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Background: Biliary rhabdomyosarcoma (RMS) is the most common biliary tumor in children. The management of affected patients contains unique challenges because of the rarity of this tumor entity and its critical location at the porta hepatis, which can make achievement of a radical resection very difficult.
Methods: In a retrospective chart analysis we analysed children suffering from biliary RMS who were registered in three different CWS trials (CWS-96, CWS-2002P, and SoTiSaR registry).
Results: Seventeen patients (12 female, 5 male) with a median age of 4.3 years were assessed. The median follow-up was 42.2 months (10.7–202.5). The 5-year overall (OS) and event free survival (EFS) rates were 58% (45–71) and 47% (34–50), respectively. Patients > 10 years of age and those with alveolar histology had the worst prognosis (OS 0%). Patients with botryoid histology had an excellent survival (OS 100%) compared to those with non-botryoid histology (OS 38%, 22–54, p = 0.047). Microscopic complete tumor resection was achieved in almost all patients who received initial tumor biopsy followed by chemotherapy and delayed surgery.
Conclusion: Positive predictive factors for survival of children with biliary RMS are age ≤ 10 years and botryoid tumor histology. Primary surgery with intention of tumor resection should be avoided.
Neuroblastoma (NB) is the most common solid extracranial tumor in childhood. Despite therapeutic progress, prognosis in high-risk NB is poor and innovative therapies are urgently needed. Therefore, we addressed the potential cytotoxic capacity of interleukin (IL)-activated natural killer (NK) cells compared to cytokine-induced killer (CIK) cells for the treatment of NB. NK cells were isolated from peripheral blood mononuclear cells (PBMCs) by indirect CD56-enrichment or CD3/CD19-depletion and expanded with different cytokine combinations, such as IL-2, IL-15, and/or IL-21 under feeder-cell free conditions. CIK cells were generated from PBMCs by ex vivo stimulation with interferon-γ, IL-2, OKT-3, and IL-15. Comparative analysis of expansion rate, purity, phenotype and cytotoxicity was performed. CD56-enriched NK cells showed a median expansion rate of 4.3-fold with up to 99% NK cell content. The cell product after CD3/CD19-depletion consisted of a median 43.5% NK cells that expanded significantly faster reaching also 99% of NK cell purity. After 10–12 days of expansion, both NK cell preparations showed a significantly higher median cytotoxic capacity against NB cells relative to CIK cells. Remarkably, these NK cells were also capable of efficiently killing NB spheroidal 3D culture in long-term cytotoxicity assays. Further optimization using a novel NK cell culture medium and a prolonged culturing procedure after CD3/CD19-depletion for up to 15 days enhanced the expansion rate up to 24.4-fold by maintaining the cytotoxic potential. Addition of an IL-21 boost prior to harvesting significantly increased the cytotoxicity. The final cell product consisted for the major part of CD16−, NCR-expressing, poly-functional NK cells with regard to cytokine production, CD107a degranulation and antitumor capacity. In summary, our study revealed that NK cells have a significantly higher cytotoxic potential to combat NB than CIK cell products, especially following the synergistic use of IL-15 and IL-21 for NK cell activation. Therefore, the use of IL-15+IL-21 expanded NK cells generated from CD3/CD19-depleted apheresis products seems to be highly promising as an immunotherapy in combination with haploidentical stem cell transplantation (SCT) for high-risk NB patients.
Rhabdomyosarcoma (RMS), the most common cancer of connective tissues in pediatrics, is often resistant to conventional therapies. One underlying mechanism of this resistance is the overexpression of Inhibitor of Apoptosis (IAP) proteins, leading to a dysfunctional cell death program within tumor cells. Smac mimetics (SM) are small molecules that can reactivate the cell death program by antagonizing IAP proteins and thereby compensating their overexpression. Here, we report that SM sensitize two RMS cell lines (RD and RH30) toward natural killer (NK) cell-mediated killing on the one hand, and increase the cytotoxic potential of NK cells on the other. The SM-induced sensitization of RH30 cells toward NK cell-mediated killing is significantly reduced through blocking tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) on NK cells prior to coculture. In addition, the presence of zVAD.fmk, a pancaspase inhibitor, rescues tumor cells from the increase in killing, indicating an apoptosis-dependent cell death. On the NK cell side, the presence of SM in addition to IL-2 during the ex vivo expansion leads to an increase in their cytotoxic activity against RH30 cells. This effect is mainly TNFα-dependent and partially mediated by NK cell activation, which is associated with transcriptional upregulation of NF-κB target genes such as IκBα and RelB. Taken together, our findings implicate that SM represent a novel double-hit strategy, sensitizing tumor and activating NK cells with one single drug.
