Pancreatic Cancer Patient-derived Organoids Can Predict Response to Neoadjuvant Chemotherapy.

OBJECTIVE: To evaluate if patient-derived organoids (PDOs) may predict response to neoadjuvant (NAT) chemotherapy in patients with pancreatic adenocarcinoma. BACKGROUND: PDOs have been explored as a biomarker of therapy response and for personalized therapeutics in patients with pancreatic cancer. METHODS: During 2017-2021, patients were enrolled into an IRB-approved protocol and PDO cultures were established. PDOs of interest were analyzed through a translational pipeline incorporating molecular profiling and drug sensitivity testing. RESULTS: One hundred thirty-six samples, including both surgical resections and fine needle aspiration/biopsy from 117 patients with pancreatic cancer were collected. This biobank included diversity in stage, sex, age, and race, with minority populations representing 1/3 of collected cases (16% Black, 9% Asian, 7% Hispanic/Latino). Among surgical specimens, PDO generation was successful in 71% (15 of 21) of patients who had received NAT prior to sample collection and in 76% (39 of 51) of patients who were untreated with chemotherapy or radiation at the time of collection. Pathological response to NAT correlated with PDO chemotherapy response, particularly oxaliplatin. We demonstrated the feasibility of a rapid PDO drug screen and generated data within 7 days of tissue resection. CONCLUSION: Herein we report a large single-institution organoid biobank, including ethnic minority samples. The ability to establish PDOs from chemotherapy-naive and post-NAT tissue enables longitudinal PDO generation to assess dynamic chemotherapy sensitivity profiling. PDOs can be rapidly screened and further development of rapid screening may aid in the initial stratification of patients to the most active NAT regimen.

Multi-disciplinary summit on genetics services for women with gynecologic cancers: A Society of Gynecologic Oncology White Paper.

OBJECTIVE: To assess current practice, advise minimum standards, and identify educational gaps relevant to genetic screening, counseling, and testing of women affected by gynecologic cancers. METHODS: The Society of Gynecologic Oncology (SGO) organized a multidisciplinary summit that included representatives from the American College of Obstetricians and Gynecologists (ACOG), the American Society Clinical Oncology (ASCO), the National Society of Genetic Counselors (NSGC), and patient advocacy groups, BrightPink and Facing our Risk of Cancer Empowered (FORCE). Three subject areas were discussed: care delivery models for genetic testing, barriers to genetic testing, and educational opportunities for providers of genetic testing. RESULTS: The group endorsed current SGO, National Comprehensive Cancer Network (NCCN), and NSGC genetic testing guidelines for women affected with ovarian, tubal, peritoneal cancers, or DNA mismatch repair deficient endometrial cancer. Three main areas of unmet need were identified: timely and universal genetic testing for women with ovarian, fallopian tube, and peritoneal cancers; education regarding minimum standards for genetic counseling and testing; and barriers to implementation of testing of both affected individuals as well as cascade testing of family members. Consensus building among all stakeholders resulted in an action plan to address gaps in education of gynecologic oncology providers and delivery of cancer genetics care.

Interplay between oxygen and Fe-S cluster biogenesis: insights from the Suf pathway.

