Halicin Is Effective Against Staphylococcus aureus Biofilms In Vitro.

BACKGROUND: Biofilms protect bacteria from the host immune system and many antibiotics, making the treatment of orthopaedic infections difficult. Halicin, a recently discovered antibiotic, has potent activity against nonorthopaedic infections in mice and the planktonic, free-living forms of many bacterial species, including Staphylococcus aureus , a common cause of orthopaedic infections. Importantly, halicin did not induce resistance in vitro and was effective against drug-resistant bacteria and proliferating and quiescent bacteria. Quiescence is an important cause of antibiotic tolerance in biofilms. However, whether halicin acts on biofilms has not been tested. QUESTIONS/PURPOSES: (1) Does halicin reduce the viability of S. aureus in less mature and more mature biofilms as it does in planktonic cultures? (2) How do the relative effects of halicin on S. aureus biofilms and planktonic cultures compare with those of conventional antibiotics (tobramycin, cefazolin, vancomycin, or rifampicin) that are commonly used in clinical orthopaedic infections? METHODS: To measure minimal biofilm eradication concentrations (MBECs) with less mature 3-day and more mature 7-day biofilms, we used 96-well peg plates that provided high throughput and excellent reproducibility. After S. aureus -Xen36 biofilm formation, planktonic bacteria were removed from the cultures, and the biofilms were exposed to various concentrations of halicin, tobramycin, cefazolin, vancomycin, or rifampicin for 20 hours. Biofilm viability was determined by measuring resazurin reduction or by counting colony-forming units after sonication. To determine effects of halicin and the conventional antibiotics on biofilm viability, we defined MBEC 75 as the lowest concentration that decreased viability by 75% or more. To determine effects on bacterial viability in planktonic cultures, minimum inhibitory concentrations (MICs) were determined with the broth dilution method. Each result was measured in four to 10 independent experiments. RESULTS: We found no differences between halicin's effectiveness against planktonic S. aureus and 3-day biofilms (MIC and MBEC 75 for 3-day biofilms was 25 µM [interquartile range 25 to 25 and 25 to 25, respectively]; p > 0.99). Halicin was eightfold less effective against more mature 7-day biofilms (MBEC 75 = 200 µM [100 to 200]; p < 0.001). Similarly, tobramycin was equally effective against planktonic culture and 3-day biofilms (MIC and MBEC 75 for 3-day biofilms was 20 µM [20 to 20 and 10 to 20, respectively]; p > 0.99). Tobramycin's MBEC 75 against more mature 7-day biofilms was 320 µM (320 to 480), which is 16-fold greater than its planktonic MIC (p = 0.03). In contrast, the MBEC 75 for cefazolin, vancomycin, and rifampicin against more mature 7-day biofilms were more than 1000-fold (> 1000; p < 0.001), 500-fold (500 to 875; p < 0.001), and 3125-fold (3125 to 5469; p = 0.004) greater than their planktonic MICs, respectively, consistent with those antibiotics' relative inactivity against biofilms. CONCLUSION: Halicin was as effective against S. aureus in less mature 3-day biofilms as those in planktonic cultures, but eightfold higher concentrations were needed for more mature 7-day biofilms. Tobramycin, an antibiotic whose effectiveness depends on biofilm maturity, was also as effective against S. aureus in less mature 3-day biofilms as those in planktonic cultures, but 16-fold higher concentrations were needed for more mature 7-day biofilms. In contrast, cefazolin, vancomycin, and rifampicin were substantially less active against both less and more mature biofilms than against planktonic cultures. CLINICAL RELEVANCE: Halicin is a promising antibiotic that may be effective against S. aureus osteomyelitis and infections on orthopaedic implants. Future studies should assess the translational value of halicin by testing its effects in animal models of orthopaedic infections; on the biofilms of other bacterial species, including multidrug-resistant bacteria; and in combination therapy with conventional antibiotics.

Failure to Cure Patients with Colorectal Liver Metastases: The Impact of the Liver Surgeon.

