Kartezio: Evolutionary Design of Explainable Pipelines for Biomedical Image Analysis

An unresolved issue in contemporary biomedicine is the overwhelming number and diversity of complex images that require annotation, analysis and interpretation. Recent advances in Deep Learning have revolutionized the field of computer vision, creating algorithms that compete with human experts in image segmentation tasks. Crucially however, these frameworks require large human-annotated datasets for training and the resulting models are difficult to interpret. In this study, we introduce Kartezio, a modular Cartesian Genetic Programming based computational strategy that generates transparent and easily interpretable image processing pipelines by iteratively assembling and parameterizing computer vision functions. The pipelines thus generated exhibit comparable precision to state-of-the-art Deep Learning approaches on instance segmentation tasks, while requiring drastically smaller training datasets, a feature which confers tremendous flexibility, speed, and functionality to this approach. We also deployed Kartezio to solve semantic and instance segmentation problems in four real-world Use Cases, and showcase its utility in imaging contexts ranging from high-resolution microscopy to clinical pathology. By successfully implementing Kartezio on a portfolio of images ranging from subcellular structures to tumoral tissue, we demonstrated the flexibility, robustness and practical utility of this fully explicable evolutionary designer for semantic and instance segmentation.Comment: 36 pages, 6 main Figures. The Extended Data Movie is available at the following link: https://www.youtube.com/watch?v=r74gdzb6hdA. The source code is available on Github: https://github.com/KevinCortacero/Kartezi

Myxoma Virus Treatment for Brain Tumour Initiating Cells: Interrogating and Enhancing Myxoma-Mediated Cell Death

Brain tumour initiating cells (BTICs) are stem-like cells hypothesized to mediate recurrence in high-grade gliomas. Preclinical success has been demonstrated in treating patient-derived BTICs with oncolytic virotherapy, using replication-competent viruses to target and kill malignant cells. Myxoma virus (MyxV) is an oncolytic candidate, which is highly effective in conventional glioma models, but only modestly effective in BTICs. The objective of this study was to improve MyxV efficacy in BTICs in vitro, combining chemotherapeutics and virotherapy. Using a pharmacoviral screen, eleven compounds that enhance MyxV-mediated cell death were identified. A lead compound, axitinib, was validated in multiple BTIC models. It was demonstrated that a virally encoded protein, M011L, prevents MyxV-induced apoptosis in BTICs, and M011L disruption was shown to greatly improve MyxV-mediated cell death through apoptosis induction. These studies have elucidated multiple strategies for improving MyxV efficacy in a preclinical glioma model, with implications for the future clinical development of MyxV

RESEARCH Open Access Rapid inflammasome activation in microglia contributes to brain disease in HIV/AIDS

Background: Human immunodeficiency virus type 1(HIV-1) infects and activates innate immune cells in the brain resulting in inflammation and neuronal death with accompanying neurological deficits. Induction of inflammasomes causes cleavage and release of IL-1β and IL-18, representing pathogenic processes that underlie inflammatory diseases although their contribution HIV-associated brain disease is unknown. Results: Investigation of inflammasome-associated genes revealed that IL-1β, IL-18 and caspase-1 were induced in brains of HIV-infected persons and detected in brain microglial cells. HIV-1 infection induced pro-IL-1β in human microglia at 4 hr post-infection with peak IL-1β release at 24 hr, which was accompanied by intracellular ASC translocation and caspase-1 activation. HIV-dependent release of IL-1β from a human macrophage cell line, THP-1, was inhibited by NLRP3 deficiency and high extracellular [K +]. Exposure of microglia to HIV-1 gp120 caused IL-1β production and similarly, HIV-1 envelope pseudotyped viral particles induced IL-1β release, unlike VSV-G pseudotyped particles. Infection of cultured feline macrophages by the related lentivirus, feline immunodeficiency virus (FIV), also resulted in the prompt induction of IL-1β. In vivo FIV infection activated multiple inflammasome-associated genes in microglia, which was accompanied by neuronal loss in cerebral cortex and neurological deficits. Multivariate analyses of data from FIV-infected and uninfected animals disclosed that IL-1β, NLRP3 and caspase-1 expression in cerebral cortex represented key molecular determinants of neurological deficits. Conclusions: NLRP3 inflammasome activation was an early and integral aspect of lentivirus infection of microglia, which was associated with lentivirus-induced brain disease. Inflammasome activation in the brain might represent a potential target for therapeutic interventions in HIV/AIDS

