Impact of comorbidities on immediate post-operative complications in oral cavity free flap patients.

PURPOSE: To examine the relationship between comorbidities and the development of immediate post-operative complications in patients undergoing oral cavity composite resection (OCCR) with free flap (FF) reconstruction. MATERIALS AND METHODS: Retrospective analysis was completed on all consecutive OCCRs with FF reconstruction performed at a single quaternary care facility between 1999 and 2020. Comorbidities, immediate post-operative complications, patient demographics, and tumor characteristics were collected. Odds ratios (OR) with 95 % confidence intervals were calculated for associations between comorbidities and immediate post-operative complications. RESULTS: 320 patients who underwent OCCR with FF reconstruction were included. One hundred twenty-one (37.8 %) patients developed a post-operative complication during their initial hospital admission. The most common complications were non-pneumonia cardiopulmonary events (14.1 %), pneumonia (9.4 %), and wound infection (8.4 %). Other complications included flap compromise, bleeding, and fistula. On multivariate analysis, patients without comorbid conditions were less likely to develop a post-operative complication (OR 0.64; 0.41-0.98). Atrial fibrillation (OR 2.94; 1.17-7.39) and cerebrovascular disease (OR 2.28; 1.08-4.84) were associated with increased odds of developing any complications. Furthermore, cerebrovascular disease (OR: 2.33; 1.04-5.39) and peripheral vascular disease (OR: 2.7; 1.2-6.08) were independently associated with pneumonia. CONCLUSION: In this retrospective review of patients undergoing OCCR with FF reconstruction for oral cavity SCC, lack of identifiable comorbidities appeared to be protective for post-operative complications while atrial fibrillation and cerebrovascular disease were associated with increased odds of any complication. Pre-existing vascular disease was also associated with an increased risk of pneumonia.

Tumor collagens predict genetic features and patient outcomes.

The extracellular matrix (ECM) is a critical determinant of tumor fate that reflects the output from myriad cell types in the tumor. Collagens constitute the principal components of the tumor ECM. The changing collagen composition in tumors along with their impact on patient outcomes and possible biomarkers remains largely unknown. The RNA expression of the 43 collagen genes from solid tumors in The Cancer Genome Atlas (TCGA) was clustered to classify tumors. PanCancer analysis revealed how collagens by themselves can identify the tissue of origin. Clustering by collagens in each cancer type demonstrated strong associations with survival, specific immunoenvironments, somatic gene mutations, copy number variations, and aneuploidy. We developed a machine learning classifier that predicts aneuploidy, and chromosome arm copy number alteration (CNA) status based on collagen expression alone with high accuracy in many cancer types with somatic mutations, suggesting a strong relationship between the collagen ECM context and specific molecular alterations. These findings have broad implications in defining the relationship between cancer-related genetic defects and the tumor microenvironment to improve prognosis and therapeutic targeting for patient care, opening new avenues of investigation to define tumor ecosystems.

Somatic mutations in collagens are associated with a distinct tumor environment and overall survival in gastric cancer.

BACKGROUND: Gastric cancer is a heterogeneous disease with poorly understood genetic and microenvironmental factors. Mutations in collagen genes are associated with genetic diseases that compromise tissue integrity, but their role in tumor progression has not been extensively reported. Aberrant collagen expression has been long associated with malignant tumor growth, invasion, chemoresistance, and patient outcomes. We hypothesized that somatic mutations in collagens could functionally alter the tumor extracellular matrix. METHODS: We used publicly available datasets including The Tumor Cancer Genome Atlas (TCGA) to interrogate somatic mutations in collagens in stomach adenocarcinomas. To demonstrate that collagens were significantly mutated above background mutation rates, we used a moderated Kolmogorov-Smirnov test along with combination analysis with a bootstrap approach to define the background accounting for mutation rates. Association between mutations and clinicopathological features was evaluated by Fisher or chi-squared tests. Association with overall survival was assessed by Kaplan-Meier and the Cox-Proportional Hazards Model. Gene Set Enrichment Analysis was used to interrogate pathways. Immunohistochemistry and in situ hybridization tested expression of COL7A1 in stomach tumors. RESULTS: In stomach adenocarcinomas, we identified individual collagen genes and sets of collagen genes harboring somatic mutations at a high frequency compared to background in both microsatellite stable, and microsatellite instable tumors in TCGA. Many of the missense mutations resemble the same types of loss of function mutations in collagenopathies that disrupt tissue formation and destabilize cells providing guidance to interpret the somatic mutations. We identified combinations of somatic mutations in collagens associated with overall survival, with a distinctive tumor microenvironment marked by lower matrisome expression and immune cell signatures. Truncation mutations were strongly associated with improved outcomes suggesting that loss of expression of secreted collagens impact tumor progression and treatment response. Germline collagenopathy variants guided interpretation of impactful somatic mutations on tumors. CONCLUSIONS: These observations highlight that many collagens, expressed in non-physiologically relevant conditions in tumors, harbor impactful somatic mutations in tumors, suggesting new approaches for classification and therapy development in stomach cancer. In sum, these findings demonstrate how classification of tumors by collagen mutations identified strong links between specific genotypes and the tumor environment.

