FLOT Versus FLOT/Trastuzumab/Pertuzumab Perioperative Therapy of Human Epidermal Growth Factor Receptor 2-Positive Resectable Esophagogastric Adenocarcinoma: A Randomized Phase II Trial of the AIO EGA Study Group.

PURPOSE: High pathologic complete response (pCR) rates and comparably good survival data were seen in a phase II trial combining perioperative fluorouracil, leucovorin, oxaliplatin, and docetaxel (FLOT) chemotherapy with trastuzumab for resectable, esophagogastric adenocarcinoma (EGA). The current trial evaluates the addition of trastuzumab and pertuzumab to FLOT as perioperative treatment for human epidermal growth factor receptor 2-positive resectable EGA. METHODS: In this multicenter, randomized phase II/III trial, patients with human epidermal growth factor receptor 2-positive, resectable EGA (≥ clinical tumor 2 or clinical nodal-positive) were assigned to four pre- and postoperative cycles of either FLOT alone (arm A) or combined with trastuzumab and pertuzumab, followed by nine cycles of trastuzumab/pertuzumab (arm B). The primary end point for the phase II part was the rate of pCR. RESULTS: The trial was closed prematurely, without transition into phase III, after results of the JACOB trial were reported. Eighty-one patients were randomly assigned (A: 41/B: 40) during the phase II part. The pCR rate was significantly improved with the trastuzumab/pertuzumab treatment (A: 12%/B: 35%; P = .02). Similarly, the rate of pathologic lymph node negativity was higher with trastuzumab/pertuzumab (A: 39%/B: 68%), whereas the R0 resection rate (A: 90%/B: 93%) and surgical morbidity (A: 43%/B: 44%) were comparable. Moreover, the inhouse mortality was equal in both arms (overall 2.5%). The median disease-free survival was 26 months in arm A and not yet reached in arm B (hazard ratio, 0.58; P = .14). After a median follow-up of 22 months, the median overall survival was not yet reached (hazard ratio, 0.56; P = .24). Disease-free survival and overall survival rates at 24 months were 54% (95% CI, 38 to 71) and 77% (95% CI, 63 to 90) in arm A and 70% (95% CI, 55 to 85) and 84% (95% CI, 72 to 96) in arm B, respectively. More ≥ grade 3 adverse events were reported with trastuzumab/pertuzumab, especially diarrhea (A: 5%/B: 41%) and leukopenia (A: 13%/B: 23%). CONCLUSION: The addition of trastuzumab/pertuzumab to perioperative FLOT significantly improved pCR and nodal negativity rates at the price of higher rates of diarrhea and leukopenia.

Perioperative chemotherapy with fluorouracil plus leucovorin, oxaliplatin, and docetaxel versus fluorouracil or capecitabine plus cisplatin and epirubicin for locally advanced, resectable gastric or gastro-oesophageal junction adenocarcinoma (FLOT4): a randomised, phase 2/3 trial.

BACKGROUND: Docetaxel-based chemotherapy is effective in metastatic gastric and gastro-oesophageal junction adenocarcinoma. This study reports on the safety and efficacy of the docetaxel-based triplet FLOT (fluorouracil plus leucovorin, oxaliplatin and docetaxel) as a perioperative therapy for patients with locally advanced, resectable tumours. METHODS: In this controlled, open-label, phase 2/3 trial, we randomly assigned 716 patients with histologically-confirmed advanced clinical stage cT2 or higher or nodal positive stage (cN+), or both, resectable tumours, with no evidence of distant metastases, via central interactive web-based-response system, to receive either three pre-operative and three postoperative 3-week cycles of 50 mg/m2 epirubicin and 60 mg/m2 cisplatin on day 1 plus either 200 mg/m2 fluorouracil as continuous intravenous infusion or 1250 mg/m2 capecitabine orally on days 1 to 21 (ECF/ECX; control group) or four preoperative and four postoperative 2-week cycles of 50 mg/m2 docetaxel, 85 mg/m2 oxaliplatin, 200 mg/m2 leucovorin and 2600 mg/m2 fluorouracil as 24-h infusion on day 1 (FLOT; experimental group). The primary outcome of the trial was overall survival (superiority) analysed in the intention-to-treat population. This trial is registered with ClinicalTrials.gov, number NCT01216644. FINDINGS: Between Aug 8, 2010, and Feb 10, 2015, 716 patients were randomly assigned to treatment in 38 German hospitals or with practice-based oncologists. 360 patients were assigned to ECF/ECX and 356 patients to FLOT. Overall survival was increased in the FLOT group compared with the ECF/ECX group (hazard ratio [HR] 0·77; 95% confidence interval [CI; 0.63 to 0·94]; median overall survival, 50 months [38·33 to not reached] vs 35 months [27·35 to 46·26]). The number of patients with related serious adverse events (including those occurring during hospital stay for surgery) was similar in the two groups (96 [27%] in the ECF/ECX group vs 97 [27%] in the FLOT group), as was the number of toxic deaths (two [<1%] in both groups). Hospitalisation for toxicity occurred in 94 patients (26%) in the ECF/ECX group and 89 patients (25%) in the FLOT group. INTERPRETATION: In locally advanced, resectable gastric or gastro-oesophageal junction adenocarcinoma, perioperative FLOT improved overall survival compared with perioperative ECF/ECX. FUNDING: The German Cancer Aid (Deutsche Krebshilfe), Sanofi-Aventis, Chugai, and Stiftung Leben mit Krebs Foundation.