Iron-sulfur (Fe-S) cluster metalloproteins conduct essential functions in nearly all contemporary forms of life. The nearly ubiquitous presence of Fe-S clusters and the fundamental requirement for Fe-S clusters in both aerobic and anaerobic Archaea, Bacteria, and Eukarya suggest that these clusters were likely integrated into central metabolic pathways early in the evolution of life prior to the widespread oxidation of Earth's atmosphere. Intriguingly, Fe-S cluster-dependent metabolism is sensitive to disruption by oxygen because of the decreased bioavailability of ferric iron as well as direct oxidation of sulfur trafficking intermediates and Fe-S clusters by reactive oxygen species. This fact, coupled with the ubiquity of Fe-S clusters in aerobic organisms, suggests that organisms evolved with mechanisms that facilitate the biogenesis and use of these essential cofactors in the presence of oxygen, which gradually began to accumulate around 2.5 billion years ago as oxygenic photosynthesis proliferated and reduced minerals that buffered against oxidation were depleted. This review highlights the most ancient of the Fe-S cluster biogenesis pathways, the Suf system, which likely was present in early anaerobic forms of life. Herein, we use the evolution of the Suf pathway to assess the relationships between the biochemical functions and physiological roles of Suf proteins, with an emphasis on the selective pressure of oxygen toxicity. Our analysis suggests that diversification into oxygen-containing environments disrupted iron and sulfur metabolism and was a main driving force in the acquisition of accessory Suf proteins (such as SufD, SufE, and SufS) by the core SufB-SufC scaffold complex. This analysis provides a new framework for the study of Fe-S cluster biogenesis pathways and Fe-S cluster-containing metalloenzymes and their complicated patterns of divergence in response to oxygen.

Resistance to therapy caused by intragenic deletion in BRCA2.

Cells with loss of BRCA2 function are defective in homologous recombination (HR) and are highly sensitive to inhibitors of poly(ADP-ribose) polymerase (PARP), which provides the basis for a new therapeutic approach. Here we show that resistance to PARP inhibition can be acquired by deletion of a mutation in BRCA2. We derived PARP-inhibitor-resistant (PIR) clones from the human CAPAN1 pancreatic cancer cell line, which carries the protein-truncating c.6174delT frameshift mutation. PIR clones could form DNA-damage-induced RAD51 nuclear foci and were able to limit genotoxin-induced genomic instability, both hallmarks of a competent HR pathway. New BRCA2 isoforms were expressed in the resistant lines as a result of intragenic deletion of the c.6174delT mutation and restoration of the open reading frame (ORF). Reconstitution of BRCA2-deficient cells with these revertant BRCA2 alleles rescued PARP inhibitor sensitivity and HR deficiency. Most of the deletions in BRCA2 were associated with small tracts of homology, and possibly arose from error-prone repair caused by BRCA2 deficiency. Similar ORF-restoring mutations were present in carboplatin-resistant ovarian tumours from c.6174delT mutation carriers. These observations have implications for understanding drug resistance in BRCA mutation carriers as well as in defining functionally important domains within BRCA2.

Analysis of autoinducer-2 quorum sensing in Yersinia pestis.

The autoinducer-2 (AI-2) quorum-sensing system has been linked to diverse phenotypes and regulatory changes in pathogenic bacteria. In the present study, we performed a molecular and biochemical characterization of the AI-2 system in Yersinia pestis, the causative agent of plague. In strain CO92, the AI-2 signal is produced in a luxS-dependent manner, reaching maximal levels of 2.5 µM in the late logarithmic growth phase, and both wild-type and pigmentation (pgm) mutant strains made equivalent levels of AI-2. Strain CO92 possesses a chromosomal lsr locus encoding factors involved in the binding and import of AI-2, and confirming this assignment, an lsr deletion mutant increased extracellular pools of AI-2. To assess the functional role of AI-2 sensing in Y. pestis, microarray studies were conducted by comparing Δpgm strain R88 to a Δpgm ΔluxS mutant or a quorum-sensing-null Δpgm ΔypeIR ΔyspIR ΔluxS mutant at 37°C. Our data suggest that AI-2 quorum sensing is associated with metabolic activities and oxidative stress genes that may help Y. pestis survive at the host temperature. This was confirmed by observing that the luxS mutant was more sensitive to killing by hydrogen peroxide, suggesting a potential requirement for AI-2 in evasion of oxidative damage. We also show that a large number of membrane protein genes are controlled by LuxS, suggesting a role for quorum sensing in membrane modeling. Altogether, this study provides the first global analysis of AI-2 signaling in Y. pestis and identifies potential roles for the system in controlling genes important to disease.

Whole exome sequence analysis of serous borderline tumors of the ovary.

