tbuckley/cabot-search huggingface dataset:
704. Enhanced Lentiviral Production Through Rational Design of Mammalian Host Cells
Authors: Jean-Francois Gelinas, Deborah R. Gill, Stephen C. Hyde
Journal: Molecular Therapy (2016)

The development of lentiviral-based therapeutics is hindered by the high costs of cGMP manufacturing, which is a particular concern for applications focused on in vivo delivery where both high titres and volumes are typically required. The key cost component of viral vector manufacturing is the production titre, which is typically several log-orders lower for lentiviral vectors than for non-enveloped vectors such as rAd or rAAV. Some progress has been made towards the generation of producer cell lines to reduce costs. However, the majority of effort to increase lentiviral production has focused on optimisation and scale-up of transient transfection-based processes, including: the selection of efficient gene transfer reagents, the ratio of plasmid DNA components, the composition of cell culture media, supplementation with growth substrates and bioreactor growth conditions (e.g. pH, O2, lactate, etc). Intriguingly, little attention has been paid to the optimisation of the mammalian host cell, with nearly all reported processes relying on commonly available and, readily transfectable, HEK 293T derivatives. We hypothesised that HEK 293T cells were not necessarily optimal for lentiviral production and have embarked on a rational design process to establish cell lines with enhanced manufacturing properties. General reviews of the HIV infectious life cycle typically describe only four host factors that limit lentiviral replication (APOBEC3G, SAMHD1, Tetherin and TRIM5α). We conducted an extensive literature review and identified a further 130 putative restriction factors active in the late phase of the lentiviral life cycle (from transcription to maturation) that are possibly relevant to lentiviral vector production. We have evaluated the effect of siRNA knock-down of these genes (3 days production, 2e5 HEK 293 cells/well, 100 nM siRNA/well in quadruplicate) on lentiviral vector production titre (3 days transduction, 5e4 HEK 293 cells/well, eGFP detection via Perkin Elmer Operetta) in a high-throughput 96-well format. Control studies with a non-target siRNA revealed that the assay variability was less than 0.2-fold (n=40). We showed that siRNA knockdown of the four canonical host restriction factors had no significant effect on lentiviral production (1.06 to 0.68-fold, n=4-23, p > 0.05). We identified 9 host factors, the knockdown of which significantly increased lentiviral production by 1.4 to 2.1-fold (threshold > 2 SD over control; p < 0.05, power > 80%). To extend the utility of these findings we are employing CRISPR/Cas9-mediated genetic disruption of these host factors. We anticipate that permanent disruption, either singly or synergistically in combination, will yield novel mammalian cell lines with a greater ability to support lentiviral production, which will be of considerable value to ongoing efforts to reduce lentiviral production costs and increase the speed of clinical development.
Adjuvant paclitaxel (P)/carboplatin (C) in stages I, II non-small cell lung cancer (NSCLC): Comparison of two schedules in phase II trials by the Minnie Pearl Cancer Research Network
Authors: C. E. McKay, J. D. Hainsworth, D. R. Spigel, H. A. Burris, S. Litchy, D. A. Yardley, G. A. Houston, J. D. Bearden, D. C. Scullin, F. A. Greco
Journal: Journal of Clinical Oncology (2005)

7123 Background: Adjuvant therapy with P (200mg/m2)/C (AUC=6) every 3 weeks for 4 cycles is associated with a 13% increase of survival in stage IB NSCLC patients (pts) (Late/breaking abstracts from the 2004 ASCO meeting. J Clin Oncol 2004;22;suppl:7019). Other recent phase III trials also support survival benefit of adjuvant chemotherapy in early stage NSCLC. In metastatic NSCLC, weekly P with q 4 week C may be superior to q 3 week P/C. We conducted two sequential phase II adjuvant trials in stages I and II NSCLC from 1996–2003 to evaluate the toxicity and feasibility of two different schedules of P/C (weekly P with q 4 week C, q 3 week P/C). Methods: Eligibility criteria for both trials included: complete surgical resection with clear margins; stages IB, IIA, IIB (T2N1, T3N0); 3–8 weeks from surgery; ECOG PS 0, 1; adequate organ function; informed consent. A total of 181 pts were enrolled in both trials. 