Changes in atherosclerotic cardiovascular disease risk over time among people living with HIV.

OBJECTIVE: To describe changes in atherosclerotic cardiovascular disease (ASCVD) risk over time among people living with HIV (PLHIV). METHODS: We used data from the TREAT Asia HIV Observational Database (TAHOD) and the Australian HIV Observational Database (AHOD). Five-year ASCVD risk was calculated using the D:A:D equation. Individuals were eligible for inclusion if they were aged ≥18 years, had started ART, had no previous history of ASCVD and had complete ASCVD risk factor data available within the first 5 years of ART initiation. RESULTS: A total of 3368 adults contributed data, 3221 were from TAHOD and 147 were from AHOD. The median age at ART initiation was 36 [IQR 31-43] years for TAHOD participants, and 42 [IQR 35-50] years for AHOD participants. Most TAHOD (70.4%) and AHOD (91.8%) participants were male. Overall, ASCVD risk increased from 0.84% (95% CI 0.81%-0.87%) at ART initiation to 1.34% (95% CI 1.29%-1.39%) after 5 years on ART. After adjusting for traditional and HIV-associated ASCVD risk factors, ASCVD risk increased at a similar rate among sub-populations defined by HIV exposure (heterosexuals, men who have sex with men, people who inject drugs), race/ethnicity (Caucasian and Asian) and nadir CD4 at ART initiation (<200 and ≥200 cells/mm3). CONCLUSIONS: These findings emphasize the growing burden of ASCVD risk among PLHIV and the need to develop interventions that are effective across a broad range of HIV sub-populations.

Projected impact of population aging on non-communicable disease burden and costs in the Kingdom of Saudi Arabia, 2020-2030.

BACKGROUND: The number of people aged greater than 65 years per 100 people aged 20-64 years is expected to almost double in The Kingdom of Saudi Arabia (KSA) between 2020 and 2030. We therefore aimed to quantify the growing non-communicable disease (NCD) burden in KSA between 2020 and 2030, and the impact this will have on the national health budget. METHODS: Ten priority NCDs were selected: ischemic heart disease, stroke, type 2 diabetes, chronic obstructive pulmonary disease, chronic kidney disease, dementia, depression, osteoarthritis, colorectal cancer, and breast cancer. Age- and sex-specific prevalence was projected for each priority NCD between 2020 and 2030. Treatment coverage rates were applied to the projected prevalence estimates to calculate the number of patients incurring treatment costs for each condition. For each priority NCD, the average cost-of-illness was estimated based on published literature. The impact of changes to our base-case model in terms of assumed disease prevalence, treatment coverage, and costs of care, coming into effect from 2023 onwards, were explored. RESULTS: The prevalence estimates for colorectal cancer and stroke were estimated to almost double between 2020 and 2030 (97% and 88% increase, respectively). The only priority NCD prevalence projected to increase by less than 60% between 2020 and 2030 was for depression (22% increase). It is estimated that the total cost of managing priority NCDs in KSA will increase from USD 19.8 billion in 2020 to USD 32.4 billion in 2030 (an increase of USD 12.6 billion or 63%). The largest USD value increases were projected for osteoarthritis (USD 4.3 billion), diabetes (USD 2.4 billion), and dementia (USD 1.9 billion). In scenario analyses, our 2030 projection for the total cost of managing priority NCDs varied between USD 29.2 billion - USD 35.7 billion. CONCLUSIONS: Managing the growing NCD burden in KSA's aging population will require substantial healthcare spending increases over the coming years.

Cost-effectiveness of screening and treating anal pre-cancerous lesions among gay, bisexual and other men who have sex with men living with HIV.

