Circulating Inflammatory Biomarkers Predict Pain Change Following Exercise-Induced Shoulder Injury: Findings From the Biopsychosocial Influence on Shoulder Pain Preclinical Trial.

Shoulder pain is a highly prevalent musculoskeletal condition that frequently leads to suboptimal clinical outcomes. This study tested the extent to which circulating inflammatory biomarkers are associated with reports of shoulder pain and upper-extremity disability for a high-risk genetic by psychological subgroup (catechol-O-methyltransferase [COMT] variation by pain catastrophizing [PCS]). Pain-free adults meeting high-risk COMT × PCS subgroup criteria completed an exercise-induced muscle injury protocol. Thirteen biomarkers were collected and analyzed from plasma 48 hours after muscle injury. Shoulder pain intensity and disability (Quick-DASH) were reported at 48 and 96 hours to calculate change scores. Using an extreme sampling technique, 88 participants were included in this analysis. After controlling for age, sex, and BMI, there were moderate positive associations between higher c-reactive protein (CRP; ߈ = .62; 95% confidence interval [CI] = -.03, 1.26), interleukin-6 (IL-6; ߈ = 3.13; CI = -.11, 6.38), and interleukin-10 (IL-10; ߈ = 2.51; CI = -.30, 5.32); and greater pain reduction from 48 to 96 hours post exercise muscle injury. Using an exploratory multivariable model to predict pain changes from 48 to 96 hours, we found participants with higher IL-10 were less likely to experience a high increase in pain (߈ = -10.77; CI = -21.25, -2.69). Study findings suggest CRP, IL-6, and IL-10 are related to shoulder pain change for a preclinical high-risk COMT × PCS subgroup. Future studies will translate to clinical shoulder pain and decipher the complex and seemingly pleiotropic interplay between inflammatory biomarkers and shoulder pain change. PERSPECTIVE: In a preclinical high-risk COMT × PCS subgroup, 3 circulating inflammatory biomarkers (CRP, IL-6, and IL-10) were moderately associated with pain improvement following exercise-induced muscle injury.

Biopsychosocial influence on shoulder pain: results from a randomized preclinical trial of exercise-induced muscle injury.

ABSTRACT: Prior cohort studies validated that a subgroup defined by a specific COMT genotype and pain catastrophizing is at increased risk for heightened responses to exercise-induced or surgically induced shoulder pain. In this clinical trial, we used our preclinical model of exercise-induced muscle injury and pain to test the efficacy of interventions matched to characteristics of this high-risk subgroup (ie, personalized medicine approach). Potential participants provided informed consent to be screened for eligibility based on subgroup membership and then, as appropriate, were enrolled into the trial. Participants (n = 261) were randomized to 1 of 4 intervention groups comprised of pharmaceutical (propranolol or placebo) and informational (general education or psychologic intervention) combinations. After muscle injury was induced, participants received randomly assigned treatment and were followed for the primary outcome of shoulder pain intensity recovery over 4 consecutive days. Recovery rates were 56.4% (placebo and psychologic intervention), 55.4% (placebo and general education), 62.9% (propranolol and psychologic intervention), and 56.1% (propranolol and general education). No statistical differences were found between intervention groups in the primary analyses. Additional analyses found no differences between these intervention groups when shoulder pain duration was an outcome, and no differential treatment responses were detected based on sex, race, or level of pain catastrophizing. This trial indicates that these treatments were not efficacious for this high-risk subgroup when shoulder pain was induced by exercise-induced muscle injury. Accordingly, this phenotype should only be used for prognostic purposes until additional trials are completed in clinical populations.

A High-Throughput Screening Platform Identifies Novel Combination Treatments for Malignant Peripheral Nerve Sheath Tumors.

