Small Gene Networks Delineate Immune Cell States and Characterize Immunotherapy Response in Melanoma.

Single-cell technologies have elucidated mechanisms responsible for immune checkpoint inhibitor (ICI) response, but are not amenable to a clinical diagnostic setting. In contrast, bulk RNA sequencing (RNA-seq) is now routine for research and clinical applications. Our workflow uses transcription factor (TF)-directed coexpression networks (regulons) inferred from single-cell RNA-seq data to deconvolute immune functional states from bulk RNA-seq data. Regulons preserve the phenotypic variation in CD45+ immune cells from metastatic melanoma samples (n = 19, discovery dataset) treated with ICIs, despite reducing dimensionality by >100-fold. Four cell states, termed exhausted T cells, monocyte lineage cells, memory T cells, and B cells were associated with therapy response, and were characterized by differentially active and cell state-specific regulons. Clustering of bulk RNA-seq melanoma samples from four independent studies (n = 209, validation dataset) according to regulon-inferred scores identified four groups with significantly different response outcomes (P < 0.001). An intercellular link was established between exhausted T cells and monocyte lineage cells, whereby their cell numbers were correlated, and exhausted T cells predicted prognosis as a function of monocyte lineage cell number. The ligand-receptor expression analysis suggested that monocyte lineage cells drive exhausted T cells into terminal exhaustion through programs that regulate antigen presentation, chronic inflammation, and negative costimulation. Together, our results demonstrate how regulon-based characterization of cell states provide robust and functionally informative markers that can deconvolve bulk RNA-seq data to identify ICI responders.

Signaling Dynamics Regulating Crosstalks between T-Cell Activation and Immune Checkpoints.

Immune checkpoint inhibitors (ICIs) targeting cytotoxic T lymphocyte-associated protein-4 (CTLA-4) and programmed cell death protein-1 (PD-1) have been hailed as major advances in cancer therapeutics; however, in many cancers response rates remain low. Extensive research efforts are underway to improve the efficacy of ICIs. The signaling pathways regulated by immune checkpoints (ICs) may be an important lever as they interfere with T-cell activation when activated by ICIs. Here, we review the current understanding of T-cell receptor signaling and their intersection with IC signaling pathways. As these signaling processes are highly dynamic and controlled by intricate spatiotemporal mechanisms, we focus on aspects of kinetic regulation that are modulated by ICs. Recent advances in computational modeling and experimental methods that can resolve spatiotemporal dynamics provide insights that reveal molecular mechanisms and new potential approaches for improving the design and application of ICIs.

High-content phenotypic assay for proliferation of human iPSC-derived cardiomyocytes identifies L-type calcium channels as targets.

Over 5 million people in the United States suffer from heart failure, due to the limited ability to regenerate functional cardiac tissue. One potential therapeutic strategy is to enhance proliferation of resident cardiomyocytes. However, phenotypic screening for therapeutic agents is challenged by the limited ability of conventional markers to discriminate between cardiomyocyte proliferation and endoreplication (e.g. polyploidy and multinucleation). Here, we developed a novel assay that combines automated live-cell microscopy and image processing algorithms to discriminate between proliferation and endoreplication by quantifying changes in the number of nuclei, changes in the number of cells, binucleation, and nuclear DNA content. We applied this assay to further prioritize hits from a primary screen for DNA synthesis, identifying 30 compounds that enhance proliferation of human induced pluripotent stem cell-derived cardiomyocytes. Among the most active compounds from the phenotypic screen are clinically approved L-type calcium channel blockers from multiple chemical classes whose activities were confirmed across different sources of human induced pluripotent stem cell-derived cardiomyocytes. Identification of compounds that stimulate human cardiomyocyte proliferation may provide new therapeutic strategies for heart failure.

Safety and Efficacy of Intraoperative Computer-Navigated Versus Non-Navigated Shoulder Arthroplasty at a Tertiary Referral.

Emerging technologies in shoulder arthroplasty, such as 3-dimensional planning software and real-time intraoperative navigation, are now available for surgeons to perform more accurate placement of the glenoid component without malposition or perforation. Using these tools, the surgeon can visualize the version, inclination, and containment of the implant and determine whether augmented components would be necessary. This review provides an updated investigation of the present literature to elucidate the role of computer navigation in modern shoulder arthroplasty.