Background: Gay, bisexual and other men who have sex with men (GBM) living with HIV have a substantially elevated risk of anal cancer (85 cases per 100,000 person-years vs 1-2 cases per 100,000 person-years in the general population). The precursor to anal cancer is high-grade squamous intraepithelial lesion (HSIL). Findings regarding the cost-effectiveness of HSIL screening and treatment in GBM are conflicting. Using recent data on HSIL natural history and treatment effectiveness, we aimed to improve upon earlier models. Methods: We developed a Markov cohort model populated using observational study data and published literature. Our study population was GBM living with HIV aged ≥35 years. We used a lifetime horizon and framed our model on the Australian healthcare perspective. The intervention was anal HSIL screening and treatment. Our primary outcome was the incremental cost-effectiveness ratio (ICER) as cost per quality-adjusted life-year (QALY) gained. Findings: Anal cancer incidence was estimated to decline by 44-70% following implementation of annual HSIL screening and treatment. However, for the most cost-effective screening method assessed, the ICER relative to current practice, Australian Dollar (AUD) 135,800 per QALY gained, remained higher than Australia's commonly accepted willingness-to-pay threshold of AUD 50,000 per QALY gained. In probabilistic sensitivity analyses, HSIL screening and treatment had a 20% probability of being cost-effective. When the sensitivity and specificity of HSIL screening were enhanced beyond the limits of current technology, without an increase in the cost of screening, ICERs improved but were still not cost-effective. Cost-effectiveness was achieved with a screening test that had 95% sensitivity, 95% specificity, and cost ≤ AUD 24 per test. Interpretation: Establishing highly sensitive and highly specific HSIL screening methods that cost less than currently available techniques remains a research priority. Funding: No specific funding was received for this analysis.

Effects of a minimum floor Price law on cigarette use in Oakland, California: A static microsimulation model.

Tobacco minimum floor price laws (MFPLs) are a non-tax price policy that set a price below which tobacco products cannot be sold, thereby raising prices. Despite their growing interest among policy makers, little is known about the effects of local MFPLs on smoking prevalence or smoking intensity. We aimed to project the impact of a local tobacco MFPL on cigarette smoking prevalence and cigarette smoking intensity in Oakland, California, including detailed analysis of several important subpopulations. We used data collected between April 2017 and December 2019 from the California Behavioral Risk Factor Surveillance System and the National Youth Tobacco Survey to construct a static microsimulation model representative of Oakland. We projected the impact of MFPLs ranging from $8.00 to $13.00 per pack. All analyses were conducted between 2019 and 2020. With the introduction of an MFPL and assuming 15% policy evasion, mean price paid per pack was projected to increase by $1.05 to $4.69, cigarette smoking prevalence was projected to drop by 0.3% to 0.8%, and smoking intensity was projected to drop by 0.7% to 2.0% among continuing smokers. Total number of cigarettes smoked per month was projected to drop by 246,000 to 734,000 cigarettes, a 3.0% to 9.0% reduction from the current level (8.2 million cigarettes). The greatest reductions in cigarette smoking prevalence were among those aged 12 to 24-years-old, of non-Hispanic black or other race/ethnicity, and living below the federal poverty level. An MFPL in Oakland may substantially reduce cigarette use and target several important subpopulations.

Cigarette Pack Prices and Sales Following Policy Changes in California, 2011-2018.

Objectives. To estimate the combined effect of California's Tobacco 21 law (enacted June 2016) and $2-per-pack cigarette excise tax increase (enacted April 2017) on cigarette prices and sales, compared with matched comparator states.Methods. We used synthetic control methods to compare cigarette prices and sales after the policies were enacted, relative to what we would have expected without the policy reforms. To estimate the counterfactual, we matched pre-reform covariate and outcome trends between California and control states to construct a "synthetic" California.Results. Compared with the synthetic control in 2018, cigarette prices in California were $1.89 higher ($7.86 vs $5.97; P < .001), and cigarette sales were 16.6% lower (19.9 vs 16.6 packs per capita; P < .001). This reduction in sales equates to 153.9 million fewer packs being sold between 2017 and 2018.Conclusions. California's new cigarette tax was largely passed on to consumers. The new cigarette tax, combined with the Tobacco 21 law, have contributed to a rapid and substantial reduction in cigarette consumption in California.

Cardiovascular disease among people living with HIV in Brazil.

