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In multiple myeloma spatial differences in the subclonal architecture, molecular signatures and composition of the microenvironment remain poorly characterized. To address this shortcoming, we perform multi-region sequencing on paired random bone marrow and focal lesion samples from 17 newly diagnosed patients. Using single-cell RNA- and ATAC-seq we find a median of 6 tumor subclones per patient and unique subclones in focal lesions. Genetically identical subclones display different levels of spatial transcriptional plasticity, including nearly identical profiles and pronounced heterogeneity at different sites, which can include differential expression of immunotherapy targets, such as CD20 and CD38. Macrophages are significantly depleted in the microenvironment of focal lesions. We observe proportional changes in the T-cell repertoire but no site-specific expansion of T-cell clones in intramedullary lesions. In conclusion, our results demonstrate the relevance of considering spatial heterogeneity in multiple myeloma with potential implications for models of cell-cell interactions and disease progression. The spatial architecture of multiple myeloma remains to be explored. Here, the authors perform bulk and single cell sequencing for samples from newly diagnosed patients and reveal gene signatures associated with focal lesions and spatial heterogeneity in the tumour microenvironment.

Transcriptome analysis by RNA sequencing (RNA-seq) has become an indispensable research tool in modern plant biology. Virtually all RNA-seq studies provide a snapshot of the steady state transcriptome, which contains valuable information about RNA populations at a given time but lacks information about the dynamics of RNA synthesis and degradation. Only a few specialized sequencing techniques, such as global run-on sequencing, have been used to provide information about RNA synthesis rates in plants. Here, we demonstrate that RNA labeling with the modified, nontoxic uridine analog 5-ethynyl uridine (5-EU) in Arabidopsis (Arabidopsis thaliana) seedlings provides insight into plant transcriptome dynamics. Pulse labeling with 5-EU revealed nascent and unstable RNAs, RNA processing intermediates generated by splicing, and chloroplast RNAs. Pulse-chase experiments with 5-EU allowed us to determine RNA stabilities without the need for chemical transcription inhibitors such as actinomycin and cordycepin. Inhibitor-free, genome-wide analysis of polyadenylated RNA stability via 5-EU pulse-chase experiments revealed RNAs with shorter half-lives than those reported after chemical inhibition of transcription. In summary, our results indicate that the Arabidopsis nascent transcriptome contains unstable RNAs and RNA processing intermediates and suggest that polyadenylated RNAs have low stability in plants. Our technique lays the foundation for easy, affordable, nascent transcriptome analysis and inhibitor-free analysis of RNA stability in plants.

Current treatment strategies for multiple myeloma (MM) are highly effective, but most patients develop relapsed/refractory disease (RRMM). The anti-CD38/CD3xCD28 trispecific antibody SAR442257 targets CD38 and CD28 on MM cells and co-stimulates CD3 and CD28 on T cells (TCs). We evaluated different key aspects such as MM cells and T cells avidity interaction, tumor killing, and biomarkers for drug potency in three distinct cohorts of RRMM patients. We found that a significantly higher proportion of RRMM patients (86%) exhibited aberrant co-expression of CD28 compared to newly diagnosed MM (NDMM) patients (19%). Furthermore, SAR442257 mediated significantly higher TC activation, resulting in enhanced MM killing compared to bispecific functional knockout controls for all relapse cohorts (Pearson’s r = 0.7). Finally, patients refractory to anti-CD38 therapy had higher levels of TGF-β (up to 20-fold) compared to other cohorts. This can limit the activity of SAR442257. Vactoserib, a TGF-β inhibitor, was able to mitigate this effect and restore sensitivity to SAR442257 in these experiments. In conclusion, SAR442257 has high potential for enhancing TC cytotoxicity by co-targeting CD38 and CD28 on MM and CD3/CD28 on T cells.