Dual-targeting triplebody 33-16-123 (SPM-2) mediates effective redirected lysis of primary blasts from patients with a broad range of AML subtypes in combination with natural killer cells.

A number of agents designed for immunotherapy of Acute Myeloid Leukemia (AML) are in preclinical and early clinical development. Most of them target a single antigen on the surface of AML cells. Here we describe the development and key biological properties of a tri-specific agent, the dual-targeting triplebody SPM-2, with binding sites for target antigens CD33 and CD123, and for CD16 to engage NK cells as cytolytic effectors. Primary blasts of nearly all AML patients carry at least one of these target antigens and the pair is particularly promising for the elimination of blasts and leukemia stem cells (LSCs) from a majority of AML patients by dual-targeting agents. The cytolytic activity of NK cells mediated by SPM-2 was analyzed in vitro for primary leukemic cells from 29 patients with a broad range of AML-subtypes. Blasts from all 29 patients, including patients with genomic alterations associated with an unfavorable genetic subtype, were lysed at nanomolar concentrations of SPM-2. Maximum susceptibility was observed for cells with a combined density of CD33 and CD123 above 10,000 copies/cell. Cell populations enriched for AML-LSCs (CD34pos and CD34pos CD38neg cells) from 2 AML patients carried an increased combined antigen density and were lysed at correspondingly lower concentrations of SPM-2 than unsorted blasts. These initial findings raise the expectation that SPM-2 may also be capable of eliminating AML-LSCs and thus of prolonging survival. In the future, patients with a broad range of AML subtypes may benefit from treatment with SPM-2.

MicroRNA-192-5p is a predictive biomarker of survival for Stage IIIB colon cancer patients.

BACKGROUND: The poor clinical prognosis of Stage IIIB colon cancer patients is due in part to the current lack of an effective diagnostic method being available and highlights a need for the identification of novel biomarkers like microRNA (miRNA). PATIENTS AND METHODS: We used microarray analysis to compare the miRNA expression profiles of eight Stage IIIB colon cancer patients with worse clinical outcome (those who developed liver metastases between 8 and 18 months after surgery) against eight 'cured' Stage IIIB colon cancer patients (those who remained disease free following surgery during the same monitoring period). In addition, quantitative real-time polymerase chain reaction (qRT-PCR) analysis was performed examining miRNAs in tumor tissue of 98 patients with Stage IIIB colon cancer. RESULTS: We found, miRNA-192-3p and miRNA-192-5p were down regulated in the patients with worsening disease compared to the control 'cured' Stage IIIB colon cancer patients (P = 0.026 and P = 0.042, respectively). Patients with higher expression of miRNA-192-5p had higher 5-year disease-free survival (DFS) (84.21%) and overall survival (OS) (89.47%) than those with lower targeted miRNA expression DFS (38.8%; hazard ratio (HR): 3.74, 95% confidence interval (CI): 1.52-9.23, P = 0.042) and OS (48.57%; HR: 5.01, 95% CI: 1.75-14.38, P = 0.033). In contrast, patients with higher expression of miRNA-192-3p did not appear to statistically impact the survival of patients in this setting (DFS 73.33% vs 64.7%, HR: 0.68, 95% CI: 0.31-1.52, P = 0.35; OS 76.67% vs 66.17%, HR: 0.62, 95% CI: 0.27-1.45, P = 0.27). CONCLUSIONS: The decreased expression of miRNA-192-5p found for patients with relapsing disease might represent a highly predictive marker to use for the prognosis of Stage IIIB colon cancer patients.