Malignant peripheral nerve sheath tumors (MPNST) are soft-tissue sarcomas that are the leading cause of mortality in patients with Neurofibromatosis type 1 (NF1). Single chemotherapeutic agents have shown response rates ranging from 18% to 44% in clinical trials, so there is still a high medical need to identify chemotherapeutic combination treatments that improve clinical prognosis and outcome. We screened a collection of compounds from the NCATS Mechanism Interrogation PlatE (MIPE) library in three MPNST cell lines, using cell viability and apoptosis assays. We then tested whether compounds that were active as single agents were synergistic when screened as pairwise combinations. Synergistic combinations in vitro were further evaluated in patient-derived orthotopic xenograft/orthoxenograft (PDOX) athymic models engrafted with primary MPNST matching with their paired primary-derived cell line where synergism was observed. The high-throughput screening identified 21 synergistic combinations, from which four exhibited potent synergies in a broad panel of MPNST cell lines. One of the combinations, MK-1775 with Doxorubicin, significantly reduced tumor growth in a sporadic PDOX model (MPNST-SP-01; sevenfold) and in an NF1-PDOX model (MPNST-NF1-09; fourfold) and presented greater effects in TP53 mutated MPNST cell lines. The other three combinations, all involving Panobinostat (combined with NVP-BGT226, Torin 2, or Carfilzomib), did not reduce the tumor volume in vivo at noncytotoxic doses. Our results support the utility of our screening platform of in vitro and in vivo models to explore new therapeutic approaches for MPNSTs and identified that combination MK-1775 with Doxorubicin could be a good pharmacologic option for the treatment of these tumors.

Targeted exon skipping of NF1 exon 17 as a therapeutic for neurofibromatosis type I.

We investigated the feasibility of utilizing an exon-skipping approach as a genotype-dependent therapeutic for neurofibromatosis type 1 (NF1) by determining which NF1 exons might be skipped while maintaining neurofibromin protein expression and GTPase activating protein (GAP)-related domain (GRD) function. Initial in silico analysis predicted exons that can be skipped with minimal loss of neurofibromin function, which was confirmed by in vitro assessments utilizing an Nf1 cDNA-based functional screening system. Skipping of exons 17 or 52 fit our criteria, as minimal effects on protein expression and GRD activity were noted. Antisense phosphorodiamidate morpholino oligomers (PMOs) were utilized to skip exon 17 in human cell lines with patient-specific pathogenic variants in exon 17, c.1885G>A, and c.1929delG. PMOs restored functional neurofibromin expression. To determine the in vivo significance of exon 17 skipping, we generated a homozygous deletion of exon 17 in a novel mouse model. Mice were viable and exhibited a normal lifespan. Initial studies did not reveal the presence of tumor development; however, altered nesting behavior and systemic lymphoid hyperplasia was noted in peripheral lymphoid organs. Alterations in T and B cell frequencies in the thymus and spleen were identified. Hence, exon skipping should be further investigated as a therapeutic approach for NF1 patients with pathogenic variants in exon 17, as homozygous deletion of exon 17 is consistent with at least partial function of neurofibromin.

HUMAN STUDY COMT and DRD3 haplotype-associated pain intensity and acute care utilization in adult sickle cell disease.

A previous exploratory analysis of a COMT gene single-nucleotide polymorphism (SNP) and a DRD3 SNP by our group suggested possible contributions to pain-related acute care utilization in people with sickle cell disease (SCD). Our aim was to extend the analysis to gene-spanning haplotypes of COMT SNPs and DRD3 SNPs to investigate possible associations with pain intensity and pain-related acute care utilization in an SCD cohort. Genotyping was conducted, and clinical data were collected, including self-reported pain intensity using PAINReportIt® (average of current pain and least and worst in past 24 hours, average pain intensity [API]) and medical record-extracted, pain-related acute care utilization data of 130 adults with SCD. Haplotype blocks were identified based on linkage disequilibria (COMT = 7 haploblocks; DRD3 = 8 haploblocks). Regression analyses were tested for association between haplotypes and API and utilization, yielding several significant findings. For COMT block 1 (rs2075507, rs4646310, rs737865), the A-G-G haplotype was associated with higher API compared to the reference A-G-A (p = 0.02), whereas the A-A-A haplotype was associated with higher utilization (p = 0.02). For DRD3 block 2 (rs9817063, rs2134655, rs963468, and rs3773679), relative to reference T-C-G-C, the T-T-G-C haplotype was associated with higher utilization (p = 0.01). For DRD3 block 4 (rs167770, rs324029, and rs324023), the A-G-T haplotype was associated with higher API (p = 0.04) and utilization (p < 0.001) relative to reference G-A-T, whereas the A-A-T haplotype was associated with higher utilization (p = 0.01). We found COMT and DRD3 haplotypes associated with pain-related SCD features, suggesting that in future studies more emphasis be placed on cis effects of SNP alleles in evaluating genetic contributions to SCD pain and acute care utilization for pain.

