Directed Differentiation of Murine and Human Small Intestinal Organoids Toward All Mature Lineages.

Intestinal organoids are three-dimensional structures derived from tissue-resident adult stem cells. These organoids recapitulate key aspects of epithelial biology and can be used to study homeostatic turnover of the corresponding tissue. Organoids can be enriched for the various mature lineages which allows studies of the respective differentiation processes and of the diverse cellular functions. Here we describe mechanisms of intestinal fate specification and how these can be exploited to drive mouse and human small intestinal organoids into each of the functionally mature lineages.

Blocking the ZZ domain of sequestosome1/p62 suppresses myeloma growth and osteoclast formation in vitro and induces dramatic bone formation in myeloma-bearing bones in vivo.

We reported that p62 (sequestosome 1) serves as a signaling hub in bone marrow stromal cells (BMSCs) for the formation of signaling complexes, including NFκB, p38MAPK and JNK, that are involved in the increased osteoclastogenesis and multiple myeloma (MM) cell growth induced by BMSCs that are key contributors to multiple myeloma bone disease (MMBD), and demonstrated that the ZZ domain of p62 (p62-ZZ) is required for BMSC enhancement of MMBD. We recently identified a novel p62-ZZ inhibitor, XRK3F2, which inhibits MM cell growth and BMSC growth enhancement of human MM cells. In the current study, we evaluate the relative specificity of XRK3F2 for p62-ZZ, characterize XRK3F2's capacity to inhibit growth of primary MM cells and human MM cell lines, and test the in vivo effects of XRK3F2 in the immunocompetent 5TGM1 MM model. We found that XRK3F2 induces dramatic cortical bone formation that is restricted to MM containing bones and blocked the effects and upregulation of tumor necrosis factor alpha (TNFα), an osteoblast (OB) differentiation inhibitor that is increased in the MM bone marrow microenvironment and utilizes signaling complexes formed on p62-ZZ, in BMSC. Interestingly, XRK3F2 had no effect on non-MM bearing bone. These results demonstrate that targeting p62 in MM models has profound effects on MMBD.

Pharmacotherapy in cancer patients with HIV/AIDS.

Antiretroviral therapy (ART) has significantly reduced morbidity and increased life expectancy of individuals infected with human immunodeficiency virus (HIV). Consequently, non-acquired immunodeficiency syndrome (AIDS)-defining malignancies are increasing in frequency, which necessitates the concurrent use of antineoplastics and ART. Although drug interactions are a major concern when combining these agents, there is currently limited guidance on dose adjustments required to maintain safe and efficacious drug exposure.

A phase 1 trial of imatinib, bevacizumab, and metronomic cyclophosphamide in advanced colorectal cancer.

BACKGROUND: This phase 1 clinical trial was conducted to determine the safety, maximum-tolerated dose (MTD), and pharmacokinetics of imatinib, bevacizumab, and metronomic cyclophosphamide in patients with advanced colorectal cancer (CRC). METHODS: Patients with refractory stage IV CRC were treated with bevacizumab 5 mg kg(-1) i.v. every 2 weeks (fixed dose) plus oral cyclophosphamide q.d. and imatinib q.d. or b.i.d. in 28-day cycles with 3+3 dose escalation. Response was assessed every two cycles. Pharmacokinetics of imatinib and cyclophosphamide and circulating tumour, endothelial, and immune cell subsets were measured. RESULTS: Thirty-five patients were enrolled. Maximum-tolerated doses were cyclophosphamide 50 mg q.d., imatinib 400 mg q.d., and bevacizumab 5 mg kg(-1) i.v. every 2 weeks. Dose-limiting toxicities (DLTs) included nausea/vomiting, neutropaenia, hyponatraemia, fistula, and haematuria. The DLT window required expansion to 42 days (1.5 cycles) to capture delayed toxicities. Imatinib exposure increased insignificantly after adding cyclophosphamide. Seven patients (20%) experienced stable disease for >6 months. Circulating tumour, endothelial, or immune cells were not associated with progression-free survival. CONCLUSION: The combination of metronomic cyclophosphamide, imatinib, and bevacizumab is safe and tolerable without significant drug interactions. A subset of patients experienced prolonged stable disease independent of dose level.

