Acquired Resistance to KRAS (G12C) Inhibition in Cancer

BACKGROUND: Clinical trials of the KRAS inhibitors adagrasib and sotorasib have shown promising activity in cancers harboring KRAS glycine-to-cysteine amino acid substitutions at codon 12 (KRAS(G12C)). The mechanisms of acquired resistance to these therapies are currently unknown. METHODS: Among patients with KRAS(G12C) -mutant cancers treated with adagrasib monotherapy, we performed genomic and histologic analyses that compared pretreatment samples with those obtained after the development of resistance. Cell-based experiments were conducted to study mutations that confer resistance to KRAS(G12C) inhibitors. RESULTS: A total of 38 patients were included in this study: 27 with non-small-cell lung cancer, 10 with colorectal cancer, and 1 with appendiceal cancer. Putative mechanisms of resistance to adagrasib were detected in 17 patients (45% of the cohort), of whom 7 (18% of the cohort) had multiple coincident mechanisms. Acquired KRAS alterations included G12D/R/V/W, G13D, Q61H, R68S, H95D/Q/R, Y96C, and high-level amplification of the KRAS(G12C) allele. Acquired bypass mechanisms of resistance included MET amplification; activating mutations in NRAS, BRAF, MAP2K1, and RET; oncogenic fusions involving ALK, RET, BRAF, RAF1, and FGFR3; and loss-of-function mutations in NF1 and PTEN. In two of nine patients with lung adenocarcinoma for whom paired tissue-biopsy samples were available, histologic transformation to squamous-cell carcinoma was observed without identification of any other resistance mechanisms. Using an in vitro deep mutational scanning screen, we systematically defined the landscape of KRAS mutations that confer resistance to KRAS(G12C) inhibitors. CONCLUSIONS: Diverse genomic and histologic mechanisms impart resistance to covalent KRAS(G12C) inhibitors, and new therapeutic strategies are required to delay and overcome this drug resistance in patients with cancer. (Funded by Mirati Therapeutics and others; ClinicalTrials.gov number, NCT03785249.)

DU Undergraduate Showcase: Research, Scholarship, and Creative Works: Abstracts

Abstracts from the DU Undergraduate Showcase

Pubertal Effects on Executive Functioning Among Autistic and Non-Autistic Youth: A Cross Sectional Study

Investigating the role of puberty on executive function (EF) development is important for understanding how maturation and its related changes can impact neural systems underlying EF in autistic (AT) adolescents. Studies document chronological age-related differences in EF among AT youth, but the impact of puberty is understudied. We examined the role of cross-sectional pubertal status (Pubertal Development Scale adrenal and gonadal indices), autism status, and assigned sex at birth on parent-reported EF (Behavior Rating Inventory of Executive Function: inhibition, shift, working memory) in AT and non-autistic (NA) youth. We hypothesized AT youth, particularly females, would show more EF problems in late puberty relative to NA youth. AT males and females and NA females in late puberty had fewer shift problems relative to pre-pubertal youth, whereas NA males in late puberty had more shift problems relative to pre-pubertal NA males. There were no significant assigned sex or puberty differences in working memory for adrenal development and in any EF domain for gonadal development. Findings suggest adrenal puberty contributes to flexibility and may promote certain EF domains in AT youth. Longitudinal research using diverse measures of puberty and EF is needed to clarify findings

Genome-wide pooled CRISPR screening in neurospheres

Spheroid culture systems have allowed in vitro propagation of cells unable to grow in canonical cell culturing conditions, and may capture cellular contexts that model tumor growth better than current model systems. The insights gleaned from genome-wide clustered regularly interspaced short palindromic repeat (CRISPR) screening of thousands of cancer cell lines grown in conventional culture conditions illustrate the value of such CRISPR pooled screens. It is clear that similar genome-wide CRISPR screens of three-dimensional spheroid cultures will be important for future biological discovery. Here, we present a protocol for genome-wide CRISPR screening of three-dimensional neurospheres. While many in-depth protocols and discussions have been published for more typical cell lines, few detailed protocols are currently available in the literature for genome-wide screening in spheroidal cell lines. For those who want to screen such cell lines, and particularly neurospheres, we provide a step-by-step description of assay development tests to be performed before screening, as well as for the screen itself. We highlight considerations of variables that make these screens distinct from, or similar to, typical nonspheroid cell lines throughout. Finally, we illustrate typical outcomes of neurosphere genome-wide screens, and how neurosphere screens typically produce slightly more heterogeneous signal distributions than more canonical cancer cell lines. Completion of this entire protocol will take 8–12 weeks from the initial assay development tests to deconvolution of the sequencing data