Analysis of AVPR1A, thermal and pressure pain thresholds, and stress in sickle cell disease.

Aim: In patients with sickle cell disease (SCD), negative physical and emotional experiences result from intense chronic and acute pain episodes, but factors underlying these, and their interactions, are not well understood. The arginine vasopressin receptor 1a gene (AVPR1A) single nucleotide polymorphism rs10877969 has been previously associated with aspects of acute pain and stress related pain. In this study, we tested for associations between this SNP, thermal and pressure pain thresholds, clinical pain, and stress in people with SCD. Methods: 150 adults enrolled with SCD completed pain intensity measures (Average Pain Intensity, API) and the Perceived Stress Questionnaire (PSQ). Thermal and pressure pain threshold data were available from quantitative sensory testing (QST), and rs10877969 genotypes were obtained. Results: In models adjusted for age and gender, between rs10877969 genotypes, we observed no significant differences in thermal (cold, p = 0.66; heat, p = 0.91) and mechanical (pressure, p = 0.33) pain thresholds. The association of rs10877969 with API (p = 0.09) was borderline, but non-significant with PSQ (p = 0.51). The correlation between clinical pain and environmental stress was significant, r = 0.18, p = 0.024, however, the interaction of genotype and PSQ was not significant (p = 0.63). Conclusion: Clinical and experimental pain were not significantly associated with the rs10877969 genotype. The rs10877969 genotype did not moderate the correlation between environmental stress and clinical pain in this population. However, a trend toward a protective T allele effect on average pain rating in SCD warrants future exploration of this SNP/gene in SCD.

Survival and NF1 Analysis in a Cohort of Orthopedics Patients with Malignant Peripheral Nerve Sheath Tumors.

Neurofibromatosis type 1 (NF1) is an autosomal dominant tumor syndrome in which benign plexiform neurofibromas are at risk of transforming into malignant peripheral nerve sheath tumors (MPNSTs), a very rare soft-tissue sarcoma. The prognosis of patients with MPNSTs is poor, with most studies reporting <50% survival at five years. However, studies evaluating MPNSTs are limited and report heterogeneous results. Because no MPNST-specific evidence-based treatment guideline exists, individual institutional experiences are very informative to the field. The main objective of this study was to investigate and report MPNST prognostic clinical and genetic biomarkers from our institution's Orthopedics service experience treating 20 cases from 1992 to 2017. Most patients were treated with resection and adjuvant radiation. Extended follow-up, averaging 11.4 years (ranging 1.1 to 25.1), revealed excellent five-year survival rates: 70% for overall and 60% for metastatic disease. An S100 B immunonegative tumor phenotype was associated with a significantly worse outcome than MPNSTs with positive S100 B stain. In addition, NF1 gene mutation analysis was performed on 27 families with NF1 in which at least one affected family member developed MPNSTs. Of the 27 NF1 germline mutations, five were large deletions spanning (or nearly spanning) the gene (18.5%), substantially more than such deletions in NF1 in general, consistent with increased risk of MPNSTs in such cases.

Effects of Patient and Surgery Characteristics on Persistent Postoperative Pain: A Mediation Analysis.

