Long-term survival after adoptive bone marrow T cell therapy of advanced metastasized breast cancer: follow-up analysis of a clinical pilot trial

Background

The bone marrow (BM) of breast cancer patients harbors tumor-reactive memory T cells (TCs) with therapeutic potential. We recently described the immunologic effects of adoptive transfer of ex vivo restimulated tumor-reactive memory TCs from the BM of 12 metastasized breast cancer patients in a clinical phase-I study. In this trial, adoptive T cell transfer resulted in the occurrence of circulating tumor antigen-reactive type-1 TCs. We here describe the long-term clinical outcome and its correlation with tumor-specific cellular immune response in 16 metastasized breast cancer patients, including 12 included in the original study.

Methods

Sixteen metastatic breast cancer patients with preexisting tumor-reactive BM memory TCs were included into the study. The study protocol involved one transfusion of TCs which were reactivated in vitro with autologous dendritic cells pulsed with lysates of MCF-7 breast cancer cells as source of tumor antigens. The presence of tumor-reactive memory TCs was analyzed by IFN-γ ELISpot assays.

Results

Tumor-reactive memory TCs in the peripheral blood were induced de novo in 7/16 patients (44 %) after adoptive TC transfer. These patients were considered immunologic responders to the therapy. Positive adoptive immunotherapy (ADI) response was observed significantly more often in patients without bone metastases (p = 0.0051), in patients with high levels of tumor-reactive BM TCs prior to therapy (p = 0.036) and correlated significantly with the estimated numbers of transferred tumor-reactive TCs (p = 0.0021). After the treatment, we observed an overall median survival of 33.8 months in the total cohort with three patients alive at last follow-up and more than 7 years after ADI. Numbers of transferred tumor-reactive TCs correlated significantly with the overall survival of patients (p = 0.017). Patients with an immunologic response to ADI in the peripheral blood had a significantly longer median survival than nonresponders (median survival 58.6 vs. 13.6 months; p = 0.009).

Conclusion

In metastasized breast cancer patients, adoptive transfer of BM TCs can induce the presence of tumor antigen-reactive type-1 TCs in the peripheral blood. Patients with immunologic response after ADI show a significantly longer overall survival. Patients with bone metastases significantly less frequently respond to the treatment and, therefore, might not be optimal candidates for ADI. Although the present study does not yet prove the therapeutic effect of ADI, these findings shed light on the relation between immune response and cancer prognosis and suggest that transfer of reactivated BM TCs might bear therapeutic potential.     

Benefit-of-doubt (BOD) scoring: A sequencing-based method for SNP candidate assessment from high to medium read number data sets

Identification of single nucleotide polymorphisms (SNPs) is a key element in sequence-based genetic analysis. Next generation sequencing offers a cost-effective basis to generate the necessary, large sequence data sets, and bioinformatic methods are being developed to process sequencing machine readouts. We were interested in detection of SNPs in a 350 kb region of an EMS-mutagenized Arabidopsis chromosome 3. The region was selectively analyzed using PCR-generated, overlapping fragments for Solexa sequencing. The ensuing reads provided a high coverage and were processed bioinformatically. In order to assess the SNP candidates obtained with a frequently used alignment program and SNP caller, we developed an additional method that allows the identification of high confidence SNP loci. The method can easily be applied to complete genome sequence data of sufficient coverage.     

Interaction of the C14-OH Group of Adriamycin with DNA Phosphate as Spectroscopically Evidenced

Abstract

Monitoring the band of the antisymmetric stretching vibration of the backbone PO₂ ^? group in DNA-anthracycline complexes demonstrates an extraordinary wavenumber shift for the adriamycin complex compared to that of daunomycin. The structures of both anthracyclines, however, are very closely related and differ only by a surplus hydroxyl group of adriamycin in the C14 position. The wavenumber shift observed for the DNA-adriamycin complex is unequivocally attributed to an additional linkage of the C14-OH of adriamycin to the phosphate group of DNA. Thus, serveral of the hypothetical structural models for the DNA-adriamycin complex for which a hydrogen bond between the C14 hydroxyl of the drug and DNA phosphate was postulated (S. Neidle, Cancer Treatment Rep. 61, 928 (1977); G. J. Quigley et. al., Proc. Natl. Acad Sci. USA 77, 7204 (1980)) get the first clear-cut experimental evidence.     