BACKGROUND: Lack of a liver surgeon (LS) may lead to failure to cure in patients with possibly resectable colorectal liver metastases (CRLM). This study aims to quantify the failure-to-cure rate due to noninclusion of an LS. PATIENTS AND METHODS: All patients who underwent chemotherapy with palliative intent for CRLM at a community oncology network between 2010 and 2018 were identified from a prospectively maintained cancer registry. Two LS blinded to patient management and outcome reviewed pretreatment imaging and assigned each scan a newly developed resectability score. Nominal group technique and independent scores were combined to determine probability of curative-intent resection. Interobserver agreement was calculated using κ testing. RESULTS: This study included 72 palliative CRLM patients. Demographic factors were: 44 (59%) male, median age 68 years (range 36-94 years), 23 (32%) rectal primary, 24 (33%) receiving oxaliplatin-based chemotherapy. Of the 72 patients with CRLM, 6 had left-sided metastases only. The median number of CRLM was 6 (1-8). Agreement on resectability was achieved in 32 (44%) patients for the entire cohort and 17 (54%) in patients without extrahepatic disease. A lower median number of CRLM was found in the group considered to be resectable by the two LS (2 versus 8; p = 0.001). Substantial agreement was found between liver surgeons in the group of patients without extrahepatic disease (κ = 0.9043). CONCLUSIONS: Over 44% of patients who were assigned palliative chemotherapy at tumor boards without an LS were considered potentially resectable upon independent LS review.

The evolution of single cell-derived colorectal cancer cell lines is dominated by the continued selection of tumor-specific genomic imbalances, despite random chromosomal instability.

Intratumor heterogeneity is a major challenge in cancer treatment. To decipher patterns of chromosomal heterogeneity, we analyzed six colorectal cancer cell lines by multiplex interphase FISH (miFISH). The mismatch-repair-deficient cell lines DLD-1 and HCT116 had the most stable copy numbers, whereas aneuploid cell lines (HT-29, SW480, SW620 and H508) displayed a higher degree of instability. We subsequently assessed the clonal evolution of single cells in two colorectal carcinoma cell lines, SW480 and HT-29, which both have aneuploid karyotypes but different degrees of chromosomal instability. The clonal compositions of the single cell-derived daughter lines, as assessed by miFISH, differed for HT-29 and SW480. Daughters of HT-29 were stable, clonal, with little heterogeneity. Daughters of SW480 were more heterogeneous, with the single cell-derived daughter lines separating into two distinct populations with different ploidy (hyper-diploid and near-triploid), morphology, gene expression and tumorigenicity. To better understand the evolutionary trajectory for the two SW480 populations, we constructed phylogenetic trees which showed ongoing instability in the daughter lines. When analyzing the evolutionary development over time, most single cell-derived daughter lines maintained their major clonal pattern, with the exception of one daughter line that showed a switch involving a loss of APC. Our meticulous analysis of the clonal evolution and composition of these colorectal cancer models shows that all chromosomes are subject to segregation errors, however, specific net genomic imbalances are maintained. Karyotype evolution is driven by the necessity to arrive at and maintain a specific plateau of chromosomal copy numbers as the drivers of carcinogenesis.

PSEUDOMARKER 2.0: efficient computation of likelihoods using NOMAD.

BACKGROUND: PSEUDOMARKER is a software package that performs joint linkage and linkage disequilibrium analysis between a marker and a putative disease locus. A key feature of PSEUDOMARKER is that it can combine case-controls and pedigrees of varying structure into a single unified analysis. Thus it maximizes the full likelihood of the data over marker allele frequencies or conditional allele frequencies on disease and recombination fraction. RESULTS: The new version 2.0 uses the software package NOMAD to maximize likelihoods, resulting in generally comparable or better optima with many fewer evaluations of the likelihood functions. CONCLUSIONS: After being modified substantially to use modern optimization methods, PSEUDOMARKER version 2.0 is more robust and substantially faster than version 1.0. NOMAD may be useful in other bioinformatics problems where complex likelihood functions are optimized.

Equilibrium landscape of ingress/egress channels and gating residues of the Cytochrome P450 3A4.

The Cytochrome P450 (CYP) enzymes metabolize a variety of drugs, which may potentially lead to toxicity or reduced efficacy when drugs are co-administered. These drug-drug interactions are often manifested by CYP3A4, the most prevalent of all CYP isozymes. We carried out multiple MD simulations employing CAVER to quantify the channels, and Hidden Markov Models (HMM) to characterize the behavior of the gating residues. We discuss channel properties, bottleneck residues with respect to their likelihood to deem the respective channel ingress or egress, gating residues regarding their open or closed states, and channel location relative to the membrane. Channels do not display coordinated motion and randomly transition between different conformations. Gateway residues also behave in a random fashion. Our findings shed light on the equilibrium behavior of the gating residues and channels in the apo state.

