CRISPR EUROPE CONGRESS 2017 AGENDA
We are currently preparing the draft agenda for . Sign up to our newsletter below to get the latest agenda, trends and updates.
Applying genome editing to Drug Discovery
The use of CRISPR/Cas9 as a valuable tool for the functional characterization of new genes associated with Prostate Cancer (PrCa) risk
Update on the CRISPR patent wars
A number of patents which cover the basic CRISPR technology have already been granted in the US and in Europe to Zhang’s group. Doudna’s group’s patent applications in the US and Europe are still pending, but they now have a granted UK patent.
In the US, the Patent & Trademark Office has recently given a decision which has awarded US patent rights to CRISPR methods in eukaryotic cells to Zhang’s group, but it is still unclear whether Doudna’s group will get granted patents covering general CRISPR methods and vectors. In Europe, 7 of Zhang’s group’s EP patents have been challenged on grounds of lack of novelty and lack of inventive step, amongst others. This presentation will first discuss what CRISPR inventions may be patented, and then provide an update on the Zhang-Doudna patent wars.
Philip Webber -Partner - Dehns
Identification of novel therapeutic targets in Acute Myeloid Leukemia using CRISPR screens
Acute myeloid leukemia (AML) is an aggressive cancer with a poor prognosis, for which the therapeutic landscape has changed little for decades. Aberrant mRNA splicing plays a key role in cancer development and genes coding for several of the major components of the spliceosome are targeted by somatic mutations in several cancers including myelodysplastic syndromes and AML. Recently, myeloid neoplasms bearing spliceosome gene mutations were shown to be preferentially susceptible to pharmacological disruption of the spliceosome. Here we report that targeting the spliceosome can also be an effective therapeutic strategy in other types of AML.
Recently, we generated a comprehensive catalogue of genetic vulnerabilities in AML using CRISPR-Cas9 genome-wide recessive screens and reported several novel intuitive and non-intuitive therapeutic candidates. Amongst these we identify SRPK1, the gene coding for a serine-threonine kinase that phosphorylates the major spliceosome protein SRSF1. Here, we demonstrate that targeted genetic disruption of SRPK1 in AML driven by MLL-fusion genes, led to differentiation and apoptosis. Additionally, mice transplanted with human AML cell lines carrying the MLL-AF9 fusion gene had a significant prolongation of survival when SRPK1 was genetically disrupted by CRISPR-gRNA. Similar effects were seen with pharmacological inhibition in vitro and in vivo using a novel SRPK1 inhibitor.
At the molecular level we show that genetic or pharmacological inhibition of SRPK1 was associated with profound changes in the splicing of multiple genes involved in the MLL leukemogenic program in association with significant changes in enzymatic modifications of core histone tails. We proceeded to perform a genome-wide CRISPR sensitisation screen to pharmacological inhibition of SRPK1 and identified, amongst other genes, BRD4 as a sensitiser. We go on to show that the BRD inhibitor iBET-151 synergizes with SRPK1 inhibition to kill MLL-AF9 both in vitro and in vivo.
Our work identifies SRPK1 as a novel therapeutic target in AML that can be used alone or in conjunctions with drugs targeting epigenetic modifications to improve their anti-leukemic effects.
Konstantinos Tzelepis - PhD Student - Sanger Institute
iTOP: a novel non-viral delivery system for gene-editing based therapeutics.
NTrans was founded in 2015, based on a proprietary platform technology for the intracellular delivery of bioactive molecules developed at the Hubrecht Institute (KNAW). The iTOP delivery technology is based on a combination of small molecule compounds which forces the uptake of large gulps of extracellular fluid (containing the bioactive molecules) by the cell. Once inside, the vesicles release their content into the cytoplasm, where the bioactive molecules (like gene editing systems) can exert their therapeutic action.
The presentation will cover the iTOP technology and our development plans for a therapy targeted at treating inheritable muscular diseases.
Marco de Boer - CEO - NTrans
CRISPR/Cas9 genome editing as a new tool to study the biology of fertilization and infertility
Egg activation is the first, vital stage of development. The sperm protein PLCzeta has been proposed to induce the Ca2+ oscillations that initiate mammalian embryogenesis. However, there has been no direct evidence that PLCzeta is the physiological trigger of these oscillations. Here we show that Plcz1–/– sperm generated by CRISPR/Cas9 genome editing fail to trigger Ca2+ oscillations in eggs and conclusively demonstrate that PLCzeta is the sole trigger of these oscillations. Surprisingly, eggs fertilized by PLCzeta-null sperm develop, albeit at greatly reduced efficiency, after a significant time-delay and display high rates of polyspermy. Consistent with this, Plcz1–/– males are subfertile but not sterile, suggesting that in the absence of PLCzeta, eventual, spontaneous egg activation occurs. This is the first demonstration that in vivo fertilization without Ca2+ oscillations can result in offspring. PLCzeta-null sperm therefore will serve to test the efficacy and safety of clinical IVF procedures and resolve long-standing questions in fertilization biology.
