyte-growth factor interactions. In exploring a sequence of B cell activation events, we discovered that SLE had no effect on the early activation event of increased phosphatidylinositol turnover. Blastogenesis, however, was inhibited in mitogen-stimulated, SLE-treated B cells. The maximum suppressive effect was observed if the SLE was added within 8-12 h of the mitogenic stimulus. SLE did not affect the viability of cells in culture. These results point to a possible unity of regulatory mechanisms between contact inhibition in fibroblasts and the processes of immunoregulation.

Stem cell transplantation for neuroblastoma.

High-risk neuroblastoma is a childhood malignancy with a poor prognosis. Gradual improvements in survival have correlated with therapeutic intensity, and the ability to harvest, process and store autologous hematopoietic stem cells has allowed for dose intensification beyond marrow tolerance. The use of high-dose chemotherapy with autologous hematopoietic stem cell rescue in consolidation has resulted in improvements in survival, although further advances are still needed. Newer approaches to SCT and supportive care, most notably the transition to PBSC, have resulted in further improvement in survival and decreases in treatment-related mortality. Research into experimental approaches to hematopoietic SCT is ongoing.

Correction to: Grading of cytokine release syndrome associated with the CAR T cell therapy tisagenlecleucel.

The original article contains several small errors. The errors & concurrent corrections are listed below [1].

Generation of a multi-antigen-directed immune response for durable control of acute lymphoblastic leukemia.

This corrects the article DOI: 10.1038/leu.2017.290.

Stabilization of flail chest injuries: minimized approach techniques to treat the core of instability.

PURPOSE: Stabilizing techniques of flail chest injuries usually need wide approaches to the chest wall. Three main regions need to be considered when stabilizing the rib cage: median-anterior with dissection of pectoral muscle; lateral-axillary with dissection of musculi (mm) serratus, externus abdominis; posterior inter spinoscapular with division of mm rhomboidei, trapezius and latissimus dorsi. Severe morbidity due to these invasive approaches needs to be considered. This study discusses possibilities for minimized approaches to the shown regions. METHOD: Fifteen patients were stabilized by locked plate osteosynthesis (MatrixRib®) between May 2012 and April 2014 and prospectively followed up. Flail chest injuries were managed through limited incisions to the anterior, the lateral, and the posterior parts of the chest wall or their combinations. Each approach was 4-10 cm using Alexis® retractor. RESULTS: One minimized approach offered sufficient access at least to four ribs posterior and laterally, four pairs of ribs anterior in all cases. There was no need to divide latissimus dorsi muscle. Trapezius und rhomboid muscles were only limited divided, whereas a subcutaneous dissection of serratus and abdominis muscles was necessary. A follow-up showed sufficient consolidation. COMPLICATIONS: pneumothorax (2) and seroma (2). CONCLUSION: Minimized approaches allow sufficient stabilization of severe dislocated rib fractures without extensive dissection or division of the important muscles. Keeping the arm and, thus, the scapula mobile is very important for providing the largest reachable surface of the rib cage through each approach.

Unrelated donor or partially matched related donor peripheral stem cell transplant with CD34+ selection and CD3+ addback for pediatric patients with leukemias.

Unmodified peripheral stem cell transplants are associated with an increased risk of extensive chronic GVHD. T depletion may reduce this risk, but the risk of graft failure or relapse may increase. To decrease the risks of both extensive chronic GVHD and graft failure, we added back a defined dose of CD3+ cells to CD34+ selected PSCs. Twenty-four patients were evaluable for outcome analysis. Donors were unrelated (23) or related (1). Conditioning was thiotepa, cyclophosphamide, and total body irradiation. Cyclosporine was used post transplant. Following CD34+ selection, a total of 5 x 10(5)/kg CD3+ cells were infused. Donors were matched for 12 patients. The median CD34+ dose infused was 7.1 x 10(6)/kg. Engraftment occurred in all patients at a median of 14 days (10-19). Twelve patients are alive in remission 15-34 months (median, 25) post PSCT. GVHD occurred in 17 patients, but was >grade II in only 2. Chronic GVHD occurred in 61.5% of evaluable patients, but was limited to skin and perioral cavity. Two patients relapsed, and 10 patients died of non-relapse causes. This study demonstrates that PSCT with CD34+ selection and a defined dose of CD3+ results in prompt engraftment and may limit development of extensive chronic GVHD.

