Endocrine and cellular physiology and pathology of the insulin-like growth factor acid-labile subunit.

The acid-labile subunit (ALS) of the insulin-like growth factor (IGF) binding protein (IGFBP) complex, encoded in humans by IGFALS, has a vital role in regulating the endocrine transport and bioavailability of IGF-1 and IGF-2. Accordingly, ALS has a considerable influence on postnatal growth and metabolism. ALS is a leucine-rich glycoprotein that forms high-affinity ternary complexes with IGFBP-3 or IGFBP-5 when they are occupied by either IGF-1 or IGF-2. These complexes constitute a stable reservoir of circulating IGFs, blocking the potentially hypoglycaemic activity of unbound IGFs. ALS is primarily synthesized by hepatocytes and its expression is lower in non-hepatic tissues. ALS synthesis is strongly induced by growth hormone and suppressed by IL-1ß, thus potentially serving as a marker of growth hormone secretion and/or activity and of inflammation. IGFALS mutations in humans and Igfals deletion in mice cause modest growth retardation and pubertal delay, accompanied by decreased osteogenesis and enhanced adipogenesis. In hepatocellular carcinoma, IGFALS is described as a tumour suppressor; however, its contribution to other cancers is not well delineated. This Review addresses the endocrine physiology and pathology of ALS, discusses the latest cell and proteomic studies that suggest emerging cellular roles for ALS and outlines its involvement in other disease states.

Augmented Reality in Interventional Radiology: Transforming Training Paradigms.

The ascent of medical technology places augmented reality (AR) at the forefront of potential advancements in interventional radiology (IR) training. This review delves into the symbiotic relationship between AR and conventional IR training, casting light on the opportunities and hurdles intrinsic to this integration. A targeted literature review was conducted using the databases PubMed, Cochrane Library, and Embase. Search terms included ((("Augmented Reality" OR "Virtual Reality")) AND ((Education OR Training))) AND (("Interventional Radiology")). Ten studies identified using the comprehensive inclusion criteria helped scrutinize the use of AR in IR training. Key outcomes include improved procedural accuracy, reduced training duration, and heightened trainee confidence. However, it also identifies limitations such as small sample sizes, reliance on simulation environments, and technological constraints in AR implementation. Despite these challenges, the review underscored the transformative potential of AR in IR education, suggesting its capacity to revolutionize training methodologies. However, it also calls for continued technological development and empirical research to address current challenges and fully leverage AR's capabilities in medical education.

Outcomes from a collaborative project developing and evaluating a community rehabilitation worker program for Northwestern Ontario First Nations.

INTRODUCTION: Major inequities exist in levels of health and wellbeing, availability, and access to healthcare services between seniors of Indigenous and non-Indigenous background in Ontario. First Nations elders are 45-55% more frail than the average senior in Ontario. Additionally, needed rehabilitation services are not easily accessible or available in the first language of most First Nations elders within their home communities. A literature review demonstrated community-based rehabilitation assistant models had been successfully developed and implemented in regions facing similar equity and access challenges. Building on these findings, a needs assessment was conducted to capture unique needs and requirements in Northwestern Ontario relating to rehabilitation among First Nations elders. METHODS: The needs assessment resulted in four First Nations, three Indigenous health organizations, three rehabilitation health organizations, and two academic institutions iteratively developing and evaluating curriculum for a Community Rehabilitation Worker (CRW) program in treaty territories 5, 9, and Robinson-Superior. The goal of the program is to train local CRWs, familiar with local languages and cultures, to provide rehabilitative services that support ageing in place, health, wellbeing, and quality of life for First Nations elders. The study employed a community participatory action research approach aligning with the OCAP® (Ownership, Control, Access, and Possession) framework for working with Indigenous populations. Seventeen community partners were active participants in the program development, evaluation, and adaptation of the CRW curriculum. Feedback was received through advisory committee meetings, surveys, and individual and group interviews. RESULTS: All 101 participants agreed, across all curriculum modules, that (1) the time allotment was realistic; (2) instructional materials, activities, and resources were appropriate and easy to understand; (3) evaluation activities accurately measured learning; and (4) participants identifying as Indigenous felt that Indigenous culture was adequately reflected. The qualitative findings highlighted the importance of incorporating culture, spirituality, traditions, local language use, and reintegration of First Nations elders into traditional activities and community activities for both the CRW curriculum and rehabilitation efforts. The need for locally available First Nations, elder-focused mental health support, transportation options, and gathering spaces such as those commonly seen in urban areas was also highlighted. CONCLUSION: The process of iteratively developing and evaluating a CRW program resulted in a Northwestern Ontario college welcoming the first cohort of students to the CRW program in March 2022. The program is co-facilitated with a First Nations Elder and includes components of local culture, language, and the reintegration of First Nations elders into community as part of the rehabilitation efforts. In addition, to appropriately support the quality of life, health, and wellbeing of First Nations elders, the project team called upon provincial and federal governments to work with First Nations to make available dedicated funding to address inequities in resources available to First Nations elders in Northwestern Ontario urban and First Nations remote communities. This included elder-focused transportation options, mental health services, and gathering places. The program implementation will be evaluated with the first cohort of CRWs for further adaptations considering potential scale and spread. As such, the project and findings may also represent a resource for others wishing to pursue similar development using participatory approaches in rural and remote communities both nationally and internationally.