Evidence for an Association of ERCC1 Expression and Mismatch Repair Status with Overall Survival in Colorectal Cancer Patients.

BACKGROUND: The aim in this study was to determine if an association of excision repair cross-complementing group 1 (ERCC1) gene and mismatch repair (MMR) status with overall survival (OS) could be found from our analysis of a large cohort of Chinese colorectal cancer patients (CRC). METHODS: In total, 2,233 tissue samples isolated from individual CRC tumors were assessed by immunohistochemistry for the expression of ERCC1 and 4 MMR genes. RESULTS: The rates of proficient MMR (pMMR) and ERCC1 expression were 89.6 and 90.7%, respectively. We found that patients with positive ERCC1 expression and deficient (d)MMR status had higher overall survival (OS) than those with either positive ERCC1 and pMMR, negative ERCC1 and dMMR, or negative ERCC1 expression and pMMR status (OS 79 vs. 69 vs. 66 vs. 61%, hazard ratio (HR) 0.90, 95% confidence interval (CI) 0.80-1.00; p = 0.043). Despite this finding, we found no statistical difference in OS between ERCC1-positive and -negative CRC patients when ERCC1 expression was considered alone (OS 70 vs. 62%, HR 0.82, 95% CI 0.65-1.04; p = 0.11). CONCLUSION: Our results indicate that the combined examination of ERCC1 expression and dMMR status can be used to aid OS assessment in CRC patients.

Systematic immunohistochemical screening for mismatch repair and ERCC1 gene expression from colorectal cancers in China: Clinicopathological characteristics and effects on survival.

BACKGROUND: We performed a systematic screening of colorectal cancer (CRC) tissues to investigate whether mismatch repair (MMR) status and ERCC1 protein expression could be predictive of clinical outcomes for these patients following the recommendation of The Evaluation of Genomic Applications in Practice of Prevention (EGAPP). METHODS: The expression of four MMR genes and ERCC1 were assessed by immunohistochemistry (IHC) from cancer tissue samples of 2233 consecutive CRC patients. RESULTS: We observed that most CRC patients with a proficient MMR (pMMR) status tended to have simultaneous ERCC1 protein expression (P< 0.001). Stage III CRC patients with deficient MMR (dMMR) had higher prognoses than the same stage patients with pMMR (DFS: 74% vs 65%, P = 0.04; OS: 79% vs 69%, P = 0.04). Here, dMMR is also associated with poorer survival for stage II patients after chemotherapy (DFS: 66% vs 78%, P = 0.04). Stage II and III patients that were shown to express ERCC1 protein had higher DFS and OS than those that were deficient in expression (stage II, DFS: 83% vs 70%, P = 0.006; OS 85% vs 73%, P = 0.02. Stage III, DFS: 67% vs56%, P = 0.03; OS: 71% vs 57%, P = 0.04). CONCLUSIONS: Our results indicate that dMMR appeared to predictive of a survival benefit for stage III CRC patients. We also found the determination of ERCC1 expression to be useful for predicting DFS or OS for stage II and III CRC patients. In addition, the expression of MMR genes and ERCC1 showed a significant relationship.

A relationship to survival is seen by combining the factors of mismatch repair status, tumor location and age of onset in colorectal cancer patients.

