Oncogenic Drivers and Therapeutic Vulnerabilities in KRAS Wild-Type Pancreatic Cancer.

PURPOSE: Approximately 8% to 10% of pancreatic ductal adenocarcinomas (PDAC) do not harbor mutations in KRAS. Understanding the unique molecular and clinical features of this subset of pancreatic cancer is important to guide patient stratification for clinical trials of molecularly targeted agents. EXPERIMENTAL DESIGN: We analyzed a single-institution cohort of 795 exocrine pancreatic cancer cases (including 785 PDAC cases) with a targeted multigene sequencing panel and identified 73 patients (9.2%) with KRAS wild-type (WT) pancreatic cancer. RESULTS: Overall, 43.8% (32/73) of KRAS WT cases had evidence of an alternative driver of the MAPK pathway, including BRAF mutations and in-frame deletions and receptor tyrosine kinase fusions. Conversely, 56.2% of cases did not harbor a clear MAPK driver alteration, but 29.3% of these MAPK-negative KRAS WT cases (12/41) demonstrated activating alterations in other oncogenic drivers, such as GNAS, MYC, PIK3CA, and CTNNB1. We demonstrate potent efficacy of pan-RAF and MEK inhibition in patient-derived organoid models carrying BRAF in-frame deletions. Moreover, we demonstrate durable clinical benefit of targeted therapy in a patient harboring a KRAS WT tumor with a ROS1 fusion. Clinically, patients with KRAS WT tumors were significantly younger in age of onset (median age: 62.6 vs. 65.7 years; P = 0.037). SMAD4 mutations were associated with a particularly poor prognosis in KRAS WT cases. CONCLUSIONS: This study defines the genomic underpinnings of KRAS WT pancreatic cancer and highlights potential therapeutic avenues for future investigation in molecularly directed clinical trials. See related commentary by Kato et al., p. 4527.

Single-cell molecular profiling using ex vivo functional readouts fuels precision oncology in glioblastoma.

BACKGROUND: Functional profiling of freshly isolated glioblastoma (GBM) cells is being evaluated as a next-generation method for precision oncology. While promising, its success largely depends on the method to evaluate treatment activity which requires sufficient resolution and specificity. METHODS: Here, we describe the 'precision oncology by single-cell profiling using ex vivo readouts of functionality' (PROSPERO) assay to evaluate the intrinsic susceptibility of high-grade brain tumor cells to respond to therapy. Different from other assays, PROSPERO extends beyond life/death screening by rapidly evaluating acute molecular drug responses at single-cell resolution. RESULTS: The PROSPERO assay was developed by correlating short-term single-cell molecular signatures using mass cytometry by time-of-flight (CyTOF) to long-term cytotoxicity readouts in representative patient-derived glioblastoma cell cultures (n = 14) that were exposed to radiotherapy and the small-molecule p53/MDM2 inhibitor AMG232. The predictive model was subsequently projected to evaluate drug activity in freshly resected GBM samples from patients (n = 34). Here, PROSPERO revealed an overall limited capacity of tumor cells to respond to therapy, as reflected by the inability to induce key molecular markers upon ex vivo treatment exposure, while retaining proliferative capacity, insights that were validated in patient-derived xenograft (PDX) models. This approach also allowed the investigation of cellular plasticity, which in PDCLs highlighted therapy-induced proneural-to-mesenchymal (PMT) transitions, while in patients' samples this was more heterogeneous. CONCLUSION: PROSPERO provides a precise way to evaluate therapy efficacy by measuring molecular drug responses using specific biomarker changes in freshly resected brain tumor samples, in addition to providing key functional insights in cellular behavior, which may ultimately complement standard, clinical biomarker evaluations.

ALK Amplification and Rearrangements Are Recurrent Targetable Events in Congenital and Adult Glioblastoma.

PURPOSE: Anaplastic lymphoma kinase (ALK) aberrations have been identified in pediatric-type infant gliomas, but their occurrence across age groups, functional effects, and treatment response has not been broadly established. EXPERIMENTAL DESIGN: We performed a comprehensive analysis of ALK expression and genomic aberrations in both newly generated and retrospective data from 371 glioblastomas (156 adult, 205 infant/pediatric, and 10 congenital) with in vitro and in vivo validation of aberrations. RESULTS: ALK aberrations at the protein or genomic level were detected in 12% of gliomas (45/371) in a wide age range (0-80 years). Recurrent as well as novel ALK fusions (LRRFIP1-ALK, DCTN1-ALK, PRKD3-ALK) were present in 50% (5/10) of congenital/infant, 1.4% (3/205) of pediatric, and 1.9% (3/156) of adult GBMs. ALK fusions were present as the only candidate driver in congenital/infant GBMs and were sometimes focally amplified. In contrast, adult ALK fusions co-occurred with other oncogenic drivers. No activating ALK mutations were identified in any age group. Novel and recurrent ALK rearrangements promoted STAT3 and ERK1/2 pathways and transformation in vitro and in vivo. ALK-fused GBM cellular and mouse models were responsive to ALK inhibitors, including in patient cells derived from a congenital GBM. Relevant to the treatment of infant gliomas, we showed that ALK protein appears minimally expressed in the forebrain at perinatal stages, and no gross effects on perinatal brain development were seen in pregnant mice treated with the ALK inhibitor ceritinib. CONCLUSIONS: These findings support use of brain-penetrant ALK inhibitors in clinical trials across infant, pediatric, and adult GBMs. See related commentary by Mack and Bertrand, p. 2567.