Use of high resolution/accurate mass full scan/data-dependent acquisition for targeted/non-targeted screening in equine doping control.

High-resolution mass spectrometry (HRMS) is a very powerful technology for equine doping control analysis. The more recently developed hybrid type of Orbitrap-based HRMS instrument allows for both targeted and non-targeted screening analyses in a single liquid chromatography-high resolution tandem mass spectrometry (LC-HRMS/MS) run. In the present study, an LC-HRMS/MS method was developed and validated to detect prohibited substances in equine sports. The substances were recovered from equine plasma by liquid-liquid extraction (LLE) using methyl tert-butyl ether and were separated on a C18 reversed-phase column using mobile phases of 5 mM ammonium formate and acetonitrile. A 7.5 min LC gradient was employed to elute substances and results indicated that the LC method generated sharp and symmetric chromatographic peaks. An in-house equine doping compound database and a spectral library were built to increase method specificity for substances of interest. Five criteria, i.e. accurate mass, retention time, isotope pattern, selected HRMS/MS fragment ions (compound database) and HRMS/MS spectra (spectral library), were employed for targeted screening. We utilized these criteria to validate targeted detection of 451 substances within our in-house equine doping compound database. By using all five criteria in screening, the false screening positive rate is significantly reduced. A screening strategy and a Microsoft Excel macro were developed to facilitate interpretation and reporting of results. As the simultaneous acquisition of the full scan HRMS data provides the opportunity for retrospective non-targeted analysis, our findings highlight the use of this novel methodology as a simple, rapid, and reliably reproducible strategy to meet the challenge of identifying an increasing number of doping substances that could potentially impact the integrity of the horse racing community.

The physical activity and nutrition-related corporate social responsibility initiatives of food and beverage companies in Canada and implications for public health.

BACKGROUND: As diet-related diseases have increased over the past decades, large food companies have come under scrutiny for contributing to this public health crisis. In response, the food industry has implemented Corporate Social Responsibility (CSR) initiatives related to nutrition and physical activity to emphasize their concern for consumers. This study sought to describe the nature and targeted demographic of physical activity and nutrition-related CSR initiatives of large food companies in Canada and to compare companies who participate in the Canadian Children's Food and Beverage Advertising Initiative (CAI), a self-regulatory initiative aimed at reducing unhealthy food advertising to children, with non-participating companies. METHODS: A cross-sectional study was conducted in 2016. Thirty-nine large food companies, including 18 participating in the CAI, were included in the study. The webpages, Facebook pages and corporate reports of these companies were surveyed to identify CSR initiatives related to nutrition and physical activity. Initiatives were then classified by type (as either philanthropic, education-oriented, research-oriented or other) and by targeted demographic (i.e. targeted at children under 18 years or the general population). Differences between CAI and non-CAI companies were tested using chi-square and Mann-Whitney U tests. RESULTS: Overall, 63 CSR initiatives were identified; 39 were nutrition-related while 24 were physical activity-related. Most (70%) initiatives were considered philanthropic activities, followed by education-oriented (20%), research-oriented (8%) and other (2%). Almost half (47%; n = 29) of initiatives targeted children. Examples of child-targeted initiatives included support of school milk programs (n = 2), the sponsorship of children's sports programs (n = 2) and the development of educational resources for teachers (n = 1). There were no statistically significant differences in the number of CSR initiatives per company (CAI: Mdn = 1, IQR = 3; non-CAI: Mdn = 0, IQR = 2; p = .183) or the proportion of child-targeted initiatives (CAI: 42%; non-CAI: 54%; p = .343) between CAI and non-CAI companies. CONCLUSION: Food companies, including many that largely sell and market unhealthy products, are heavily involved in physical activity and nutrition-related initiatives in Canada, many of which are targeted to children. Government policies aimed at protecting children from unhealthy food marketing should consider including CSR initiatives that expose children to food company branding.