OBJECTIVE: Serous borderline tumor (SBT) is a unique histopathologic entity of the ovary, believed to be intermediate between benign cystadenoma and invasive low-grade serous carcinoma. While somatic mutations in the KRAS or BRAF, and rarely ERBB2, genes have been well characterized in SBTs, other genetic alterations have not been described. Toward a more comprehensive understanding of the molecular genetic architecture of SBTs, we undertook whole exome sequencing of this tumor type. METHODS: Following pathologic review and laser capture microdissection to enrich for tumor cells, whole exomes were prepared from DNA of two independent SBTs and subjected to massively parallel DNA sequencing. RESULTS: Both tumors contained an activating mutation of the BRAF gene. A total of 15 additional somatic mutations were identified, nine in one tumor and six in the other. Eleven were missense mutations and four were nonsense or deletion mutations. Fourteen of the 16 genes found to be mutated in this study have been reported to be mutated in other cancers. Furthermore, 12 of these genes are mutated in ovarian cancers. The FBXW7 and KIAA1462 genes are noteworthy candidates for a pathogenic role in serous borderline tumorigenesis. CONCLUSIONS: These findings suggest that a very small number of somatic genetic mutations are characteristic of SBTs of the ovary, thus supporting their classification as a relatively genetically stable tumor type. The mutant genes described herein represent novel candidates for the pathogenesis of ovarian SBT.

Host subversion of bacterial metallophore usage drives copper intoxication.

IMPORTANCE: During infection, bacteria must overcome the dual threats of metal starvation and intoxication. This work reveals that the zinc-withholding response of the host sensitizes S. aureus to copper intoxication. In response to zinc starvation, S. aureus utilizes the metallophore staphylopine. The current work revealed that the host can leverage the promiscuity of staphylopine to intoxicate S. aureus during infection. Significantly, staphylopine-like metallophores are produced by a wide range of pathogens, suggesting that this is a conserved weakness that the host can leverage to toxify invaders with copper. Moreover, it challenges the assumption that the broad-spectrum metal binding of metallophores is inherently beneficial to bacteria.

Comparison of avalanche survival patterns in Canada and Switzerland.

BACKGROUND: Current recommendations for rescue and resuscitation of people buried in avalanches are based on Swiss avalanche survival data. We analyzed Canadian survival patterns and compared them with those from Switzerland. METHODS: We extracted relevant data for survivors and nonsurvivors of complete avalanche burials from Oct. 1, 1980, to Sept. 30, 2005, from Canadian and Swiss databases. We calculated survival curves for Canada with and without trauma-related deaths as well as for different outdoor activities and snow climates. We compared these curves with the Swiss survival curve. RESULTS: A total of 301 people in the Canadian database and 946 in the Swiss database met the inclusion criteria. The overall proportion of people who survived did not differ significantly between the two countries (46.2% [139/301] v. 46.9% [444/946]; p = 0.87). Significant differences were observed between the overall survival curves for the two countries (p = 0.001): compared with the Swiss curve, the Canadian curve showed a quicker drop at the early stages of burial and poorer survival associated with prolonged burial. The probability of survival fell quicker with trauma-related deaths and in denser snow climates. Poorer survival probabilities in the Canadian sample were offset by significantly quicker extrication (median duration of burial 18 minutes v. 35 minutes in the Swiss sample; p < 0.001). INTERPRETATION: Observed differences in avalanche survival curves between the Canadian and Swiss samples were associated with the prevalence of trauma and differences in snow climate. Although avoidance of avalanches remains paramount for survival, the earlier onset of asphyxia, especially in maritime snow climates, emphasizes the importance of prompt extrication, ideally within 10 minutes. Protective devices against trauma and better clinical skills in organized rescue may further improve survival.

Genome-wide association study provides evidence for a breast cancer risk locus at 6q22.33.