OBJECTIVE: Acute postoperative pain intensity is associated with persistent postsurgical pain (PPP) risk. However, it remains unclear whether acute postoperative pain intensity mediates the relationship between clinical factors and persistent pain. MATERIALS AND METHODS: Participants from a mixed surgical population completed the Brief Pain Inventory and Pain Catastrophizing Scale before surgery, and the Brief Pain Inventory daily after surgery for 7 days and at 30 and 90 days after surgery. We considered mediation models using the mean of the worst pain intensities collected daily on each of postoperative days (PODs) 1 to 7 against outcomes of worst pain intensity at the surgical site endpoints reflecting PPP (POD 90) and subacute pain (POD 30). RESULTS: The analyzed cohort included 284 participants for the POD 90 outcome. For every unit increase of maximum acute postoperative pain intensity through PODs 1 to 7, there was a statistically significant increase of mean POD 90 pain intensity by 0.287 after controlling for confounding effects. The effects of female versus male sex (m=0.212, P=0.034), pancreatic/biliary versus colorectal surgery (m=0.459, P=0.012), thoracic cardiovascular versus colorectal surgery (m=0.31, P=0.038), every minute increase of anesthesia time (m=0.001, P=0.038), every unit increase of preoperative average pain score (m=0.012, P=0.015), and every unit increase of catastrophizing (m=0.044, P=0.042) on POD 90 pain intensity were mediated through acute PODs 1 to 7 postoperative pain intensity. DISCUSSION: Our results suggest the mediating relationship of acute postoperative pain on PPP may be predicated on select patient and surgical factors.

Slow Dynamics of Acute Postoperative Pain Intensity Time Series Determined via Wavelet Analysis Are Associated With the Risk of Severe Postoperative Day 30 Pain.

BACKGROUND: Evidence suggests that increased early postoperative pain (POP) intensities are associated with increased pain in the weeks following surgery. However, it remains unclear which temporal aspects of this early POP relate to later pain experience. In this prospective cohort study, we used wavelet analysis of clinically captured POP intensity data on postoperative days 1 and 2 to characterize slow/fast dynamics of POP intensities and predict pain outcomes on postoperative day 30. METHODS: The study used clinical POP time series from the first 48 hours following surgery from 218 patients to predict their mean POP on postoperative day 30. We first used wavelet analysis to approximate the POP series and to represent the series at different time scales to characterize the early temporal profile of acute POP in the first 2 postoperative days. We then used the wavelet coefficients alongside demographic parameters as inputs to a neural network to predict the risk of severe pain 30 days after surgery. RESULTS: Slow dynamic approximation components, but not fast dynamic detailed components, were linked to pain intensity on postoperative day 30. Despite imbalanced outcome rates, using wavelet decomposition along with a neural network for classification, the model achieved an F score of 0.79 and area under the receiver operating characteristic curve of 0.74 on test-set data for classifying pain intensities on postoperative day 30. The wavelet-based approach outperformed logistic regression (F score of 0.31) and neural network (F score of 0.22) classifiers that were restricted to sociodemographic variables and linear trajectories of pain intensities. CONCLUSIONS: These findings identify latent mechanistic information within the temporal domain of clinically documented acute POP intensity ratings, which are accessible via wavelet analysis, and demonstrate that such temporal patterns inform pain outcomes at postoperative day 30.

Patient and Procedural Determinants of Postoperative Pain Trajectories.

BACKGROUND: The primary goal of this study was to evaluate patterns in acute postoperative pain in a mixed surgical patient cohort with the hypothesis that there would be heterogeneity in these patterns. METHODS: This study included 360 patients from a mixed surgical cohort whose pain was measured across postoperative days 1 through 7. Pain was characterized using the Brief Pain Inventory. Primary analysis used group-based trajectory modeling to estimate trajectories/patterns of postoperative pain. Secondary analysis examined associations between sociodemographic, clinical, and behavioral patient factors and pain trajectories. RESULTS: Five distinct postoperative pain trajectories were identified. Many patients (167 of 360, 46%) were in the moderate-to-high pain group, followed by the moderate-to-low (88 of 360, 24%), high (58 of 360, 17%), low (25 of 360, 7%), and decreasing (21 of 360, 6%) pain groups. Lower age (odds ratio, 0.94; 95% CI, 0.91 to 0.99), female sex (odds ratio, 6.5; 95% CI, 1.49 to 15.6), higher anxiety (odds ratio, 1.08; 95% CI, 1.01 to 1.14), and more pain behaviors (odds ratio, 1.10; 95% CI, 1.02 to 1.18) were related to increased likelihood of being in the high pain trajectory in multivariable analysis. Preoperative and intraoperative opioids were not associated with postoperative pain trajectories. Pain trajectory group was, however, associated with postoperative opioid use (P < 0.001), with the high pain group (249.5 oral morphine milligram equivalents) requiring four times more opioids than the low pain group (60.0 oral morphine milligram equivalents). CONCLUSIONS: There are multiple distinct acute postoperative pain intensity trajectories, with 63% of patients reporting stable and sustained high or moderate-to-high pain over the first 7 days after surgery. These postoperative pain trajectories were predominantly defined by patient factors and not surgical factors.