In order to meet the challenges in data evaluation and comparability between studies in multiple myeloma (MM) minimal residual disease (MRD) assessment, the goal of the current study was to provide a step-by-step evaluation of next-generation sequencing (NGS) and multicolor flow cytometry (MFC) data. Bone marrow (BM) sample pairs from 125 MM patients were analyzed by NGS and MFC MM MRD methods. Tumor load (TL) and limit of detection (LOD) and quantification (LOQ) were calculated. The best-fit MRD cut-off was chosen as 1 × 10−5, resulting in an overall 9.6% (n overall = 12 (NGS n = 2, MFC n = 10)) nonassessable cases. The overall concordance rate between NGS and MFC was 68.0% (n = 85); discordant results were found in 22.4% (11.2% (n = 14) of cases in each direction. Overall, 55.1% (n = 60/109) and 49.5% (n = 54/109) of patients with a serological response ≥ very good partial response (VGPR) showed BM MRD negativity by NGS and MFC, respectively. A good correlation in the TL assessed by both techniques was found (correlation coefficient = 0.8, n = 40, p < 0.001). Overall, our study shows good concordance between MM BM MRD status and TL when comparing NGS and MFC at a threshold of 10–5. However, a sufficient number of analyzed events and calculation of MRD key metrics are essential for the comparison of methods and evaluability of data at a specific MRD cut-off.

To comprehensively unravel the temporal relationship between initiating and driver events and its impact on clinical outcomes, we analyzed 421 whole-genome sequencing profiles from 382 patients. Using clock-like mutational signatures, we estimated a time lag of 2-4 decades between initiating events and diagnosis. In patients with hyperdiploidy, we demonstrate that trisomies of odd-numbered chromosomes can be acquired simultaneously with other chromosomal gains, such as 1q gain. We provide evidence that hyperdiploidy is acquired after canonical IGH translocation when both events are present. Finally, patients with early 1q gain had adverse outcomes similar to those with 1q amplification (>1 extra-copies), but faring worse than those with late 1q gain. This underscores that the prognostic impact of 1q gain/amp depends more on the timing of acquisition than on the number of extra copies gained. Overall, this study contributes to a better understanding of the life history of MM and may have prognostic implications.

Transcriptome analysis by RNA sequencing (RNA-seq) has become an indispensable core research tool in modern plant biology. Virtually all RNA-seq studies provide a snapshot of the steady-state transcriptome, which contains valuable information about RNA populations at a given time, but lacks information about the dynamics of RNA synthesis and degradation. Only a few specialized sequencing techniques, such as global run-on sequencing (GRO-seq), have been applied in plants and provide information about RNA synthesis rates. Here, we demonstrate that RNA labeling with a modified, non-toxic uridine analog, 5-ethynyl uridine (5-EU), in Arabidopsis thaliana seedlings provides insight into the dynamic nature of a plant transcriptome. Pulse-labeling with 5-EU allowed the detection and analysis of nascent and unstable RNAs, of RNA processing intermediates generated by splicing, and of chloroplast RNAs. We also conducted pulse-chase experiments with 5-EU, which allowed us to determine RNA stabilities without the need for chemical inhibition of transcription using compounds such as actinomycin and cordycepin. Genome-wide analysis of RNA stabilities by 5-EU pulse-chase experiments revealed that this inhibitor-free RNA stability measurement results in RNA half-lives much shorter than those reported after chemical inhibition of transcription. In summary, our results show that the Arabidopsis nascent transcriptome contains unstable RNAs and RNA processing intermediates, and suggest that half-lives of plant RNAs are largely overestimated. Our results lay the ground for an easy and affordable nascent transcriptome analysis and inhibitor-free analysis of RNA stabilities in plants.

This thesis is a study of the link between logistics organization structures and corporate strategy, including corporate organization.To analyze the link between the three topics, frameworks for analysis were developed for each. Corporate strategy was divided into three types: cost leadership, differentiation, and focus. Corporate organization structures were divided into two frameworks: amount of decentralization and types of structures (functional, product, area, and matrix). Seven logistics organization structures were discussed: traditional, functional, program, matrix, channel management, partnership and third-part or contract logistics.Four basic hypotheses on the organizational fit between the three topics were discussed: the fit between logistics and corporate organization structures, and the fit between the logistics organization structure and each of the three strategic options.The hypotheses were tested, covering all strategic options, through five case studies of firms engaged in different industries. The evidence from the cases support all of the hypotheses.