Resistance to Oncolytic Myxoma Virus Therapy in Nf1^−/−/Trp53^−/− Syngeneic Mouse Glioma Models Is Independent of Anti-Viral Type-I Interferon

<div><p>Despite promising preclinical studies, oncolytic viral therapy for malignant gliomas has resulted in variable, but underwhelming results in clinical evaluations. Of concern are the low levels of tumour infection and viral replication within the tumour. This discrepancy between the laboratory and the clinic could result from the disparity of xenograft versus syngeneic models in determining <i>in vivo</i> viral infection, replication and treatment efficacy. Here we describe a panel of primary mouse glioma lines derived from <i>Nf1</i>^+/−<i>Trp53</i>^+/− mice in the C57Bl/6J background for use in the preclinical testing of the oncolytic virus Myxoma (<b>MYXV</b>). These lines show a range of susceptibility to MYXV replication <i>in vitro</i>, but all succumb to viral-mediated cell death. Two of these lines orthotopically grafted produced aggressive gliomas. Intracranial injection of MYXV failed to result in sustained viral replication or treatment efficacy, with minimal tumour infection that was completely resolved by 7 days post-infection. We hypothesized that the stromal production of Type-I interferons (<b>IFNα/β</b>) could explain the resistance seen in these models; however, we found that neither the cell lines <i>in vitro</i> nor the tumours <i>in vivo</i> produce any IFNα/β in response to MYXV infection. To confirm IFNα/β did not play a role in this resistance, we ablated the ability of tumours to respond to IFNα/β via IRF9 knockdown, and generated identical results. Our studies demonstrate that these syngeneic cell lines are relevant preclinical models for testing experimental glioma treatments, and show that IFNα/β is not responsible for the MYXV treatment resistance seen in syngeneic glioma models.</p></div

K1492 and K1861 form aggressive intracranial tumours and MYXV treatment results in no efficacy and minimal viral infection with no viral replication.

<p><b>A</b> – Histopathology of 14 day K1492 and K1861 by H&E (first column 25×, second column 200×) and astrocytic markers S100b (200×) and GFAP (200×). 5×10⁴ cells of K1492 (<b>B</b>) and K1861 (<b>C</b>) were intracranially implanted in C57Bl/6J mice and received 5×10⁶ PFUs vMyx-FLuc (MYXV), UV-inactivated virus (DV), PBS, or no treatment (NT) on day 14. <b>D</b> – Luciferase measured (Total FLUX) from region-of-interest around the entire mouse skull following 5×10⁶ PFUs vMyx-FLuc in K1492 (n = 8), K1861 (n = 10) or no tumour (n = 5). Error bars represent standard error. <b>E</b> – Viral recovery from K1492 (n = 4) and K1861 (n = 4) tumours following intracranial treatment with vMyx-FLuc. Input virus represents mice where virus was recovered 1 hour post-injection. Error bars represent standard error. <b>F</b> – Immunohistochemical staining for early MYXV protein MT-7e in 14 day K1492 at 1, 3 and 7 days post-treatment (Top row 25×; Bottom row 100×).</p

IRF9 knockdown results in loss of protection of IFNβ on K1492 in vitro.

<p><b>A</b> – RT-PCR of IRF9 message level after stable transduction of IRF9 shRNA (IRF9kd), scrambled control (Scram) or wildtype (WT) K1492. <b>B</b> – Luminescence activity of an inducible luciferase construct stably transfected into IRF9 knockdown (IRF9kd), scrambled control (Scram) or wildtype cells (WT) K1492 cells. Error bars represent standard error and asterisks p<0.05 when compared to WT K1492. <b>C</b> – K1492 knockdown construct or controls 48 hpi with 1.0 MOI vMyx-GFP with or without the pretreated with 1.0 units of mouse IFNβ as measured by Alamar blue. Error bars represent standard error and asterisks p<0.05 when compared MYXV alone.</p