A predictive microfluidic model of human glioblastoma to assess trafficking of blood–brain barrier-penetrant nanoparticles

The blood–brain barrier represents a significant challenge for the treatment of high-grade gliomas, and our understanding of drug transport across this critical biointerface remains limited. To advance preclinical therapeutic development for gliomas, there is an urgent need for predictive in vitro models with realistic blood–brain-barrier vasculature. Here, we report a vascularized human glioblastoma multiforme (GBM) model in a microfluidic device that accurately recapitulates brain tumor vasculature with self-assembled endothelial cells, astrocytes, and pericytes to investigate the transport of targeted nanotherapeutics across the blood–brain barrier and into GBM cells. Using modular layer-by-layer assembly, we functionalized the surface of nanoparticles with GBM-targeting motifs to improve trafficking to tumors. We directly compared nanoparticle transport in our in vitro platform with transport across mouse brain capillaries using intravital imaging, validating the ability of the platform to model in vivo blood–brain-barrier transport. We investigated the therapeutic potential of functionalized nanoparticles by encapsulating cisplatin and showed improved efficacy of these GBM-targeted nanoparticles both in vitro and in an in vivo orthotopic xenograft model. Our vascularized GBM model represents a significant biomaterials advance, enabling in-depth investigation of brain tumor vasculature and accelerating the development of targeted nanotherapeutics.

High-grade gliomas are the most common primary malignant brain tumors in adults (1). These include grade IV astrocytomas, commonly known as glioblastoma multiforme (GBM), which account for more than 50% of all primary brain cancers and have dismal prognoses, with a 5-y survival rate of less than 5% (2). Due to their infiltrative growth into the healthy brain tissue, surgery often fails to eradicate all tumor cells (3). While chemotherapy and radiation modestly improve median survival (4), most patients ultimately succumb to their tumors. This is primarily due to the presence of a highly selective and regulated endothelium between blood and brain parenchyma known as the blood–brain barrier (BBB) (5), which limits the entry of therapeutics into the brain tissue where tumors are located. The BBB, characterized by a unique cellular architecture of endothelial cells (ECs), pericytes (PCs), and astrocytes (ACs) (6, 7), displays up-regulated expression of junctional proteins and reduced paracellular and transcellular transports compared to other endothelia (8). While this barrier protects the brain from toxins and pathogens, it also severely restricts the transport of many therapeutics, as evidenced by the low cerebrospinal fluid (CSF)-to-plasma ratio of most chemotherapeutic agents (9). There is thus an important need to develop new delivery strategies to cross the BBB and target tumors, enabling sufficient drug exposure (10).Despite rigorous research efforts to develop effective therapies for high-grade gliomas, the majority of trialed therapeutics have failed to improve outcomes in the clinic, even though the agents in question are effective against tumor cells in preclinical models (11). This highlights the inability of current preclinical models to accurately predict the performance of therapeutics in human patients. To address these limitations, we developed an in vitro microfluidic model of vascularized GBM tumors embedded in a realistic human BBB vasculature. This BBB-GBM platform features brain microvascular networks (MVNs) in close contact with a GBM spheroid, recapitulating the infiltrative properties of gliomas observed in the clinic (12) and those of the brain tumor vasculature, with low permeability, small vessel diameter, and increased expression of relevant junctional and receptor proteins (7). This platform is well suited for quantifying vascular permeability of therapeutics and simultaneously investigating modes of transport across the BBB and into GBM tumor cells.There is strong rationale for developing therapeutic nanoparticles (NPs) for GBM and other brain tumors, as they can be used to deliver a diverse range of therapeutic agents and, with appropriate functionalization, can be designed to exploit active transport mechanisms across the BBB (13, 14). Liposomal NPs have been employed in the oncology clinic to improve drug half-life and decrease systemic toxicity (15), but, to date, no nanomedicines have been approved for therapeutic indications in brain tumors. We hypothesize that a realistic BBB-GBM model composed entirely of human cells can accelerate preclinical development of therapeutic NPs. Using our BBB-GBM model, we investigated the trafficking of layer-by-layer NPs (LbL-NPs) and ultimately designed a GBM-targeted NP. The LbL approach leverages electrostatic assembly to generate modular NP libraries with highly controlled architecture. We have used LbL-NPs to deliver a range of therapeutic cargos in preclinical tumor models (16, 17) and have recently demonstrated that liposomes functionalized with BBB-penetrating ligands improved drug delivery across the BBB to GBM tumors (18). Consistent with clinical data (19), we observed that the low-density lipoprotein receptor-related protein 1 (LRP1) was up-regulated in the vasculature near GBM spheroids in the BBB-GBM model and leveraged this information to design and iteratively test a library of NPs. We show that the incorporation of angiopep-2 (AP2) peptide moieties on the surface of LbL-NPs leads to increased BBB permeability near GBM tumors through LRP1-mediated transcytosis. With intravital imaging, we compared the vascular permeabilities of dextran and LbL-NPs in the BBB-GBM platform to those in mouse brain capillaries and validated the predictive potential of our in vitro model. Finally, we show the capability of the BBB-GBM platform to screen therapeutic NPs and predict in vivo efficacy, demonstrating improved efficacy of cisplatin (CDDP) when encapsulated in GBM-targeting LbL-NPs both in vitro and in vivo.     