The brain-penetrant clinical ATM inhibitor AZD1390 radiosensitizes and improves survival of preclinical brain tumor models.

Poor survival rates of patients with tumors arising from or disseminating into the brain are attributed to an inability to excise all tumor tissue (if operable), a lack of blood-brain barrier (BBB) penetration of chemotherapies/targeted agents, and an intrinsic tumor radio-/chemo-resistance. Ataxia-telangiectasia mutated (ATM) protein orchestrates the cellular DNA damage response (DDR) to cytotoxic DNA double-strand breaks induced by ionizing radiation (IR). ATM genetic ablation or pharmacological inhibition results in tumor cell hypersensitivity to IR. We report the primary pharmacology of the clinical-grade, exquisitely potent (cell IC50, 0.78 nM), highly selective [>10,000-fold over kinases within the same phosphatidylinositol 3-kinase-related kinase (PIKK) family], orally bioavailable ATM inhibitor AZD1390 specifically optimized for BBB penetration confirmed in cynomolgus monkey brain positron emission tomography (PET) imaging of microdosed 11C-labeled AZD1390 (Kp,uu, 0.33). AZD1390 blocks ATM-dependent DDR pathway activity and combines with radiation to induce G2 cell cycle phase accumulation, micronuclei, and apoptosis. AZD1390 radiosensitizes glioma and lung cancer cell lines, with p53 mutant glioma cells generally being more radiosensitized than wild type. In in vivo syngeneic and patient-derived glioma as well as orthotopic lung-brain metastatic models, AZD1390 dosed in combination with daily fractions of IR (whole-brain or stereotactic radiotherapy) significantly induced tumor regressions and increased animal survival compared to IR treatment alone. We established a pharmacokinetic-pharmacodynamic-efficacy relationship by correlating free brain concentrations, tumor phospho-ATM/phospho-Rad50 inhibition, apoptotic biomarker (cleaved caspase-3) induction, tumor regression, and survival. On the basis of the data presented here, AZD1390 is now in early clinical development for use as a radiosensitizer in central nervous system malignancies.

Orally Bioavailable and Blood-Brain Barrier-Penetrating ATM Inhibitor (AZ32) Radiosensitizes Intracranial Gliomas in Mice.

Inhibition of ataxia-telangiectasia mutated (ATM) during radiotherapy of glioblastoma multiforme (GBM) may improve tumor control by short-circuiting the response to radiation-induced DNA damage. A major impediment for clinical implementation is that current inhibitors have limited central nervous system (CNS) bioavailability; thus, the goal was to identify ATM inhibitors (ATMi) with improved CNS penetration. Drug screens and refinement of lead compounds identified AZ31 and AZ32. The compounds were then tested in vivo for efficacy and impact on tumor and healthy brain. Both AZ31 and AZ32 blocked the DNA damage response and radiosensitized GBM cells in vitro AZ32, with enhanced blood-brain barrier (BBB) penetration, was highly efficient in vivo as radiosensitizer in syngeneic and human, orthotopic mouse glioma model compared with AZ31. Furthermore, human glioma cell lines expressing mutant p53 or having checkpoint-defective mutations were particularly sensitive to ATMi radiosensitization. The mechanism for this p53 effect involves a propensity to undergo mitotic catastrophe relative to cells with wild-type p53. In vivo, apoptosis was >6-fold higher in tumor relative to healthy brain after exposure to AZ32 and low-dose radiation. AZ32 is the first ATMi with oral bioavailability shown to radiosensitize glioma and improve survival in orthotopic mouse models. These findings support the development of a clinical-grade, BBB-penetrating ATMi for the treatment of GBM. Importantly, because many GBMs have defective p53 signaling, the use of an ATMi concurrent with standard radiotherapy is expected to be cancer-specific, increase the therapeutic ratio, and maintain full therapeutic effect at lower radiation doses. Mol Cancer Ther; 17(8); 1637-47. ©2018 AACR.

Short- to mid-term outcomes of anatomic MCL reconstruction with Achilles tendon allograft after multiligament knee injury.