Patient emergency health-care use before hospital admission for COVID-19 and long-term outcomes in Scotland: a national cohort study.

BACKGROUND: It is unclear what effect the pattern of health-care use before admission to hospital with COVID-19 (index admission) has on the long-term outcomes for patients. We sought to describe mortality and emergency readmission to hospital after discharge following the index admission (index discharge), and to assess associations between these outcomes and patterns of health-care use before such admissions. METHODS: We did a national, retrospective, complete cohort study by extracting data from several national databases and linking the databases for all adult patients admitted to hospital in Scotland with COVID-19. We used latent class trajectory modelling to identify distinct clusters of patients on the basis of their emergency admissions to hospital in the 2 years before the index admission. The primary outcomes were mortality and emergency readmission up to 1 year after index admission. We used multivariable regression models to explore associations between these outcomes and patient demographics, vaccination status, level of care received in hospital, and previous emergency hospital use. FINDINGS: Between March 1, 2020, and Oct 25, 2021, 33 580 patients were admitted to hospital with COVID-19 in Scotland. Overall, the Kaplan-Meier estimate of mortality within 1 year of index admission was 29·6% (95% CI 29·1-30·2). The cumulative incidence of emergency hospital readmission within 30 days of index discharge was 14·4% (95% CI 14·0-14·8), with the number increasing to 35·6% (34·9-36·3) patients at 1 year. Among the 33 580 patients, we identified four distinct patterns of previous emergency hospital use: no admissions (n=18 772 [55·9%]); minimal admissions (n=12 057 [35·9%]); recently high admissions (n=1931 [5·8%]), and persistently high admissions (n=820 [2·4%]). Patients with recently or persistently high admissions were older, more multimorbid, and more likely to have hospital-acquired COVID-19 than patients with no or minimal admissions. People in the minimal, recently high, and persistently high admissions groups had an increased risk of mortality and hospital readmission compared with those in the no admissions group. Compared with the no admissions group, mortality was highest in the recently high admissions group (post-hospital mortality HR 2·70 [95% CI 2·35-2·81]; p<0·0001) and the risk of readmission was highest in the persistently high admissions group (3·23 [2·89-3·61]; p<0·0001). INTERPRETATION: Long-term mortality and readmission rates for patients hospitalised with COVID-19 were high; within 1 year, one in three patients had died and a third had been readmitted as an emergency. Patterns of hospital use before index admission were strongly predictive of mortality and readmission risk, independent of age, pre-existing comorbidities, and COVID-19 vaccination status. This increasingly precise identification of individuals at high risk of poor outcomes from COVID-19 will enable targeted support. FUNDING: Chief Scientist Office Scotland, UK National Institute for Health Research, and UK Research and Innovation.

Growing dendrites enhance a neuron's computational power and memory capacity.