OBJECTIVES: There is a paucity of data on cardiovascular disease (CVD) among people living with HIV (PLHIV) in resource-limited countries. We assessed factors associated with CVD and the impact of prevalent CVD on all-cause mortality in PLHIV on antiretroviral therapy in Brazil. METHODS: Competing risk regression to assess factors associated with CVD and all-cause mortality in the HIV-Brazil Cohort Study between 2003 and 2014. RESULTS: Among 5614 patients, the rate of CVD was 3.5 (95% confidence interval [95% CI] 2.9-4.3) per 1000 person-years. CVD was associated with older age (adjusted hazard ratio [aHR] 6.4 for ≥55 years vs. <35 years, 95% CI: 2.5-16.3, P < 0.01), black race (aHR 1.8 vs. white race, 95% CI: 1.0-3.1, P = 0.04), past CVD (aHR 3.0 vs. no past CVD, 95% CI: 1.4-6.2, P < 0.01), hypertension (aHR 1.8 vs. no hypertension, 95% CI: 1.0-3.1, P = 0.04), high-grade dyslipidemia (aHR 9.3 vs. no high-grade dyslipidemia, 95% CI: 6.0-14.6, P < 0.01), ever smoking (aHR 2.4 vs. never, 95% CI: 1.2-5.0, P = 0.02) and low nadir CD4 cell count (aHR 1.8 for 100-250 cells/mm3 vs. >250 cells/mm3 , 95% CI: 1.0-3.2, P = 0.05). The rate of death was 16.6 (95% CI: 15.1-18.3) per 1000 person-years. Death was strongly associated with having had a past CVD event (aHR 1.7 vs. no past CVD event, 95% CI: 1.1-2.7, P = 0.01). CONCLUSIONS: Traditional and HIV-specific factors associated with CVD among PLHIV in Brazil are similar to those identified among PLHIV in high-income countries. PLHIV in Brazil with a history of CVD have a high risk of death. CVD care and treatment remain priorities for PLHIV in Brazil as this population ages and antiretroviral therapy use expands.

OBJECTIFS: Il existe peu de données sur les maladies cardiovasculaires (MCV) chez les personnes vivant avec le VIH (PVVIH) dans les pays à ressources limitées. Nous avons évalué les facteurs associés aux MCV et l'impact des MCV prévalentes sur la mortalité toutes causes confondues des PVVIH sous le traitement antirétroviral au Brésil. MÉTHODES: Régression des risques concurrente pour évaluer les facteurs associés aux MCV et à la mortalité toutes causes confondues dans l'étude de cohorte VIH-Brésil entre 2003 et 2014. RÉSULTATS: Parmi 5.614 patients, le taux de MCV était de 3,5 (intervalle de confiance à 95% [IC95%] 2,9-4,3) pour 1.000 personnes-années. Les MCV étaient associées à un âge plus avancé (rapport de risque ajusté [aHR] 6,4 chez les ≥55 ans versus chez les <35 ans, IC95%: 2,5-16,3 ; p <0,01), race noire (aHR: 1,8 versus race blanche, IC95%: 1,0-3,1 ; p = 0,04), MCV passée (aHR: 3,0 versus pas de MCV passée, IC95%: 1,4-6,2 ; p <0,01), hypertension (aHR: 1,8 versus pas d'hypertension, IC95%: 1,0-3,1 ; p = 0,04), dyslipidémie de grade élevé (aHR 9,3 versus absence de dyslipidémie de grade élevé, IC95%: 6,0-14,6 ; p <0,01), tabagisme (aHR 2,4 versus n'avoir jamais fumé, IC95%: 1,2-5,0 ; p = 0,02) et faible nombre de CD4 au nadir (aHR: 1,8 pour 100-250 cellules/mm3 versus >250 cellules/mm3 , IC95%: 1,0-3,2 ; p = 0,05). Le taux de décès était de 16,6 (IC95%: 15,1-18,3) pour 1.000 personnes-années. Le décès était fortement associé à un événement MCV antérieur (aHR: 1,7 versus aucun événement MCV antérieur, IC95%: 1,1-2,7 ; p = 0,01). CONCLUSIONS: Les facteurs traditionnels et spécifiques au VIH associés aux MCV chez les PVVIH au Brésil sont similaires à ceux identifiés chez les PVVIH dans les pays à revenu élevé. Les PVVIH au Brésil ayant des antécédents de MCV ont un risque élevé de décès. Les soins et le traitement des MCV restent des priorités pour les PVVIH au Brésil à mesure que cette population vieillit et que l'utilisation des thérapies antirétrovirales augmente.

Can Australia Reach the World Health Organization Hepatitis C Elimination Goal by 2025 Among Human Immunodeficiency Virus-positive Gay and Bisexual Men?

