The gene for spinocerebellar ataxia type 2 (SCA2) has been mapped to 12q24.1. A 1.1-megabase contig in the candidate region was assembled in P1 artificial chromosome and bacterial artificial chromosome clones. Using this contig, we identified a CAG trinucleotide repeat with CAA interruptions that was expanded in patients with SCA2. In contrast to other unstable trinucleotide repeats, this CAG repeat was not highly polymorphic in normal individuals. In SCA2 patients, the repeat was perfect and expanded to 36-52 repeats. The most common disease allele contained (CAG)37, one of the shortest expansions seen in a CAG expansion syndrome. The repeat occurs in the 5'-coding region of SCA2 which is a member of a novel gene family.

To study mechanisms underlying resistance to the BCL2 inhibitor venetoclax in acute myeloid leukemia (AML), we used a genome-wide CRISPR/Cas9 screen to identify gene knockouts resulting in drug resistance. We validated TP53 , BAX , and PMAIP1 as genes whose inactivation results in venetoclax resistance in AML cell lines. Resistance to venetoclax resulted from an inability to execute apoptosis driven by BAX loss, decreased expression of BCL2, and/or reliance on alternative BCL2 family members such as BCL2L1. The resistance was accompanied by changes in mitochondrial homeostasis and cellular metabolism. Evaluation of TP53 knockout cells for sensitivities to a panel of small-molecule inhibitors revealed a gain of sensitivity to TRK inhibitors. We relate these observations to patient drug responses and gene expression in the Beat AML dataset. Our results implicate TP53 , the apoptotic network, and mitochondrial functionality as drivers of venetoclax response in AML and suggest strategies to overcome resistance. SIGNIFICANCE: AML is challenging to treat due to its heterogeneity, and single-agent therapies have universally failed, prompting a need for innovative drug combinations. We used a genetic approach to identify genes whose inactivation contributes to drug resistance as a means of forming preferred drug combinations to improve AML treatment.

Epithelial tubes represent fundamental building blocks of metazoan organisms; however, the mechanisms responsible for their formation and maintenance are not well understood. Here, we show that the evolutionarily conserved coiled-coil MAGUK protein Dlg5 is required for epithelial tube maintenance in mammalian brain and kidneys. We demonstrate that Dlg5(-/-) mice develop fully penetrant hydrocephalus and kidney cysts caused by a deficiency in membrane delivery of cadherin-catenin adhesion complexes and loss of cell polarity. Dlg5 travels with cadherin-containing vesicles and binds to syntaxin 4, a t-SNARE protein that regulates fusion of transport vesicles with the lateral membrane domain. We propose that Dlg5 functions in plasma membrane delivery of cadherins by linking cadherin-containing transport vesicles with the t-SNARE targeting complex. These findings show that Dlg5 is causally involved in hydrocephalus and renal cysts and reveal that targeted membrane delivery of cadherin-catenin adhesion complexes is critical for cell polarity and epithelial tube maintenance.

FLT3 mutations are prevalent in AML patients and confer poor prognosis. Crenolanib, a potent type I pan-FLT3 inhibitor, is effective against both internal tandem duplications and resistance-conferring tyrosine kinase domain mutations. While crenolanib monotherapy has demonstrated clinical benefit in heavily pretreated relapsed/refractory AML patients, responses are transient and relapse eventually occurs. Here, to investigate the mechanisms of crenolanib resistance, we perform whole exome sequencing of AML patient samples before and after crenolanib treatment. Unlike other FLT3 inhibitors, crenolanib does not induce FLT3 secondary mutations, and mutations of the FLT3 gatekeeper residue are infrequent. Instead, mutations of NRAS and IDH2 arise, mostly as FLT3-independent subclones, while TET2 and IDH1 predominantly co-occur with FLT3-mutant clones and are enriched in crenolanib poor-responders. The remaining patients exhibit post-crenolanib expansion of mutations associated with epigenetic regulators, transcription factors, and cohesion factors, suggesting diverse genetic/epigenetic mechanisms of crenolanib resistance. Drug combinations in experimental models restore crenolanib sensitivity.

The characterization of cancer genomes has provided insight into somatically altered genes across tumors, transformed our understanding of cancer biology, and enabled tailoring of therapeutic strategies. However, the function of most cancer alleles remains mysterious, and many cancer features transcend their genomes. Consequently, tumor genomic characterization does not influence therapy for most patients. Approaches to understand the function and circuitry of cancer genes provide complementary approaches to elucidate both oncogene and non-oncogene dependencies. Emerging work indicates that the diversity of therapeutic targets engendered by non-oncogene dependencies is much larger than the list of recurrently mutated genes. Here we describe a framework for this expanded list of cancer targets, providing novel opportunities for clinical translation.