Neocortical pyramidal neurons have many dendrites, and such dendrites are capable of, in isolation of one-another, generating a neuronal spike. It is also now understood that there is a large amount of dendritic growth during the first years of a humans life, arguably a period of prodigious learning. These observations inspire the construction of a local, stochastic algorithm based on an earlier stochastic, homeostatic, Hebbian developmental theory. Here we investigate the neurocomputational advantages and limits on this novel algorithm that combines dendritogenesis with supervised adaptive synaptogenesis. Neurons created with this algorithm have enhanced memory capacity, can avoid catastrophic interference (forgetting), and have the ability to unmix mixture distributions. In particular, individual dendrites develop within each class, in an unsupervised manner, to become feature-clusters that correspond to the mixing elements of class-conditional mixture distribution. Error-free classification is demonstrated with input perturbations up to 40%. Although discriminative problems are used to understand the capabilities of the stochastic algorithm and the neuronal connectivity it produces, the algorithm is in the generative class, it thus seems ideal for decisions that require generalization, i.e., extrapolation beyond previous learning.

Signaling Pathways of the Insulin-like Growth Factor Binding Proteins.

The 6 high-affinity insulin-like growth factor binding proteins (IGFBPs) are multifunctional proteins that modulate cell signaling through multiple pathways. Their canonical function at the cellular level is to impede access of insulin-like growth factor (IGF)-1 and IGF-2 to their principal receptor IGF1R, but IGFBPs can also inhibit, or sometimes enhance, IGF1R signaling either through their own post-translational modifications, such as phosphorylation or limited proteolysis, or by their interactions with other regulatory proteins. Beyond the regulation of IGF1R activity, IGFBPs have been shown to modulate cell survival, migration, metabolism, and other functions through mechanisms that do not appear to involve the IGF-IGF1R system. This is achieved by interacting directly or functionally with integrins, transforming growth factor ß family receptors, and other cell-surface proteins as well as intracellular ligands that are intermediates in a wide range of pathways. Within the nucleus, IGFBPs can regulate the diverse range of functions of class II nuclear hormone receptors and have roles in both cell senescence and DNA damage repair by the nonhomologous end-joining pathway, thus potentially modifying the efficacy of certain cancer therapeutics. They also modulate some immune functions and may have a role in autoimmune conditions such as rheumatoid arthritis. IGFBPs have been proposed as attractive therapeutic targets, but their ubiquity in the circulation and at the cellular level raises many challenges. By understanding the diversity of regulatory pathways with which IGFBPs interact, there may still be therapeutic opportunities based on modulation of IGFBP-dependent signaling.

Noncoding RNA actions through IGFs and IGF binding proteins in cancer.

The insulin-like growth factors (IGFs) and their regulatory proteins-IGF receptors and binding proteins-are strongly implicated in cancer progression and modulate cell survival and proliferation, migration, angiogenesis and metastasis. By regulating the bioavailability of the type-1 IGF receptor (IGF1R) ligands, IGF-1 and IGF-2, the IGF binding proteins (IGFBP-1 to -6) play essential roles in cancer progression. IGFBPs also influence cell communications through pathways that are independent of IGF1R activation. Noncoding RNAs (ncRNAs), which encompass a variety of RNA types including microRNAs (miRNAs) and long-noncoding RNAs (lncRNAs), have roles in multiple oncogenic pathways, but their many points of intersection with IGF axis functions remain to be fully explored. This review examines the functional interactions of miRNAs and lncRNAs with IGFs and their binding proteins in cancer, and reveals how the IGF axis may mediate ncRNA actions that promote or suppress cancer. A better understanding of the links between ncRNA and IGF pathways may suggest new avenues for prognosis and therapeutic intervention in cancer. Further, by exploring examples of intersecting ncRNA-IGF pathways in non-cancer conditions, it is proposed that new opportunities for future discovery in cancer control may be generated.

Abundance and accessibility of forage for reindeer in forests of Northern Sweden: Impacts of landscape and winter climate regime.