69 pts were treated in the first trial (IB 38, IIA/B 31) with P (200mg/m2)/C (AUC=6) repeated every 3 weeks for 3 courses and 112 pts were treated in the second trial (IB 38, IIA/B 74) with weekly P 100mg/m2 days 1, 8, 15, and carboplatin (AUC=6) day 1, repeated every 28 days for 3 cycles. Pts were followed every 2–3 months subsequently. Results: Toxicity of both regimens was mild with no treatment-related deaths. There were only 2 hospitalizations for neutropenia/fever with each schedule, but there was more nadir grade 3 neutropenia detected in the weekly schedule. Other grade 3/4 toxicities were unusual and similar in both schedules. Survival at 4 years for all pts with stages IB and IIA/IIB was 70% and 50%, respectively. There was no difference with comparing P schedules (p=0.87 log rank). 90% of the pts completed the every 3-week P/C (82% full doses) and 85% completed the weekly P/C therapy (75% full doses). Conclusions: Both P/C regimens are feasible and well tolerated. The survival of pts in these two phase II trials with 3 cycles appears similar to the recently reported positive phase III trials of 4 cycles of adjuvant chemotherapy. Author Disclosure Employment or Leadership Consultant or Advisory Role Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Bristol-Myers Squibb
Interim Analysis of EFC6663, a Multicenter Phase 2 Study of Alvocidib (flavopiridol), Demonstrates Clinical Responses Among Patients with Fludarabine Refractory CLL
Authors: Mark C Lanasa, Leslie Andritsos, Jennifer R Brown, Janice Gabrilove, Federico Caligaris-Cappio, Richard Larson, Thomas J. Kipps, Veronique Leblond, Donald Milligan, Ann Janssens, Nyla A. Heerema, Stephan Stilgenbauer, John C. Byrd, Michael R. Grever
Journal: Blood (2010)
Diagnosis of Human Prion Disease Using Real-Time Quaking-Induced Conversion Testing of Olfactory Mucosa and Cerebrospinal Fluid Samples
Authors: Matilde Bongianni, Christina Orrù, Bradley R. Groveman, Luca Sacchetto, Michele Fiorini, Giovanni Tonoli, Giorgio Triva, Stefano Capaldi, Silvia Testi, Sergio Ferrari, Annachiara Cagnin, Anna Ladogana, Anna Poleggi, Elisa Colaizzo, Dorina Tiple, Luana Vaianella, Santina Castriciano, Daniele Marchioni, Andrew G. Hughson, Daniele Imperiale, Tatiana Cattaruzza, Gian Maria Fabrizi, Maurizio Pocchiari, Salvatore Monaco, Byron Caughey, Gianluigi Zanusso
Journal: JAMA Neurology (2016)

Early and accurate in vivo diagnosis of Creutzfeldt-Jakob disease (CJD) is necessary for quickly distinguishing treatable from untreatable rapidly progressive dementias and for future therapeutic trials. This early diagnosis is becoming possible using the real-time quaking-induced conversion (RT-QuIC) seeding assay, which detects minute amounts of the disease-specific pathologic prion protein in cerebrospinal fluid (CSF) or olfactory mucosa (OM) samples.To develop an algorithm for accurate and early diagnosis of CJD by using the RT-QuIC assay on CSF samples, OM samples, or both.In this case-control study, samples of CSF and OM were collected from 86 patients with a clinical diagnosis of probable (n = 51), possible (n = 24), or suspected (n = 11) CJD and 104 negative control samples (54 CSF and 50 OM). The CSF and OM samples were analyzed using conventional RT-QuIC. The CSF samples underwent further testing using improved RT-QuIC conditions. In addition, the diagnostic performance of a novel, easy-to-use, gentle flocked swab for sampling of OM was evaluated. Data were collected from January 1 to June 30, 2015.Correlations between RT-QuIC results and the final diagnosis of recruited patients.Among the 86 patients (37 men [43%] and 49 women [57%]; mean [SD] age, 65.7 [11.5] years) included for analysis, all 61 patients with sporadic CJD had positive RT-QuIC findings using OM or CSF samples or both for an overall RT-QuIC diagnostic sensitivity of 100% (95% CI, 93%-100%). All patients with a final diagnosis of non-prion disease (71 CSF and 67 OM samples) had negative RT-QuIC findings for 100% specificity (95% CI, 94%-100%). Of 8 symptomatic patients with various mutations causing CJD or Gerstmann-Sträussler-Scheinker syndrome, 6 had positive and 2 had negative RT-QuIC findings for a sensitivity of 75% (95% CI, 36%-96%).A proposed diagnostic algorithm for sporadic CJD combines CSF and OM RT-QuIC testing to provide virtually 100% diagnostic sensitivity and specificity in the clinical phase of the disease.