PURPOSE: Multiple techniques have been described in the literature for reconstruction of the medial collateral ligament. The purpose of this study is to describe functional outcome, range of motion, and knee stability following anatomic MCL reconstruction utilizing an Achilles tendon bone allograft after multiligament knee injury. METHODS: A comprehensive search of a single-hospital multiligament knee injury (MLKI) procedural database was conducted to identify all patients that underwent reconstruction of the MCL utilizing an Achilles tendon bone allograft and with 2-year clinical follow-up. Medical charts were retrospectively reviewed to determine each patient's knee dislocation (KD) grade, final range of motion, stability on clinical examination, and the incidence of complications and reoperations. KOOS, IKDC, and Marx scores were also collected. RESULTS: Thirty-two knees in 32 patients (21 males and 11 females) with a mean age of 30 years (range 15-51) were followed for an average of 40 months (range 28-87 months) following MCL reconstruction with Achilles tendon bone allograft. For patients with multiligament knee injuries, there were 14 KD-I (11 ACL/MCL; 3 MCL/PCL; 1 MCL/ACL/LCL; 1 MCL/PCL/LCL), 12 KD 3-M, and 3 KD-IV. One patient underwent isolated revision MCL reconstruction. At final follow-up, clinically significant valgus laxity was observed in only 1 patient (3%). All patients were able to achieve full extension of the knee and the average flexion was 121.1 ± 19.6. The average IKDC score was 67.6 ± 19.9 (range 27.7-98.9), the average KOOS score 77.1 ± 16.8 (range 31-100). The average Marx score was 4.9 (range 0-16, SD 5.2). Thirty-one of 32 (96%) patients reported being satisfied with results of the surgery. Knee dislocation grades were significantly correlated with post-operative outcome measures. CONCLUSION: In a series utilizing a modified Marx Achilles tendon, MCL reconstruction in the setting of MLKI demonstrated satisfactory clinical and functional outcomes, as well as patient satisfaction at short- to mid-term follow-up. Furthermore, knee dislocation grades were demonstrated to correlate with post-operative IKDC, KOOS, and Marx scores. LEVEL OF EVIDENCE: Type IV.

Construct Rigidity: Keystone for Treating Pelvic Discontinuity.

BACKGROUND: Pelvic discontinuity is uncommon and presents the surgeon with complex reconstructive challenges. The objective of this study is to report the results of current strategies used in the treatment of pelvic discontinuity. METHODS: We retrospectively analyzed prospectively collected data on 113 consecutive revision total hip arthroplasties performed for the treatment of unilateral pelvic discontinuity at a single institution. The study included 18 male and 95 female patients with a mean age of 63 years at the time of revision surgery. Preoperative, immediate postoperative, and latest follow-up radiographs were reviewed to assess healing of the discontinuity as well as acetabular component stability. Treatment modalities included an uncemented cup with a posterior column plate (50 hips; 44%), a cup-cage construct (27 hips; 24%), an antiprotrusio cage with or without a posterior column plate (26 hips; 23%), and an uncemented cup alone (10 hips; 9%). The average duration of follow-up for each of these types of surgical reconstruction was similar (range, 3.9 to 7.2 years). RESULTS: Five-year revision-free survivorship of the implant was best with a cup-cage construct (100%) and worst with an uncemented cup with a posterior column plate (80%) and a cup alone (80%). Healing of the discontinuity was achieved in 50% of the hips with an uncemented cup alone, 74% of the hips with an uncemented cup and a posterior column plate, 74% of the hips with a cup-cage construct, and 88% of the hips with an antiprotrusio cage construct (91% of these hips when structural allograft was used). The overall complication rate was 26.5%. The average Harris hip score improved from 54 preoperatively to 69 postoperatively (95% confidence interval: 50 to 57 preoperatively and 65 to 72 postoperatively; p = 0.017). CONCLUSIONS: Improved survivorship and healing rates were seen in this series when a reconstruction cage was used as an adjunct to an uncemented cup (cup-cage) or in combination with structural allograft bone that bridged the discontinuity. LEVEL OF EVIDENCE: Therapeutic Level III. See Instructions for Authors for a complete description of levels of evidence.

Soft Tissue Neoplasms Causing Apparent Venous Thoracic Outlet Syndrome.