BACKGROUND: Human immunodeficiency virus (HIV)-positive gay and bisexual men (GBM) in Australia are well engaged in care. The World Health Organization's (WHO) hepatitis C virus (HCV) elimination target of an 80% reduction in incidence by 2030 may be reachable ahead of time in this population. METHODS: We predicted the effect of treatment and behavioral changes on HCV incidence among HIV-positive GBM up to 2025 using a HCV transmission model parameterized with Australian data. We assessed the impact of changes in behavior that facilitate HCV transmission in the context of different rates of direct-acting antiviral (DAA) use. RESULTS: HCV incidence in our model increased from 0.7 per 100 person-years in 2000 to 2.5 per 100 person-years in 2016 and had the same trajectory as previously reported clinical data. If the proportion of eligible (HCV RNA positive) patients using DAAs stays at 65% per year between 2016 and 2025, with high-risk sexual behavior and injecting drug use remaining at current levels, HCV incidence would drop to 0.4 per 100 person-years (85% decline from 2016). In the same treatment scenario but with substantial increases in risk behavior, HCV incidence would drop to 0.6 per 100 person-years (76% decline). If the proportion of eligible patients using DAAs dropped from 65% per year in 2016 to 20% per year in 2025 and risk behavior did not change, HCV incidence would drop to 0.7 per 100 person-years (70% reduction). CONCLUSIONS: Reaching the WHO HCV elimination target by 2025 among HIV-positive GBM in Australia is achievable.

Changes in renal function with long-term exposure to antiretroviral therapy in HIV-infected adults in Asia.

PURPOSE: Renal disease is common among people living with human immunodeficiency virus (HIV). However, there is limited information on the incidence and risk factors associated with renal dysfunction among this population in Asia. METHODS: We used data from the TREAT Asia HIV Observational Database. Patients were included if they started antiretroviral therapy during or after 2003, had a serum creatinine measurement at antiretroviral therapy initiation (baseline), and had at least 2 follow-up creatinine measurements taken ≥3 months apart. Patients with a baseline estimated glomerular filtration rate (eGFR) ≤60 mL/min/1.73 m2 were excluded. Chronic kidney disease was defined as 2 consecutive eGFR values ≤60 mL/min/1.73 m2 taken ≥3 months apart. Generalized estimating equations were used to identify factors associated with eGFR change. Competing risk regression adjusted for study site, age and sex, and cumulative incidence plots were used to evaluate factors associated with chronic kidney disease (CKD). RESULTS: Of 2547 patients eligible for this analysis, tenofovir was being used by 703 (27.6%) at baseline. Tenofovir use, high baseline eGFR, advanced HIV disease stage, and low nadir CD4 were associated with a decrease in eGFR during follow-up. Chronic kidney disease occurred at a rate of 3.4 per 1000 patient/years. Factors associated with CKD were tenofovir use, old age, low baseline eGFR, low nadir CD4, and protease inhibitor use. CONCLUSIONS: There is an urgent need to enhance renal monitoring and management capacity among at-risk groups in Asia and improve access to less nephrotoxic antiretrovirals.

Incidence of Postsuppression Virologic Rebound in Perinatally HIV-Infected Asian Adolescents on Stable Combination Antiretroviral Therapy.

PURPOSE: To assess the incidence and predictors of postsuppression virologic rebound (VR) among adolescents on stable combination antiretroviral therapy in Asia. METHODS: Perinatally HIV-infected Asian adolescents (10-19 years) with documented virologic suppression (two consecutive viral loads [VLs] <400 copies/mL ≥6 months apart) were included. Baseline was the date of the first VL <400 copies/mL at age ≥10 years or the 10th birthday for those with prior suppression. Cox proportional hazards models were used to identify predictors of postsuppression VR (VL >1,000 copies/mL). RESULTS: Of 1,379 eligible adolescents, 47% were males. At baseline, 22% were receiving protease inhibitor-containing regimens; median CD4 cell count (interquartile range [IQR]) was 685 (448-937) cells/mm3; 2% had preadolescent virologic failure (VF) before subsequent suppression. During adolescence, 180 individuals (13%) experienced postsuppression VR at a rate of 3.4 (95% confidence interval: 2.9-3.9) per 100 person-years, which was consistent over time. Median time to VR during adolescence (IQR) was 3.3 (2.1-4.8) years. Wasting (weight-for-age z-score <-2.5), being raised by grandparents, receiving second-line protease inhibitor-based regimens, starting combination antiretroviral therapy after 2005, and having preadolescent VF were independent predictors of adolescent VR. At VR, median age, CD4 cell count, and VL (IQR) were 14.8 (13.2-16.4) years, 507 (325-723) cells/mm3, and 4.1 (3.5-4.7) log10 copies/mL, respectively. CONCLUSIONS: A modest and consistent incidence of postsuppression VR was documented during adolescence in our cohort. Having poor weight, receiving second-line regimens, and prior VF were associated with an increased VR rate. Adolescents at higher risk of VR may benefit from more intensive VL monitoring to enhance adherence management.