Predictors of and Trends in High-Intensity End-of-Life Care Among Children With Cancer: A Population-Based Study Using Health Services Data
Authors: Alisha Kassam, Rinku Sutradhar, Kimberley Widger, Adam Rapoport, Jason D. Pole, Katherine Nelson, Joanne Wolfe, Craig C. Earle, Sumit Gupta
Journal: Journal of Clinical Oncology (2017)

Purpose Children with cancer often receive high-intensity (HI) medical care at the end-of-life (EOL). Previous studies have been limited to single centers or lacked detailed clinical data. We determined predictors of and trends in HI-EOL care by linking population-based clinical and health-services databases. Methods A retrospective decedent cohort of patients with childhood cancer who died between 2000 and 2012 in Ontario, Canada, was assembled using a provincial cancer registry and linked to population-based health-care data. Based on previous studies, the primary composite measure of HI-EOL care comprised any of the following: intravenous chemotherapy < 14 days from death; more than one emergency department visit; and more than one hospitalization or intensive care unit admission < 30 days from death. Secondary measures included those same individual measures and measures of the most invasive (MI) EOL care (eg, mechanical ventilation < 14 days from death). We determined predictors of outcomes with appropriate regression models. Sensitivity analysis was restricted to cases of cancer-related mortality, excluding treatment-related mortality (TRM) cases. Results The study included 815 patients; of these, 331 (40.6%) experienced HI-EOL care. Those with hematologic malignancies were at highest risk (odds ratio, 2.5; 95% CI, 1.8 to 3.6; P < .001). Patients with hematologic cancers and those who died after 2004 were more likely to experience the MI-EOL care (eg, intensive care unit, mechanical ventilation, odds ratios from 2.0 to 5.1). Excluding cases of TRM did not substantively change the results. Conclusion Ontario children with cancer continue to experience HI-EOL care. Patients with hematologic malignancies are at highest risk even when excluding TRM. Of concern, rates of the MI-EOL care have increased over time despite increased palliative care access. Linking health services and clinical data allows monitoring of population trends in EOL care and identifies high-risk populations for future interventions.
Role of matrix metalloproteinase-8 in the modulation of systemic 5-fluorouracil pharmacokinetics by local irradiation.