Selective imaging of solid tumours via the calcium-dependent high-affinity binding of a cyclic octapeptide to phosphorylated Annexin A2.

The heterogeneity and continuous genetic adaptation of tumours complicate their detection and treatment via the targeting of genetic mutations. However, hallmarks of cancer such as aberrant protein phosphorylation and calcium-mediated cell signalling provide broadly conserved molecular targets. Here, we show that, for a range of solid tumours, a cyclic octapeptide labelled with a near-infrared dye selectively binds to phosphorylated Annexin A2 (pANXA2), with high affinity at high levels of calcium. Because of cancer-cell-induced pANXA2 expression in tumour-associated stromal cells, the octapeptide preferentially binds to the invasive edges of tumours and then traffics within macrophages to the tumour's necrotic core. As proof-of-concept applications, we used the octapeptide to detect tumour xenografts and metastatic lesions, and to perform fluorescence-guided surgical tumour resection, in mice. Our findings suggest that high levels of pANXA2 in association with elevated calcium are present in the microenvironment of most solid cancers. The octapeptide might be broadly useful for selective tumour imaging and for delivering drugs to the edges and to the core of solid tumours.

Inhibition of Ubiquitin-Specific Protease 14 Suppresses Cell Proliferation and Synergizes with Chemotherapeutic Agents in Neuroblastoma.

Neuroblastoma is the most common extracranial malignant solid tumor in children, and drug resistance is a major reason for poor outcomes. Elevated proteasome activity plays an important role in neuroblastoma tumor development and resistance to conventional chemotherapy. Ubiquitin-specific protease 14 (USP14), one of three deubiquitinases associated with the regulatory subunit of the proteasome, is emerging as a potential therapeutic target in multiple tumor types. However, the role of USP14 in neuroblastoma is yet to be elucidated. We found that USP14 inhibition in neuroblastoma via knockdown or a specific inhibitor such as b-AP15 suppressed cell proliferation by inducing cell apoptosis. Furthermore, b-AP15 significantly inhibited neuroblastoma tumor growth in NGP and SH-SY5Y xenograft mouse models. For combination treatment, b-AP15 plus conventional chemotherapeutic agents such as doxorubicin or VP-16 resulted in synergistic antitumor effects on neuroblastoma. Our study demonstrates that USP14 is required for cell viability and is a novel therapeutic target in neuroblastoma. Moreover, USP14 inhibition may add value in combination therapy due to its powerful synergistic effects in treating neuroblastoma.

MELK is a novel therapeutic target in high-risk neuroblastoma.

Maternal embryonic leucine zipper kinase (MELK) is known to modulate intracellular signaling and control cellular processes. However, the role of MELK in oncogenesis is not well defined. In this study, using two microarray datasets of neuroblastoma (NB) patients, we identified that MELK expression is significantly correlated to poor overall survival, unfavorable prognosis, and high-risk status. We found that MELK is a direct transcription target of MYCN and MYC in NB, and MYCN increases MELK expression via direct promoter binding. Interestingly, knockdown of MELK expression significantly reduced the phosphorylation of target protein Retinoblastoma (pRb) and inhibited NB cell growth. Furthermore, pharmacological inhibition of MELK activity by small-molecule inhibitor OTSSP167 significantly inhibited cell proliferation, anchorage-independent colony formation, blocked cell cycle progression, and induced apoptosis in different NB cell lines including a drug-resistant cell line. Additionally, OTSSP167 suppressed NB tumor growth in an orthotopic xenograft mouse model. Overall, our data suggest that MELK is a novel therapeutic target for NB and its inhibitor OTSSP167 is a promising drug for further clinical development.

Novel proteasome inhibitor delanzomib sensitizes cervical cancer cells to doxorubicin-induced apoptosis via stabilizing tumor suppressor proteins in the p53 pathway.

Cervical cancer, the third most commonly occurring cancer, is the second leading cause of cancer related mortality among women. Aberrant ubiquitination and proteasome activity, both human papillomavirus and tumor derived, have been shown to contribute to tumor angiogenesis, proliferation, and invasion in many cancers, including cervical cancer. Thus, small molecule proteasome inhibitors are a potential and strategic treatment option for cervical cancer. In this study, novel proteasome inhibitor delanzomib (CEP-18770) exhibited potent pro-apoptotic and cytotoxic effects on a panel of cervical cancer cell lines by blocking proteasomal activity. Delanzomib also significantly sensitized cervical cancer cells to treatment of doxorubicin (Dox), a traditional chemotherapeutic agent. Furthermore, proteasome inhibition revealed stabilization of p53 and p53 transcriptional targets and induction of p38/JNK phosphorylation. Additionally, delanzomib worked synergistically with Dox to further upregulate p53 and its downstream targets and enhanced Dox-induced p38 phosphorylation. Our study strongly supports the 26S proteasome as a potential therapeutic target in cervical cancer and proteasome inhibition by delanzomib may be a potential treatment strategy for cervical cancer patients.