BACKGROUND: The progression of colorectal cancer (CRC) may differ depending on the location of the tumor and the age of onset of the disease. Previous studies also suggested that the molecular basis of CRC varies with tumor location, which could affect the clinical management of patients. Therefore, we performed survival analysis looking at different age groups and mismatch repair status (MMR) of CRC patients according to primary tumor location in an attempt to identify subgroups of CRC that might help in the prognosis of disease. METHODS: A group of 2233 patients operated on to remove their CRC tumors were analyzed (521 with right colon cancer, 740 with left colon cancer and 972 with rectal cancer). The expression of four MMR genes was assessed by immunohistochemistry (IHC), independent of clinical criteria. From the data collected, a predictive model for overall survival (OS) could be constructed for some associations of tumor location and age of onset using Kaplan-Meier, logistic and Cox regression analysis. RESULTS: When tumor location was considered as the lone factor, we found no statistical difference in overall survival (OS) between right cancer (68%), left cancer (67%) or rectal cancer tumor locations (71%) (HR: 1.17, 95%CI (confidence interval): 0.97-1.43, P = 0.057). When age of onset was considered, middle age (40-59 years) and older (60-85 years) patients were found to have higher OS than younger onset cancer (20-39 years) patients (69% vs 71% vs 59%, HR: 1.07, 95% confidence interval (CI): 0.91-1.25, P = 0.008). When both age of onset and tumor location were considered in combination as disease factors, we found that the subgroup of patients with left colon cancer from middle age (69%) and older (67%) aged patients had higher OS than younger (54%) patients (HR: 0.89, 95%CI: 0.68-1.16, P = 0.048). However in patients with right colon cancers, we found no statistical difference is OS between younger, middle age or older grouped patients (60% vs 71% vs 67%, HR: 0.84, 95% CI: 0.61-1.16, P = 0.194). With regard to rectal located cancers, we found that younger (62%) and middle age (68) patients had lower OS than older (77%) patients (HR:1.46, 95%CI: 1.13-1.88, P = 0.004). The rates of deficient MMR (dMMR) was 10.4%. We found no statistical difference in OS stratified by tumor locations. However, right colon cancer patients with dMMR (86%) had higher OS than those with proficient MMR (pMMR) (63%) (HR: 3.01, 95% CI: 1.82-4.97, P<0.001). Left colon cancer patients with dMMR (76%) also had higher OS than those with pMMR (66%) (HR: 1.67, 95% CI: 0.95-2.92, P = 0.01). Oppositely, rectal cancer patients with dMMR (60%) had lower OS than those pMMR (68%) (HR: 0.77, 95% CI: 0.51-1.17, P = 0.04). CONCLUSIONS: These data demonstrate that primary tumor location can be an important factor when considered along with age of onset for the prognosis of CRC. Primary tumor location is also an important factor to evaluate the predictive effect of MMR status for the prognosis of CRC.

Impact of age and mismatch repair status on survival in colorectal cancer.

Previous studies have suggested that deficiencies in mismatch repair genes (dMMR) often occur in patients with colorectal cancer (CRC) and contribute to disease etiology. Here, we looked for a correlation of MMR status to disease outcomes from a large number of Chinese CRC patients stratified by the age of onset of disease. A total of 2233 CRC patients were analyzed and tissue biopsies of surgically removed tumors scored for MMR gene status. The patient distribution after classification consisted of 188 younger aged patients (20-39 years of age), 1024 middle aged patients (40-59 years of age), and 1020 older aged patients (60-85 years of age). In this analysis, the expression of four MMR genes was assessed by immunohistochemistry (IHC). We found that the young group of CRC patients with dMMR had higher overall survival (OS) than the young group of patients with proficient MMR (pMMR) (77% vs. 56%, P = 0.03). Middle-aged patients with dMMR also had higher OS than middle-aged group patients with pMMR (78% vs. 68%, P = 0.012). However, we found no statistical difference in OS between dMMR and pMMR status in the older group of patients (75% vs. 71%, P = 0.224). Finally, the middle- and older-aged group set of patients had higher OS than the young group of patients (69% vs. 71% vs. 59%, P = 0.008). These data demonstrated that the age of disease onset can be an important factor to help evaluate the prognosis of CRC when combined with the analysis of MMR status within tumor biopsied tissue.

CD19-specific triplebody SPM-1 engages NK and γδ T cells for rapid and efficient lysis of malignant B-lymphoid cells.