As the biology of mesenchymal stromal cells (MSCs) in patients with non-malignant hematological diseases (NMHD) is poorly understood, in the current study we performed a basic characterization of the phenotype and functional activity of NMHD-MSCs. Bone marrow (BM) of patients with thalassemia major (TM) possessed a significantly higher number of nucleated cells (BM-MNCs)/mL BM than healthy donors (P < 0.0001), which however did not result in a higher number of colony forming units-fibroblast (CFU-F) per milliliter BM. In contrast, from 1 × 106 BM-MNCs of patients with sickle cell disease (SCD) were generated significantly more CFU-Fs than from TM-BM-MNCs (P < 0.013) and control group (P < 0.02). In addition, NMHD-MSCs expressed significantly lower levels of CD146 molecule, demonstrated an equal proliferation potential and differentiated along three lineages (osteoblasts, chondrocytes and adipocytes) as healthy donors’ MSCs, with exception of TM-MSCs which differentiated weakly in adipocytes. In contrast to other NMHD-MSCs and healthy donors’ MSCs, TM-MSCs demonstrated an impaired in vitro immunosuppressive potential, either. Noteworthy, the majority of the immunosuppressive effect of NMHD-MSCs was mediated through prostaglandin-E2 (PGE2), because indomethacin (an inhibitor of PGE2 synthesis) was able to significantly reverse this effect. Our results indicate therefore that NMHD-MSCs, except TM-MSCs, may be used as an autologous cell-based therapy for post-transplant complications such as graft failure, graft-versus-host disease (GvHD) and osteonecrosis.
Chromosomal rearrangements of the human MLL (mixed lineage leukemia) gene are associated with high-risk infant, pediatric, adult and therapy-induced acute leukemias. We used long-distance inverse-polymerase chain reaction to characterize the chromosomal rearrangement of individual acute leukemia patients. We present data of the molecular characterization of 1590 MLL-rearranged biopsy samples obtained from acute leukemia patients. The precise localization of genomic breakpoints within the MLL gene and the involved translocation partner genes (TPGs) were determined and novel TPGs identified. All patients were classified according to their gender (852 females and 745 males), age at diagnosis (558 infant, 416 pediatric and 616 adult leukemia patients) and other clinical criteria. Combined data of our study and recently published data revealed a total of 121 different MLL rearrangements, of which 79 TPGs are now characterized at the molecular level. However, only seven rearrangements seem to be predominantly associated with illegitimate recombinations of the MLL gene (~ 90%): AFF1/AF4, MLLT3/AF9, MLLT1/ENL, MLLT10/AF10, ELL, partial tandem duplications (MLL PTDs) and MLLT4/AF6, respectively. The MLL breakpoint distributions for all clinical relevant subtypes (gender, disease type, age at diagnosis, reciprocal, complex and therapy-induced translocations) are presented. Finally, we present the extending network of reciprocal MLL fusions deriving from complex rearrangements.
Ataxia-telangiectasia (A-T) is a hereditary immune system disorder with neurodegeneration. Its first neurologic symptoms include ataxic gait in early childhood, with slowly progressive cerebellar ataxia, oculomotor apraxia, oculocutaneous telangiectasia, and progressive muscle weakness. Neonatal screening for severe T-cell deficiency was recently found to diagnose A-T patients with a significantly reduced naïve T-cell pool. Our study includes 69 A-T patients between 8 January 2002 and 1 December 2019. Nineteen cases of cancer were diagnosed in 17 patients (25%), with a median overall survival [OS; 95% cumulative indcidence (CI)] of 26·9 years for the entire cohort. The 15-year OS of 82·5% (72–95%) was significantly decreased among A-T patients with malignancies, who had a median OS of 2·11 years, with a two-year-estimated OS of 50·7% (31–82%). Haematological malignancies were the major causes of death within the initial years of life with a 15 times increased risk for death [HR (95% CI): 6·9 (3·1–15.2), P < 0·001] upon malignancy diagnosis. Male patients with A-T are at a higher cancer risk than their female counterparts. This manuscript highlights the need for cancer surveillance and prevention, as well as optimal treatment in this cohort.
Background: The benefit of adjuvant therapy in synovial sarcoma (SS) treatment is under debate. Long-term follow-up data are missing.
Methods: SS patients treated in the consecutive trials CWS-81, CWS-86, CWS-91, CWS-96, CWS-2002-P, and the SoTiSaR-registry till 2013 were analyzed.
Results: Median age of 185 patients was 13.9 years (0.1–56)—with median follow-up of 7.4 years for 163 survivors. Most tumors (76%) were located in extremities. Size was < 3 cm in 58 (31%), 3–5 cm in 59 (32%), 5–10 cm in 42 (23%), and > 10 cm in 13 (7%) (13 missing). In 84 (45%) tumors, first excision was complete (R0 corresponding to IRS-I-group) and in 101 (55%) marginal (R1 corresponding to IRS-II-group). In a subsequent surgical intervention during chemotherapy, R0-status was accomplished in 23 additional IRS-II-group patients with secondary surgery. Radiotherapy was administered to 135 (73%), thereof 62 with R0-status and 67 R1-status (6 missing information). Adjuvant chemotherapy was administered to all but six patients. 5-year event-free (EFS) and overall survival (OS) was 82.9% ± 5.7 (95%CI) and 92.5% ± 3.9. Local and metastatic relapse-free survival was 91.3% ± 4.3 and 92.3% ± 4.1 at 5 years, respectively. In the multivariate analysis, tumor size and no chemotherapy were independently associated with EFS. Size and site were associated with OS. In a detailed analysis of local and metastatic events, tumor size was associated with an independent risk for developing metastases. No independent factor for suffering local recurrence could be identified.