The survival of reindeer during winter, their period of greatest food stress, depends largely on the abundance and accessibility of forage in their pastures. In Northern Sweden, realized availability of forage is notably affected by snow conditions and the impacts of forestry. While these factors have been examined in isolation, their combined effect has, to the best of our knowledge to date, not been researched. In this study, vegetation surveys and analysis of snow conditions were undertaken in forest stands at various stages of recovery from clear-cutting. The variation in abundance and growth of understory species edible by reindeer, such as lichen, was noted as forests matured. The barrier effect of ice lenses in the snow was also measured in these stands. Lichen biomass was significantly affected by a combination of stand maturity, understory vegetation height, and lichen height. Soil disturbance from the processes of felling and competition in the vegetation communities recovering from this disturbance were identified as key drivers of change in lichen biomass. Overall, clear-cut forests had some of the greatest prevalence of ice lenses in the snow column, and forage availability at these sites was up to 61% less than in mature stands over 58 years in age. It is suggested that alternative silviculture methods are investigated for use in this reindeer herding region, as frequent clear-cutting and consequent reduction in the average forest stand age and maturity class may be detrimental to reindeer grazing, reducing both abundance of forage, and access to it during winter.

Displacement of Native FXYD Protein From Na+/K+-ATPase With Novel FXYD Peptide Derivatives: Effects on Doxorubicin Cytotoxicity.

The seven mammalian FXYD proteins associate closely with α/ß heterodimers of Na+/K+-ATPase. Most of them protect the ß1 subunit against glutathionylation, an oxidative modification that destabilizes the heterodimer and inhibits Na+/K+-ATPase activity. A specific cysteine (Cys) residue of FXYD proteins is critical for such protection. One of the FXYD proteins, FXYD3, confers treatment resistance when overexpressed in cancer cells. We developed two FXYD3 peptide derivatives. FXYD3-pep CKCK retained the Cys residue that can undergo glutathionylation and that is critical for protecting the ß1 subunit against glutathionylation. FXYD3-pep SKSK had all Cys residues mutated to Serine (Ser). The chemotherapeutic doxorubicin induces oxidative stress, and suppression of FXYD3 with siRNA in pancreatic- and breast cancer cells that strongly express FXYD3 increased doxorubicin-induced cytotoxicity. Exposing cells to FXYD3-pep SKSK decreased co-immunoprecipitation of FXYD3 with the α1 Na+/K+-ATPase subunit. FXYD3-pep SKSK reproduced the increase in doxorubicin-induced cytotoxicity seen after FXYD3 siRNA transfection in pancreatic- and breast cancer cells that overexpressed FXYD3, while FXYD3-pep CKCK boosted the native protein's protection against doxorubicin. Neither peptide affected doxorubicin's cytotoxicity on cells with no or low FXYD3 expression. Fluorescently labeled FXYD3-pep SKSK was detected in a perinuclear distribution in the cells overexpressing FXYD3, and plasmalemmal Na+/K+-ATPase turnover could not be implicated in the increased sensitivity to doxorubicin that FXYD3-pep SKSK caused. FXYD peptide derivatives allow rapid elimination or amplification of native FXYD protein function. Here, their effects implicate the Cys residue that is critical for countering ß1 subunit glutathionylation in the augmentation of cytotoxicity with siRNA-induced downregulation of FXYD3.

Climate change, fire return intervals and the growing risk of permanent forest loss in boreal Eurasia.

Climate change has driven an increase in the frequency and severity of fires in Eurasian boreal forests. A growing number of field studies have linked the change in fire regime to post-fire recruitment failure and permanent forest loss. In this study we used four burned area and two forest loss datasets to calculate the landscape-scale fire return interval (FRI) and associated risk of permanent forest loss. We then used machine learning to predict how the FRI will change under a high emissions scenario (SSP3-7.0) by the end of the century. We found that there are currently 133,000 km2 forest at high, or extreme, risk of fire-induced forest loss, with a further 3 M km2 at risk by the end of the century. This has the potential to degrade or destroy some of the largest remaining intact forests in the world, negatively impact the health and economic wellbeing of people living in the region, as well as accelerate global climate change.

Amphiregulin increases migration and proliferation of epithelial ovarian cancer cells by inducing its own expression via PI3-kinase signaling.