Toxicities of busulfan/melphalan versus carboplatin/etoposide/melphalan for high-dose chemotherapy with stem cell rescue for high-risk neuroblastoma.

The optimal autologous stem cell rescue (HDC-SCR) regimen for children with high-risk neuroblastoma (HR-NBL) is not defined. Carboplatin/etoposide/melphalan (CEM) is the current US standard; however, European data suggest busulfan/melphalan (Bu/Mel) may have less toxicity. Published data regarding toxicities associated with CEM and Bu/Mel are limited. We conducted a single-institution retrospective cohort study of children with HR-NBL who received CEM or Bu/Mel preparative regimens. Toxicity data were analyzed using χ(2) or Fisher's exact, Wilcoxon two-sample or log-rank tests. Sinusoidal obstruction syndrome (SOS) was observed in 7/44 CEM (15.9%) and 5/21 (24%) Bu/Mel patients (P=0.50). Median time to SOS was longer following Bu/Mel than CEM (20 versus 9 days, P=0.02). Pulmonary hypertension (PHTN) was observed in ~20% of children after Bu/Mel and none after CEM (P=0.01). CEM patients had more nephrotoxicity (P=0.001), packed red blood cell (P=0.02) and platelet transfusions (P=0.008), and days on maximum pain support (P=0.0007). Time to engraftment, length of stay, documented infection rates and HDC-SCR-related mortality were similar. Nephrotoxicity and resource utilization associated with cytopenias and mucositis were greater after CEM. Pulmonary toxicities were more severe after Bu/Mel, and increased vigilance for PHTN may be warranted, particularly in children with hypoxemia out of proportion to respiratory distress.

Improved anti-leukemia activities of adoptively transferred T cells expressing bispecific T-cell engager in mice.

Despite the impressive clinical efficacy of T cells engineered to express chimeric antigen receptors (CAR-Ts), the current applications of CAR-T cell therapy are limited by major treatment-related toxicity. Thus, safer yet effective alternative approaches must be developed. In this study, we compared CD19 bispecific T-cell engager (BiTE)-transferred T cells that had been transfected by RNA electroporation with CD19 CAR RNA-transferred T cells both in vitro and in an aggressive Nalm6 leukemia mouse model. BiTEs were secreted from the transferred T cells and enabled both the transferred and bystander T cells to specifically recognize CD19(+) cell lines, with increased tumor killing ability, prolonged functional persistence, increased cytokine production and potent proliferation compared with the CAR-T cells. More interestingly, in comparison with CD3/CD28 bead-stimulated T cells, T cells that were expanded by a rapid T-cell expansion protocol (REP) showed enhanced anti-tumor activities for both CAR and BiTE RNA-electroporated T cells both in vitro and in a Nalm6 mouse model (P<0.01). Furthermore, the REP T cells with BiTE RNAs showed greater efficacy in the Nalm6 leukemia model compared with REP T cells with CAR RNA (P<0.05) and resulted in complete leukemia remission.

Tandem high-dose therapy in rapid sequence for children with high-risk neuroblastoma.