Triplebodies are antibody-derived recombinant proteins carrying 3 antigen-binding domains in a single polypeptide chain. Triplebody SPM-1 was designed for lysis of CD19-bearing malignant B-lymphoid cells through the engagement of CD16-expressing cytolytic effectors, including NK and γδ T cells.SPM-1 is an optimized version of triplebody ds(19-16-19) and includes humanization, disulfide stabilization and the removal of potentially immunogenic sequences. A three-step chromatographic procedure yielded 1.7 - 5.5 mg of purified, monomeric protein per liter of culture medium. In cytolysis assays with NK cell effectors, SPM-1 mediated potent lysis of cancer-derived B cell lines and primary cells from patients with various B-lymphoid malignancies, which surpassed the ADCC activity of the therapeutic antibody Rituximab. EC50-values ranged from 3 to 86 pM. Finally, in an impedance-based assay, SPM-1 mediated a particularly rapid lysis of CD19-bearing target cells by engaging and activating both primary and expanded human γδ T cells from healthy donors as effectors.These data establish SPM-1 as a useful tool for a kinetic analysis of the cytolytic reactions mediated by γδ T and NK cells and as an agent deserving further development towards clinical use for the treatment of B-lymphoid malignancies.

Association between Ki67 Index and Clinicopathological Features in Colorectal Cancer.

BACKGROUND: Conflicting results have been reported about the association between the Ki67 labeling index (Ki67-Li) and clinical outcome in patients with colorectal cancer (CRC). PATIENTS AND METHODS: Ki67 expression was assessed by immunohistochemistry (IHC) in 2,233 consecutive CRC cases. RESULTS: We determined 992 cases to have a low and 1,241 cases to have a high Ki67-Li (representing an approximately 44-56% breakdown in distribution between low versus high patients designated by phenotype). Stage III patients with a high Ki67-Li had higher 3-year disease-free survival (DFS) and overall survival (OS) than those with a low Ki67-Li (DFS 70 vs. 61%; p = 0.02 and OS 75 vs. 64%; p = 0.008). We also found significantly improved 3-year progression-free survival (PFS) for stage IV patients in the high versus the low Ki67-Li group (PFS 14 vs. 10%; p = 0.02). Yet, we found no statistical differences in prognosis for stage I and II patients and in OS for stage IV patients between high versus low Ki67-Li (p > 0.05). CONCLUSION: Our results suggest that high Ki67-Li can be an independent prognostic biomarker to aid the assessment of patient outcomes in both stage III and IV CRC.

Chip-based platform for dynamic analysis of NK cell cytolysis mediated by a triplebody.

Cancer therapy via redirected lysis mediated by antibodies and antibody-derived agents relies on the availability of substantial numbers of sufficiently active immune effector cells. To monitor antitumor responses before and during therapy, sensitive methods are needed, capable of quantitating specific lysis of target cells. Here we present a chip-based single-cell cytometric assay, which uses adherent human target cells arrayed in structured micro-fields. Using a fluorescent indicator of cell death and time-lapse microscopy in an automated high-throughput mode, we measured specific target cell lysis by activated human NK cells, mediated by the therapeutic single chain triplebody SPM-2 (33-16-123). This antibody-derived tri-specific fusion protein carries binding sites for the myeloid antigens CD33 and CD123 and recruits NK cells via a binding site for the Fc-receptor CD16. Specific lysis increased with increasing triplebody concentration, and the single-cell assay was validated by direct comparison with a standard calcein-release assay. The chip-based approach allowed measurement of lysis events over 16 hours (compared to 4 hours for the calcein assay) and required far smaller numbers of primary cells. In addition, dynamic properties inaccessible to conventional methods provide new details about the activation of cytolytic effector cells by antibody-derived agents. Thus, the killing rate exhibited a dose-dependent maximum during the reaction interval. In clinical applications ex vivo monitoring of NK activity of patient's endogenous cells will likely help to choose appropriate therapy, to detect impaired or recovered NK function, and possibly to identify rare subsets of cancer cells with particular sensitivity to effector-cell mediated lysis.

Dual-targeting triplebody 33-3-19 mediates selective lysis of biphenotypic CD19+ CD33+ leukemia cells.