John Parrington - Associate Professor - University of Oxford
Modelling Estrogen Receptor Mutations in Breast Cancer
Estrogen receptor (ER) is a key driver of breast cancer development and progression and is the target for endocrine therapies. Use of endocrine agents has contributed greatly to reductions in breast cancer mortality over the last three decades. However, many patients develop resistance to these therapies, for reasons that remain unclear. Recent studies have identified ESR1 (ER) mutations in metastatic breast cancer that are predicted to encode ER proteins that are constitutively active and resistant to anti-estrogens. However, functional studies to determine the action of these mutations is impeded by the lack of breast cancer cell lines encoding ESR1 mutations. Here, we have developed the engineered CRISPR-Cas9 genome editing system to incorporate ESR1 mutations in the genomically encoded gene in the well-studied estrogen responsive and ER-positive MCF7 breast cancer cell line. These lines provide a new resource for understanding the functional importance of ESR1 mutations and avenues for treatment of breast cancer patients featuring these mutations.
Laki Buluwela - Professor of Cancer Medicine - Imperial College London
Genome Editing of the Pig Genome using TALENs and CRISPR
Genetically modified (GM) pigs hold great promise for agriculture, disease modeling, and xenotransplantation applications. However, porcine embryonic stem cells or induced pluripotent stem cells with germline chimeric ability have not been reported. GM- pig models are primarily generated by nuclear transfer following genome-editing in somatic cells. By taking the advantage of latest genome-editing tools such as TALENs and CRISPR, we have generated dozens of GM-pigs in the last five years. I will first introduce several porcine disease models, including severe combined immunodeficiency (SCID, Knockout of porcine RAG1 or RAG2 with TALENs), autosomal recessive early- onset Parkinson’s disease (Double-knockout of porcine PARK2/PINK1 with CRISPR/Cas9), Duchenne muscular dystrophy (knockout of porcine DMD with TALAEs, CRISPR/Cas9 and Cpf1). I will then describe humanizing two porcine genes INS and ALB. Finally, we have generated a Cre-dependent Cas9-expressing pig line where Cre expression induces Cas9 expression from the ROSA26 locus, which enables efficient in vivo genome-editing such as lineage tracing andsimultaneous inactivation of five tumour suppressor genes (TP53, PTEN, APC, BRCA1, and BRCA2) and activation of KRAS. These results show that genome-editing in the pig is efficient and can provide valuable resources for basic and clinical applications.
Dr Xiaoping Li - Staff Scientist - The Wellcome Trust Sanger Institute
CRISPR Genome Editing to Understand Autophagy Gene Function
Use of CRISPR genome editing to assess gene function
Use of CRISPR genome editing to tag endogenous genes
Use of CRISPR genome editing in arrayed screening
Dr Robin Ketteler - Group Lead - UCL
Development of gene editing as a therapy for Duchenne muscular dystrophy (DMD)
Linda Popplewell - Lecturer in Biomedical Sciences - Royal Holloway University of London
Engineering CRISPR Gene Drives for Population Control in the Malaria Mosquito
Andrew Hammond - Post Doc - Imperial College London
CRISPR/Cas9 gene editing to dissect the mechanisms of microRNA regulation and function
MicroRNAs (miRNAs) have emerged as key regulators of cellular function. MiRNA families and miRNA clusters are two key features in miRNA biology. MiRNA families consist of members that share an identical seed region and extensive sequence homology, a property that hampers miRNA research as there is a lack of elegant tools for specific miRNA manipulation. On the other hand, miRNA clusters consist of a set of miRNAs located in close proximity in a genomic locus and transcribed as a single primary transcript. MiRNA clusters are highly conserved indicating an evolutionary pressure to maintain such organization.
Our aim was to explore the use of gene editing for clustered miRNA inhibition. We employed the CRISPR/Cas9 system and determined the effect of gene editing in four miRNA clusters composed of miRNAs belonging to the same or distinct families. Using CRISPR/Cas9 editing we show that (1) miRNA clusters constitute transcriptional units exhibiting strict tertiary requirements. Further processing of the primary transcript depends on intact structural features mainly on the miRNA stem loops. (2) In spite of the sequence diversity of primiRNAs, mutations are well tolerated provided that they don’t disrupt critical elements such as sequence motifs at the terminal loops in hairpins, recognized by downstream processing complexes. (3) MiRNAs in a cluster form co-transcriptional units but their expression is not always interdependent.
In conclusion, CRISPR/Cas9 emerged as a powerful tool to dissect the regulation and function of clustered miRNAs. The technology provides a unique platform to establish mutant cell lines or mouse models and elucidate the specific contribution of miRNA families in cellular responses both at baseline and following injury.
Anna Zampetaki - Lecturer in Cardiovascular Biology - Kings College London
Can genome editing find novel synthetic lethal targets for cancer therapy?
CRISPR-Cas9 has radically increased the accessibility of gene editing and as just a 100 nucleotide RNA is required to guide Cas9 to the target gene, it has proved possible to adapt the infrastructure of short-hairpin RNA interference screen to genome-wide knock out screens. Several proof of principle positive selection screens have identified genes whose knock-out leads to drug or toxin resistance, gain of metastatic potential or altered signalling in immune cells. Furthermore, a small set of papers have attempted to define the “essentialome” – the set of genes required for viability in culture.
The identification of genetic-context-dependent essential genes could open up new avenues to target undruggable cancer driver genes such as KRAS, TP53 and other tumour suppressors. Can novel synthetic lethal targets be identified by CRISPR-Cas9? Horizon has screened some 30 cell lines with an sgRNA library targeting around 3000 genes. My presentation will outline the general landscape of our results and provide a more specific indication of the technology’s ability to recover known genetic interactions or those suggested by RNA interference approaches. I will also discuss our experience with technically-demanding whole genome negative-selection screens where we found 4 kinesins whose knock-out sensitise cells to low doses of paclitaxel.
Jon Moore - CSO - Horizon Discovery