Venous thoracic outlet syndrome (vTOS) usually results from compression of the subclavian vein classically as a result of narrowing of the costoclavicular space. We report 2 rare cases of soft tissue neoplasms resulting in apparent vTOS. The first case is a 46-year-old female with a 2-year history of intermittent unilateral shoulder pain, who was initially diagnosed with intervertebral disk herniation. Cervical fusion was performed; however, her symptoms progressed and she additionally developed paresthesias and venous congestion. Computed tomography (CT) angiogram demonstrated a 13-cm-encapsulated mass within the subscapularis muscle compressing the axillary vein. Radiological findings suggested lipoma. She subsequently underwent complete resection via a transaxillary approach with extension along the lateral border of the latissimus. Final pathology confirmed an intramuscular lipoma. The second case is a 21-year-old female who presented with acute onset of unilateral chest wall pain, palpable nodularity, and venous congestion. CT chest showed pulmonary embolism and an anterior chest wall mass. An initial attempt at resection was aborted due to proximity of the mass to the subclavian vein. The mass enlarged on serial imaging, measuring 3.8 cm in greatest dimension. Additionally, tumor thrombus was seen, and a subsequent ultrasound-guided biopsy was positive for high-grade synovial sarcoma. Positron emission tomography scan showed a pulmonary nodule that was resected thoracoscopically with pathology confirming metastatic synovial sarcoma. Subsequently, she underwent neoadjuvant chemoradiation followed by successful resection of the chest wall mass. An extended infraclavicular approach with a secondary transaxillary incision was utilized to achieve adequate exposure and margins. Final pathology was consistent with preoperative biopsy. Venous reconstruction was not needed. Although rare, an extrinsic mass as a cause of apparent TOS should be in the differential diagnosis. Surgical approach is based on tumor type, location, and proximity to the neurovascular bundle.

Incidence and long-term follow-up of isolated posterior cruciate ligament tears.

PURPOSE: Isolated posterior cruciate ligament (PCL) tears are an uncommon injury. The goals of this study are to (1) determine the population-based incidence of isolated PCL tears, (2) compare the occurrence of secondary meniscal tears or arthritis in patients with PCL deficiency to patients without PCL tears, and (3) evaluate factors associated with long-term sequelae among patients with PCL deficiency. METHODS: This retrospective study included a population-based incidence cohort of 48 patients with new-onset, isolated PCL tears between 1990 and 2010, as well as an age and sex-matched cohort of individuals without PCL tears. A chart review was performed to collect information related to the initial injury, treatment, and outcomes. Subjects were retrospectively followed to determine the development of subsequent meniscal tears, arthritis, or total knee arthroplasty (TKA). RESULTS: The age- and sex-adjusted annual incidence of isolated, complete PCL tears was 1.8 (95 % CI 1.3, 2.3) per 100,000. During a mean 12.2-year follow-up, patients with isolated PCL tears had a significantly higher likelihood (HR 6.2, 95 % CI 1.8, 21.2) of symptomatic arthritis compared to individuals without PCL tears. The likelihood of subsequent meniscal tears (HR 2.1, 95 % CI 0.4, 10.7) and TKA (HR 3.2, 95 % CI 0.5, 19.6) was more frequent among patients with PCL tears compared to subjects without PCL tears. Older age at injury was significantly associated with future arthritis (P = 0.003) and TKA (P = 0.02). CONCLUSION: Isolated PCL tears remain a rare injury with an estimated annual incidence of 2 per 100,000 persons. Patients with isolated PCL tears have a significantly higher risk of symptomatic arthritis than patients without PCL tears. Older age at injury is associated with a higher risk of arthritis and the need for TKA. The results of this study can be used to educate patients about the natural history of isolated PCL tears and provide a baseline of expectations for the future development of arthritis and subsequent meniscal injury following isolated PCL injury. LEVEL OF EVIDENCE: Retrospective comparative study, Level III.

Kinase Inhibition Leads to Hormesis in a Dual Phosphorylation-Dephosphorylation Cycle.

Many antimicrobial and anti-tumour drugs elicit hormetic responses characterised by low-dose stimulation and high-dose inhibition. While this can have profound consequences for human health, with low drug concentrations actually stimulating pathogen or tumour growth, the mechanistic understanding behind such responses is still lacking. We propose a novel, simple but general mechanism that could give rise to hormesis in systems where an inhibitor acts on an enzyme. At its core is one of the basic building blocks in intracellular signalling, the dual phosphorylation-dephosphorylation motif, found in diverse regulatory processes including control of cell proliferation and programmed cell death. Our analytically-derived conditions for observing hormesis provide clues as to why this mechanism has not been previously identified. Current mathematical models regularly make simplifying assumptions that lack empirical support but inadvertently preclude the observation of hormesis. In addition, due to the inherent population heterogeneities, the presence of hormesis is likely to be masked in empirical population-level studies. Therefore, examining hormetic responses at single-cell level coupled with improved mathematical models could substantially enhance detection and mechanistic understanding of hormesis.