Authors: C. Hsieh, C. Liu, Y. Hsieh, H. Tai, L. Wang, T. Tsai, Y. Chen
Journal: Journal of Clinical Oncology (2011)

466 Background: Concurrent chemoradiation with 5-fluorouracil (5-FU) is widely accepted for cancer treatment. However, the interactions between radiation and 5-FU remain unclear. We evaluated the influence of local irradiation on the pharmacokinetics of 5-FU in rats. Methods: The single-fraction radiation was delivered to the whole pelvic fields of Sprague-Dawley rats from a linear accelerator after computerized tomography-based planning. 5-FU at 100 mg/kg was intravenously infused 24 hours after radiation. A high-performance liquid chromatography system equipped with a UV detector was used to measure 5-FU in the blood. Matrix metalloproteinase-8 (MMP-8) inhibitor I was administered to the rats to examine whether or not RT modulation of 5-FU pharmacokinetic parameters could be blocked. Results: Compared with sham-irradiated controls, whole pelvic irradiation reduced the area under the concentration versus time curve (AUC) of 5-FU in plasma and, in contrast, increased the AUC in bile in a radiation dose-dependent manner. Based on protein array analysis, the amount of plasma MMP-8 was increased by whole pelvic irradiation (2.8-fold by 0.5 Gy and 5.3-fold by 2 Gy) in comparison with controls. Pretreatment with MMP-8 inhibitor reversed the effect of irradiation on AUC of 5-FU in plasma. Conclusions: Local irradiation may modulate the systemic pharmacokinetics of 5-FU through stimulating the release of MMP-8. This unexpected influence is worthy of further investigation and for consideration in clinical practice of radiotherapy for cancer. No significant financial relationships to disclose.
583. Nonionic Block Copolymers Guide Macrophages To Transfect Skeletal Muscle Cells Upon Direct i. m. Injections
Authors: Vivek Mahajan, Zagit Gaymalov, Richa Gupta, Alexander V. Kabanov
Journal: Molecular Therapy (2015)

Purpose: Skeletal muscle offers a clinically feasible site for i. m. delivery of genes for therapeutic proteins expression and prophylactic DNA vaccine purposes. Though much research has been focused on development of cationic lipids and polymers for encapsulating plasmid DNA (pDNA) for in vitro transfections, direct i. m. injection of pDNA still results in higher muscle transfection levels. In comparison to existing technologies such as viral vectors there still is a need to further improve muscle transfection levels with pDNA injections. Non-ionic block copolymers, Pluronic P85 and SP1017 (mixture of L61 and F127), when mixed with pDNA, enhance skeletal muscle transfection in euthymic (BALB/c and C57BL/6) mouse and not athymic (BALB/c nu/nu) mouse. Therefore, we studied role of immune cells in muscle transfection. Methods and Results: pDNA alone or an admix of pDNA+Pluronic were injected into tibialis anterior (TA) muscle in presence or absence of local inflammation to demonstrate the effect of immune cells in muscle transfection. Local-inflammation drastically enhanced DNA, RNA and gene expression levels when pDNA was co-delivered with P85 or SP1017 and not pDNA alone. Pluronics enhanced in vitro gene expression in MPs to upto 4 orders in magnitude and not muscle cells. Moreover, adoptive transfer of ex vivo transfected MPs with GFP DNA to ischemic mice resulted in GFP expression in muscle fibers along with MPs. Therefore, in vitro muscle inflammation model was developed by co-culturing DNA transfected macrophages (MPs) with myotubes (MTs) to see the effect of P85 on gene transfer to MTs. Treatment of co-culture (pDNA transfected MPs with MTs) with P85 resulted in increase in total and muscle specific gene expression ca. 30 and 120 times respectively, confirming pDNA transfer from MPs to MTs. Moreover, in vivo depletion of MPs resulted in significant decrease in gene expression confirming the role of MPs in Pluronic based muscle transfection. Finally, pre-injection with P85 on day 1.5 and not day 5 at subsequent sites of DNA injections, recruited MPs to the site of injections and further enhanced the gene expression levels of muscles injected with pDNA+P85 and not pDNA alone. Conclusions: Pluronic block copolymers utilize immune cells, specifically MPs, to deliver genes to otherwise very hard to transfect skeletal muscle fibers and hence constitute a novel class of biopharmaceuticals or a platform technology for in vivo gene delivery.