A functional interaction between the MAGUK protein hDlg and the gap junction protein connexin 43 in cervical tumour cells.

Gap junctions, composed of Cxs (connexins), allow direct intercellular communication. Gap junctions are often lost during the development of malignancy, although the processes behind this are not fully understood. Cx43 is a widely expressed Cx with a long cytoplasmic C-terminal tail that contains several potential protein-interaction domains. Previously, in a model of cervical carcinogenesis, we showed that the loss of gap junctional communication correlated with relocalization of Cx43 to the cytoplasm late in tumorigenesis. In the present study, we demonstrate a similar pattern of altered expression for the hDlg (human discs large) MAGUK (membrane-associated guanylate kinase) family tumour suppressor protein in cervical tumour cells, with partial co-localization of Cx43 and hDlg in an endosomal/lysosomal compartment. Relocalization of these proteins is not due to a general disruption of cell membrane integrity or Cx targeting. Cx43 (via its C-terminus) and hDlg interact directly in vitro and can form a complex in cells. This novel interaction requires the N- and C-termini of hDlg. hDlg is not required for Cx43 internalization in W12GPXY cells. Instead, hDlg appears to have a role in maintaining a cytoplasmic pool of Cx43. These results demonstrate that hDlg is a physiologically relevant regulator of Cx43 in transformed epithelial cells.

Solubility of ammonium metal fluorides in aqueous ethanol mixtures - implications for scandium recovery by antisolvent crystallization.

The recovery of scandium from waste streams of other mining and metallurgical processing industries is gaining research interest due to the scarcity of scandium-containing ores. Hydrometallurgical techniques such as leaching, solvent extraction and crystallization amongst others have been successfully applied to recover scandium salts from such waste streams. Scandium can be recovered as (NH4)3ScF6 by antisolvent crystallization from NH4F strip liquors obtained after solvent extraction. The coextraction of metal impurities such as Fe, Al, Zr and Ti causes contamination of the final solid product. The extent of coprecipitation of ammonium metal fluorides depends on their initial concentration in the strip liquor and their solubility in the NH4F-antisolvent mixtures. Here, the solubility of ammonium metal fluorides of Sc, Zr, Fe, Al and Ti is reported separately in 3 mol L-1 NH4F-ethanol mixtures at 25 °C as well as in a system containing all five solid phases. The solubility of (NH4)3ZrF7 is slightly higher than that of (NH4)3ScF6, while the solubilities of (NH4)3FeF6 and (NH4)3AlF6 are significantly lower in comparison to (NH4)3ScF6. The solubility of (NH4)2TiF6 is 1-2 orders of magnitude higher than those of other ammonium metal fluorides. When a mixture of ammonium metal fluoride salts is dissolved in the same 3 mol L-1 NH4F-ethanol mixture as for the individual salts, the resultant solubility of the ammonium metal fluorides of Sc, Zr and Fe decreases significantly, while the resultant solubility of ammonium aluminum hexafluoride increases. This is likely due to changes in solution speciation with increased NH4F concentration and ionic strength.

From Trace to Pure: Pilot-Scale Scandium Recovery from TiO2 Acid Waste.

Scandium (Sc), declared a critical raw material in the European Union (EU), could face further supply issues as the EU depends almost entirely on imports from China, Russia, and Ukraine. In this study, a tandem nanofiltration-solvent extraction procedure for Sc recovery from titania (TiO2) acid waste was piloted and then augmented by antisolvent crystallization. The new process, comprising advanced filtration (hydroxide precipitation, micro-, ultra-, and nanofiltration), solvent extraction, and antisolvent crystallization, was assessed in relation to material and energy inputs and benchmarked on ScF3 production. From ∼1 m3 of European acid waste containing traces of Sc (81 mg L-1), ∼13 g of Sc (43% yield, nine stages) was recovered as (NH4)3ScF6 with a purity of approximately 95%, demonstrating the technical feasibility of the approach. The production costs per kilogram of ScF3 were lower than reported market prices, which underscores a competitive process at scale. Although a few technical bottlenecks (e.g., S/L separation and electricity consumption) need to be overcome, combining advanced filtration with solvent extraction and antisolvent crystallization promises a future supply of this critical raw material from European secondary sources.

Melanoma cell-derived factors stimulate hyaluronan synthesis in dermal fibroblasts by upregulating HAS2 through PDGFR-PI3K-AKT and p38 signaling.