A phase I trial of isolated hepatic perfusion (IHP) using 5-FU and oxaliplatin in patients with unresectable isolated liver metastases from colorectal cancer.

BACKGROUND: Isolated hepatic perfusion (IHP) with melphalan is an established approach for patients with unresectable metastatic liver lesions. This study determined the safety and maximum tolerated dose (MTD) of 5-FU with oxaliplatin via IHP. METHODS: Standard 3 × 3 Phase I design. Subjects with unresectable isolated CRC liver metastases scheduled for HAI pump were eligible. IHP used fixed-dose oxaliplatin with escalating 5-FU doses. Toxicity (CTCAE v 4.0) and response (RECIST), progression-free survival, and overall survival (OS) were assessed. Systemic and IHP plasma PK of 5-FU, anabolites, and platinum were determined. RESULTS: All 12 patients had received ≥ 1 line of pre-IHP chemotherapy. There were 4 grade 3 serious adverse events (33.3 %) and 1 grade 4 event (8.3 %). Also, 2 dose-limiting toxicities occurred at DL2 at 300 mg/m(2), resulting in expansion of DL1 at 200 mg/m(2) 5-FU, the eventual MTD. At 6-month follow-up, 9 patients (82 %) demonstrated partial response, while 2 (18 %) exhibited stable disease. Also, 64 % of patients demonstrated a >50 % decrease in CEA. The 1- and 2-year OS probabilities were 90.9 and 71.6 %, respectively, with median follow-up of 24 months. IHP exposures (AUC0-60 min) were 10.9 ± 4.5 µgPt h/mL, 49.3 ± 30.7 µg h/mL 5-FU (DL1), and 70.5 ± 35.5 µg h/mL 5-FU (DL2). Systemic exposure (AUC0-inf) relative to IHP exposure was negligible for both platinum (1.1 ± 1.5 %) and 5-FU (0.09 ± 0.10 %). CONCLUSIONS: The MTD for IHP was 200 mg/m(2) 5-FU with 40 mg/m(2) oxaliplatin. Systemic exposure to the agents was minimal during IHP. The response and survival observed warrants assessment in a larger phase II trial.

Safety and efficacy of decitabine in combination with temozolomide in metastatic melanoma: a phase I/II study and pharmacokinetic analysis.

BACKGROUND: Temozolomide (TMZ) is widely used for chemotherapy of metastatic melanoma. We hypothesized that epigenetic modulators will reverse chemotherapy resistance, and in this article, we report studies that sought to determine the recommended phase 2 dose (RP2D), safety, and efficacy of decitabine (DAC) combined with TMZ. PATIENTS AND METHODS: In phase I, DAC was given at two dose levels: 0.075 and 0.15 mg/kg intravenously daily × 5 days/week for 2 weeks, TMZ orally 75 mg/m(2) qd for weeks 2-5 of a 6-week cycle. The phase II portion used a two-stage Simon design with a primary end point of objective response rate (ORR). RESULTS: The RP2D is DAC 0.15 mg/kg and TMZ 75 mg/m(2). The phase II portion enrolled 35 patients, 88% had M1c disease; 42% had history of brain metastases. The best responses were 2 complete response (CR), 4 partial response (PR), 14 stable disease (SD), and 13 progressive disease (PD); 18% ORR and 61% clinical benefit rate (CR + PR + SD). The median overall survival (OS) was 12.4 months; the 1-year OS rate was 56%. Grade 3/4 neutropenia was common but lasted >7 days in six patients. CONCLUSIONS: The combination of DAC and TMZ is safe, leads to 18% ORR and 12.4-month median OS, suggesting possible superiority over the historical 1-year OS rate, and warrants further evaluation in a randomized setting.

Phase I study investigating the safety and feasibility of combining imatinib mesylate (Gleevec) with sorafenib in patients with refractory castration-resistant prostate cancer.