The epidermal growth factor receptor (EGFR) is overexpressed in many types of cancer, including epithelial ovarian cancer (EOC), and its expression has been found to correlate with advanced stage and poor prognosis. The EGFR ligand amphiregulin (AREG) has been investigated as a target for human cancer therapy and is known to have an autocrine role in many cancers. A cytokine array identified AREG as one of several cytokines upregulated by EGF in a phosphatidylinositol 3-kinase (PI3-K) dependent manner in EOC cells. To investigate the functional role of AREG in EOC, its effect on cellular migration and proliferation was assessed in two EOC cells lines, OV167 and SKOV3. AREG increased both migration and proliferation of EOC cell line models through activation of PI3-K signaling, but independent of mitogen activated protein kinase (MAPK) signaling. Through an AREG autocrine loop mediated via PI3-K, upregulation of AREG led to increased levels of both AREG transcript and secreted AREG, while downregulation of endogenous AREG decreased the ability of exogenous AREG to induce cell migration and proliferation. Further, inhibition of endogenous AREG activity or metalloproteinase activity decreased EGF-induced EOC migration and proliferation, indicating a role for soluble endogenous AREG in mediating the functional effects of EGFR in inducing migration and proliferation in EOC.

Constructing multilayered neural networks with sparse, data-driven connectivity using biologically-inspired, complementary, homeostatic mechanisms.

The immense complexity of the brain requires that it be built and controlled by intrinsic, self-regulating mechanisms. One such mechanism, the formation of new connections via synaptogenesis, plays a central role in neuronal connectivity and, ultimately, performance. Adaptive synaptogenesis networks combine synaptogenesis, associative synaptic modification, and synaptic shedding to construct sparse networks. Here, inspired by neuroscientific observations, novel aspects of brain development are incorporated into adaptive synaptogenesis. The extensions include: (i) multiple layers, (ii) neuron survival and death based on information transmission, and (iii) bigrade growth factor signaling to control the onset of synaptogenesis in succeeding layers and to control neuron survival and death in preceding layers. Also guiding this research is the assumption that brains must achieve a compromise between good performance and low energy expenditures. Simulations of the network model demonstrate the parametric and functional control of both performance and energy expenditures, where performance is measured in terms of information loss and classification errors, and energy expenditures are assumed to be a monotonically increasing function of the number of neurons. Major insights from this study include (a) the key role a neural layer between two other layers has in controlling synaptogenesis and neuron elimination, (b) the performance and energy-savings benefits of delaying the onset of synaptogenesis in a succeeding layer, and (c) how the elimination of neurons in a preceding layer provides energy savings, code compression, and can be accomplished without significantly degrading information transfer or classification performance.

Novel Prognostic Markers in Triple-Negative Breast Cancer Discovered by MALDI-Mass Spectrometry Imaging.

There are no widely-accepted prognostic markers currently available to predict outcomes in patients with triple-negative breast cancer (TNBC), and no targeted therapies with confirmed benefit. We have used MALDI mass spectrometry imaging (MSI) of tryptic peptides to compare regions of cancer and benign tissue in 10 formalin-fixed, paraffin-embedded sections of TNBC tumors. Proteins were identified by reference to a peptide library constructed by LC-MALDI-MS/MS analyses of the same tissues. The prognostic significance of proteins that distinguished between cancer and benign regions was estimated by Kaplan-Meier analysis of their gene expression from public databases. Among peptides that distinguished between cancer and benign tissue in at least 3 tissues with a ROC area under the curve >0.7, 14 represented proteins identified from the reference library, including proteins not previously associated with breast cancer. Initial network analysis using the STRING database showed no obvious functional relationships except among collagen subunits COL1A1, COL1A2, and COL63A, but manual curation, including the addition of EGFR to the analysis, revealed a unique network connecting 10 of the 14 proteins. Kaplan-Meier survival analysis to examine the relationship between tumor expression of genes encoding the 14 proteins, and recurrence-free survival (RFS) in patients with basal-like TNBC showed that, compared to low expression, high expression of nine of the genes was associated with significantly worse RFS, most with hazard ratios >2. In contrast, in estrogen receptor-positive tumors, high expression of these genes showed only low, or no, association with worse RFS. These proteins are proposed as putative markers of RFS in TNBC, and some may also be considered as possible targets for future therapies.

Ibudilast sensitizes glioblastoma to temozolomide by targeting Macrophage Migration Inhibitory Factor (MIF).

