Implementing a comprehensive translational oncology platform: from molecular testing to actionability.

BACKGROUND: In order to establish the workflows required to implement a real-time process involving multi-omic analysis of patient samples to support precision-guided therapeutic intervention, a tissue acquisition and analysis trial was implemented. This report describes our findings to date, including the frequency with which mutational testing led to precision-guided therapy and outcome for those patients. METHODS: Eligible patients presenting to Oregon Health and Science University Knight Cancer Institute were enrolled on the study. Patients with biopsy proven metastatic or locally advanced unresectable prostate cancer, breast cancer, pancreatic adenocarcinoma, or refractory acute myelogenous leukemia receiving standard of care therapy were eligible. Metastatic site biopsies were collected and analyzed using the Knight Diagnostic Lab GeneTrails comprehensive solid tumor panel (124 genes). CLIA certified genomic information was made available to the treating physician. RESULTS: Between 1/26/2017 and 5/30/2018, 38 patients were enrolled, with 28 successfully undergoing biopsy. Of these, 25 samples yielded sufficient tumor for analysis. The median biopsy cellularity and number of cores collected were 70% (15-90%) and 5 (2-20), respectively. No procedure-related complications occurred. GeneTrails analysis revealed that 22 of 25 (88%) tumor samples harbored at least one potentially actionable mutation, and 18 (72%) samples harbored 2 or more potentially actionable mutations. The most common genetic alterations identified involved: DNA damage repair genes, cell cycle regulating genes, PIK3CA/Akt/mTOR pathway, and FGF gene family. To date, CLIA certified genomic results were used by treating physicians for precision-guided therapy in 5 (23%) patients. CONCLUSION: We report the feasibility of real-time tissue acquisition and analysis to support a successful translational oncology platform. The workflow will provide the foundation to improve access and accrual to biomarker driven precision oncology trials.

Exceptional Response to Trastuzumab Deruxtecan in a Patient With Recurrent Ovarian Clear Cell Carcinoma With Human Epidermal Growth Factor Receptor 2 Expression.

Case report of a HER2-expressed ovarian clear cell carcinoma with exceptional response to trastuzumab deruxtecan.

Cyclin E overexpression impairs progression through mitosis by inhibiting APC(Cdh1).

Overexpression of cyclin E, an activator of cyclin-dependent kinase 2, has been linked to human cancer. In cell culture models, the forced expression of cyclin E leads to aneuploidy and polyploidy, which is consistent with a direct role of cyclin E overexpression in tumorigenesis. In this study, we show that the overexpression of cyclin E has a direct effect on progression through the latter stages of mitotic prometaphase before the complete alignment of chromosomes at the metaphase plate. In some cases, such cells fail to divide chromosomes, resulting in polyploidy. In others, cells proceed to anaphase without the complete alignment of chromosomes. These phenotypes can be explained by an ability of overexpressed cyclin E to inhibit residual anaphase-promoting complex (APC(Cdh1)) activity that persists as cells progress up to and through the early stages of mitosis, resulting in the abnormal accumulation of APC(Cdh1) substrates as cells enter mitosis. We further show that the accumulation of securin and cyclin B1 can account for the cyclin E-mediated mitotic phenotype.

An omic and multidimensional spatial atlas from serial biopsies of an evolving metastatic breast cancer.

Mechanisms of therapeutic resistance and vulnerability evolve in metastatic cancers as tumor cells and extrinsic microenvironmental influences change during treatment. To support the development of methods for identifying these mechanisms in individual people, here we present an omic and multidimensional spatial (OMS) atlas generated from four serial biopsies of an individual with metastatic breast cancer during 3.5 years of therapy. This resource links detailed, longitudinal clinical metadata that includes treatment times and doses, anatomic imaging, and blood-based response measurements to clinical and exploratory analyses, which includes comprehensive DNA, RNA, and protein profiles; images of multiplexed immunostaining; and 2- and 3-dimensional scanning electron micrographs. These data report aspects of heterogeneity and evolution of the cancer genome, signaling pathways, immune microenvironment, cellular composition and organization, and ultrastructure. We present illustrative examples of how integrative analyses of these data reveal potential mechanisms of response and resistance and suggest novel therapeutic vulnerabilities.