Is nelfinavir exposure associated with cancer incidence in HIV-positive individuals?

OBJECTIVE: Nelfinavir exhibits potent anticancer properties against a range of tumours. However, in 2006/2007, nelfinavir supplies were accidently contaminated with a carcinogen. This analysis investigated the association between nelfinavir use and cancer risk in HIV-positive persons. DESIGN: Observational cohort study. METHODS: D:A:D study data was analysed using Poisson regression models to examine associations between cancer incidence and cumulative nelfinavir exposure, current nelfinavir exposure, and exposure to nelfinavir between 1 July 2006-30 June 2007. RESULTS: A total of 42 006 individuals (50% white, 73% male) contributed 303 005 person-years of follow-up between 1 January 2004 and 1 February 2014. At study enrolment, median age was 40 [interquartile range (IQR) 33-46] years and 8305 individuals had a history of nelfinavir use [median duration 1.7 (IQR 0.7-3.4) years]. During follow-up, nelfinavir was used by 2476 individuals for a median of 1.7 (IQR 0.7-3.8) years; 1063 were exposed to nelfinavir between 1 July 2006 and 30 June 2007. Overall, 2279 cancers were diagnosed at a rate of 0.75 [95% confidence interval (95% CI) 0.72-0.78] per 100 person-years. Neither greater cumulative exposure to nelfinavir [adjusted risk ratio (aRR) 0.93 for every additional 5 years, 95% CI 0.82-1.06, P = 0.26] nor current use of nelfinavir (aRR 0.98 vs other protease inhibitor use, 95% CI 0.68-1.41, P = 0.92) were associated with cancer risk. The adjusted risk of cancer for participants exposed to nelfinavir between 1 July 2006 and 30 June 2007 compared to those receiving other treatment over this period was 1.07 (95% CI 0.78-1.46, P = 0.68). CONCLUSION: Nelfinavir use was not associated with a lower cancer incidence than other protease inhibitor regimens. As of February 2014, exposure to the 2006/2007 contamination of nelfinavir does not appear to be associated with increased cancer incidence.

Trends in first-line antiretroviral therapy in Asia: results from the TREAT Asia HIV observational database.

BACKGROUND: Antiretroviral therapy (ART) has evolved rapidly since its beginnings. This analysis describes trends in first-line ART use in Asia and their impact on treatment outcomes. METHODS: Patients in the TREAT Asia HIV Observational Database receiving first-line ART for ≥ 6 months were included. Predictors of treatment failure and treatment modification were assessed. RESULTS: Data from 4662 eligible patients was analysed. Patients started ART in 2003-2006 (n = 1419), 2007-2010 (n = 2690) and 2011-2013 (n = 553). During the observation period, tenofovir, zidovudine and abacavir use largely replaced stavudine. Stavudine was prescribed to 5.8% of ART starters in 2012/13. Efavirenz use increased at the expense of nevirapine, although both continue to be used extensively (47.5% and 34.5% of patients in 2012/13, respectively). Protease inhibitor use dropped after 2004. The rate of treatment failure or modification declined over time (22.1 [95%CI 20.7-23.5] events per 100 patient/years in 2003-2006, 15.8 [14.9-16.8] in 2007-2010, and 11.6 [9.4-14.2] in 2011-2013). Adjustment for ART regimen had little impact on the temporal decline in treatment failure rates but substantially attenuated the temporal decline in rates of modification due to adverse event. In the final multivariate model, treatment modification due to adverse event was significantly predicted by earlier period of ART initiation (hazard ratio 0.52 [95%CI 0.33-0.81], p = 0.004 for 2011-2013 versus 2003-2006), older age (1.56 [1.19-2.04], p = 0.001 for ≥ 50 years versus <30 years), female sex (1.29 [1.11-1.50], p = 0.001 versus male), positive hepatitis C status (1.33 [1.06-1.66], p = 0.013 versus negative), and ART regimen (11.36 [6.28-20.54], p<0.001 for stavudine-based regimens versus tenofovir-based). CONCLUSIONS: The observed trends in first-line ART use in Asia reflect changes in drug availability, global treatment recommendations and prescriber preferences over the past decade. These changes have contributed to a declining rate of treatment modification due to adverse event, but not to reductions in treatment failure.