The development of unit activity in the lateral geniculate nucleus of the kitten

The spontaneous unit activity in the lateral geniculate nucleus of 15 kittens has been analyzed in terms of the sleep-wake state. Lateral geniculate units in animals ranging from 2 days to 5 weeks of age were recorded under chronic conditions by means of a hydraulically movable glass micropipet. During the first postnatal month, the mean frequency of firing increased progressively in the awake state and in paradoxical sleep but was not modified in quiet or slow wave sleep. However, as early as the third postnatal week, the pattern of modulation of the frequency of unit discharge in response to changes in state reached an organization close to that seen in the adult cat. The most dramatic alteration in the cellular discharge configuration was represented by the introduction of the “burst-pause” pattern in slow wave sleep. Serial interspike interval distributions demonstrated the appearance of this pattern at the end of the second week. The organization of unit firing in the 1-month-old animal was qualitatively similar to that in the adult, though quantitatively less intense. The pattern of development of unit firing is discussed in light of the appearance of other important developmental features such as cortical synchronization, ponto-geniculo-occipital spikes, and maturation of certain monamine-containing cells in the central nervous system. Some structural and neurochemical factors in development are mentioned that may be responsible for the ontogenetic organization of unit discharges in the lateral geniculate nucleus.     

Lung cancer and occupation in nonsmokers: a multicenter case-control study in Europe

BACKGROUND: Tobacco smoking is the main cause for lung cancer worldwide, making it difficult to examine the carcinogenic role of other risk factors because of possible confounding by smoking. Therefore, the present study aimed to investigate the association between lung cancer and occupation independent of smoking. METHODS: A case-control study of lung cancer was carried out between March 1998 and January 2002 in 16 centers from 7 European countries, including 223 never-smoking cases and 1039 controls. Information on lifestyle and occupation was obtained through detailed questionnaires. Job and industries were classified as entailing exposure to known or suspected carcinogens; in addition, expert assessment provided exposure estimates to specific agents. RESULTS: The odds ratio of lung cancer among women employed for more than 12 years in suspected high-risk occupations was 1.75 (95% confidence interval = 0.63-4.85). A comparable increase in risk was not detected for employment in established high-risk occupations or among men. Increased risk of lung cancer was suggested among individuals exposed to nonferrous metal dust and fumes, crystalline silica, and organic solvents. CONCLUSION: Occupations were found to play a limited role in lung cancer risk among never-smokers. Jobs entailing exposure to suspected lung carcinogens should receive priority in future studies among women. Nonferrous metal dust and fumes and silica may exert a carcinogenic effect independently from smoking.     

Comparison of metabolic monitors in critically ill, ventilated patients

OBJECTIVE: We compared the Deltatrac II, the M-COVX, and the Evita 4 metabolic monitoring devices under clinical conditions. METHODS: A prospective simultaneous clinical comparison was performed in a general intensive care department of a tertiary university hospital in 43 ventilated, critically ill patients. The monitors were compared simultaneously. After 30 min of steady state, oxygen consumption per unit time, carbon dioxide consumption per unit time, resting energy expenditure, and respiratory quotient were recorded for the Deltatrac II; the same parameters in addition to end-tidal carbon dioxide and fraction of inspired oxygen were recorded for the M-COVX; and carbon dioxide consumption per unit time, end-tidal carbon dioxide, and fraction of inspired oxygen were recorded for the Evita 4. Pulmonary gas-exchange measurements from the Deltatrac II and resting energy expenditure and respiratory quotient from the M-COVX were obtained after 30 min. The other parameters were calculated from the last five measurements obtained at the end of the study period. RESULTS: A good correlation was found between oxygen consumption per unit time and resting energy expenditure as obtained from the Deltatrac II and the M-COVX (r = 0.76 and 0.75, respectively; P < 0.001), but the correlation was lower between carbon dioxide consumption per unit time as obtained from the Deltatrac II and the M-COVX or Evita 4 (r = 0.67 and 0.48, respectively). Agreement between the different methods did not reach clinical acceptability, exceeding a 20% difference using the Bland-Altman statistical methods. CONCLUSION: Poor agreement was found between the Deltatrac II and M-COVX or Evita 4 metabolic monitors, despite a good correlation between measurements, leading to the conclusion that the M-COVX and Evita 4 provide less accurate measurements of metabolic gas exchange in stable ventilated patients. These devices can be used for daily nutritional assessment and continuous monitoring, but the Deltatrac II remains the method of choice for metabolic measurement.     