Genomic and biological study of fusion genes as resistance mechanisms to EGFR inhibitors.

The clinical significance of gene fusions detected by DNA-based next generation sequencing remains unclear as resistance mechanisms to EGFR tyrosine kinase inhibitors in EGFR mutant non-small cell lung cancer. By studying EGFR inhibitor-resistant patients treated with a combination of an EGFR inhibitor and a drug targeting the putative resistance-causing fusion oncogene, we identify patients who benefit and those who do not from this treatment approach. Through evaluation including RNA-seq of potential drug resistance-imparting fusion oncogenes in 504 patients with EGFR mutant lung cancer, we identify only a minority of them as functional, potentially capable of imparting EGFR inhibitor resistance. We further functionally validate fusion oncogenes in vitro using CRISPR-based editing of EGFR mutant cell lines and use these models to identify known and unknown drug resistance mechanisms to combination therapies. Collectively, our results partially reveal the complex nature of fusion oncogenes as potential drug resistance mechanisms and highlight approaches that can be undertaken to determine their functional significance.

Microenvironment drives cell state, plasticity, and drug response in pancreatic cancer.

Prognostically relevant RNA expression states exist in pancreatic ductal adenocarcinoma (PDAC), but our understanding of their drivers, stability, and relationship to therapeutic response is limited. To examine these attributes systematically, we profiled metastatic biopsies and matched organoid models at single-cell resolution. In vivo, we identify a new intermediate PDAC transcriptional cell state and uncover distinct site- and state-specific tumor microenvironments (TMEs). Benchmarking models against this reference map, we reveal strong culture-specific biases in cancer cell transcriptional state representation driven by altered TME signals. We restore expression state heterogeneity by adding back in vivo-relevant factors and show plasticity in culture models. Further, we prove that non-genetic modulation of cell state can strongly influence drug responses, uncovering state-specific vulnerabilities. This work provides a broadly applicable framework for aligning cell states across in vivo and ex vivo settings, identifying drivers of transcriptional plasticity and manipulating cell state to target associated vulnerabilities.

BRCA1/Trp53 heterozygosity and replication stress drive esophageal cancer development in a mouse model.

BRCA1 germline mutations are associated with an increased risk of breast and ovarian cancer. Recent findings of others suggest that BRCA1 mutation carriers also bear an increased risk of esophageal and gastric cancer. Here, we employ a Brca1/Trp53 mouse model to show that unresolved replication stress (RS) in BRCA1 heterozygous cells drives esophageal tumorigenesis in a model of the human equivalent. This model employs 4-nitroquinoline-1-oxide (4NQO) as an RS-inducing agent. Upon drinking 4NQO-containing water, Brca1 heterozygous mice formed squamous cell carcinomas of the distal esophagus and forestomach at a much higher frequency and speed (∼90 to 120 d) than did wild-type (WT) mice, which remained largely tumor free. Their esophageal tissue, but not that of WT control mice, revealed evidence of overt RS as reflected by intracellular CHK1 phosphorylation and 53BP1 staining. These Brca1 mutant tumors also revealed higher genome mutation rates than those of control animals; the mutational signature SBS4, which is associated with tobacco-induced tumorigenesis; and a loss of Brca1 heterozygosity (LOH). This uniquely accelerated Brca1 tumor model is also relevant to human esophageal squamous cell carcinoma, an often lethal tumor.

Debate: Limitations on universality: the "right to health" and the necessity of legal nationality.

BACKGROUND: The "right to health," including access to basic healthcare, has been recognized as a universal human right through a number of international agreements. Attempts to protect this ideal, however, have relied on states as the guarantor of rights and have subsequently ignored stateless individuals, or those lacking legal nationality in any nation-state. While a legal nationality alone is not sufficient to guarantee that a right to healthcare is accessible, an absence of any legal nationality is almost certainly an obstacle in most cases. There are millions of so-called stateless individuals around the globe who are, in effect, denied medical citizenship in their countries of residence. A central motivating factor for this essay is the fact that statelessness as a concept is largely absent from the medical literature. The goal for this discussion, therefore, is primarily to illustrate the need for further monitoring of health access issues by the medical community, and for a great deal more research into the effects of statelessness upon access to healthcare. This is important both as a theoretical issue, in light of the recognition by many of healthcare as a universal right, as well as an empirical fact that requires further exploration and amelioration. DISCUSSION: Most discussions of the human right to health assume that every human being has legal nationality, but in reality there are at least 11 to 12 million stateless individuals worldwide who are often unable to access basic healthcare. The examples of the Roma in Europe, the hill tribes of Thailand, and many Palestinians in Israel highlight the negative health impacts associated with statelessness. SUMMARY: Stateless individuals often face an inability to access the most basic healthcare, much less the "highest attainable standard of health" outlined by international agreements. Rather than presuming nationality, statelessness must be recognized by the medical community. Additionally, it is imperative that stateless populations be recognized, the health of these populations be tracked, and more research conducted to further elaborate upon the connection between statelessness and access to healthcare services, and hence a universal right to health.

Children, ADHD, and citizenship.

The diagnosis of attention-deficit hyperactivity disorder is a subject of controversy, for a host of reasons. This paper seeks to explore the manner in which children's interests may be subsumed to those of parents, teachers, and society as a whole in the course of diagnosis, treatment, and labeling, utilizing a framework for children's citizenship proposed by Elizabeth Cohen. Additionally, the paper explores aspects of discipline associated with the diagnosis, as well as distributional pathologies resulting from the application of the diagnosis in potentially biased ways.