Bilateral pelvic discontinuity: a unique condition characterized by high failure rates of current treatment.

BACKGROUND: Bilateral pelvic discontinuity is characterized by complete dissociation of the superior and inferior pelvis secondary to bone loss or fracture. The end result is a freely mobile inferior pelvis at the level of each discontinuity which presents a significant reconstruction challenge. This clinical entity has not been described previously, and the results of surgical treatment are not known. METHODS: We retrospectively reviewed all identified cases of pelvic discontinuity (PD) treated with revision THA at one institution. We identified 133 pelvic discontinuities. Within this group, 6 patients had bilateral simultaneous PDs. Preoperative, intraoperative, and postoperative data and radiographic imaging were reviewed preoperatively and postoperatively for the characteristics of the dissociation and assessing PD healing and fixation of components after surgery. RESULTS: There were no preoperative factors that could distinguish these patients from the rest of the group of discontinuities (3 rheumatoid arthritis, 2 osteonecrosis of the femoral head, 1 developmental dysplasia). The reconstructions performed included 2 cup/cage, 5 posterior plating and uncemented cup, 3 cage alone, and 2 cups only. Ten of 12 hips had at least 1 complication postoperatively. At final follow-up, only 1 patient (17%) had radiographic evidence that both discontinuities had healed (posterior plate with uncemented cup). CONCLUSIONS: Bilateral pelvic discontinuity is rare but presents the surgeon with a major reconstructive challenge. Only 1 patient went on to radiographic healing with current treatment strategies. Continued motion of the contralateral pelvic dissociation may account for the high failure rates. Surgeons should be aware of the challenges presented by this diagnosis and develop strategies to improve outcomes.

Automatic extraction of angiogenesis bioprocess from text.

MOTIVATION: Understanding key biological processes (bioprocesses) and their relationships with constituent biological entities and pharmaceutical agents is crucial for drug design and discovery. One way to harvest such information is searching the literature. However, bioprocesses are difficult to capture because they may occur in text in a variety of textual expressions. Moreover, a bioprocess is often composed of a series of bioevents, where a bioevent denotes changes to one or a group of cells involved in the bioprocess. Such bioevents are often used to refer to bioprocesses in text, which current techniques, relying solely on specialized lexicons, struggle to find. RESULTS: This article presents a range of methods for finding bioprocess terms and events. To facilitate the study, we built a gold standard corpus in which terms and events related to angiogenesis, a key biological process of the growth of new blood vessels, were annotated. Statistics of the annotated corpus revealed that over 36% of the text expressions that referred to angiogenesis appeared as events. The proposed methods respectively employed domain-specific vocabularies, a manually annotated corpus and unstructured domain-specific documents. Evaluation results showed that, while a supervised machine-learning model yielded the best precision, recall and F1 scores, the other methods achieved reasonable performance and less cost to develop. AVAILABILITY: The angiogenesis vocabularies, gold standard corpus, annotation guidelines and software described in this article are available at http://text0.mib.man.ac.uk/~mbassxw2/angiogenesis/ CONTACT: xinglong.wang@gmail.com.

Cancer genome analysis informatics.

The analysis of cancer genomes has benefited from the advances in technology that enable data to be generated on an unprecedented scale, describing a tumour genome's sequence and composition at increasingly high resolution and reducing cost. This progress is likely to increase further over the coming years as next-generation sequencing approaches are applied to the study of cancer genomes, in tandem with large-scale efforts such as the Cancer Genome Atlas and recently announced International Cancer Genome Consortium efforts to complement those already established such as the Sanger Institute Cancer Genome Project. This presents challenges for the cancer researcher and the research community in general, in terms of analysing the data generated in one's own projects and also in coordinating and interrogating data that are publicly available. This review aims to provide a brief overview of some of the main informatics resources currently available and their use, and some of the informatics approaches that may be applied in the study of cancer genomes.