In many cancers hyaluronan content is increased, either by tumor cells or the surrounding stromal cells and this increased hyaluronan content correlates with unfavorable clinical prognosis. In the present work, we studied the effects of melanoma cell (aggressive melanoma cell line C8161)-derived factors on fibroblast hyaluronan synthesis, intracellular signaling, MMP expression and invasion. Treatment of the fibroblast cultures with melanoma cell conditioned medium (CM) caused accumulation of hyaluronan in the culture medium and formation of thick pericellular hyaluronan coat and hyaluronan cables. The expression of Has2 was increased approximately 20-fold by the C8161 melanoma cell CM, while Has1 and Has3 were increased twofold. Knock-down of Has2 expression with siRNA showed that Has2 was responsible for the increased hyaluronan synthesis induced by the melanoma cell CM. To find out the signaling routes, which led to Has2 upregulation, the phosphorylation profiles of 46 kinases were screened with phosphokinase array kit. Melanoma cell CM treatment strongly induced a rapid phosphorylation of p38, JNK, AKT, CREB, HSP27, STAT3 and cJUN. Treatment of the fibroblasts with specific inhibitors of PI3K, AKT and p38 reduced the melanoma cell CM-induced hyaluronan secretion, while the inhibitor of PDGFR totally blocked it. In addition, siRNA for PDGFRα/ß inhibited Has2 upregulation in melanoma cell CM-treated fibroblasts. In parallel with the increased hyaluronan synthesis the melanoma cell CM-treated fibroblasts showed spindle shape, numerous long cell protrusions, enhanced MMP expression and increased invasion into collagen-Cultrex matrix. siRNA blocking of Has2 or PDGFRα/ß expression reversed the stimulatory effect of melanoma cell CM on fibroblast invasion. PDGF secreted by melanoma cells thus mediated fibroblasts activation, with HAS2 upregulation as a major factor in the fibroblast response. This effect on stromal matrix is suggested to favor tumor growth.

Fatal Allograft Rejection and Cardiac Allograft Vasculopathy After Treatment With Pembrolizumab for Metastatic Melanoma in a Heart Transplant Recipient: A Case Report.

Checkpoint inhibitors decrease the progression of many cancers. However, the experience in immunosuppressed patients is limited, with reports of possible serious adverse events. We present a heart transplant recipient treated with pembrolizumab for metastatic melanoma who developed fatal rejection. The patient was a 29 year-old man who underwent heart transplantation at the age of 10 years for congenital heart disease. Seventeen years after transplant, he was diagnosed with scalp melanoma pT3a, N2a, M0, Stage IIIA, positive for BRAF V600E mutation treated with excision, which metastasized to his lungs and brain a year later. Dabrafenib and trametinib were started with transient response. Additional options and their risks were discussed, and pembrolizumab was started 4 months later due to the incomplete response to previous therapy. Five days after initiation the patient presented with moderate cellular rejection and possible antibody mediated rejection (ISHLT Grade 2R, pAMR 1H). Pembrolizumab was discontinued, and he was treated with steroids. Seven months later he presented in cardiogenic shock and severe coronary allograft vasculopathy. Biopsy was negative for cellular rejection, but suspicious for antibody mediated rejection (ISHLT Grade 0R, pAMR 1H), and he had a new serum alloantibody. Despite steroids and plasmapheresis he remained in refractory cardiogenic shock and died of cardiac arrest.

Keratinocytes stimulate fibroblast hyaluronan synthesis through the release of stratifin: a possible role in the suppression of scar tissue formation.

Scar tissue formation during healing of extensive skin wounds may be related to delayed reepithelialization, while elevated levels of hyaluronan may suppress scar tissue formation. This study investigates the expression of hyaluronan by human skin fibroblasts in response to keratinocyte-conditioned medium (KCM), and attempts to identify any active factors within the conditioned medium. Serum-free KCM was assessed for its ability to stimulate the incorporation of (3) H-glucosamine into fibroblast glycosaminoglycans, and hyaluronan synthesis. Conditioned medium was concentrated with an ultrafiltration membrane with a 30 kDa cutoff. Stratifin was assessed for its ability to stimulate hyaluronan synthesis and its role in KCM. KCM stimulated fibroblast glycosaminoglycan synthesis up to a 3.3-fold increase and a 6.5-fold increase in hyaluronan synthesis compared with serum-free controls. Preliminary characterization of the active factors showed that they are retained by a >30 kDa ultrafiltration membrane, and are protease sensitive and heat resistant. Emmprin and stratifin were shown to stimulate fibroblast hyaluronan synthesis, and the hyaluronan-stimulating activity of KCM was removed following stratifin depletion and to a lesser extent by emmprin depletion. Keratinocytes release soluble factors, including stratifin, that stimulate fibroblast hyaluronan synthesis, and this stimulation of fibroblast hyaluronan may contribute to the suppression of scar tissue formation.