Simultaneous targeting of multiple tumor-associated antigens (TAAs) in cancer immunotherapy is presumed to enhance tumor cell selectivity and to reduce immune escape.The combination of B lymphoid marker CD19 and myeloid marker CD33 is exclusively present on biphenotypic B/myeloid leukemia cells. Triplebody 33-3-19 binds specifically to both of these TAAs and activates T cells as immune effectors. Thereby it induces specific lysis of established myeloid (MOLM13, THP-1) and B-lymphoid cell lines (BV173, SEM, Raji, ARH77) as well as of primary patient cells. EC50 values range from 3 pM to 2.4 nM. In accordance with our hypothesis, 33-3-19 is able to induce preferential lysis of double- rather than single-positive leukemia cells in a target cell mixture: CD19/CD33 double-positive BV173 cells were eliminated to a significantly greater extent than CD19 single-positive SEM cells (36.6% vs. 20.9% in 3 hours, p = 0.0048) in the presence of both cell lines. In contrast, equivalent elimination efficiencies were observed for both cell lines, when control triplebody 19-3-19 or a mixture of the bispecific single chain variable fragments 19-3 and 33-3 were used. This result highlights the potential of dual-targeting agents for efficient and selective immune-intervention in leukemia patients.

The cost-effectiveness of UGT1A1 genotyping before colorectal cancer treatment with irinotecan from the perspective of the German statutory health insurance.

BACKGROUND: The evidence concerning the cost-effectiveness of UGT1A1*28 genotyping is ambiguous and does not allow drawing valid conclusions for Germany. This study evaluates the cost-effectiveness of UGT1A1 genotyping in patients with metastatic colorectal cancer undergoing irinotecan-based chemotherapy compared to no testing from the perspective of the German statutory health insurance. MATERIAL AND METHODS: A decision-analytic Markov model with a life time horizon was developed. No testing was compared to two genotype-dependent therapy strategies: 1) dose reduction by 25%; and 2) administration of a prophylactic G-CSF growth factor analog for homozygous and heterozygous patients. Probability, quality of life and cost parameters used in this study were based on published literature. Deterministic and probabilistic sensitivity analyses were performed to account for parameter uncertainties. RESULTS: Strategy 1 dominated all remaining strategies. Compared to no testing, it resulted in only marginal QALY increases (0.0002) but a cost reduction of €580 per patient. Strategy 2 resulted in the same health gains but increased costs by €10 773. In the probabilistic analysis, genotyping and dose reduction was the optimal strategy in approximately 100% of simulations at a threshold of €50 000 per QALY. Deterministic sensitivity analysis shows that uncertainty for this strategy originated primarily from costs for irinotecan-based chemotherapy, from the prevalence of neutropenia among heterozygous patients, and from whether dose reduction is applied to both homozygotes and heterozygotes or only to the former. CONCLUSION: This model-based synthesis of the most recent evidence suggests that pharmacogenetic UGT1A1 testing prior to irinotecan-based chemotherapy dominates non-personalized colon cancer care in Germany. However, as structural uncertainty remains high, these results require validation in clinical practice, e.g. based on a managed-entry agreement.

Hematologic Malignancies in Pregnancy: Management Guidelines From an International Consensus Meeting.

PURPOSE: The incidence of hematologic malignancies during pregnancy is 0.02%. However, this figure is increasing, as women delay conception until a later age. Systemic symptoms attributed to the development of a hematologic cancer may overlap with physiologic changes of pregnancy. A favorable prognosis is contingent upon early diagnosis and treatment. Therefore, a high index of suspicion is required by health care providers. Although timely, accurate diagnosis followed by appropriate staging is essential and should not be delayed due to pregnancy, management guidelines are lacking due to insufficient evidence-based research. Consequently, treatment is delayed, posing significant risks to maternal and fetal health, and potential pregnancy termination. This report provides guidelines for clinical management of hematologic cancers during the perinatal period, which were developed by a multidisciplinary team including an experienced hematologist/oncologist, a high-risk obstetrics specialist, a neonatologist, and experienced nurses, social workers, and psychologists. METHODS: These guidelines were developed by experts in the field during the first International Consensus Meeting of Prenatal Hematologic Malignancies, which took place in Leuven, Belgium, on May 23, 2014. RESULTS AND CONCLUSION: This consensus summary equips health care professionals with novel diagnostic and treatment methodologies that aim for optimal treatment of the mother, while protecting fetal and pediatric health.

The ethics of deep brain stimulation (DBS).