Recurrence in patients with glioblastoma (GBM) is inevitable resulting in short survival times, even in patients with O-6-Methylguanine-DNA Methyltransferase (MGMT) methylation. Other pathways must be activated to escape from temozolomide (TMZ) treatment, however acquired resistance mechanisms to TMZ are not well understood. Herein, frozen tumors from 36 MGMT methylated patients grouped according to overall survival were extracted and proteins were profiled using surface-enhanced laser desorption/ionization (SELDI) with time-of flight (TOF) proteomics to identify low molecular weight proteins that associated with poor survival outcomes. Overexpression of macrophage migration inhibitory factor (MIF) was identified in human GBM specimens that were MGMT methylated but showed poor survival. This correlation was confirmed in an independent cohort of human GBM. MIF overexpression has been reported in several cancer types, including GBM. We repurposed ibudilast, a specific MIF inhibitor, and treated patient derived cell lines. Ibudilast showed modest anti-proliferative activity however, when combined with TMZ, significant synergism was observed, resulting in cell cycle arrest and apoptosis. In vivo, combined ibudilast and TMZ treatment of a patient derived xenograft (PDX) model resulted in significantly longer overall survival. Our findings have significant clinical implications for people with GBM. Since clinical trials involving ibudilast have shown no adverse side effects and the drug readily penetrates the blood brain barrier, treatment of GBM with this combination is clinically achievable.

IGFBP-3 interacts with NONO and SFPQ in PARP-dependent DNA damage repair in triple-negative breast cancer.

Women with triple-negative breast cancer (TNBC) are generally treated by chemotherapy but their responsiveness may be blunted by DNA double-strand break (DSB) repair. We previously reported that IGFBP-3 forms nuclear complexes with EGFR and DNA-dependent protein kinase (DNA-PKcs) to modulate DSB repair by non-homologous end-joining (NHEJ) in TNBC cells. To discover IGFBP-3 binding partners involved in chemoresistance through stimulation of DSB repair, we analyzed the IGFBP-3 interactome by LC-MS/MS and confirmed interactions by coimmunoprecipitation and proximity ligation assay. Functional effects were demonstrated by DNA end-joining in vitro and measurement of γH2AX foci. In response to 20 µM etoposide, the DNA/RNA-binding protein, non-POU domain-containing octamer-binding protein (NONO) and its dimerization partner splicing factor, proline/glutamine-rich (SFPQ) formed complexes with IGFBP-3, demonstrated in basal-like TNBC cell lines HCC1806 and MDA-MB-468. NONO binding to IGFBP-3 was also shown in a cell-free biochemical assay. IGFBP-3 complexes with NONO and SFPQ were blocked by inhibiting EGFR with gefitinib or DNA-PKcs with NU7026, and by the PARP inhibitors veliparib and olaparib, which also reduced DNA end-joining activity and delayed the resolution of the γH2AX signal (i.e. inhibited DNA DSB repair). Downregulation of the long noncoding RNA in NHEJ pathway 1 (LINP1) by siRNA also blocked IGFBP-3 interaction with NONO-SFPQ. These findings suggest a PARP-dependent role for NONO and SFPQ in IGFBP-3-dependent DSB repair and the involvement of LINP1 in the complex formation. We propose that targeting of the DNA repair function of IGFBP-3 may enhance chemosensitivity in basal-like TNBC, thus improving patient outcomes.

Contrasting effects of IGF binding protein-3 expression in mammary tumor cells and the tumor microenvironment.

IGFBP-3 has both stimulatory and inhibitory effects on cancer progression. The growth of EO771 mammary carcinoma cells as syngeneic tumors in C57BL/6 mice is reduced in Igfbp3-null (BP3KO) mice, suggesting that systemic IGFBP-3 enhances tumor progression. In this study we assessed the growth of EO771 cells expressing human IGFBP-3 in BP3KO mice. Cells expressing hIGFBP-3 showed decreased proliferation in vitro and increased levels of IGF-1 receptor (IGF1R) protein but not mRNA, consistent with sequestration of endogenous IGF by IGFBP-3. The growth rate of these cells was restored by exposure to IGF-1 or analogues with reduced affinity for IGFBP-3 (long Arg3-IGF-1) or IGF1R (Leu24-IGF-1). In EO771 cells implanted orthotopically into mice, hIGFBP-3 expression by the cells inhibited tumor establishment in BP3KO but not wild-type mice. For tumors that successfully established, final weight was not affected significantly by hIGFBP-3 expression. However, final tumor weight was inversely related to intratumoral T cell counts, and sera from BP3KO mice with tumors showed low-titer immunoreactivity against IGFBP-3. The contrasting effects on tumor establishment and progression of IGFBP-3 expressed by mammary carcinoma cells, compared to systemic stromal and circulating IGFBP-3, highlights the complexity of growth regulation by IGFBP-3 in mammary tumors.