Sensory and Psychological Factors Predict Exercise-Induced Shoulder Injury Responses in a High-Risk Phenotype Cohort.

Our prior studies identified a high-risk phenotype (ie, high pain sensitivity variant of the catechol-O-methyltransferase gene (Single Nucleotide Polymorphism [SNP] rs6269) and pain catastrophizing scores) for shoulder pain. The current study identified sensory and psychological predictors of heightened pain responses following exercise-induced shoulder injury. Healthy participants (N = 131) with the SNP rs6269 catechol-O-methyltransferase gene and Pain Catastrophizing Scale scores ≥5 underwent baseline sensory and psychological testing followed by an established shoulder fatigue protocol, to induce muscle injury. Movement-evoked pain, pain intensity, disability, and strength were assessed 24 hours postinjury. Demographic, sensory, and psychological variables were included as predictors in full and parsimonious models for each outcome. The highest variance explained was for the shoulder disability outcome (full model R2 = .20, parsimonious R2 = .13). In parsimonious models, the individual predictors identified were: 1) 1st pulse heat pain sensitivity for isometric shoulder movement-evoked pain and pain intensity; 2) pressure pain threshold for shoulder disability; 3) fear of pain for active shoulder movement-evoked pain and shoulder disability; and 4) depressive symptoms for shoulder strength. Findings indicate specific pain sensitivity and psychological measures may have additional prognostic value for self-reported disability within a high-risk phenotype. These findings should be tested in a clinical cohort for validation. PERSPECTIVE: The current study extends previous work by providing insight regarding how poor shoulder outcomes may develop within a high-risk phenotype. Specifically, 1st pulse heat pain sensitivity and pressure pain threshold were sensory measures, and fear of pain and depressive symptoms were psychological measures, that improved prediction of different shoulder outcomes.

Genetics of human malignant peripheral nerve sheath tumors.

Malignant peripheral nerve sheath tumors (MPNSTs) are heterogeneous, highly aggressive tumors with no widely effective treatment other than surgery. Genomic architecture of MPNST is similar to other soft tissue sarcomas, with a relatively modest burden of single nucleotide variants and an elevated frequency of copy-number alterations. Recent advances in genomic studies identified previously unrecognized critical involvement of polycomb repressor complex 2 (PRC2) core components SUZ12 and EED in transition to malignancy. Notably, somatic changes in NF1, CDKN2A/B, and PRC2 are found in most MPNST regardless of their etiology (e.g. neurofibromatosis type 1-associated vs. sporadic vs. radiation-induced), indicating that similar molecular mechanisms impact pathogenesis in these neoplasms. The timing and specific order of genetic or epigenetic changes may, however, explain the typically poorer prognosis of NF1-associated MPNSTs. Studies that reveal genes and regulatory pathways uniquely altered in malignancies are essential to development of targeted tumor therapies. Characterization of MPNST molecular profiles may also contribute to tools for earlier detection, and prediction of prognosis or drug response. Here we review the genetic discoveries and their implications in understanding MPNST biology.

Association of P2RX7 functional variants with localized aggressive periodontitis.