We performed a three-phase genome-wide association study (GWAS) using cases and controls from a genetically isolated population, Ashkenazi Jews (AJ), to identify loci associated with breast cancer risk. In the first phase, we compared allele frequencies of 150,080 SNPs in 249 high-risk, BRCA1/2 mutation-negative AJ familial cases and 299 cancer-free AJ controls using chi(2) and the Cochran-Armitage trend tests. In the second phase, we genotyped 343 SNPs from 123 regions most significantly associated from stage 1, including 4 SNPs from the FGFR2 region, in 950 consecutive AJ breast cancer cases and 979 age-matched AJ controls. We replicated major associations in a third independent set of 243 AJ cases and 187 controls. We obtained a significant allele P value of association with AJ breast cancer in the FGFR2 region (P = 1.5 x 10(-5), odds ratio (OR) 1.26, 95% confidence interval (CI) 1.13-1.40 at rs1078806 for all phases combined). In addition, we found a risk locus in a region of chromosome 6q22.33 (P = 2.9 x 10(-8), OR 1.41, 95% CI 1.25-1.59 at rs2180341). Using several SNPs at each implicated locus, we were able to verify associations and impute haplotypes. The major haplotype at the 6q22.33 locus conferred protection from disease, whereas the minor haplotype conferred risk. Candidate genes in the 6q22.33 region include ECHDC1, which encodes a protein involved in mitochondrial fatty acid oxidation, and also RNF146, which encodes a ubiquitin protein ligase, both known pathways in breast cancer pathogenesis.

Microarray analysis of early stage serous ovarian cancers shows profiles predictive of favorable outcome.

PURPOSE: Although few women with advanced serous ovarian cancer are cured, detection of the disease at an early stage is associated with a much higher likelihood of survival. We previously used gene expression array analysis to distinguish subsets of advanced cancers based on disease outcome. In the present study, we report on gene expression of early-stage cancers and validate our prognostic model for advanced-stage cancers. EXPERIMENTAL DESIGN: Frozen specimens from 39 stage I/II, 42 stage III/IV, and 20 low malignant potential cancers were obtained from four different sites. A linear discriminant model was used to predict survival based upon array data. RESULTS: We validated the late-stage survival model and show that three of the most differentially expressed genes continue to be predictive of outcome. Most early-stage cancers (38 of 39 invasive, 15 of 20 low malignant potential) were classified as long-term survivors (median probabilities 0.97 and 0.86). MAL, the most differentially expressed gene, was further validated at the protein level and found to be an independent predictor of poor survival in an unselected group of advanced serous cancers (P = 0.0004). CONCLUSIONS: These data suggest that serous ovarian cancers detected at an early stage generally have a favorable underlying biology similar to advanced-stage cases that are long-term survivors. Conversely, most late-stage ovarian cancers seem to have a more virulent biology. This insight suggests that if screening approaches are to succeed it will be necessary to develop approaches that are able to detect these virulent cancers at an early stage.

Triaging multiple victims in an avalanche setting: the Avalanche Survival Optimizing Rescue Triage algorithmic approach.

As winter backcountry activity increases, so does exposure to avalanche danger. A complicated situation arises when multiple victims are caught in an avalanche and where medical and other rescue demands overwhelm resources in the field. These mass casualty incidents carry a high risk of morbidity and mortality, and there is no recommended approach to patient care specific to this setting other than basic first aid principles. The literature is limited with regard to triaging systems applicable to avalanche incidents. In conjunction with the development of an electronic avalanche rescue training module by the Canadian Avalanche Association, we have designed the Avalanche Survival Optimizing Rescue Triage algorithm to address the triaging of multiple avalanche victims to optimize survival and disposition decisions.

Differential exoprotease activities confer tumor-specific serum peptidome patterns.