Detection of Metabolic Key Enzymes of Methane Turnover Processes in Cold Seep Microbial Biofilms

In anoxic environments, methane oxidation is conducted in a syntrophic process between methanotrophic archaea (ANME) and sulfate reducing bacteria (SRB). Microbial mats consisting of ANME, SRB and other microorganisms form methane seep-related carbonate buildups in the anoxic bottom waters of the Black Sea Crimean shelf. To shed light on the localization of the biochemical processes at the level of single cells in the Black Sea microbial mats, we applied antibody-based markers for key enzymes of the relevant metabolic pathways. The dissimilatory adenosine-5′-phosphosulfate (APS) reductase, methyl-coenzyme M reductase (MCR) and methanol dehydrogenase (MDH) were selected to localize sulfate respiration, reverse methanogenesis and aerobic methane oxidation, respectively. The key enzymes could be localized by double immunofluorescence and immunocytochemistry at light- and electron microscopic levels. In this study we show that sulfate reduction is conducted synchronized and in direct proximity to reverse methanogenesis of ANME archaea. Microcolonies in interspaces between ANME/SRB express methanol dehydrogenase, which is indicative for oxidation of C₁ compounds by methylotrophic or methanotrophic bacteria. Thus, in addition to syntrophic AOM, oxygen-dependent processes are also conducted by a small proportion of the microbial population.     

Search for α3β2/3γ2 subtype selective ligands that are stable on human liver microsomes

Selective modulation of specific benzodiazepine receptor (BzR) gamma amino butyric acid-A (GABA_A) receptor ion channels has been identified as an important method for separating out the variety of pharmacological effects elicited by BzR-related drugs. Importantly, it has been demonstrated that both α2β_(2/3)γ2 (α2BzR) and α3BzR (and/or α2/α3) BzR subtype selective ligands exhibit anxiolytic effects with little or no sedation. Previously we have identified several such ligands; however, three of our parent ligands exhibited significant metabolic liability in rodents in the form of a labile ester group. Here eight analogs are reported which were designed to circumvent this liability by utilizing a rational replacement of the ester moiety based on medicinal chemistry precedents. In a metabolic stability study using human liver microsomes, four compounds were found to undergo slower metabolic transformation, as compared to their corresponding ester analogs. These compounds were also evaluated in in vitro efficacy assays. Additionally, bioisostere 11 was evaluated in a rodent model of anxiety. It exhibited anxiolytic activity at doses of 10 and 100 mg/kg and was devoid of sedative properties.     

The impact of prehistoric mining activities on the environment: a multidisciplinary study at the fen Schwarzenbergmoos (Brixlegg, Tyrol, Austria)

The exploitation of copper ore deposits of the northern Greywacke Zone was initiated by the implementation of metallurgic technologies in the Eastern Alps thousands of years ago. This multi-proxy study aimed to detect prehistoric mining phases in the vicinity of a prominent copper ore deposit in the Lower Inn Valley. Therefore we studied a peat core from a fen using pollen, micro charcoal and geochemical analyses. In the same fen, an archaeological investigation revealed an ore beneficiation site, well dated by dendrochronological analysis to the Late Bronze Age (9th century b.c.). First hints of mining activities reflected by the occurrence of anthropogenic indicators in the pollen diagram, associated with elevated metal values, at the beginning of the Bronze Age might result from early mineral prospecting and metallurgical experiments around the use of fahlore. The local ore deposit was then abandoned until during the Bronze Age mining activities started to increase. This is reflected by an expansion of the pioneer species Pinus and Larix on mine spoil heaps in the proximity. Concomitantly metal ratios and micro charcoal increase. From about 1000 to 850 b.c. a strong impact of mining activities is displayed in the multi-proxy data. The local forest was partly cleared on and in the vicinity of the fen. According to dendrochronological data the ore beneficiation plant was in use from about 900 to 870 b.c. Until about 700 b.c. another period with moderate impact by mining activities in the further vicinity of the fen shows up.