Niche anchorage and signaling through membrane-bound Kit-ligand/c-kit receptor are kinase independent and imatinib insensitive.

Kit ligand (KitL) and its tyrosine kinase receptor c-kit are critical for germ cells, melanocytes, mastocytes, and hematopoietic stem cells. Alternative splicing of KitL generates membrane-bound KitL (mb-KitL) or soluble KitL, providing survival or cell migration, respectively. Here we analyzed whether c-kit can function both as an adhesion and signaling receptor to mb-KitL presented by the environmental niche. At contacts between fibroblasts and MC/9 mast cells, mb-KitL, and c-kit formed ligand/receptor clusters that formed stable complexes, which resisted dissociation by c-kit blocking mAbs and provided cell anchorage under physiological shear stresses. Clusters recruited tyrosine-phosphorylated proteins and induced spatially restricted F-actin polymerization. Mutational analysis of c-kit demonstrated kinase-independent mb-KitL/c-kit clustering, anchorage, F-actin polymerization, and Tyr567-dependent cluster phosphorylation. Kinase inhibition of c-kit by imatinib reduced cluster coalescence, but allowed cluster phosphorylation and F-actin polymerization, which required PI3K recruitment and a newly identified juxtamembrane residue. Synergies between integrin and c-kit-mediated spreading and adhesion of MC/9 cells were studied in vitro on immobilized-KitL/fibronectin surfaces. While c-kit blocking antibodies prevented spreading, imatinib blocked spreading induced by soluble- but not immobilized KitL. Thus, "mechanical" activation of c-kit provides signaling, niche-anchorage, and synergies with integrin-mediated adhesion, which is independent of kinase function and resistant to c-kit kinase inhibitors.-

A novel mode of cell detachment from fibrillar fibronectin matrix under shear.

Cells within tissues are surrounded by fibrillar extracellular matrix (ECM) that supports cell adhesion via integrin receptors. The strength of cell interactions with fibrillar matrix and the effects of force on these interactions have not been quantified. To this end, we used a spinning disc device to apply radially increasing shear to human HT1080 fibrosarcoma cells attached to a cell-derived fibrillar fibronectin (FN) matrix. The shear required to detach 50% of HT1080 cells was eight times greater on a FN-coated, rigid glass substrate than on fibrillar FN matrix. Covalent crosslinking of the FN matrix increased its stiffness tenfold and produced a modest increase in shear detachment force for these cells. On FN-coated surfaces, cells detach by releasing interactions between alpha5beta1 integrin and FN. By contrast, cell detachment from fibrillar matrix occurred through a novel mechanism of fibril breakage, which left holes in the matrix visible by fluorescence microscopy. These results show that cells require less force to detach from fibrillar matrix than from FN adsorbed on glass and that detachment occurs through breaking fibrils instead of by release of integrin-matrix bonds. Thus, ECM fibril breakage is another molecular feature to consider when understanding cell and tissue homeostasis.

Mechanically activated integrin switch controls alpha5beta1 function.

The cytoskeleton, integrin-mediated adhesion, and substrate stiffness control a common set of cell functions required for development and homeostasis that are often deranged in cancer. The connection between these mechanical elements and chemical signaling processes is not known. Here, we show that alpha(5)beta(1) integrin switches between relaxed and tensioned states in response to myosin II-generated cytoskeletal force. Force combines with extracellular matrix stiffness to generate tension that triggers the integrin switch. This switch directly controls the alpha(5)beta(1)-fibronectin bond strength through engaging the synergy site in fibronectin and is required to generate signals through phosphorylation of focal adhesion kinase. In the context of tissues, this integrin switch connects cytoskeleton and extracellular matrix mechanics to adhesion-dependent motility and signaling pathways.

Rab-coupling protein coordinates recycling of alpha5beta1 integrin and EGFR1 to promote cell migration in 3D microenvironments.