BACKGROUND: Determining the maximum tolerated dose (MTD) and the dose-limiting toxicity (DLT) of sorafenib (S) plus imatinib (IM) in castration-resistant prostate cancer (CRPC) patients. METHODS: Refractory CRPC patients were enrolled onto this 3+3 dose escalation designed study. Imatinib pharmacokinetics (PK) were determined on day 15, 4 h post dose with a validated LC-MS assay. RESULTS: Seventeen patients were enrolled; 10 evaluable (6 at 400 mg S qd with 300 mg IM qd (DL0) and 4 at 400 mg S bid with 300 mg IM qd (DL1)); inevaluable patients received <1 cycle. The median age was 73 (57-89); median prostatic serum antigen was 284 ng ml(-1) (11.7-9027). Median number of prior non-hormonal therapies was 3 (1-12). Dose-limiting toxicities were diarrhoea and hand-foot syndrome. Maximum tolerated dose was 400 mg S and 300 mg IM both daily. No biochemical responses were observed. Two patients had stable disease by RECIST. Median time to progression was 2 months (1-5). Median OS was 6 months (1-30+) with 3/17 patients (17%) alive at 21 months median follow-up. Ten patients had PK data suggesting that S reduced IM clearance by 55%, resulting in 77% increased exposure (P=0.005; compared with historical data). CONCLUSION: This is the first report showing that S+IM can be administered in CRPC at a dose of 400 mg S and 300 mg IM, daily.

Role of PARP inhibitors in cancer biology and therapy.

Deeper understanding of DNA repair mechanisms and their potential value as therapeutic targets in oncology heralded the clinical development of poly(ADP-ribose) polymerase (PARP) inhibitors. Although initially developed to exploit synthetic lethality in models of cancer associated with defective DNA repair, our burgeoning knowledge of PARP biology has resulted in these agents being exploited both in cancer with select chemotherapeutic agents and in non-malignant diseases. In this review article, we briefly review the mechanisms of DNA repair and pre-clinical development of PARP inhibitors before discussing the clinical development of the various PARP inhibitors in depth.

Disposition and toxicity of trabectedin (ET-743) in wild-type and mdr1 gene (P-gp) knock-out mice.

Trabectedin is a novel anticancer drug active against soft tissue sarcomas. Trabectedin is a substrate for P-glycoprotein (P-gp), which is encoded by mdr1a/1b in rodents. Plasma and tissue distribution, and excretion of [(14)C]-trabectedin were evaluated in wild-type and mdr1a/1b(-/-) mice. In parallel, we investigated the toxicity profile of trabectedin by serial measurements of blood liver enzymes and general pathology. [(14)C]-trabectedin was extensively distributed into tissues, and rapidly converted into a range of unknown metabolic products. The excretion of radioactivity was similar in both genotypes. The plasma clearance of unchanged trabectedin was not reduced when P-gp was absent, but organs under wild type circumstances protected by P-gp showed increased trabectedin concentrations in mdr1a/1b(-/-) mice. Although hepatic trabectedin concentrations were not increased when P-gp was absent, mdr1a/1b(-/-) mice experienced more severe liver toxicity. P-gp plays a role in the in vivo disposition and toxicology of trabectedin.

Human mass balance study of the novel anticancer agent ixabepilone using accelerator mass spectrometry.

Ixabepilone (BMS-247550) is a semi-synthetic, microtubule stabilizing epothilone B analogue which is more potent than taxanes and has displayed activity in taxane-resistant patients. The human plasma pharmacokinetics of ixabepilone have been described. However, the excretory pathways and contribution of metabolism to ixabepilone elimination have not been determined. To investigate the elimination pathways of ixabepilone we initiated a mass balance study in cancer patients. Due to autoradiolysis, ixabepilone proved to be very unstable when labeled with conventional [14C]-levels (100 microCi in a typical human radio-tracer study). This necessitated the use of much lower levels of [14C]-labeling and an ultra-sensitive detection method, Accelerator Mass Spectrometry (AMS). Eight patients with advanced cancer (3 males, 5 females; median age 54.5 y; performance status 0-2) received an intravenous dose of 70 mg, 80 nCi of [14C]ixabepilone over 3 h. Plasma, urine and faeces were collected up to 7 days after administration and total radioactivity (TRA) was determined using AMS. Ixabepilone in plasma and urine was quantitated using a validated LC-MS/MS method. Mean recovery of ixabepilone-derived radioactivity was 77.3% of dose. Fecal excretion was 52.2% and urinary excretion was 25.1%. Only a minor part of TRA is accounted for by unchanged ixabepilone in both plasma and urine, which indicates that metabolism is a major elimination mechanism for this drug. Future studies should focus on structural elucidation of ixabepilone metabolites and characterization of their activities.