OBJECTIVE: The purpose of this study was to investigate involvement of the P2X7 receptor in the rare condition, localized aggressive periodontitis. MATERIAL AND METHODS: Peripheral blood from 220 African Americans (103 with localized aggressive periodontitis and 117 healthy unrelated controls) was stimulated with lipopolysaccharide from E coli and Porphyromonas gingivalis. P2RX7 single nucleotide polymorphisms rs208294 (H155Y), rs1718119 (T348A), rs2230911 (T357S) and rs3751143 (E496A) were genotyped in 103 localized aggressive periodontitis patients and 117 healthy unrelated subjects. We examined genetic association between four P2RX7 single nucleotide polymorphisms and localized aggressive periodontitis, and tested for correlations between the single nucleotide polymorphisms and inflammatory response to lipopolysaccharide in blood samples from these patients. RESULTS: A significant association with localized aggressive periodontitis was observed with rs1718119 A (Thr) allele (P = 0.0063, odds ratio = 1.904) and with a haplotype containing this allele (P = 0.0075). Additionally, significant correlations with these data were found: the rs1718119 G allele correlated with greater production of IL-6, IL-2 and GM-CSF; the C (His) allele of rs208294 correlated with lower levels of IL-12p40; and the C (Thr) allele of rs2230911 correlated with greater levels of G-CSF. CONCLUSION: The data from these analyses support a possible biological relationship between P2RX7 genetic variants and inflammatory response in localized aggressive periodontitis patients.

NF1 patient missense variants predict a role for ATM in modifying neurofibroma initiation.

In Neurofibromatosis type 1, NF1 gene mutations in Schwann cells (SC) drive benign plexiform neurofibroma (PNF), and no additional SC changes explain patient-to-patient variability in tumor number. Evidence from twin studies suggests that variable expressivity might be caused by unidentified modifier genes. Whole exome sequencing of SC and fibroblast DNA from the same resected PNFs confirmed biallelic SC NF1 mutations; non-NF1 somatic SC variants were variable and present at low read number. We identified frequent germline variants as possible neurofibroma modifier genes. Genes harboring variants were validated in two additional cohorts of NF1 patients and by variant burden test. Genes including CUBN, CELSR2, COL14A1, ATR and ATM also showed decreased gene expression in some neurofibromas. ATM-relevant DNA repair defects were also present in a subset of neurofibromas with ATM variants, and in some neurofibroma SC. Heterozygous ATM G2023R or homozygous S707P variants reduced ATM protein expression in heterologous cells. In mice, genetic Atm heterozygosity promoted Schwann cell precursor self-renewal and increased tumor formation in vivo, suggesting that ATM variants contribute to neurofibroma initiation. We identify germline variants, rare in the general population, overrepresented in NF1 patients with neurofibromas. ATM and other identified genes are candidate modifiers of PNF pathogenesis.

OPRM1, OPRK1, and COMT genetic polymorphisms associated with opioid effects on experimental pain: a randomized, double-blind, placebo-controlled study.

Genetic polymorphisms have been shown to affect opioid requirement for pain relief. However, true genetic effect is often difficult to assess due to underlying pain conditions and placebo effects. The goal of this study was to understand how common polymorphisms affect opioid effects while controlling for these factors. A randomized, double-blind, placebo-controlled study was implemented to assess how opioid effects are modulated by COMT (rs6269, rs4633, rs4848, rs4680), OPRM1 (A118G), and OPRK1 (rs1051660, rs702764, rs16918875). One hundred and eight healthy subjects underwent experimental pain testing before and after morphine, butorphanol, and placebo (saline). Association analysis was performed between polymorphisms/haplotypes and opioid response, while correcting for race, gender, placebo effects, and multiple comparisons. Pressure pain was significantly associated with rs6269 and rs4633 following butorphanol. The AA genotype of rs4680 or A_T_C_A/ A_T_C_A (rs6269_rs4633_ rs4818_rs4680) diplotype of COMT, combined with the AG genotype of OPRM1 A118G, showed significantly increased pressure pain threshold from butorphanol. Opioid effects on pressure, ischemic, heat pain, and side effects were nominally associated with several SNPs and haplotypes. Effects were often present in one opioid but not the other. This indicates that these polymorphisms affect pain relief from opioids, and that their effects are opioid and pain modality specific.