Here we show that blocking the adhesive function of alphavbeta3 integrin with soluble RGD ligands, such as osteopontin or cilengitide, promoted association of Rab-coupling protein (RCP) with alpha5beta1 integrin and drove RCP-dependent recycling of alpha5beta1 to the plasma membrane and its mobilization to dynamic ruffling protrusions at the cell front. These RCP-driven changes in alpha5beta1 trafficking led to acquisition of rapid/random movement on two-dimensional substrates and to a marked increase in fibronectin-dependent migration of tumor cells into three-dimensional matrices. Recycling of alpha5beta1 integrin did not affect its regulation or ability to form adhesive bonds with substrate fibronectin. Instead, alpha5beta1 controlled the association of EGFR1 with RCP to promote the coordinate recycling of these two receptors. This modified signaling downstream of EGFR1 to increase its autophosphorylation and activation of the proinvasive kinase PKB/Akt. We conclude that RCP provides a scaffold that promotes the physical association and coordinate trafficking of alpha5beta1 and EGFR1 and that this drives migration of tumor cells into three-dimensional matrices.

alpha6beta4 integrin activates Rac-dependent p21-activated kinase 1 to drive NF-kappaB-dependent resistance to apoptosis in 3D mammary acini.

Malignant transformation and multidrug resistance are linked to resistance to apoptosis, yet the molecular mechanisms that mediate tumor survival remain poorly understood. Because the stroma can influence tumor behavior by regulating the tissue phenotype, we explored the role of extracellular matrix signaling and tissue organization in epithelial survival. We report that elevated (alpha6)beta4 integrin-dependent Rac-Pak1 signaling supports resistance to apoptosis in mammary acini by permitting stress-dependent activation of the p65 subunit of NF-kappaB through Pak1. We found that inhibiting Pak1 through expression of N17Rac or PID compromises NF-kappaB activation and renders mammary acini sensitive to death, but that resistance to apoptosis could be restored to these structures by overexpressing wild-type NF-kappaB p65. We also observed that acini expressing elevated levels of Pak1 can activate p65 and survive death treatments, even in the absence of activated Rac, yet will die if activation of NF-kappaB is simultaneously inhibited through expression of IkappaBalphaM. Thus, mammary tissues can resist apoptotic stimuli by activating NF-kappaB through alpha6beta4 integrin-dependent Rac-Pak1 signaling. Our data emphasize the importance of the extracellular matrix stroma in tissue survival and suggest that alpha6beta4 integrin-dependent Rac stimulation of Pak1 could be an important mechanism mediating apoptosis-resistance in some breast tumors.

Effect of biomaterial surface properties on fibronectin-alpha5beta1 integrin interaction and cellular attachment.

The ability of fibronectin (Fn) to mediate cell adhesion through binding to alpha(5)beta(1) integrins is dependent on the conditions of its adsorption to the surface. Using a model system of alkylsilane SAMs with different functional groups (X=OH, COOH, NH(2) and CH(3)) and an erythroleukemia cell line expressing a single integrin (alpha(5)beta(1)), the effect of surface properties on the cellular adhesion with adsorbed Fn layers was investigated. (125)I-labeled Fn, a modified biochemical cross-linking/extraction technique and a spinning disc apparatus were combined to quantify the Fn adsorption, integrin binding and adhesion strength, respectively. This methodology allows for a binding equilibrium analysis that more closely reflects cellular adhesion found in stable tissue constructs in vivo. Differences in detachment strength and integrin binding were explained in terms of changes in the adhesion constant (psi, related to affinity) and binding efficiency of the adsorbed Fn for the alpha(5)beta(1) integrins (CH(3) approximately NH(2)

Tensional homeostasis and the malignant phenotype.

Tumors are stiffer than normal tissue, and tumors have altered integrins. Because integrins are mechanotransducers that regulate cell fate, we asked whether tissue stiffness could promote malignant behavior by modulating integrins. We found that tumors are rigid because they have a stiff stroma and elevated Rho-dependent cytoskeletal tension that drives focal adhesions, disrupts adherens junctions, perturbs tissue polarity, enhances growth, and hinders lumen formation. Matrix stiffness perturbs epithelial morphogenesis by clustering integrins to enhance ERK activation and increase ROCK-generated contractility and focal adhesions. Contractile, EGF-transformed epithelia with elevated ERK and Rho activity could be phenotypically reverted to tissues lacking focal adhesions if Rho-generated contractility or ERK activity was decreased. Thus, ERK and Rho constitute part of an integrated mechanoregulatory circuit linking matrix stiffness to cytoskeletal tension through integrins to regulate tissue phenotype.