Nuclear Insulin-Like Growth Factor Binding Protein-3 As a Biomarker in Triple-Negative Breast Cancer Xenograft Tumors: Effect of Targeted Therapy and Comparison With Chemotherapy.

Triple-negative breast cancer (TNBC) typically has a worse outcome than other breast cancer subtypes, in part owing to a lack of approved therapeutic targets or prognostic markers. We have previously described an oncogenic pathway in basal-like TNBC cells, initiated by insulin-like growth factor binding protein-3 (IGFBP-3), in which the epidermal growth factor receptor (EGFR) is transactivated by sphingosine-1-phosphate (S1P) resulting from sphingosine kinase (SphK)-1 activation. Oncogenic IGFBP-3 signaling can be targeted by combination treatment with the S1P receptor modulator and SphK inhibitor, fingolimod, and the EGFR kinase inhibitor, gefitinib (F + G). However, the interaction of this treatment with chemotherapy has not been documented. Since we observed nuclear localization of IGFBP-3 in some TNBC tumors, this study aimed to evaluate the prognostic significance of nuclear IGFBP-3 in pre-clinical models of basal-like TNBC treated with F + G and doxorubicin. Orthotopic xenograft tumors were grown in nude mice from the human basal-like TNBC cell lines MDA-MB-468 and HCC1806, and were treated with gefitinib, 25 mg/Kg, plus fingolimod, 5 mg/Kg, 3-times weekly. In some studies, doxorubicin was also administered once weekly for 6 weeks. Tumor tissue proteins were quantitated by immunohistochemistry (IHC). Interaction between doxorubicin and F + G was also studied in proliferation assays in vitro. In both tumor models, tissue staining for IGFBP-3 was predominantly nuclear. Combination of F + G significantly enhanced mouse survival, decreased nuclear IGFBP-3 and Ki67 staining, and increased apoptosis (cleaved caspase-3) staining. Kaplan-Meier survival analysis showed that a high tumor IGFBP-3 IHC score (>median), like a high Ki67 score, was significantly associated with shorter survival time, whereas a high apoptosis score was associated with prolonged survival. Studied in vitro in both cell lines, low-dose doxorubicin that had little effect alone, strongly enhanced the cytostatic effect of low-dose F + G combination. However, in both in vivo models, doxorubicin at maximum-tolerated dose neither inhibited tumor growth when administered alone, nor enhanced the significant inhibitory effect of F + G. We conclude that doxorubicin may not add benefit to the inhibitory effect of F + G unless its dose-limiting toxicity can be overcome. Nuclear IGFBP-3 appears to have potential as a prognostic marker in TNBC and could be evaluated for clinical utility.

Enhancement of mammary tumour growth by IGFBP-3 involves impaired T cell accumulation.

Epidemiological studies show an association between obesity and poor breast cancer prognosis. We previously demonstrated that global IGFBP-3 deficiency, in IGFBP-3-null mice, resulted in a 50% reduction in mammary tumour growth over 3 weeks relative to tumours in wild-type (WT) C57BL/6 mice. This growth reduction was ameliorated by high fat feeding-induced obesity. This study aimed to examine how IGFBP-3 promotes tumour growth by influencing the immune tumour microenvironment in healthy and obese mice. Syngeneic EO771 cells, which lack detectable IGFBP-3 expression, were grown as orthotopic tumours in WT and IGFBP-3-null C57BL/6 mice placed on either a control chow or a high-fat diet (HFD), and examined by quantitative PCR and immunohistochemistry. In WT mice, increased stromal expression of IGFBP-3 was positively associated with tumour growth, supporting the hypothesis that IGFBP-3 in the microenvironment promotes tumour progression. Examining markers of immune cell subsets, gene expression of Ifng, Cd8a, Cd8b1 and Tnf and CD8 measured by immunohistochemistry were elevated in tumours of IGFBP-3-null mice compared to WT, indicating an accumulation of CD8+ T cells, but this increase was absent if the IGFBP-3-null mice had been exposed to HFD. Expression of these genes was negatively associated with tumour growth. Although similar among groups overall, Nkg2d and Tnfsf10 tumoural expression was associated with decreased tumour growth. Overall, the results of this study provide an immune-based mechanism by which host IGFBP-3 may promote breast tumour growth in the EO771 murine breast cancer model, and suggest that targeting IGFBP-3 might make a novel contribution to immune therapy for breast cancer.