Assessment of nociception and related quality-of-life measures in a porcine model of neurofibromatosis type 1.

Neurofibromatosis type 1 (NF1) is an autosomal dominant genetic disorder resulting from germline mutations in the NF1 gene, which encodes neurofibromin. Patients experience a variety of symptoms, but pain in the context of NF1 remains largely underrecognized. Here, we characterize nociceptive signaling and pain behaviors in a miniswine harboring a disruptive NF1 mutation (exon 42 deletion). We present the first characterization of pain-related behaviors in a pig model of NF1, identifying unchanged agitation scores, lower tactile thresholds (allodynia), and decreased response latencies to thermal laser stimulation (hyperalgesia) in NF1 (females only) pigs. Male NF1 pigs with tumors showed reduced sleep quality and increased resting, 2 health-related quality-of-life symptoms found to be comorbid in people with NF1 pain. We explore these phenotypes in relationship to suppression of the increased activity of the N-type voltage-gated calcium (CaV2.2) channel by pharmacological antagonism of phosphorylation of a regulatory protein-the collapsin response mediator protein 2 (CRMP2), a known interactor of neurofibromin, and by targeting the interface between the α subunit of CaV2.2 and the accessory ß-subunits with small molecules. Our data support the use of NF1 pigs as a large animal model for studying NF1-associated pain and for understanding the pathophysiology of NF1. Our findings demonstrate the translational potential of 2 small molecules in reversing ion channel remodeling seen in NF1. Interfering with CaV2.2, a clinically validated target for pain management, might also be a promising therapeutic strategy for NF1-related pain management.

EZH1/2 function mostly within canonical PRC2 and exhibit proliferation-dependent redundancy that shapes mutational signatures in cancer.

Genetic mutations affecting chromatin modifiers are widespread in cancers. In malignant peripheral nerve sheath tumors (MPNSTs), Polycomb repressive complex 2 (PRC2), which plays a crucial role in gene silencing, is inactivated through recurrent mutations in core subunits embryonic ectoderm development (EED) and suppressor of zeste 12 homolog (SUZ12), but mutations in PRC2's main catalytic subunit enhancer of zeste homolog 2 (EZH2) have never been found. This is in contrast to myeloid and lymphoid malignancies, which harbor frequent loss-of-function mutations in EZH2. Here, we investigated whether the absence of EZH2 mutations in MPNST is due to a PRC2-independent (i.e., noncanonical) function of the enzyme or to redundancy with EZH1. We show that, in the absence of SUZ12, EZH2 remains bound to EED but loses its interaction with all other core and accessory PRC2 subunits. Through genetic and pharmacological analyses, we unambiguously establish that EZH2 is functionally inert in this context, thereby excluding a PRC2-independent function. Instead, we show that EZH1 and EZH2 are functionally redundant in the slowly proliferating MPNST precursors. We provide evidence that the compensatory function of EZH1 is alleviated upon higher proliferation. This work reveals how context-dependent redundancies can shape tumor-type specific mutation patterns in chromatin regulators.

Low mutation burden and frequent loss of CDKN2A/B and SMARCA2, but not PRC2, define premalignant neurofibromatosis type 1-associated atypical neurofibromas.