Trabectedin (Yondelis, formerly ET-743), a mass balance study in patients with advanced cancer.

Trabectedin (Yondelis, formerly ET-743) is an anti-cancer drug currently undergoing phase II development. Despite extensive pharmacokinetic studies, the human disposition and excretory pathways of trabectedin remain largely unknown. Our objective was to determine the mass balance of trabectedin in humans. To this aim, we intravenously administered [(14)C]trabectedin to 8 cancer patients, followed by collection of whole blood, urine and faeces samples. A 24-h infusion was administered to 2 patients, whereas the other 6 patients received a 3-h infusion. Levels of total radioactivity and unchanged trabectedin were determined and used for calculation of pharmacokinetic parameters. No schedule dependency of pharmacokinetic parameters was observed apart from C(max). Plasma and whole blood concentrations of [(14)C]trabectedin related radioactivity were comparable. Only 8% of the plasma exposure to [(14)C]trabectedin related compounds is accounted for by trabectedin, indicating the importance of metabolism in trabectedin elimination. Trabectedin displays a large volume of distribution (+/-1700 L), relative to total radioactivity (+/-220 L). [(14)C]trabectedin related radioactivity is mainly excreted in the faeces (mean: 55.5% of the dose). Urinary excretion accounts for 5.9% of the dose on average resulting in a mean overall recovery of 61.4% (3-h administration schedule). The excretion of unchanged trabectedin is very low both in faeces and in urine (< 1% of dose). In conclusion, trabectedin is extensively metabolised and principally excreted in the faeces.

Abutment load transfer by removable partial denture obturator frameworks in different acquired maxillary defects.

STATEMENT OF PROBLEM: Excessive stress on abutment teeth adjacent to a maxillary resection defect during loading of partial denture obturator frameworks may shorten the life of the teeth. PURPOSE: The aim of this study was to photoelastically compare the forces exerted on the supporting structures of abutment teeth in 3 differently sized surgical resections with removable partial denture designs used to restore such maxillectomy defects. MATERIAL AND METHODS: Composite photoelastic models were constructed of a human maxilla that had undergone each of 3 maxillectomies: partial, radical, and radical involving the contralateral premaxilla. The abutment teeth included all remaining anterior teeth, the first premolar, and second molar, except the radical maxillectomy, which included the contralateral premaxilla where all remaining teeth were used as abutment teeth. All abutment teeth were restored with complete metal crowns, and removable partial denture frameworks were fabricated. Loading zones were selected according to the resection, and a 10-lb load was applied at each load point. The resulting stresses were observed and recorded photographically in a circular polariscope. The 2 teeth adjacent to the resection were then splinted, and the loading regimens were repeated. RESULTS: Without splinting, loads closer to the defect produced lingual tipping of the teeth adjacent to the resection and a mesial tipping tendency of the second molar. The tipping effects were greatest in the model with the largest resection. Splinting reduced tipping of the teeth adjacent to the resection and produced more uniform stress around these 2 abutment tooth roots for all of the models. CONCLUSION: The results of this in vitro study suggest that splinting the 2 teeth adjacent to a resection defect improves stress distribution around the roots during loading. This could increase the clinical life of the abutment teeth.

Hepatotoxicity and metabolism of trabectedin: a literature review.

Trabectedin is a promising anticancer drug currently undergoing phase II evaluation. In preclinical studies, trabectedin was found to cause hepatotoxicity and in patients it reversibly increases plasma levels of liver enzymes. On the basis of preclinical work, it was suggested that metabolism of trabectedin contributed to the pharmacological effects of trabectedin, including hepatotoxicity in rats and increases in liver enzymes in humans. Our aim was to review the current literature on the metabolism of trabectedin and its role in the increases in liver enzymes and hepatotoxicity. We conclude that the trabectedin metabolic profile appears to predict the reversible nature of hepatotoxicity. The rat may not be the best species to investigate trabectedin hepatotoxicity because both trabectedin metabolic profile and reversibility of hepatotoxicity differs from humans. Humans and monkeys display a similar metabolic profile of trabectedin and in both species hepatotoxicity is reversible. Trabectedin is a drug with predictable hepatotoxic effects. Monitoring of plasma levels of liver enzymes ensures safe use of trabectedin in the clinic. Future investigations must be aimed at elucidating the mechanism of trabectedin hepatotoxicity.