PURPOSE: Advances in chemotherapy and supportive care have slowly improved survival rates for patients with high-risk neuroblastoma. The focus of many of these chemotherapeutic advances has been dose intensification. In this phase II trial involving children with advanced neuroblastoma, we used a program of induction chemotherapy followed by tandem high-dose, myeloablative treatments (high-dose therapy) with stem-cell rescue (HDT/SCR) in rapid sequence. PATIENTS AND METHODS: Patients underwent induction chemotherapy during which peripheral-blood stem and progenitor cells were collected and local control measures undertaken. Patients then received tandem courses of HDT/SCR, 4 to 6 weeks apart. Thirty-nine patients (age 1 to 12 years) were assessable, and 70 cycles of HDT/SCR were completed. RESULTS: Pheresis was possible in the case of all patients, despite their young ages, with an average of 7.2 x 10(6) CD34(+) cells/kg available to support each cycle. Engraftment was rapid; median time to neutrophil engraftment was 11 days. Four patients who completed the first HDT course did not complete the second, and there were three deaths due to toxicity. With a median follow-up of 22 months (from diagnosis), 26 of 39 patients remained event-free. The 3-year event-free survival rate for these patients was 58%. CONCLUSION: A tandem HDT/SCR regimen for high-risk neuroblastoma is a feasible treatment strategy for children and may improve disease-free survival.

Outcomes of transplantation with partial T-cell depletion of matched or mismatched unrelated or partially matched related donor bone marrow in children and adolescents with leukemias.

Graft-versus-host disease (GVHD) remains a major barrier to successful hematopoietic stem cell transplant for patients who lack a matched related donor. Partial T-cell depletion (TCD) of the graft may decrease the risk of severe GVHD with unrelated donors (URD) and partially matched related donors (PMRD) while retaining an antileukemic effect. We analyzed our experience using URD and PMRD for pediatric patients with leukemias from 1990 to 2001. A subgroup of 'matched' URD donor pairs was retrospectively analyzed for high-resolution class I. Partial TCD was accomplished with monoclonal antibody T10B9 or OKT3 and complement. There were 76 URD (45% matched) and 28 PMRD recipients. Event-free survival (EFS) was 38.3%, and overall survival (OS) 45.1% at 3 years. On multivariate analysis, there was no difference in survival based upon marrow source, but nonrelapse mortality was higher with the use of PMRD. Relapse occurred in 6% of ALL patients, and 22.8% of AML/MDS patients. Grades III-IV GVHD was observed in only 6.7% of patients. Partial TCD allows use of matched or mismatched URD, or PMRD with little mortality from GVHD, durable engraftment, and no increase in relapse risk.

Unrelated donor bone marrow transplantation for children with severe aplastic anemia: minimal GVHD and durable engraftment with partial T cell depletion.

Both increased graft rejection and increased graft vs host disease (GVHD) remain obstacles to success for unrelated donor (URD) BMT for patients with SAA. Partial T cell depletion (PTCD) may decrease the risk of severe GVHD, while still maintaining sufficient donor T lymphocytes to ensure engraftment. We report on 12 patients with SAA who underwent PTCD URD BMT. All patients had failed medical therapy or relapsed following initial responses, and were transfusion dependent. The median age was 6 years, and there were five males. Donors were matched for four patients, and mismatched for eight. All patients received total body irradiation with either Ara-C or thiotepa and cyclophosphamide. PTCD was accomplished using monoclonal antibody T10B9 or OKT3 and complement. All patients engrafted, with a median time of 18 days to ANC >500. Only one patient had greater than grade II acute GVHD; two patients had limited and one patient extensive chronic GVHD. Nine patients are alive and transfusion independent at a median months post BMT. Three patients died from infection or renal failure. This series suggests that an aggressive immunosuppressive conditioning regimen with PTCD results in successful engraftment and minimal GVHD in pediatric patients with SAA, even with HLA mismatched donors.

Safety and efficacy of allogeneic PBSC collection in normal pediatric donors: the pediatric blood and marrow transplant consortium experience (PBMTC) 1996-2003.