BACKGROUND: Neurofibromatosis type 1 (NF1) is a tumor-predisposition disorder caused by germline mutations in NF1. NF1 patients have an 8-16% lifetime risk of developing a malignant peripheral nerve sheath tumor (MPNST), a highly aggressive soft-tissue sarcoma, often arising from preexisting benign plexiform neurofibromas (PNs) and atypical neurofibromas (ANFs). ANFs are distinct from both PN and MPNST, representing an intermediate step in malignant transformation. METHODS: In the first comprehensive genomic analysis of ANF originating from multiple patients, we performed tumor/normal whole-exome sequencing (WES) of 16 ANFs. In addition, we conducted WES of 3 MPNSTs, copy-number meta-analysis of 26 ANFs and 28 MPNSTs, and whole transcriptome sequencing analysis of 5 ANFs and 5 MPNSTs. RESULTS: We identified a low number of mutations (median 1, range 0-5) in the exomes of ANFs (only NF1 somatic mutations were recurrent), and frequent deletions of CDKN2A/B (69%) and SMARCA2 (42%). We determined that polycomb repressor complex 2 (PRC2) genes EED and SUZ12 were frequently mutated, deleted, or downregulated in MPNSTs but not in ANFs. Our pilot gene expression study revealed upregulated NRAS, MDM2, CCND1/2/3, and CDK4/6 in ANFs and MPNSTs, and overexpression of EZH2 in MPNSTs only. CONCLUSIONS: The PN-ANF transition is primarily driven by the deletion of CDKN2A/B. Further progression from ANF to MPNST likely involves broad chromosomal rearrangements and frequent inactivation of the PRC2 genes, loss of the DNA repair genes, and copy-number increase of signal transduction and cell-cycle and pluripotency self-renewal genes.

Genetic and psychological factors interact to predict physical impairment phenotypes following exercise-induced shoulder injury.

BACKGROUND: We investigated interactions between genetic and psychological factors in predicting shoulder impairment phenotypes. We hypothesized that pro-inflammatory genes would display stronger relationships compared with pain-related genes when combined with psychological factors for predicting phenotypic changes. SUBJECTS AND METHODS: Altogether, 190 participants completed a 5-day experimental protocol. An experimental shoulder injury model was used to induce physical impairment, and a priori selected genetic (pain-related, pro-inflammatory) and psychological (anxiety, depressive symptoms, pain catastrophizing, fear of pain, kinesiophobia) factors were included as predictors of interest. Impairment phenotypes were injury-induced deficits in range of motion (ROM) and strength. After controlling for age, sex, and race, genetic and psychological predictors were entered separately as main effects and interaction terms in regression models for each phenotype. RESULTS: Strong statistical evidence was provided for interactions between: 1) IL-1ß (rs1143634) and fear of pain for predicting loss of shoulder flexion and abduction, 2) IL-1ß (rs1143634) and anxiety for predicting loss of flexion, and 3) IL-1ß (rs1143634) and depressive symptoms for predicting loss of internal rotation. In addition, the interaction between OPRM1 (rs1799971) and fear of pain as well as COMT (rs4818) and pain catastrophizing provided strong statistical evidence for predicting strength loss. CONCLUSION: Pro-inflammatory gene variants contributed more to physical impairment with two single nucleotide polymorphisms (SNPs; IL-1ß [rs1143634] and TNF/LTA [rs2229094]) interacting with psychological factors to predict six shoulder impairment phenotypes. In comparison, two pain-related gene SNPs (OPRM1 [rs1799971] and COMT [rs4818]) interacted with psychological factors to predict four shoulder impairment phenotypes (abduction: 5-day average loss; strength loss: 5-day average, peak, and relative loss).

Pharmacological and genomic profiling of neurofibromatosis type 1 plexiform neurofibroma-derived schwann cells.

Neurofibromatosis type I (NF1) is an autosomal dominant genetic condition characterized by peripheral nervous system tumors (PNSTs), including plexiform neurofibromas (pNFs) that cause nerve dysfunction, deformity, pain damage to adjacent structures, and can undergo malignant transformation. There are no effective therapies to prevent or treat pNFs. Drug discovery efforts are slowed by the 'benign' nature of the Schwann cells that are the progenitor cells of pNF. In this work we characterize a set of pNF-derived cell lines at the genomic level (via SNP Arrays, RNAseq, and Whole Exome- Sequencing), and carry out dose response-based quantitative high-throughput screening (qHTS) with a collection of 1,912 oncology-focused compounds in a 1536-well microplate cell proliferation assays. Through the characterization and screening of NF1-/-, NF1+/+ and NF1+/- Schwann cell lines, this resource introduces novel therapeutic avenues for the development for NF1 associated pNF as well as all solid tumors with NF1 somatic mutations. The integrated data sets are openly available for further analysis at http://www.synapse.org/pnfCellCulture.