Our first product is aimed at research labs across Universities and drug companies before we develop a more hi-tech and fully automated clinical scale product. KAREL will help such labs achieve single-cell manipulation that can be coupled with measurements of phenotype response using downstream multiplexed omics analyses.

Example 1 – Cancer research

In the last several years, scientists have revealed that we might be composed of millions of unique and unrepeatable cells. In cancer, a single-cell can evolve into a malignant mass of tumor cells with divergent lineages and forming distinct sub-populations leading to intra-tumor heterogeneity, in addition to heterogeneity across tumors from different patients. Therefore while treatments may be effective against the majority of a given tumor, minor sub-populations of cells often acquire resistance to further treatment leading to the recurrence of cancer. Single-cell analyses have begun to reveal the vast extent of genetic and phenotypic heterogeneity among tumor cells and its role in cancer biology. Both DNA and RNA single-cell sequencing (SCS) are powerful tools for resolving clonal substructure within a single tumor and can reveal rare subclonal (<1%) mutations that may play a critical role in tumor evolution and therapy resistance. In studies applying single cell exome sequencing to study clonal diversity in kidney tumors, muscle-invasive bladder cancer and colon cancer, it has been discovered that single cells share common founder mutations, suggesting evolution from a common genetic lineage. Moreover, analyzing bulk populations only generates an averaged gene expression level in all cells identifying limited mutations. In contrast, single-cell RNA-seq, via sequencing the transcriptome of individual cells in depth can detect low abundance mutations, cell-to-cell transcript variation at the single nucleotide level and gene expression heterogeneity among single-cells.

Cas9 bound to target DNA. Crystal structure of Cas9-guide RNA complex primed for DNA cleavage.

Example 2 – Massively parallel scale gene editing of individual cells using CRISPR-Cas technology

The unprecedented ease and efficacy of the clustered, regularly interspaced, short palindromic repeat (CRISPR) technology presents an attractive option for massive scale single-cell gene editing to be used for repair of pathogenic mutations or engineering of cells such as therapeutic T cells. CRISPR-Cas9 genome editing entails introducing DNA double-strand breaks (DSBs) with the Cas9 nuclease at a chromosomal locus of interest targeted by the sgRNA (single guide RNA); these DSBs are repaired via non-homologous end-joining (NHEJ) or homologous recombination (HR). Targeting genes with this approach, aims at maintaining the endogenous gene regulation and the gene of interest can be delivered right at its physiological site on the genome. Compared to currently used viruses, our technology is a giant leap toward the true promise of gene editing in a safe way in individual target cells.

An early prototype of the KAREL microchip with the nanoneedle.

A little snapshot of what some of the most prominent scientists of our time have to say about KAREL. 

This technology aims at solving a fundamental problem in genomics and medicine. Ronald W. Davis, Ph.D., Director of Stanford Genome Technology Center, “Today’s Greatest Inventors”, Atlantic Magazine (2013), Genetics pioneer.

This robotic chip technology can make studying single cells on a high-throughput scale a standard norm for drug research, sequencing and development. Michael Snyder, Ph.D., Stanford W. Ascherman Professor and Chair, Department of Genetics; Director of Center for Genomics and Personalized Medicine, Stanford University School of Medicine, Personalized Medicine pioneer.

The major applications of this technology will be in controlling gene expressions (over expression, under expression, knockdowns) in cells and dosage of the delivery of drug molecules to cells. Ben Barres, M.D., Ph.D., Chair & Professor, Department of Neurobiology, Stanford University School of Medicine, Neuroscience pioneer.

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