Association of differential expression of immunoregulatory molecules and presence of targetable mutations may inform rational design of clinical trials.

BACKGROUND: Immune checkpoint inhibitors (ICIs) and genomic biomarker-driven targeted therapies have revolutionized the modern oncologic treatment arsenal. The next step has been to combine targeted agents and ICIs. In doing so, some combination regimens may be more logical than others. PATIENTS AND METHODS: Whole-exome and whole-transcriptome sequencing were performed on 2739 unselected later-stage clinical cases from 24 solid tumor subtypes in the NantHealth database, and data were also curated from 5746 similarly sequenced patients across 28 solid tumor subtypes in The Cancer Genome Atlas (TCGA). Significant differential expression of 10 immunoregulatory molecules [IRMs (genes)] was analyzed for association with mutant versus wild-type genes. RESULTS: Twenty-three significant associations between currently actionable variants and RNA-expressed checkpoint genes were identified in the TCGA cases; 10 were validated in the external cohort of 2739 clinical cases from NantHealth (P values were adjusted using Benjamini-Hochberg multiple hypothesis correction to reduce false-discovery rate). Within the same 5746 TCGA profiles, 2740 TCGA patients were identified as having one or more potentially oncogenic single-nucleotide variant (SNV) mutation within an established 50-gene hotspot panel. Of the 50 genes, SNVs within 15 were found to be significantly associated with differential expression of at least one IRM after adjusting for tissue enrichment; six were confirmed significant associations in an independent set of 2739 clinical cases from NantHealth. CONCLUSIONS: Logically combining ICIs with targeted therapies may offer unique treatment strategies for patients with cancer. The presence of specific mutations impacts the expression of IRMs, an observation of potential importance for selecting combinations of gene- and immune-targeted therapeutics.

GCP5 and GCP6: two new members of the human gamma-tubulin complex.

The gamma-tubulin complex is a large multiprotein complex that is required for microtubule nucleation at the centrosome. Here we report the purification and characterization of the human gamma-tubulin complex and the identification of its subunits. The human gamma-tubulin complex is a ring of ~25 nm, has a subunit structure similar to that reported for gamma-tubulin complexes from other species, and is able to nucleate microtubule polymerization in vitro. Mass spectrometry analysis of the human gamma-tubulin complex components confirmed the presence of four previously identified components (gamma-tubulin and gamma-tubulin complex proteins [GCPs] 2, 3, and 4) and led to the identification of two new components, GCP5 and GCP6. Sequence analysis revealed that the GCPs share five regions of sequence similarity and define a novel protein superfamily that is conserved in metazoans. GCP5 and GCP6, like other components of the gamma-tubulin complex, localize to the centrosome and associate with microtubules, suggesting that the entire gamma-tubulin complex takes part in both of these interactions. Stoichiometry experiments revealed that there is a single copy of GCP5 and multiple copies of gamma-tubulin, GCP2, GCP3, and GCP4 within the gamma-tubulin complex. Thus, the gamma-tubulin complex is conserved in structure and function, suggesting that the mechanism of microtubule nucleation is conserved.

Extragenic bypass suppressors of mutations in the essential gene BLD2 promote assembly of basal bodies with abnormal microtubules in Chlamydomonas reinhardtii.

bld2-1 mutant Chlamydomonas reinhardtii strains assemble basal bodies with singlet microtubules; bld2-1 cells display flagellar assembly defects as well as positioning defects of the mitotic spindle and cleavage furrow. To further understand the role of the BLD2 gene, we have isolated three new bld2 alleles and three partially dominant extragenic suppressors, rgn1-1, rgn1-2, and rgn1-3. bld2 rgn1-1 strains have phenotypes intermediate between those of bld2 and wild-type strains with respect to flagellar number, microtubule rootlet organization, cleavage furrow positioning, and basal body structural phenotypes. Instead of the triplet microtubules of wild-type cells, bld2 rgn1-1 basal bodies have mixtures of no, singlet, doublet, and triplet microtubules. The bld2-4 allele was made by insertional mutagenesis and identified in a noncomplementation screen in a diploid strain. The bld2-4 allele has a lethal phenotype based on mitotic segregation in diploid strains and in haploid strains generated by meiotic recombination. The lethal phenotype in haploid strains is suppressed by rgn1-1; these suppressed strains have similar phenotypes to other bld2 rgn1-1 double mutants. It is likely that BLD2 is an essential gene that is needed for basal body assembly and function.