Discussion: Omission of chemotherapy in a non-stratified way seems not justified. Size governs survival due to high linear association with risk of suffering metastatic recurrence in a granular classification.
Heute erkranken in Deutschland etwa 750 Kinder und Jugendliche Jahr für Jahr an Leukämien und anderen bösartigen Bluterkrankungen, davon rund 600 an einer Akuten Lymphatischen Leukämie (ALL) und 150 an einer Akuten Myeloischen Leukämie (AML). Seit der Einführung moderner Chemotherapieprotokolle im Jahr 1970 ist aus dieser einst tödlichen eine jetzt heilbare Erkrankung geworden. Allerdings erleiden immer noch zirka 20 Prozent der Kinder mit einer ALL einen Rückfall. Die meisten der Kinder benötigen eine Stammzelltransplantation. In Deutschland sind dies pro Jahr etwa 100; in Frankfurt werden im Jahr gegenwärtig 20 Patienten mit Stammzelltransplantationen behandelt, davon rund 55 Prozent mit akuten Leukämien. Eine Stammzell-transplantation kostet bei Kindern gegenwärtig etwa 120000 Euro.
Second mitochondria-derived activator of caspase (Smac) mimetics are considered as promising anticancer therapeutics that are currently under investigation in early clinical trials. They induce apoptosis by antagonizing inhibitor of apoptosis proteins, which are frequently overexpressed in cancer. We previously reported that Smac mimetics, such as BV6, additionally exert non-apoptotic functions in glioblastoma (GBM) cells by stimulating migration and invasion in a nuclear factor kappa B (NF-κB)-dependent manner. Because NF-κB target genes mediating these effects are largely unknown, we performed whole-genome expression analyses. Here, we identify chemokine (C-C motif) ligand 2 (CCL2) as the top-listed NF-κB-regulated gene being upregulated upon BV6 treatment in GBM cells. BV6-induced upregulation and secretion of CCL2 are required for migration and invasion of GBM cells because knockdown of CCL2 in GBM cells abolishes these effects. Co-culture experiments of GBM cells with non-malignant astroglial cells reveal that BV6-stimulated secretion of CCL2 by GBM cells into the supernatant triggers migration of astroglial cells toward GBM cells because CCL2 knockdown in BV6-treated GBM cells impedes BV6-stimulated migration of astroglial cells. In conclusion, we identify CCL2 as a BV6-induced NF-κB target gene that triggers migration and invasion of GBM cells and exerts paracrine effects on the GBM's microenvironment by stimulating migration of astroglial cells. These findings provide novel insights into the biological functions of Smac mimetics with important implications for the development of Smac mimetics as cancer therapeutics.
Background: Reconstitution of cytomegalovirus-specific CD3+CD8+ T cells (CMV-CTLs) after allogeneic hematopoietic stem cell transplantation (HSCT) is necessary to bring cytomegalovirus (CMV) reactivation under control. However, the parameters determining protective CMV-CTL reconstitution remain unclear to date.
Design and Methods: In a prospective tri-center study, CMV-CTL reconstitution was analyzed in the peripheral blood from 278 patients during the year following HSCT using 7 commercially available tetrameric HLA-CMV epitope complexes. All patients included could be monitored with at least CMV-specific tetramer.
Results: CMV-CTL reconstitution was detected in 198 patients (71%) after allogeneic HSCT. Most importantly, reconstitution with 1 CMV-CTL per µl blood between day +50 and day +75 post-HSCT discriminated between patients with and without CMV reactivation in the R+/D+ patient group, independent of the CMV-epitope recognized. In addition, CMV-CTLs expanded more daramtaically in patients experiencing only one CMV-reactivation than those without or those with multiple CMV reactivations. Monitoring using at least 2 tetramers was possible in 63% (n = 176) of the patients. The combinations of particular HLA molecules influenced the numbers of CMV-CTLs detected. The highest CMV-CTL count obtained for an individual tetramer also changed over time in 11% of these patients (n = 19) resulting in higher levels of HLA-B*0801 (IE-1) recognizing CMV-CTLs in 14 patients.
Conclusions: Our results indicate that 1 CMV-CTL per µl blood between day +50 to +75 marks the beginning of an immune response against CMV in the R+/D+ group. Detection of CMV-CTL expansion thereafter indicates successful resolution of the CMV reactivation. Thus, sequential monitoring of CMV-CTL reconstitution can be used to predict patients at risk for recurrent CMV reactivation.