Biology subjects

Clifford, S. C.

Publications and source records attributed to Clifford, S. C..

3 recordsLinked to original sources

bioRxiv2024

Structural mimicry of UM171 and neomorphic cancer mutants co-opts E3 ligase KBTBD4 for HDAC1/2 recruitment

Neomorphic mutations and drugs can elicit unanticipated effects that hinder mechanistic understanding for clinical practice. Recurrent indel mutations in the Kelch domain of the KBTBD4 E3 ligase rewire epigenetic programs for stemness in medulloblastoma by recruiting LSD1-CoREST-HDAC1/2 complexes as neo-substrates for ubiquitination and degradation. Remarkably, UM171, an investigational drug for haematopoietic stem cell transplantation, was found to degrade LSD1-CoREST-HDAC1/2 complexes in a wild-type KBTBD4-dependent manner, suggesting a potential common mode of action. We identified that these neomorphic interactions were mediated by the HDAC deacetylase domain. Cryo-EM studies of both wild-type and mutant KBTBD4 captured 2:1 and 2:2 KBTBD4-HDAC2 complexes at resolutions spanning 2.7 [A] to 3.1 [A]. The mutant and drug-induced complexes adopted similar structural assemblies requiring both Kelch domains in the KBTBD4 dimer for each HDAC2 interaction. UM171 was identified as a bona fide molecular glue binding across the ternary interface. Most strikingly, the indel mutation reshaped the same surface of KBTBD4 providing the first example of a natural mimic of a molecular glue. Together, the structures provide mechanistic understanding of neomorphic KBTBD4 and help to explain the structure-activity relationships of UM171 derivatives for future drug design.

molecular biology↗

bioRxiv2022

Medulloblastoma Group 3 and 4 Tumors Comprise a Clinically and Biologically Significant Expression Continuum Reflecting Human Cerebellar Development

Medulloblastoma is currently sub-classified into distinct DNA methylation subgroups/subtypes with particular clinico-molecular features. Using RNA-seq in large well annotated cohorts of medulloblastoma we show that transcriptionally Group3 and Group4 medulloblastomas exist not as discrete types but as intermediates on a bipolar continuum between archetypal Group3 and Group4 entities. Continuum position is prognostic, reflects propensity for specific DNA copy-number changes, key switches in isoform/enhancer usage and RNA-editing. Examining scRNA-seq profiles we show intra-tumoral transcriptional heterogeneity along the continuum is limited in a subtype-dependent manner. By integrating with a human scRNA-seq reference atlas we show this continuum is mirrored by an equivalent continuum of transcriptional cell types in early fetal cerebellar development. We identify unique developmental niches for all four major subgroups and link each to a common developmental antecedent. Our findings show a transcriptional continuum arising from oncogenic disruption of highly specific fetal cerebellar cell types, linked to almost every aspect of Group3/Group4 molecular biology and clinico-pathology.

cancer biology↗

bioRxiv2017

Development of a targeted sequencing approach to identify prognostic, predictive and diagnostic markers in paediatric solid tumours

The implementation of personalised medicine in childhood cancers has been limited by a lack of clinically validated multi-target sequencing approaches specific for paediatric solid tumours. In order to support innovative clinical trials in high-risk patients with unmet need, we have developed a clinically relevant targeted sequencing panel spanning 311 kb and comprising 78 genes involved in childhood cancers. A total of 132 samples were used for the validation of the panel, including Horizon Discovery cell blends (n=4), cell lines (n=15), formalin-fixed paraffin embedded (FFPE, n=83) and fresh frozen tissue (FF, n=30) patient samples. Cell blends containing known single nucleotide variants (SNVs, n=528) and small insertion-deletions (indels n=108) were used to define panel sensitivities of [≥]98% for SNVs and [≥]83% for indels [95% CI] and panel specificity of [≥]98% [95% CI] for SNVs. FFPE samples performed comparably to FF samples (n=15 paired). Of 95 well-characterised genetic abnormalities in 33 clinical specimens and 13 cell lines (including SNVs, indels, amplifications, rearrangements and chromosome losses), 94 (98.9%) were detected by our approach. We have validated a robust and practical methodology to guide clinical management of children with solid tumours based on their molecular profiles. Our work demonstrates the value of targeted gene sequencing in the development of precision medicine strategies in paediatric oncology.

genomics↗