Deep brain stimulation (DBS) is an invasive technique designed to stimulate certain deep brain regions for therapeutic purposes and is currently used mainly in patients with neurodegenerative disorders, such as Parkinson's disease. However, DBS is also used increasingly for other experimental applications, such as the treatment of psychiatric disorders (e.g. severe depression), weight reduction. Apart from its therapeutic potential, DBS can cause severe adverse effects, some that might also have a significant impact on the patient's personality and autonomy by the external stimulation of DBS which effects lie beyond the individual's control and free will. The article's purpose is to outline the procedures of DBS currently used in therapeutic and experimental applications and to discuss the ethical concerns regarding this procedure. It will address the clinical benefit-risk-ratio, the particular ethics of research in this field, and the ethical issues raised by affecting a patient's or an individual's personality and autonomous behaviour. Moreover, a potential ethical guideline, the Ulysses contract is discussed for the field of clinical application as well as the question of responsibility.

T cell-recruiting triplebody 19-3-19 mediates serial lysis of malignant B-lymphoid cells by a single T cell.

Triplebody 19-3-19, an antibody-derived protein, carries three single chain fragment variable domains in tandem in a single polypeptide chain. 19-3-19 binds CD19-bearing lymphoid cells via its two distal domains and primary T cells via its CD3-targeting central domain in an antigen-specific manner. Here, malignant B-lymphoid cell lines and primary cells from patients with B cell malignancies were used as targets in cytotoxicity tests with pre-stimulated allogeneic T cells as effectors. 19-3-19 mediated up to 95 % specific lysis of CD19-positive tumor cells and, at picomolar EC50 doses, had similar cytolytic potency as the clinically successful agent Blinatumomab. 19-3-19 activated resting T cells from healthy unrelated donors and mediated specific lysis of both autologous and allogeneic CD19-positive cells. 19-3-19 led to the elimination of 70 % of CD19-positive target cells even with resting T cells as effectors at an effector-to-target cell ratio of 1 : 10. The molecule is therefore capable of mediating serial lysis of target cells by a single T cell. These results highlight that central domains capable of engaging different immune effectors can be incorporated into the triplebody format to provide more individualized therapy tailored to a patient's specific immune status.

Phage idiotype vaccination: first phase I/II clinical trial in patients with multiple myeloma.

BACKGROUND: Multiple myeloma is characterized by clonal expansion of B cells producing monoclonal immunoglobulins or fragments thereof, which can be detected in the serum and/or urine and are ideal target antigens for patient-specific immunotherapies. METHODS: Using phage particles as immunological carriers, we employed a novel chemically linked idiotype vaccine in a clinical phase I/II trial including 15 patients with advanced multiple myeloma. Vaccines composed of purified paraproteins linked to phage were manufactured successfully for each patient. Patients received six intradermal immunizations with phage idiotype vaccines in three different dose groups. RESULTS: Phage idiotype was well tolerated by all study participants. A subset of patients (80% in the middle dose group) displayed a clinical response indicated by decrease or stabilization of paraprotein levels. Patients exhibiting a clinical response to phage vaccines also raised idiotype-specific immunoglobulins. Induction of a cellular immune response was demonstrated by a cytotoxicity assay and delayed type hypersensitivity tests. CONCLUSION: We present a simple, time- and cost-efficient phage idiotype vaccination strategy, which represents a safe and feasible patient-specific therapy for patients with advanced multiple myeloma and produced promising anti-tumor activity in a subset of patients.

A dual-targeting triplebody mediates preferential redirected lysis of antigen double-positive over single-positive leukemic cells.

The single-chain triplebody HLA-ds16-hu19 consists of three single-chain Fv (scFv) antibody fragments connected in a single polypeptide chain. This protein with dual-targeting capacity mediated preferential lysis of antigen double positive(dp) over single-positive (sp) leukemic cells by recruitment of natural killer (NK) cells as effectors. The two distal scFv modules were specific for the histocompatibility protein HLA-DR and the lymphoid antigen CD19, the central one for the Fc gamma receptor CD16. In antibody-dependent cellular cytotoxicity (ADCC) experiments with a mixture of leukemic target cells comprising both HLA-DR sp HuT-78 or Kasumi-1 cells and (HLA-DR plus CD19) dp SEM cells, the triplebody mediated preferential lysis of the dp cells even when the sp cells were present in ≤ 20-fold numerical excess.The triplebody promoted equal lysis of SEM cells at 2.5-fold and 19.5-fold lower concentrations than the parental antibodies specific for HLA-DR and CD19, respectively. Finally, the triplebody also eliminated primary leukemic cells at lower concentrations than an equimolar mixture of bispecific single-chain Fv fragments (bsscFvs) separately addressing each target antigen (hu19-ds16 and HLA-ds16). The increased selectivity of targeting and the preferential lysis of dp over sp cells achieved by dual-targeting open attractive new perspectives for the use of dual-targeting agents in cancer therapy.