Single Cell Genetic Profiling of Tumors of Breast Cancer Patients Aged 50 Years and Older Reveals Enormous Intratumor Heterogeneity Independent of Individual Prognosis.

PURPOSE: Older breast cancer patients are underrepresented in cancer research even though the majority (81.4%) of women dying of breast cancer are 55 years and older. Here we study a common phenomenon observed in breast cancer which is a large inter- and intratumor heterogeneity; this poses a tremendous clinical challenge, for example with respect to treatment stratification. To further elucidate genomic instability and tumor heterogeneity in older patients, we analyzed the genetic aberration profiles of 39 breast cancer patients aged 50 years and older (median 67 years) with either short (median 2.4 years) or long survival (median 19 years). The analysis was based on copy number enumeration of eight breast cancer-associated genes using multiplex interphase fluorescence in situ hybridization (miFISH) of single cells, and by targeted next-generation sequencing of 563 cancer-related genes. RESULTS: We detected enormous inter- and intratumor heterogeneity, yet maintenance of common cancer gene mutations and breast cancer specific chromosomal gains and losses. The gain of COX2 was most common (72%), followed by MYC (69%); losses were most prevalent for CDH1 (74%) and TP53 (69%). The degree of intratumor heterogeneity did not correlate with disease outcome. Comparing the miFISH results of diploid with aneuploid tumor samples significant differences were found: aneuploid tumors showed significantly higher average signal numbers, copy number alterations (CNAs) and instability indices. Mutations in PIKC3A were mostly restricted to luminal A tumors. Furthermore, a significant co-occurrence of CNAs of DBC2/MYC, HER2/DBC2 and HER2/TP53 and mutual exclusivity of CNAs of HER2 and PIK3CA mutations and CNAs of CCND1 and PIK3CA mutations were revealed. CONCLUSION: Our results provide a comprehensive picture of genome instability profiles with a large variety of inter- and intratumor heterogeneity in breast cancer patients aged 50 years and older. In most cases, the distribution of chromosomal aneuploidies was consistent with previous results; however, striking exceptions, such as tumors driven by exclusive loss of chromosomes, were identified.

Effect of different classes of gadolinium-based contrast agents on control and nephrogenic systemic fibrosis-derived fibroblast proliferation.

PURPOSE: To determine the ability of different types of gadolinium-based contrast agents (GBCAs) to stimulate fibroblast proliferation in monolayer cell culture. MATERIALS AND METHODS: The National Health Service West Glasgow Ethics Committee granted approval for this study. Fibroblasts established from healthy volunteers (control subjects) and from lesional skin of patients with nephrogenic systemic fibrosis were exposed to a range of concentrations of ionic and nonionic linear and macrocyclic contrast agents over 4 days, and the effect on growth was determined. The lowest concentration of contrast agent that stimulated the maximum effect on fibroblast growth was selected for determination of its effect on fibroblast growth over 8 days. The effect of contrast agents on hyaluronan and collagen synthesis was determined with an enzyme-linked immunosorbent assay. Responses were assessed with analysis of variance (general linear model). RESULTS: The linear gadolinium contrast agents (gadodiamide, gadoversetamide, gadopentetate dimeglumine, and gadobenate dimeglumine) produced a maximum stimulation of fibroblast proliferation at a concentration of 0.1 mmol/L, with cell numbers increasing up to 2.3-fold. The macrocyclic contrast agents (gadoteric acid and gadoteridol) produced a maximum stimulation of fibroblast proliferation at a concentration of 5 mmol/L. The reference gadolinium agents (N-methylglucamine gadolinium ethylenediaminetetraacetic acid and gadolinium trichloride) stimulated fibroblast proliferation at a concentration of 0.01 mmol/L and were toxic at a concentration greater than 1 mmol/L. Growth curves supported the dose-response observations. Hyaluronan synthesis was stimulated by gadoversetamide, gadobenate dimeglumine, gadodiamide, and gadopentetate dimeglumine at a concentration of 0.1 mmol/L and by gadolinium trichloride at a concentration of 0.01 mmol/L, whereas collagen synthesis was unaffected. CONCLUSION: This study provides evidence that different classes of gadolinium chelates stimulate human fibroblast proliferation.

