Hema-seq reveals genomic aberrations in a rare simultaneous occurrence of hematological malignancies.

Co-occurrence of multiple myeloma and acute myelogenous leukemia is rare, with both malignancies often tracing back to multipotent hematopoietic stem cells. Cytogenetic techniques are the established baseline for diagnosis and characterization of complex hematological malignancies. In this study, we develop a workflow called Hema-seq to delineate clonal changes across various hematopoietic lineages through the integration of whole-genome sequencing, copy-number variations, cell morphology, and cytogenetic aberrations. In Hema-seq, cells are selected from Wright-stained slides and fluorescent probe-stained slides for sequencing. This technique therefore enables direct linking of whole-genome sequences to cytogenetic profiles. Through this method, we mapped sequential clonal alterations within the hematopoietic lineage, identifying critical shifts leading to myeloma and acute myeloid leukemia (AML) cell formations. By synthesizing data from each cell lineage, we provided insights into the hematopoietic tree's clonal evolution. Overall, this study highlights Hema-seq's capability in deciphering genomic heterogeneity in complex hematological malignancies, which can enable better diagnosis and treatment strategies.

Barcoded multiple displacement amplification for high coverage sequencing in spatial genomics.

Determining mutational landscapes in a spatial context is essential for understanding genetically heterogeneous cell microniches. Current approaches, such as Multiple Displacement Amplification (MDA), offer high genome coverage but limited multiplexing, which hinders large-scale spatial genomic studies. Here, we introduce barcoded MDA (bMDA), a technique that achieves high-coverage genomic analysis of low-input DNA while enhancing the multiplexing capabilities. By incorporating cell barcodes during MDA, bMDA streamlines library preparation in one pot, thereby overcoming a key bottleneck in spatial genomics. We apply bMDA to the integrative spatial analysis of triple-negative breast cancer tissues by examining copy number alterations, single nucleotide variations, structural variations, and kataegis signatures for each spatial microniche. This enables the assessment of subclonal evolutionary relationships within a spatial context. Therefore, bMDA has emerged as a scalable technology with the potential to advance the field of spatial genomics significantly.

Spatial epitranscriptomics reveals A-to-I editome specific to cancer stem cell microniches.

Epitranscriptomic features, such as single-base RNA editing, are sources of transcript diversity in cancer, but little is understood in terms of their spatial context in the tumour microenvironment. Here, we introduce spatial-histopathological examination-linked epitranscriptomics converged to transcriptomics with sequencing (Select-seq), which isolates regions of interest from immunofluorescence-stained tissue and obtains transcriptomic and epitranscriptomic data. With Select-seq, we analyse the cancer stem cell-like microniches in relation to the tumour microenvironment of triple-negative breast cancer patients. We identify alternative splice variants, perform complementarity-determining region analysis of infiltrating T cells and B cells, and assess adenosine-to-inosine base editing in tumour tissue sections. Especially, in triple-negative breast cancer microniches, adenosine-to-inosine editome specific to different microniche groups is identified.

Advances in Tumor Sampling and Sequencing in Breast Cancer and their Application in Precision Diagnostics and Therapeutics.

Intra- and Inter-tumoral heterogeneity is one of the main hurdles in diagnosing and treating breast cancer. Selecting, sampling, and sequencing the samples appropriately provide unique opportunities in realizing precision medicine. This chapter reviews some of the past landmarks, state-of-the-art technologies, and future directions of translational research in terms of tumor sampling technologies and sequencing in breast cancer. In the state-of-the-art technologies section, the technologies are categorized in terms of scientific, precision diagnostic, and precision therapeutic tools. Finally, limitations and future directions regarding various translational research for clinical applications using these technologies will be discussed.

Direct 2D-to-3D transformation of pen drawings.

Pen drawing is a method that allows simple, inexpensive, and intuitive two-dimensional (2D) fabrication. To integrate such advantages of pen drawing in fabricating 3D objects, we developed a 3D fabrication technology that can directly transform pen-drawn 2D precursors into 3D geometries. 2D-to-3D transformation of pen drawings is facilitated by surface tension-driven capillary peeling and floating of dried ink film when the drawing is dipped into an aqueous monomer solution. Selective control of the floating and anchoring parts of a 2D precursor allowed the 2D drawing to transform into the designed 3D structure. The transformed 3D geometry can then be fixed by structural reinforcement using surface-initiated polymerization. By transforming simple pen-drawn 2D structures into complex 3D structures, our approach enables freestyle rapid prototyping via pen drawing, as well as mass production of 3D objects via roll-to-roll processing.

OPENchip: an on-chip in situ molecular profiling platform for gene expression analysis and oncogenic mutation detection in single circulating tumour cells.

Liquid biopsy holds promise towards practical implementation of personalized theranostics of cancer. In particular, circulating tumour cells (CTCs) can provide clinically actionable information that can be directly linked to prognosis or therapy decisions. In this study, gene expression patterns and genetic mutations in single CTCs are simultaneously analysed by strategically combining microfluidic technology and in situ molecular profiling technique. Towards this, the development and demonstration of the OPENchip (On-chip Post-processing ENabling chip) platform for single CTC analysis by epithelial CTC enrichment and subsequent in situ molecular profiling is reported. For in situ molecular profiling, padlock probes that identify specific desired targets to examine biomarkers of clinical relevance in cancer diagnostics were designed and used to create libraries of rolling circle amplification products. We characterize the OPENchip in terms of its capture efficiency and capture purity, and validate the probe design using different cell lines. By integrating the obtained results, molecular analyses of CTCs from metastatic breast cancer (HER2 (ERBB2) gene expression and PIK3CA mutations) and metastatic pancreatic cancer (KRAS gene mutations) patients were demonstrated without any off-chip processes. The results substantiate the potential implementation of early molecular detection of cancer through sequencing-free liquid biopsy.