The use of peripheral blood stem cells (PBSC) for allogeneic transplants in adults has greatly increased. This trend is reflected in pediatrics, where healthy children increasingly are donating PBSC or donor lymphocyte infusion (DLI) via apheresis for use by ill siblings. There is a potential concern that the risks of PBSC collection may differ for pediatric donors. However, no large studies have assessed safety issues in this population. To address this need, we reviewed 218 (213 PBSC, five DLI) collections in 201 normal pediatric donors (8 months to 17 years, median 11.8 years) at 22 institutions in the Pediatric Blood and Marrow Transplant Consortium. Donors received a median of 4 days of growth factor, and mean collection yield was 9.1 x 10(6) CD34+ cells/kg recipient weight. Younger age, days of apheresis, and male gender predicted increased yield of CD34+ cells/kg donor weight. Growth factor-induced pain was mild and reported in less than 15% of patients. Most donors <20 kg (23/25, 92%) required PRBC priming of the apheresis machine. This experience with over 200 collections demonstrates that PBSC collection is safe in normal pediatric donors and desired CD34 cell yields are easily achieved. Younger children utilize more medical resources and children <20 kg usually require a single blood product exposure.

The angiogenesis inhibitor tnp-470 effectively inhibits human neuroblastoma xenograft growth, especially in the setting of subclinical disease.

Tumor vascularity is highly correlated with disease outcome in neuroblastoma. Thus, novel therapeutics that target the vascular endothelium are candidates for incorporation into clinical trials. We therefore examined the effect of TNP-470 on human neuroblastoma growth in mouse models reflecting both clinically evident and minimal disease. Mice were inoculated s.c. or by tail vein injection with 10(7) human neuroblastoma-derived CHP-134 cells and treated with TNP-470 (100 mg/kg/dose s.c. three times a week or by continuous infusion) or saline. Treatment was given as a single agent in established xenografts, 10 days after 450 mg/kg of cyclophosphamide, or 12 h after tumor inoculation. Tumor growth rate was markedly inhibited in mice receiving TNP-470 administered alone both s.c. and by continuous infusion with a treatment to control ratio (T:C) at day 16 of 0.3 (P < 0.001) and a T:C at day 30 of 0.4 (P = 0.029) for each dosing method, respectively. TNP-470 also significantly inhibited tumor growth when administered following cyclophosphamide (T:C at day 30 = 0.2, P < 0.001) and inhibited disease establishment when given shortly after xenograft inoculation (T:C at day 30 = 0.1, P < 0.001) or tail vein injection. TNP-470 was shown to directly inhibit angiogenesis by Matrigel assay (P =.010) and to increase the apoptotic index in treated tumors. These data show that TNP-470 is a potent inhibitor of human neuroblastoma growth rate and tumorigenicity. We speculate that TNP-470 may be a useful adjuvant therapy for high-risk neuroblastoma patients, particularly when used in settings of minimal disease status.

Angiogenesis inhibitor TNP-470 during bone marrow transplant: safety in a preclinical model.

High-dose therapy with stem cell rescue is a treatment option for patients with advanced solid tumors. Although this approach has promise for some pediatric cancers, especially neuroblastoma, it is limited by the risk of relapse posttransplant as well as concern about possible reinfused tumor cells in autologous stem cell products. Antiangiogenic agents given during and after recovery from high-dose therapy with stem cell rescue may decrease the risk of relapse. TNP-470 is an antiangiogenic agent now in clinical trials. Although it inhibits the growth of bone marrow (BM) colony-forming cells in vitro, no significant hematological toxicity has been seen in Phase I trials. To assess the feasibility of using antiangiogenic agents during the period of posttransplant hematopoietic engraftment, we have developed a model of stem cell transplant in mice. Mice were lethally irradiated and then rescued with stem cells containing a transgene expressed in the hematopoietic lineage. Mice were then treated with TNP-470 or placebo, and assessed for survival, successful engraftment, and kinetics of engraftment. Both treated and control mice demonstrated reliable multilineage engraftment as well as normal lymphoid maturation with no excess mortality in the treated group. WBCs were lower but still within the normal range at d+28 in mice treated with bolus TNP-470, but not in those treated with continuous infusion TNP-470, compared with controls. These data indicate that inhibitors of angiogenesis do not adversely impact engraftment after stem cell transplantation.