Bilateral giant retinal tear and retinal detachment in a young emmetropic man after Jarish-Herxheimer reaction in ocular syphilis

BACKGROUND: Giant retinal tears were previously described in patients affected by panuveitis. We report the case of a patient presenting giant retinal tears in both eyes affected by ocular syphilis. PATIENT AND METHOD: A 45-year-old patient presented a 5 days history of sudden bilateral drop of vision, two weeks after penicillin therapy for secondary syphilis. The best visual acuity was 0.5 RE and light perception LE. Biomicroscopy showed an intense vitritis associated with bilateral giant tear and retinal detachment. Both TPHA and VDRL were positive. Lumbar puncture showed lymphoplasmocytosis with intrathecal synthesis of IgM. RESULTS: Topical steroids treatment was applied and intravenous penicillin was given during 14 days. Pars plana vitrectomy with silicon oil tamponade was performed in association with endophotocoagulation in the left eye and cryocoagulation in the right eye. Vitreous PCR was negative. Evolution was successful with an attached retina. CONCLUSIONS: This represents the first reported case of giant retinal tear with retinal detachment in a patient presenting a syphilitic panuveitis.     

The effect of cyclo-DfKRG peptide immobilization on titanium on the adhesion and differentiation of human osteoprogenitor cells

This study takes place in the field of development of a bioactive surface of titanium alloys. In this paper, titanium was functionalized with cyclo-DfKRG peptide by coating or grafting using different anchors (thiol or phosphonate) as spacers between the surface and the peptide. Cell adhesion, and differentiation of human osteoprogenitor (HOP) cells arising from human bone marrow were investigated. Our results seem to demonstrate that cyclo-DfKRG peptide coating with a phosphonate anchor and grafting procedure contributes to higher cell adhesion and a strong ALP and Cbfa1 mRNA expression, after 10 days of cell seeding. At the contrary, this peptide coated with a thiol anchor stimulates differentiation of HOP within 3 days of culture.     

Surgical Management of Metastases to the Thyroid Gland

Background

Metastases to the thyroid gland are uncommon, with rates reported between 0.02% and 1.4% of surgically resected thyroid specimens. Our goal was to present our experience with surgical management of metastases to the thyroid gland.

Methods

Twenty-one patients with metastatic disease to the thyroid were identified from a database of 1,992 patients with thyroid cancer who had surgery during 1986–2005. Patient, tumor, treatment, and outcome details were recorded by analysis of charts. The median age at time of surgery was 68 (range, 39–83) years; 12 were men and 9 were women.

Results

All patients were managed by surgery, including lobectomy in ten patients, total thyroidectomy in six, completion thyroidectomy in two, and subtotal thyroidectomy in one. In two patients, the thyroid lesion was found to be unresectable at the time of surgery. Histopathology revealed renal cell carcinoma in ten, malignant melanoma in three, gastrointestinal adenocarcinoma in three, breast cancer in one, sarcoma in one, and adenocarcinoma from an unknown primary site in three patients. Seventeen patients have died. The cause of death in all 17 was widespread metastatic disease from their respective primary tumors. The median survival from surgery to death or last follow-up was 26.5 (range, 2–114) months.

Conclusions

In patients with metastases to the thyroid gland, local control of metastatic disease in the central compartment of the neck can be successfully achieved with minimal morbidity with surgical resection in selected patients.     

Select screening in a specific high-risk population of patients suggests a stage migration toward detection of non-muscle-invasive bladder cancer

Background

More than 25% of bladder cancer (BC) cases are still muscle-invasive at first diagnosis. Screening is unproven to enable the detection of more non–muscle-invasive tumors. BC association with aristolochic acid nephropathy (AAN) was reported after intake of slimming pills containing Chinese herbs.

Objective

We evaluated whether a BC screening protocol in a high-risk and unique patient population had an impact on the stage of tumor presentation.

Design, setting, and participants

Forty-eight AAN-affected patients were enrolled in a screening program, establishing BC incidence during prospective screening cystoscopies and biopsies biannually for up to 10 yr. Two patients were lost to follow-up, and three refused screening after consenting.

Measurements

Patients were evaluated for presence of BC and tumor stage at diagnosis.

Results and limitations

BC was diagnosed in 25 patients (52%). Among 43 patients who underwent screening cystoscopies (median follow-up: 94 mo), 22 were first diagnosed with non–muscle-invasive BC but none with muscle-invasive tumors and none died of BC. Three women who declined follow-up were diagnosed and died with advanced metastatic disease. The limitations of our findings include the small sample size of this case series, the absence of a real control group, and the particular risk factor in these patients that differs from the usual risk factors, such as smoking or industrial chemicals.

Conclusions

BC screening in high-risk groups may allow identification of tumors before muscle invasion. The optimal screening schedule and the relevance of the present findings in smoking-related BC remain to be defined.