Oro-facial fibrosis in systemic sclerosis: a reconstructive journey.

Oro-facial fibrosis presents a significant disease burden in patients with systemic sclerosis, but there remains no established treatment modality. Autologous fat grafting is a minimally invasive surgical procedure that is now increasingly recognised for its regenerative capacity, propagating an expansion of heterogeneous indications beyond volume restoration, including fibrotic diseases such as systemic sclerosis. We present a 42-year-old woman with oro-facial involvement of systemic sclerosis leading to severe limitation in mouth opening and closure, with marked retraction of the lower lip and gingival display. We describe the reconstructive journey over a 12-year period, where the antifibrotic effect of autologous fat grafting served as the basis on which a series of surgical procedures were performed to achieve functional and aesthetic improvement. Autologous fat grafting provides a novel treatment modality for oro-facial skin fibrosis, previously considered a non-treatable disease manifestation of systemic sclerosis.

Feasibility study of stem-cell enriched autologous lipotransfer to treat oro-facial fibrosis in systemic sclerosis (Sys-Stem): Protocol for open-label randomised controlled trial.

INTRODUCTION: Oro-facial fibrosis is a common and disabling manifestation of systemic sclerosis (SSc), causing a plethora of functional, aesthetic and social compromise, yet is without effective treatment. Autologous lipotransfer is an established minimally invasive surgical procedure that is postulated to exert anti-fibrotic effects by adipose-derived stem cells, and presents a novel method in the treatment of fibrotic conditions. This study aims to assess the safety and efficacy of autologous lipotransfer for facial involvement in SSc. METHODS AND ANALYSIS: This is the first randomised controlled study with an open label design to assess autologous lipotransfer for oro-facial involvement in systemic sclerosis. The goals of this study are to assess the feasibility of using a range of quantitative and qualitative outcome measures to effectively measure disease severity and treatment outcome, and to assess patient acceptability for future multi-centre trials. A total of 50 participants will be randomised to a treatment or control group. The treatment group will receive autologous fat transfer to the peri-oral region by a single surgeon. Dermal fibroblasts and adipose-derived stem cells will be isolated from tissue samples. All outcome measures will be taken at baseline, then at 6 weeks, 3 months and 6 months from the time of intervention in the treatment arm, or from baseline in the control arm. ETHICS AND DISSEMINATION: The study has ethical approval (REC reference 19/LO/0718). Results will be available to patients, patient user groups, clinicians and the public through presentations at national and international rheumatology conferences and published in peer reviewed journals. TRIAL REGISTRATION: Registered on ISRCTN registry (ISRCTN17793055).

Lipotransfer provides effective soft tissue replacement for acquired partial lipodystrophy.

We present a 48-year-old female patient who presented with features consistent with acquired partial lipodystrophy (APL) also known as 'Barraquer-Simons syndrome'. It is a rare disease characterised by a gradual and progressive onset of lipoatrophy limited to the face, neck, upper limbs, thorax and abdomen and sparing the lower extremities. The resultant physical appearance can have significant psychosocial sequelae, further compounded by misdiagnosis and delay in recognition and management. Treatment is aimed at surgical correction of soft tissue destruction. Autologous fat transfer is an established plastic and reconstructive procedure that is safe and minimally invasive and can be used to reconstruct a variety of soft tissue defects and has shown to be an effective treatment modality in patients with APL.

Moderately High Tenofovir Diphosphate in Dried Blood Spots Indicates Drug Resistance in Viremic Persons Living with HIV.