Prognostic relevance of tryptase and chymase positive mast cells and neoangiogenesis in colorectal carcinomas
Authors: M. S. Varela, L. Mauro, L. Puricelli, J. M. Lastiri, M. G. Pallotta, S. Specterman, L. Lauría
Journal: Journal of Clinical Oncology (2004)

3745 Background: Human mast cells (MC) are subdivided into two subsets according to the expression of proteases: MCt positive only for tryptase and MCtc positive for both tryptase and chymase. Their biological role in tumor progression is almost unknown. Methods: We studied the expression patterns of MCt and MCtc in relation to microvessel density (MVD) and other known clinicopathological parameters in 87 primary colorectal carcinomas (CCR) to determine their prognostic significance. Intratumoral and normal peritumoral MC and MVD (CD31+) were analyzed by immunohystochemistry. Positivity was assessed in five areas of 1.2 mm2. Areas of high number of MC and MVD positivity (“hotspot”) were also counted. Overall survival, at five years follow-up was determined. Results: A very high correlation between the two methods to evaluate MC or MDV positivity was found (p<0,01). MCt and MCtc were less frequent inside the tumor than in the normal adjacent mucosa, being MCt the predominant phenotype. A positive correlation between the number of MCt and MVD was found (p<0.01). When we analyzed the relationship between MCt, MCtc or MVD and the main clinicopathological features, we found differences in MCt and MVD according to Duke's stage classification. Thus, patients classified as Duke A always presented the lowest values for both types of MC and MCV. Besides, we found that the number of intratumoral MCt correlated positively with lymphatic and /or blood vessel invasion (p<0.05). To assess prognosis, patients were divided into two groups according to high and low MCt, MCtc or MVD expression. Univariate study indicates a non significant association between high intratumoral MCt and low MCtc. Unexpectedly, a significant association between low MVD expression and bad outcome was found. By multivariate analysis, intratumoral MVD was found to be an independent prognostic indicator. Conclusions: MCt, MCtc and MVD in CCR, is less frequent in tumors from patients with low stage. Although, high intratumoral MCt and low MCtc was observed in tumors from patients with bad outcome, only MVD showed to be an independent prognostic marker. No significant financial relationships to disclose.
Neuropsychological deficits in children with brain tumors at diagnosis: influence of tumor site and tumor-related factors
Authors: L. Iuvone, L. Peruzzi, C. Di Rocco, G. Tamburrini, M. Caldarelli, I. Lazzareschi, P. Maurizi, R. Riccardi
Journal: Journal of Clinical Oncology (2005)

1571 Background Children with Central Nervous System (CNS) tumors are frequently affected by neurocognitive sequelae which deeply influence the quality of life. Surgery, radiotherapy and chemotherapy have been indicated as causes of neurocognitive impairment. We have investigated the role of tumor location and tumor-related factors on neuropsychological deficits before any therapy or surgical intervention in order to obtain baseline values useful to evaluate possible neuropsychological sequelae of subsequent treatments. Methods 54 children with brain tumors were evaluated early after radiological diagnosis. Children with previous neurological disturbances and with severe neurological conditions were excluded. Age at diagnosis: 8,3 y (range: 5m - 18 y); Males/ females: 3/2; Tumor location: posterior fossa: 38,6%; supratentorial: 15,9%; cerebral hemispheres: 45,5%; hydrocephalus: 31,8%. Early signs or symptoms of the tumor and the duration of clinical manifestations at diagnosis were investigated. We measured: total intelligence quotient (TIQ), verbal intelligence quotient (VIQ) and performance intelligence quotient (PIQ); language comprehension and production; short and long term verbal and visual memory; visual motor integration abilities; attention; frontal functions; lateralization. Results Mean IQ values were within the average (TIQ: 97,71 +/- 17,02; VIQ: 103,15 +/- 16,69; PIQ: 95,54+/-15,62). However 22,7% of children had IQ values lower than 85 and half of them were mentally retarded. 80% of them had cortical tumors involving temporo-mesial structures. Lower values of IQ were associated with long lasting clinical signs. Specific cognitive deficits were present in 65,8%, frequently mild and coherent with tumor site. Major neurological signs were related to deficits in memory span; no correlation was found between neurocognitive performances and hydrocephalus. Conclusions Cognitive deficits are already present at the time of diagnosis in a relatevely high percentage and are mainly related to tumor site; children with cortical tumor, mainly in temporal lobes, are at higher risk for severe alterations of cognitive development. (Supported by FOP; Ali Di Scorta) No significant financial relationships to disclose.