Insulin-like growth factor receptor and sphingosine kinase are prognostic and therapeutic targets in breast cancer.

BACKGROUND: Targeting the type 1 insulin-like growth factor receptor (IGF1R) in breast cancer remains an ongoing clinical challenge. Oncogenic IGF1R-signaling occurs via activation of PI3K/AKT/MAPK downstream mediators which regulate cell proliferation and protein synthesis. To further understand IGF1R signaling we have investigated the involvement of the oncogenic IGF1R-related sphingosine kinase (SphK) pathway. METHODS: The prognostic (overall survival, OS) and therapeutic (anti-endocrine therapy) co-contribution of IGF1R and SphK1 were investigated using breast cancer patient samples (n = 236) for immunohistochemistry to measure total and phosphorylated IGF1R and SphK1. Kaplan-Meier and correlation analyses were performed to determine the contribution of high versus low IGF1R and/or SphK1 expression to OS in patients treated with anti-endocrine therapy. Cell viability and colony formation in vitro studies were completed using estrogen receptor (ER) positive and negative breast cancer cell-lines to determine the benefit of IGF1R inhibitor (OSI-906) and SphK inhibitor (SKI-II) co-therapy. Repeated measures and 1-way ANOVA were performed to compare drug treatments groups and the Chou-Talalay combination index (CI) was calculated to estimate drug synergism in vitro (CI < 1). RESULTS: High IGF1R and SphK1 protein co-expression in tumor tissue was associated with improved OS specifically in ER-positive disease and stratified for anti-endocrine therapy. A significant synergistic inhibition of cell viability and/or colony formation following OSI-906 and SKI-II co-treatment in vitro was evident (p < 0.05, CI < 1). CONCLUSION: We conclude that high IGF1R and SphK1 co-expression act together as prognostic indicators and are potentially, dual therapeutic targets for the development of a more effective IGF1R-directed combination breast cancer therapy.

Limitations of galactose therapy in phosphoglucomutase 1 deficiency.

INTRODUCTION: Phosphoglucomutase 1 deficiency (PGM1 deficiency) has been identified as both, glycogenosis and congenital disorder of glycosylation (CDG). The phenotype includes hepatopathy, myopathy, oropharyngeal malformations, heart disease and growth retardation. Oral galactose supplementation at a dosage of 1 g per kg body weight per day is regarded as the therapy of choice. RESULTS: We report on a patient with a novel disease causing mutation, who was treated for 1.5 years with oral galactose supplementation. Initially, elevated transaminases were reduced and protein glycosylation of serum transferrin improved rapidly. Long-term surveillance however indicated limitations of galactose supplementation at the standard dose: 1 g per kg body weight per day did not achieve permanent correction of protein glycosylation. Even increased doses of up to 2.5 g per kg body weight did not result in complete normalization. Furthermore, we described for the first time heart rhythm abnormalities, i.e. long QT Syndrome associated with a glycosylation disorder. Mass spectrometry of IGFBP3, which was assumed to play a major role in growth retardation associated with PGM1 deficiency, revealed no glycosylation abnormalities. Growth rate did not improve under galactose supplementation. CONCLUSIONS: The results of our study indicate that the current standard dose of galactose might be too low to achieve normal glycosylation in all patients. In addition, growth retardation in PGM1 deficiency is complex and multifactorial. Furthermore, heart rhythm abnormalities must be considered when treating patients with PGM1 deficiency.