Characterizing advanced breast cancer heterogeneity and treatment resistance through serial biopsies and comprehensive analytics.

Molecular heterogeneity in metastatic breast cancer presents multiple clinical challenges in accurately characterizing and treating the disease. Current diagnostic approaches offer limited ability to assess heterogeneity that exists among multiple metastatic lesions throughout the treatment course. We developed a precision oncology platform that combines serial biopsies, multi-omic analysis, longitudinal patient monitoring, and molecular tumor boards, with the goal of improving cancer management through enhanced understanding of the entire cancer ecosystem within each patient. We describe this integrative approach using comprehensive analytics generated from serial-biopsied lesions in a metastatic breast cancer patient. The serial biopsies identified remarkable heterogeneity among metastatic lesions that presented clinically as discordance in receptor status and genomic alterations with mixed treatment response. Based on our study, we highlight clinical scenarios, such as rapid progression or mixed response, that indicate consideration for repeat biopsies to evaluate intermetastatic heterogeneity (IMH), with the objective of refining targeted therapy. We present a framework for understanding the clinical significance of heterogeneity in breast cancer between metastatic lesions utilizing multi-omic analyses of serial biopsies and its implication for effective personalized treatment.

Activation of the yeast Hippo pathway by phosphorylation-dependent assembly of signaling complexes.

Scaffold-assisted signaling cascades guide cellular decision-making. In budding yeast, one such signal transduction pathway called the mitotic exit network (MEN) governs the transition from mitosis to the G1 phase of the cell cycle. The MEN is conserved and in metazoans is known as the Hippo tumor-suppressor pathway. We found that signaling through the MEN kinase cascade was mediated by an unusual two-step process. The MEN kinase Cdc15 first phosphorylated the scaffold Nud1. This created a phospho-docking site on Nud1, to which the effector kinase complex Dbf2-Mob1 bound through a phosphoserine-threonine binding domain, in order to be activated by Cdc15. This mechanism of pathway activation has implications for signal transmission through other kinase cascades and might represent a general principle in scaffold-assisted signaling.

Cell cycle phosphorylation of mitotic exit network (MEN) proteins.

Phosphorylation of proteins is an important mechanism used to regulate most cellular processes. Recently, we completed an extensive phosphoproteomic analysis of the core proteins that constitute the Saccharomyces cerevisiae centrosome. Here, we present a study of phosphorylation sites found on the mitotic exit network (MEN) proteins, most of which are associated with the cytoplasmic face of the centrosome. We identified 55 sites on Bfa1, Cdc5, Cdc14 and Cdc15. Eight sites lie in cyclin-dependent kinase motifs (Cdk, S/T-P), and 22 sites are completely conserved within fungi. More than half of the sites were found in centrosomes from mitotic cells, possibly in preparation for their roles in mitotic exit. Finally, we report phosphorylation site information for other important cell cycle and regulatory proteins.

A cell cycle phosphoproteome of the yeast centrosome.

Centrosomes organize the bipolar mitotic spindle, and centrosomal defects cause chromosome instability. Protein phosphorylation modulates centrosome function, and we provide a comprehensive map of phosphorylation on intact yeast centrosomes (18 proteins). Mass spectrometry was used to identify 297 phosphorylation sites on centrosomes from different cell cycle stages. We observed different modes of phosphoregulation via specific protein kinases, phosphorylation site clustering, and conserved phosphorylated residues. Mutating all eight cyclin-dependent kinase (Cdk)-directed sites within the core component, Spc42, resulted in lethality and reduced centrosomal assembly. Alternatively, mutation of one conserved Cdk site within γ-tubulin (Tub4-S360D) caused mitotic delay and aberrant anaphase spindle elongation. Our work establishes the extent and complexity of this prominent posttranslational modification in centrosome biology and provides specific examples of phosphorylation control in centrosome function.