Hair as a biological specimen for therapeutic drug monitoring.

To study the potential of hair as a biological specimen in therapeutic drug monitoring (TDM), a review of the correlation between hair concentration, blood levels, dose and therapeutic effects of various drugs is presented. The results indicate there is a fair correlation between plasma and hair for several medicines. Moreover, hair samples have the advantage of providing information of more prolonged drug intake, thus allowing a better evaluation of patient compliance. Using segmental hair analysis, only a single sample is necessary. We conclude that hair has potential as a biological specimen in TDM--at least as far as compliance is concerned and possibly as a longer term record of drug concentrations--and that correlation between hair concentration, blood levels and clinical efficacy should be investigated for all drugs where TDM is indicated.

Restoration of partially edentulous patients using customized implant abutments.

The ideal placement of implants is not always possible in partially edentulous patients. The diverse and unique implant positions that occur in clinical practice may be difficult or impossible to restore through the use of conventional abutments. Customized abutments permit the fabrication of aesthetic restorations that correct deficiencies in implant angulation, alignment, and position. These abutments also enhance the soft tissue emergence profile of the restorations and allow the prosthetic margins to be properly positioned in all dimensions. Additional benefits include ease of treatment delivery and comparative expense.

Restoration of irradiated patients using osseointegrated implants: current perspectives.

STATEMENT OF PROBLEM: The success rates of osseointegrated implants used to restore patients who were irradiated for head and neck tumors are influenced by radiation-induced changes in the hard and soft tissues. PURPOSE: This article examined, by review of the literature, current perspectives on the restoration of irradiated patients using osseointegrated implants. RESULTS: In published reports that investigated both intraoral and extraoral applications, irradiation decreased implant success rates and the amount of reduction was dependent on the location within the craniofacial skeleton. The limited number of implants and patients in these studies precludes definitive conclusions regarding the efficacy of placing implants into irradiated tissues. The implants placed into the irradiated anterior mandible have demonstrated an acceptable implant success rate of 94% to 100% with a minimal risk of osteoradionecrosis. The efficacy of implants in the posterior mandible has not been examined. Implant success rates ranged from 69% to 95% in the irradiated maxilla for intraoral applications. Extraoral applications demonstrated excellent implant success rates in the temporal bone (91% to 100%). The rates in the anterior nasal floor have varied from 50% to 100%. The implant success rates in the frontal bone decreased as the length of the studies increased (96% to 33%). The long-term efficacy of implants in the irradiated frontal bone is poor.

Implants in the partially edentulous patient: restorative considerations.

Restorative considerations are critical to the long-term success of fixed implant-supported prostheses, especially in the posterior quadrants of the partially edentulous patient. The parafunctional habit of bruxism must be identified and addressed. The restoration should dictate implant placement. Control of forces directed upon the prosthesis and implants is critical to long-term success. Anatomic limitations to implant placement and surgical procedures to correct these deficiencies must be considered for their impact on the prosthetic restoration. Nonaxial forces or bending moments should be minimized by the use of an adequate number, position and alignment of implants; by control of the occlusion; and by design of the prosthesis. The patient must understand the risks, limitations, costs and time commitments of implant restorations prior to treatment.

An implant-supported fixed-removable prosthesis with a milled tissue bar and Hader clip retention as a restorative option for the edentulous maxilla.

Restoration of the edentulous maxilla with implant retained and supported prosthesis is challenging because of inherent anatomic limitations present after the loss of teeth. A fixed-removable prosthesis is one treatment choice for restoration of the edentulous maxilla with implants. This prosthesis meets the requirements for esthetics, phonetics, comfort, and hygiene, as well as favorable biomechanical stress distribution to the implants. This article presents a procedure for fabrication of a fixed-removable prosthesis with a precision milled bar, Hader clip attachments, and a superstructure prosthesis. The criteria for patient selection and the advantages and disadvantages of this prosthesis are discussed.