Targeted therapy for LIMD1-deficient non-small cell lung cancer subtypes.

An early event in lung oncogenesis is loss of the tumour suppressor gene LIMD1 (LIM domains containing 1); this encodes a scaffold protein, which suppresses tumorigenesis via a number of different mechanisms. Approximately 45% of non-small cell lung cancers (NSCLC) are deficient in LIMD1, yet this subtype of NSCLC has been overlooked in preclinical and clinical investigations. Defining therapeutic targets in these LIMD1 loss-of-function patients is difficult due to a lack of 'druggable' targets, thus alternative approaches are required. To this end, we performed the first drug repurposing screen to identify compounds that confer synthetic lethality with LIMD1 loss in NSCLC cells. PF-477736 was shown to selectively target LIMD1-deficient cells in vitro through inhibition of multiple kinases, inducing cell death via apoptosis. Furthermore, PF-477736 was effective in treating LIMD1-/- tumours in subcutaneous xenograft models, with no significant effect in LIMD1+/+ cells. We have identified a novel drug tool with significant preclinical characterisation that serves as an excellent candidate to explore and define LIMD1-deficient cancers as a new therapeutic subgroup of critical unmet need.

miR-132 suppresses transcription of ribosomal proteins to promote protective Th1 immunity.

Determining the mechanisms that distinguish protective immunity from pathological chronic inflammation remains a fundamental challenge. miR-132 has been shown to play largely immunoregulatory roles in immunity; however, its role in CD4+ T cell function is poorly understood. Here, we show that CD4+ T cells express high levels of miR-132 and that T cell activation leads to miR-132 up-regulation. The transcriptomic hallmark of splenic CD4+ T cells lacking the miR-132/212 cluster during chronic infection is an increase in mRNA levels of ribosomal protein (RP) genes. BTAF1, a co-factor of B-TFIID and novel miR-132/212-3p target, and p300 contribute towards miR-132/212-mediated regulation of RP transcription. Following infection with Leishmania donovani, miR-132-/- CD4+ T cells display enhanced expression of IL-10 and decreased IFNγ. This is associated with reduced hepatosplenomegaly and enhanced pathogen load. The enhanced IL-10 expression in miR-132-/- Th1 cells is recapitulated in vitro following treatment with phenylephrine, a drug reported to promote ribosome synthesis. Our results uncover that miR-132/212-mediated regulation of RP expression is critical for optimal CD4+ T cell activation and protective immunity against pathogens.

A HIF-LIMD1 negative feedback mechanism mitigates the pro-tumorigenic effects of hypoxia.

The adaptive cellular response to low oxygen tensions is mediated by the hypoxia-inducible factors (HIFs), a family of heterodimeric transcription factors composed of HIF-α and HIF-ß subunits. Prolonged HIF expression is a key contributor to cellular transformation, tumorigenesis and metastasis. As such, HIF degradation under hypoxic conditions is an essential homeostatic and tumour-suppressive mechanism. LIMD1 complexes with PHD2 and VHL in physiological oxygen levels (normoxia) to facilitate proteasomal degradation of the HIF-α subunit. Here, we identify LIMD1 as a HIF-1 target gene, which mediates a previously uncharacterised, negative regulatory feedback mechanism for hypoxic HIF-α degradation by modulating PHD2-LIMD1-VHL complex formation. Hypoxic induction of LIMD1 expression results in increased HIF-α protein degradation, inhibiting HIF-1 target gene expression, tumour growth and vascularisation. Furthermore, we report that copy number variation at the LIMD1 locus occurs in 47.1% of lung adenocarcinoma patients, correlates with enhanced expression of a HIF target gene signature and is a negative prognostic indicator. Taken together, our data open a new field of research into the aetiology, diagnosis and prognosis of LIMD1-negative lung cancers.

MicroRNA-155 induction via TNF-α and IFN-γ suppresses expression of programmed death ligand-1 (PD-L1) in human primary cells.

Programmed death ligand-1 (PD-L1) is a critical regulator of T cell function contributing to peripheral immune tolerance. Although it has been shown that posttranscriptional regulatory mechanisms control PD-L1 expression in cancer, it remains unknown whether such regulatory loops operate also in non-transformed cells. Here we studied PD-L1 expression in human dermal lymphatic endothelial cells (HDLECs), which play key roles in immunity and cancer. Treatment of HDLECs with the pro-inflammatory cytokines IFN-γ and TNF-α synergistically up-regulated PD-L1 expression. IFN-γ and TNF-α also affected expression of several microRNAs (miRNAs) that have the potential to suppress PD-L1 expression. The most highly up-regulated miRNA following IFN-γ and TNF-α treatment in HDLECs was miR-155, which has a central role in the immune system and cancer. Induction of miR-155 was driven by TNF-α, the effect of which was significantly enhanced by IFN-γ. The PD-L1 3'-UTR contains two functional miR-155-binding sites. Endogenous miR-155 controlled the kinetics and maximal levels of PD-L1 induction upon IFN-γ and TNF-α treatments. We obtained similar findings in dermal fibroblasts, demonstrating that the IFN-γ/TNF-α/miR-155/PD-L1 pathway is not restricted to HDLECs. These results reveal miR-155 as a critical component of an inflammation-induced regulatory loop controlling PD-L1 expression in primary cells.

