Abstract PR02: Mutations in RABL3 alter KRAS prenylation and are associated with hereditary pancreatic cancer
Authors: Nissim Sahar , Leshchiner Ignaty , Mancias Joseph , Greenblatt Matthew , Maertens Ophélia , Cassa Christopher , Rosenfeld Jill , Cox Andrew , Hedgepeth John , Wucherpfennig Julia , Kim Andrew , Henderson Jake , Gonyo Patrick , Brandt Anthony , Lorimer Ellen , Unger Bethany , Getz Gad , Sunyaev Shamil , Harper Wade , Cichowski Karen , Kimmelman Alec , Houvras Yariv , Syngal Sapna , Williams Carol , Goessling Wolfram 
Published: June 2021

Abstract
Beginning with discovery of a new genetic cause of familial pancreatic ductal adenocarcinoma (PDAC), this work uncovers a mechanism for regulating KRAS intracellular trafficking with implications for hereditary cancer, sporadic cancer, and RASopathy syndromes. PDAC is one of the deadliest solid cancers, with limited treatment options despite intensive research efforts. Familial predisposition to PDAC is thought to occur in ~10% of cases, but causative genes have not been identified in most of these families. Uncovering the genetic basis for PDAC susceptibility has immediate prognostic implications for families and can provide precious mechanistic clues to PDAC pathogenesis. Here, we perform whole-genome sequence analysis in a family with five cases of PDAC and identify a germline nonsense mutation in the member of RAS oncogene family-like 3 (RABL3) gene that has never before been directly associated with hereditary cancer. The truncated mutant allele (RABL3_p.Ser36*) co-segregates with cancer occurrence. To evaluate the contribution of the RABL3 mutant allele in hereditary cancer, we generated rabl3 heterozygous mutant zebrafish and found increased susceptibility to cancer formation in two independent cancer models. Complementary unbiased approaches implicate RABL3 in RAS pathway regulation. RNA-Seq and genome-set enrichment analysis of juvenile rabl3 mutants reveals a KRAS upregulation signature. Furthermore, affinity-purification mass spectrometry for proteins associated with RABL3 or RABL3_p.Ser36* identifies Rap1 GTPase-GDP Dissociation Stimulator 1 (RAP1GDS1, SmgGDS), a chaperone that regulates prenylation of RAS GTPases. Indeed, in vitro studies demonstrate that RABL3_p.Ser36* accelerates KRAS prenylation, and this impact is lost in the absence of H/N/KRAS proteins. Whereas heterozygous rabl3 mutant zebrafish exhibit cancer predisposition, homozygous rabl3 mutant zebrafish develop severe craniofacial, skeletal, and growth defects consistent with human RASopathies, and these defects are partially rescued with the MEK inhibitor trametinib. Finally, we identify additional germline mutations in RABL3 that impact RAS activity in vivo and have a significant burden in a cohort of patients with developmental disorders, suggesting a role in undiagnosed RASopathies. Moreover, RABL3 is upregulated in multiple human PDAC cell lines and knockdown abrogates proliferation, consistent with a broader role for RABL3 in PDAC. The discovered causative RABL3 germline mutation provides new diagnostic opportunities for genetic testing in other cancer families and uncovers an alternative mechanism for dysregulated RAS signaling in development and cancer.
This abstract is also being presented as Poster A12.
Citation Format: Sahar Nissim, Ignaty Leshchiner, Joseph Mancias, Matthew Greenblatt, Ophélia Maertens, Christopher Cassa, Jill Rosenfeld, Andrew Cox, John Hedgepeth, Julia Wucherpfennig, Andrew Kim, Jake Henderson, Patrick Gonyo, Anthony Brandt, Ellen Lorimer, Bethany Unger, Gad Getz, Shamil Sunyaev, Wade Harper, Karen Cichowski, Alec Kimmelman, Yariv Houvras, Sapna Syngal, Carol Williams, Wolfram Goessling. Mutations in RABL3 alter KRAS prenylation and are associated with hereditary pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference on Targeting RAS-Driven Cancers; 2018 Dec 9-12; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(5_Suppl):Abstract nr PR02.