EPITHELIOMA OF THE LACRIMAL SAC
Authors: RALPH A. FENTON
Journal: JAMA (1925)

True neoplasms of the lacrimal sac are somewhat unusual. Many reported tumors are granulomatous or degenerative products of long standing inflammation, while others arise intrinsically from various elements in the sac wall. The following cases of true epithelioma are noted in recent literature: Guibert<sup>1</sup>(in 1905), mentioned a limited epithelioma in which cure was effected by the roentgen ray, and following him Rollet<sup>2</sup>and Lafon<sup>3</sup>reported cases of primary epithelioma of the sac, the latter recurrent. Bistis<sup>4</sup>reported a primary cancer in a man of 60, in which the sac wall became 1 cm. thick. Many of the cases reported have come from Italian clinics, and De Vincentiis,<sup>5</sup>as long ago as 1877, pointed out the danger of confusion in diagnosis with ordinary thickening of the sac wall in a chronic dacryocystitis. Maggi,<sup>6</sup>Pasetti,<sup>7</sup>and Denti<sup>8</sup>have contributed several cases. Hermann<sup>9</sup>
4. Developing an Engineered Nipah Virus Glycoprotein Based Lentiviral Vector System Retargeted To Cell Surface Receptors of Choice
Authors: Ruben R. Bender, Anke Muth, Irene C. Schneider, Andrea Maisner, Christian J. Buchholz
Journal: Molecular Therapy (2015)

Delivery of genes selectively into specific cell types is a major bottleneck in successful gene therapy. Receptor-targeted lentiviral vectors (RT-LVs) can be an effective tool to achieve selective transfer of genes to cell types of choice, ex vivo and in vivo. So far, many different target receptors specific for cell types present in many different tissue types have been successfully addressed. RT-LVs based on engineered measles virus (MV) glycoproteins are most selective but can only be produced at moderate vector titers and suffer from susceptibility towards neutralising antibodies induced by vaccination against measles. In this study, LVs have been pseudotyped with modified Nipah virus (NiV) glycoproteins in order to generate LVs with high specificity for target cell populations and enhanced titers. The glycoproteins were engineered to make them deficient for the use of EphrinB2/3, the natural NiV receptor, and truncated in their cytoplasmic tails to allow efficient incorporation into LV particles. Designed ankyrin repeat proteins (DARPins) or single chain antibodies (scFv) specific for the cell surface receptor of choice were fused to the G glycoprotein to mediate receptor binding and subsequent cell entry. All G-DARPin/-scFv fusion proteins were efficiently expressed at the cell surface and incorporated into LV particles. The particles exhibited selective entry into their particular target cells and showed stable transgene expression. Importantly, NiV-LVs were at least 250-fold less effectively neutralized than MV glycoprotein pseudotyped LVs if incubated with pooled human intravenous immunoglobulin (Intratect®). So far, EpCAM, CD4, and CD8 served as functional receptors for NiV glycoprotein based RT-LVs. The CD8-targeted vector selectively transduced CD8+ lymphocytes when added to human PBMCs in vitro. Remarkably, these NiV-based RT-LVs could be produced at 8- to 68-fold higher titers compared to the corresponding MV glycoprotein-based RT-LVs. Current efforts focus on the quantification of these observations and identification of the underlying mechanisms. In addition, the capability of NiV-based RT-LVs to transduce target cell populations in vivo is being analyzed.