Recent studies have established distinctive serum polypeptide patterns through mass spectrometry (MS) that reportedly correlate with clinically relevant outcomes. Wider acceptance of these signatures as valid biomarkers for disease may follow sequence characterization of the components and elucidation of the mechanisms by which they are generated. Using a highly optimized peptide extraction and matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) MS-based approach, we now show that a limited subset of serum peptides (a signature) provides accurate class discrimination between patients with 3 types of solid tumors and controls without cancer. Targeted sequence identification of 61 signature peptides revealed that they fall into several tight clusters and that most are generated by exopeptidase activities that confer cancer type-specific differences superimposed on the proteolytic events of the ex vivo coagulation and complement degradation pathways. This small but robust set of marker peptides then enabled highly accurate class prediction for an external validation set of prostate cancer samples. In sum, this study provides a direct link between peptide marker profiles of disease and differential protease activity, and the patterns we describe may have clinical utility as surrogate markers for detection and classification of cancer. Our findings also have important implications for future peptide biomarker discovery efforts.

Patterns of death among avalanche fatalities: a 21-year review.

BACKGROUND: Avalanches are a significant cause of winter recreational fatalities in mountain regions. The purpose of this study was to determine the relative contributions of trauma and asphyxia to avalanche deaths. METHODS: We reviewed all avalanche fatalities between 1984 and 2005 that had been investigated by the offices of the British Columbia Coroners Service and the Chief Medical Examiner of Alberta. In addition, we searched the database of the Canadian Avalanche Centre for fatal avalanche details. We calculated injury severity scores for all victims who underwent autopsy. RESULTS: There were 204 avalanche fatalities with mortality information over the 21-year study period. Of these, 117 victims underwent autopsy, and 87 underwent forensic external examination. Asphyxia caused 154 (75%) deaths. Trauma caused 48 (24%) deaths, with the rate of death from trauma ranging from 9% (4/44) for snowmobilers to 42% (5/12) for ice climbers. In addition, 13% (12/92) of the asphyxia victims who underwent autopsy had major trauma, defined as an injury severity score of greater than 15. Only 48% (23/48) of victims for whom trauma was the primary cause of death had been completely buried. INTERPRETATION: Asphyxia and severe trauma caused most avalanche fatalities in western Canada. The relative rates differed between snowmobilers and those engaged in other mountain activities. Our findings should guide recommendations for safety devices, safety measures and resuscitation.

A phase II trial of erlotinib in combination with bevacizumab in patients with metastatic breast cancer.

PURPOSE: To evaluate the efficacy and toxicity of erlotinib plus bevacizumab in patients with metastatic breast cancer (MBC), targeting the epidermal growth factor receptor (EGFR/HER1) and the vascular endothelial growth factor (VEGF) pathway. EXPERIMENTAL DESIGN: Thirty-eight patients with MBC were enrolled and treated at two institutions with erlotinib, a small molecule EGFR tyrosine kinase inhibitor (150 mg p.o. daily) plus bevacizumab, an anti-VEGF antibody (15 mg/kg i.v. every 3 weeks). Patients had one to two prior chemotherapy regimens for metastatic disease. The primary end point was response rate by Response Evaluation Criteria in Solid Tumors criteria using a Simon 2-stage design. Secondary end points included toxicity, time to progression, response duration, and stabilization of disease of > or = 26 weeks. Correlative studies were done on tumor tissue, including EGFR expression and mutation analysis. RESULTS: One patient achieved a partial response for 52+ months. Fifteen patients had stable disease at first evaluation at 9 weeks; 4 of these patients had stable disease beyond 26 weeks. Median time to progression was 11 weeks (95% confidence interval, 8-18 weeks). Diarrhea of any grade was observed in 84% of patients (grade 3 in 3%); 76% experienced grade 1 or 2 skin rash, and 18% developed hypertension (grade 3 in 11%). The level of EGFR expression was not predictive of response to therapy. CONCLUSIONS: The combination of erlotinib and bevacizumab was well-tolerated but had limited activity in unselected patients with previously treated MBC. Biomarkers are needed to identify those MBC patients likely to respond to anti-EGFR/HER1 plus anti-VEGF therapy.