NK cells from an AML patient have recovered in remission and reached comparable cytolytic activity to that of a healthy monozygotic twin mediated by the single-chain triplebody SPM-2.

BACKGROUND: The capacity of patient's Natural Killer cells (NKs) to be activated for cytolysis is an important prerequisite for the success of antibody-derived agents such as single-chain triplebodies (triplebodies) in cancer therapy. NKs recovered from AML patients at diagnosis are often found to be reduced in peripheral blood titers and cytolytic activity. Here, we had the unique opportunity to compare blood titers and cytolytic function of NKs from an AML patient with those of a healthy monozygotic twin. The sibling's NKs were compared with the patient's drawn either at diagnosis or in remission after chemotherapy. The cytolytic activities of NKs from these different sources for the patient's autologous AML blasts and other leukemic target cells in conjunction with triplebody SPM-2, targeting the surface antigens CD33 and CD123 on the AML cells, were compared. METHODS: Patient NKs drawn at diagnosis were compared to NKs drawn in remission after chemotherapy and a sibling's NKs, all prepared from PBMCs by immunomagnetic beads (MACS). Redirected lysis (RDL) assays using SPM-2 and antibody-dependent cellular cytotoxicity (ADCC) assays using the therapeutic antibody RituximabTM were performed with the enriched NKs. In addition, MACS-sorted NKs were analyzed for NK cell activating receptors (NCRs) by flow cytometry, and the release of TNF-alpha and IFN-gamma from blood samples of both siblings after the addition of the triplebody were measured in ELISA-assays. RESULTS: Patient NKs isolated from peripheral blood drawn in remission produced comparable lysis as NKs from the healthy twin against the patient's autologous bone marrow (BM) blasts, mediated by SPM-2. The NCR receptor expression profiles on NKs from patient and twin were similar, but NK cell titers in peripheral blood were lower for samples drawn at diagnosis than in remission. CONCLUSIONS: Peripheral blood NK titers and ex vivo cytolytic activities mediated by triplebody SPM-2 were comparable for cells drawn from an AML patient in remission and a healthy twin. If these results can be generalized, then NKs from AML patients in remission are sufficient in numbers and cytolytic activity to make triplebodies promising new agents for the treatment of AML.

Priority-setting, rationing and cost-effectiveness in the German health care system.

Germany has just started a public debate on priority-setting, rationing and cost-effectiveness due to the cost explosion within the German health care system. To date, the costs for German health care run at 11,6% of its Gross Domestic Product (GDP, 278,3 billion €) that represents a significant increase from the 5,9 % levels present in 1970. In response, the German Parliament has enacted several major and minor legal reforms over the last three decades for the sake of cost containment and maintaining stability of the health care system. The Statutory Health Insurance--SHI (Gesetzliche Krankenversicherung--GKV) is based on the fundamental principle of solidarity and provides an ethical and legal framework for implementing equity, comprehensiveness and setting the principles and rules for financing and providing health care services and benefits. Within the SHI system, several major actors can be identified: the Federal Ministry of Health, the 16 state ministries of health, the Federal Joint Committee (G-BA), the physicians (with their associations) and the hospitals (with their organizations) on the provider side, and the sickness funds with their associations on the purchasers' side. This article reviews the structure and complexities of the German health care system with its major players and participants. The focus will be put on relevant ethical, legal and economic aspects for prioritization, rationalization, rationing and cost-effectiveness of medical benefits and services. In conclusion, this article pleads for open discussion on the challenging subject of priority-setting instead of accepting the implicit and non-transparent rationing of medical services that currently occurs at many different levels within the health care system, as it stands today.