BACKGROUND: Tenofovir diphosphate (TFV-DP) in dried blood spots (DBS) is a strong predictor of viral suppression in persons living with HIV (PLWH). Its association with antiretroviral therapy (ART) resistance remains unknown. METHODS: Blood was collected in PLWH receiving TDF-containing ART enrolled in a 48-week study. Tenofovir diphosphate/emtricitabine triphosphate (FTC-TP) were quantified from the same sample as HIV viral load (VL) in PLWH who developed resistance within ≤12 months. RESULTS: The study enrolled 807 participants, of whom 10 had new resistance-conferring mutations. Among these, median (interquartile range) TFV-DP and HIV VL were 956 (407-1510) fmol/punch and 9840 (513-68,200) copies/mL, respectively. Five had quantifiable FTC-TP in DBS. Based on previously published data, a TFV-DP concentration of 956 fmol/punch would have an adjusted odds of virologic suppression of 32.8 versus TFV-DP <350 fmol/punch, making viremia of ∼10,000 copies/mL an unexpected outcome. CONCLUSION: Moderately high TFV-DP in DBS (700-1249 fmol/punch) in PLWH with high viremia suggest that antiretroviral drug resistance might be present.

Role of Stereotactic Body Radiation Therapy Before Orthotopic Liver Transplantation: Retrospective Evaluation of Pathologic Response and Outcomes.

PURPOSE: To analyze the results of stereotactic body radiation therapy (SBRT) in patients with early-stage, localized hepatocellular carcinoma who underwent definitive orthotopic liver transplantation (OLT). METHODS AND MATERIALS: The subjects of this retrospective report are 38 patients diagnosed with hepatocellular carcinoma who underwent SBRT per institutional phase 1 to 2 eligibility criteria, before definitive OLT. Pre-OLT radiographs were compared with pathologic gold standard. Analysis of treatment failures and deaths was undertaken. RESULTS: With median follow-up of 4.8 years from OLT, 9 of 38 patients (24%) recurred, whereas 10 of 38 patients (26%) died. Kaplan-Meier estimates of 3-year overall survival and disease-free survival are 77% and 74%, respectively. Sum longest dimension of tumors was significantly associated with disease-free survival (hazard ratio 1.93, P=.026). Pathologic response rate (complete plus partial response) was 68%. Radiographic scoring criteria performed poorly; modified Response Evaluation Criteria in Solid Tumors produced highest concordance (κ = 0.224). Explants revealed viable tumor in 74% of evaluable patients. Treatment failures had statistically larger sum longest dimension of tumors (4.0 cm vs 2.8 cm, P=.014) and non-statistically significant higher rates of lymphovascular space invasion (44% vs 17%), cT2 disease (44% vs 21%), ≥pT2 disease (67% vs 34%), multifocal tumors at time of SBRT (44% vs 21%), and less robust mean α-fetoprotein response (-25 IU/mL vs -162 IU/mL). CONCLUSIONS: Stereotactic body radiation therapy before to OLT is a well-tolerated treatment providing 68% pathologic response, though 74% of explants ultimately contained viable tumor. Radiographic response criteria poorly approximate pathology. Our data suggest further stratification of patients according to initial disease burden and treatment response.

Phase 1 study of abemaciclib, an inhibitor of CDK 4 and 6, as a single agent for Japanese patients with advanced cancer.

PURPOSE: To confirm the safety and tolerability, evaluate the pharmacokinetics (PK), and investigate the antitumor activity of abemaciclib in Japanese patients with advanced cancer. METHODS: We conducted a non-randomized, single-arm, open-label, dose-escalation phase 1 study of abemaciclib administered orally every 12 h (Q12H) on a 28-day cycle at doses of 100 mg (Cohort 1, n = 3), 150 mg (Cohort 2, n = 3), or 200 mg [Cohort 3, n = 6, maximum tolerated dose (MTD)]. Dose escalation was based on the frequency of dose-limiting toxicity (DLT). MTD, as established in the previous phase 1 study in non-Japanese patients, was the highest dose level at which <33 % of patients experienced DLT. RESULTS: Eleven of the 12 patients who received treatment with abemaciclib discontinued: 10 patients due to progressive disease, and 1 due to a DLT (Cohort 3, grade 2 nausea). Diarrhea, the most common treatment-emergent adverse event (AE), was managed supportively and did not require study treatment discontinuation. There were no drug-related serious AEs and no patients with corrected QT (QTc) > 480 ms or QTc change of >60 ms from baseline. The abemaciclib PK profile was characterized by slow absorption and high PK variability after single or repeated doses. Two patients, one with breast cancer and one with neuroendocrine tumor, experienced >30 % decrease in tumor size from baseline. CONCLUSIONS: In Japanese patients with advanced cancer, single-agent abemaciclib has an acceptable safety profile and demonstrates antitumor activity at a dose of 200 mg Q12H. These findings support ongoing development of abemaciclib for diverse populations with advanced cancer.