Detection of enzyme activity in orthotopic murine breast cancer by fluorescence lifetime imaging using a fluorescence resonance energy transfer-based molecular probe.

Cancer-related enzyme activity can be detected noninvasively using activatable fluorescent molecular probes. In contrast to "always-on" fluorescent molecular probes, activatable probes are relatively nonfluorescent at the time of administration due to intramolecular fluorescence resonance energy transfer (FRET). Enzyme-mediated hydrolysis of peptide linkers results in reduced FRET and increase of fluorescence yield. Separation of signal from active and inactive probe can be difficult with conventional intensity-based fluorescence imaging. Fluorescence lifetime (FLT) measurement is an alternative method to detect changes in FRET. Thus, we investigate FLT imaging for in vivo detection of FRET-based molecular probe activation in an orthotopic breast cancer model. Indeed, the measured FLT of the enzyme-activatable molecular probe increases from 0.62 ns just after injection to 0.78 ns in tumor tissue after 4 h. A significant increase in FLT is not observed for an always-on targeted molecular probe with the same fluorescent reporter. These results show that FLT contrast is a powerful addition to preclinical imaging because it can report molecular activity in vivo due to changes in FRET. Fluorescence lifetime imaging exploits unique characteristics of fluorescent molecular probes that can be further translated into clinical applications, including noninvasive detection of cancer-related enzyme activity.

Noninvasive photoacoustic and fluorescence sentinel lymph node identification using dye-loaded perfluorocarbon nanoparticles.

The contrast mechanisms used for photoacoustic tomography (PAT) and fluorescence imaging differ in subtle, but significant, ways. The design of contrast agents for each or both modalities requires an understanding of the spectral characteristics as well as intra- and intermolecular interactions that occur during formulation. We found that fluorescence quenching that occurs in the formulation of near-infrared (NIR) fluorescent dyes in nanoparticles results in enhanced contrast for PAT. The ability of the new PAT method to utilize strongly absorbing chromophores for signal generation allowed us to convert a highly fluorescent dye into an exceptionally high PA contrast material. Spectroscopic characterization of the developed NIR dye-loaded perfluorocarbon-based nanoparticles for combined fluorescence and PA imaging revealed distinct dye-dependent photophysical behavior. We demonstrate that the enhanced contrast allows detection of regional lymph nodes of rats in vivo with time-domain optical and photoacoustic imaging methods. The results further show that the use of fluorescence lifetime imaging, which is less dependent on fluorescence intensity, provides a strategic approach to bridge the disparate contrast reporting mechanisms of fluorescence and PA imaging methods.

Targeting of alpha(nu)beta(3)-integrins expressed on tumor tissue and neovasculature using fluorescent small molecules and nanoparticles.

AIM: Receptor-specific small molecules and nanoparticles are widely used in molecular imaging of tumors. Although some studies have described the relative strengths and weaknesses of the two approaches, reports of a direct comparison and analysis of the two strategies are lacking. Herein, we compared the tumor-targeting characteristics of a small near-infrared fluorescent compound (cypate-peptide conjugate) and relatively large perfluorocarbon-based nanoparticles (250 nm diameter) for imaging alpha(nu)beta(3)-integrin receptor expression in tumors. MATERIALS & METHODS: Near-infrared fluorescent small molecules and nanoparticles were administered to living mice bearing subcutaneous or intradermal syngeneic tumors and imaged with whole-body and high-resolution optical imaging systems. RESULTS: The nanoparticles, designed for vascular constraint, remained within the tumor vasculature while the small integrin-avid ligands diffused into the tissue to target integrin expression on tumor and endothelial cells. Targeted small-molecule and nanoparticle contrast agents preferentially accumulated in tumor tissue with tumor-to-muscle ratios of 8 and 7, respectively, compared with 3 for nontargeted nanoparticles. CONCLUSION: Fluorescent small molecular probes demonstrate greater overall early tumor contrast and rapid visualization of tumors, but the vascular-constrained nanoparticles are more selective for detecting cancer-induced angiogenesis. A combination of both imaging agents provides a strategy to image and quantify integrin expression in tumor tissue and tumor-induced neovascular systems.