Argonaute Utilization for miRNA Silencing Is Determined by Phosphorylation-Dependent Recruitment of LIM-Domain-Containing Proteins.

As core components of the microRNA-induced silencing complex (miRISC), Argonaute (AGO) proteins interact with TNRC6 proteins, recruiting other effectors of translational repression/mRNA destabilization. Here, we show that LIMD1 coordinates the assembly of an AGO-TNRC6 containing miRISC complex by binding both proteins simultaneously at distinct interfaces. Phosphorylation of AGO2 at Ser 387 by Akt3 induces LIMD1 binding, which in turn enables AGO2 to interact with TNRC6A and downstream effector DDX6. Conservation of this serine in AGO1 and 4 indicates this mechanism may be a fundamental requirement for AGO function and miRISC assembly. Upon CRISPR-Cas9-mediated knockout of LIMD1, AGO2 miRNA-silencing function is lost and miRNA silencing becomes dependent on a complex formed by AGO3 and the LIMD1 family member WTIP. The switch to AGO3 utilization occurs due to the presence of a glutamic acid residue (E390) on the interaction interface, which allows AGO3 to bind to LIMD1, AJUBA, and WTIP irrespective of Akt signaling.

Is patient assessment of asthma care delivery associated with publicly reported performance measures?

OBJECTIVE: To determine the association between patient assessment of healthcare delivery and publicly reported asthma performance measures. METHODS: We identified individuals 5-50 years old who had asthma quality measures reported to the Minnesota Community Measurement© (MCM©) by their clinic and sent them a postal survey which included within it the Patient Assessment of Chronic Illness care (PACIC), a validated measure of patient perception of the quality of healthcare delivery. We performed a multivariable analysis to examine the association between PACIC scores and achievement of asthma care quality measures. RESULTS: The response rate for the ACS was 102/367 (28%); a non-response bias analysis revealed no differences between responders and non-responders for age, gender and asthma control. Most responders (73%) reported taking asthma medications daily and most (71%) had poorly controlled asthma. The PACIC score was not associated with any of the asthma quality measures based on the data reported to MCM© by the clinic. A higher PACIC score was, however, associated with having an asthma action plan based on patient-reported data in the ACS (p < 0.0001) but not with patient-reported asthma control or emergency department/hospitalizations for asthma. CONCLUSIONS: Patient assessment of high quality asthma care delivery was associated with patient self-report of having an asthma action plan but was not associated with any of the publicly reported asthma performance measures.

Predicting which medication classes interfere with allergy skin testing.

Medications often interfere with allergy skin test interpretation. This study was performed to determine which medications interfere with allergy skin tests. We retrospectively reviewed skin-prick test results from patients who had discontinued H(1)-antagonists, tricyclic antidepressants (TCAs), selective serotonin reuptake inhibitors (SSRIs), selective norepinephrine reuptake inhibitors (SNRIs), benzodiazepines, atypical antidepressants, antipsychotics, hypnotics, sedatives, proton pump inhibitors (PPIs), and H(2)-antagonists between 0 and 7 days before allergy skin testing. Ninety-seven subjects had taken second-generation H(1)-antihistamines within 7 days of skin testing; all patients who had stopped 3 days before testing had positive histamine controls. Two hundred sixty-eight skin tests performed on patients taking a single medication of interest showed that patients had the following percentages of a positive histamine control: TCAs, 56.5%; SNRIs, 100%; H(2)-blockers, 100%; SSRIs, 97%; PPIs, 97%; benzodiazepines, 85.7%; and atypical antidepressants/sedatives, 92.6%. The 580 patients taking multiple medications of interest showed that the odds ratio and 95% confidence intervals of a negative histamine test for patients taking TCAs were 6.33 (2.11-20.5), for H(1)-blockers were 4.95 (1.78-15.1), for benzodiazepines were 5.01 (1.72-15.80), for atypical antidepressants/sedatives were 3.11 (1.09-9.61), and for H(2)-blockers were 2.91 (0.97-9.37). The odds of a negative histamine test for SSRIs, SNRIs, or PPIs were not significantly increased. SSRIs, SNRIs, and PPIs are unlikely to interfere with skin testing. TCAs, H(1)-blockers, benzodiazepines, quetiapine, and mirtazapine should be discontinued temporarily if clinically able. H(2)-antagonists, bupropion, eszopiclone, trazodone, or zolpidem showed minimal interference with immediate hypersensitivity skin test histamine response.