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Antibody-dependent cellular cytotoxicity (ADCC) is a key effector mechanism of natural killer (NK) cells that is mediated by therapeutic monoclonal antibodies (mAbs). This process is facilitated by the Fc receptor CD16a on human NK cells. CD16a appears to be the only activating receptor on NK cells that is cleaved by the metalloprotease a disintegrin and metalloproteinase-17 upon stimulation. We previously demonstrated that a point mutation of CD16a prevents this activation-induced surface cleavage. This noncleavable CD16a variant is now further modified to include the high-affinity noncleavable variant of CD16a (hnCD16) and was engineered into human induced pluripotent stem cells (iPSCs) to create a renewable source for human induced pluripotent stem cell-derived NK (hnCD16-iNK) cells. Compared with unmodified iNK cells and peripheral blood-derived NK (PB-NK) cells, hnCD16-iNK cells proved to be highly resistant to activation-induced cleavage of CD16a. We found that hnCD16-iNK cells were functionally mature and exhibited enhanced ADCC against multiple tumor targets. In vivo xenograft studies using a human B-cell lymphoma demonstrated that treatment with hnCD16-iNK cells and anti-CD20 mAb led to significantly improved regression of B-cell lymphoma compared with treatment utilizing anti-CD20 mAb with PB-NK cells or unmodified iNK cells. hnCD16-iNK cells, combined with anti-HER2 mAb, also mediated improved survival in an ovarian cancer xenograft model. Together, these findings show that hnCD16-iNK cells combined with mAbs are highly effective against hematologic malignancies and solid tumors that are typically resistant to NK cell-mediated killing, demonstrating the feasibility of producing a standardized off-the-shelf engineered NK cell therapy with improved ADCC properties to treat malignancies that are otherwise refractory., (© 2020 by The American Society of Hematology.)

The aryl hydrocarbon receptor (AHR) plays an important physiological role in hematopoiesis. AHR is highly expressed in hematopoietic stem and progenitor cells (HSPCs) and inhibition of AHR results in a marked expansion of human umbilical cord blood-derived HSPCs following cytokine stimulation. It is unknown whether AHR also contributes earlier in human hematopoietic development. To model hematopoiesis, human embryonic stem cells (hESCs) were allowed to differentiate in defined conditions in the presence of the AHR antagonist StemReginin-1 (SR-1) or the AHR agonist 2,3,7,8-tetrachlorodibenzo- p- dioxin (TCDD). We demonstrate a significant increase in CD34 + CD31 + hematoendothelial cells in SR-1-treated hESCs, as well as a twofold expansion of CD34 + CD45 + hematopoietic progenitor cells. Hematopoietic progenitor cells were also significantly increased by SR-1 as quantified by standard hematopoietic colony-forming assays. Using a clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9)-engineered hESC- RUNX1c -tdTomato reporter cell line with AHR deletion, we further demonstrate a marked enhancement of hematopoietic differentiation relative to wild-type hESCs. We also evaluated whether AHR antagonism could promote innate lymphoid cell differentiation from hESCs. SR-1 increased conventional natural killer (cNK) cell differentiation, whereas TCDD treatment blocked cNK development and supported group 3 innate lymphoid cell (ILC3) differentiation. Collectively, these results demonstrate that AHR regulates early human hematolymphoid cell development and may be targeted to enhance production of specific cell populations derived from human pluripotent stem cells., (© 2017 by The American Society of Hematology.)

Background Aims: There is an urgent need for novel therapeutic strategies for relapsed ovarian cancer. Dramatic clinical anti-tumor effects have been observed with interleukin (IL)-2 activated natural killer (NK) cells; however, intravenous delivery of NK cells in patients with ovarian cancer has not been successful in ameliorating disease. We investigated in vivo engraftment of intraperitoneally (IP) delivered NK cells in an ovarian cancer xenograft model to determine if delivery mode can affect tumor cell killing and circumvent lack of NK cell expansion., Methods: An ovarian cancer xenograft mouse model was established to evaluate efficacy of IP-delivered NK cells. Tumor burden was monitored by bioluminescent imaging of luciferase-expressing ovarian cancer cells. NK cell persistence, tumor burden and NK cell trafficking were evaluated. Transplanted NK cells were evaluated by flow cytometry and cytotoxicity assays., Results: IP delivery of human NK cells plus cytokines led to high levels of circulating NK and was effective in clearing intraperitoneal ovarian cancer burden in xenografted mice. NK cells remained within the peritoneal cavity 54 days after injection and had markers of maturation. Additionally, surviving NK cells were able to kill ovarian cancer cells at a rate similar to pre-infusion levels, supporting that in vivo functionality of human NK cells can be maintained after IP infusion., Conclusions: IP delivery of NK cells leads to stable engraftment and antitumor response in an ovarian cancer xenograft model. These data support further pre-clinical and clinical evaluation of IP delivery of allogeneic NK cells in ovarian cancer., (Copyright © 2013 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.)

Background: Weekly gemcitabine with GC every 3-4 weeks is considered conventional first-line chemotherapy for advanced urothelial carcinoma (UC). Weekly split-dose cisplatin with wGC might be less toxic and have similar activity, but has not been compared with GC. We pooled published phase II trials of GC and wGC to compare efficacy and safety., Patients and Methods: Two trials of wGC and 3 trials of GC were identified. Because the data were not derived from randomized trials, GC and wGC were not formally compared, and exact 95% quasi-binomial confidence intervals (CI), for response rates and grade ≥ 3 toxicities were calculated., Results: The 95% CI overlapped, suggesting agreement between wGC and GC. No clear difference in response rates and grade ≥ 3 toxicities between GC and wGC were observed., Conclusion: Gemcitabine combined with day 1 cisplatin and wGC yielded similar responses and grade ≥ 3 toxicities in advanced UC in a hypothesis-generating pooled analysis of phase II trials. Considering the probable lower nephrotoxicity of fractionated cisplatin, prospective evaluation of wGC might be warranted across cisplatin-eligible and -ineligible patients to develop a single chemotherapy template for the development of combinations with biological agents in a broad population of patients., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Advancements in human pluripotent stem cell (hPSC) research have potential to revolutionize therapeutic transplantation. It has been demonstrated that transcription factors may play key roles in regulating maintenance, expansion, and differentiation of hPSCs. In addition to its regulatory functions in hematopoiesis and blood-related disorders, the transcription factor RUNX1 is also required for the formation of definitive blood stem cells. In this study, we demonstrated that expression of endogenous RUNX1a, an isoform of RUNX1, parallels with lineage commitment and hematopoietic emergence from hPSCs, including both human embryonic stem cells and inducible pluripotent stem cells. In a defined hematopoietic differentiation system, ectopic expression of RUNX1a facilitates emergence of hematopoietic progenitor cells (HPCs) and positively regulates expression of mesoderm and hematopoietic differentiation-related factors, including Brachyury, KDR, SCL, GATA2, and PU.1. HPCs derived from RUNX1a hPSCs show enhanced expansion ability, and the ex vivo-expanded cells are capable of differentiating into multiple lineages. Expression of RUNX1a in embryoid bodies (EBs) promotes definitive hematopoiesis that generates erythrocytes with β-globin production. Moreover, HPCs generated from RUNX1a EBs possess ≥9-week repopulation ability and show multilineage hematopoietic reconstitution in vivo. Together, our results suggest that RUNX1a facilitates the process of producing therapeutic HPCs from hPSCs.

Human pluripotent stem cells represent an accessible cell source for novel cell-based clinical research and therapies. With the realization of induced pluripotent stem cells (iPSCs), it is possible to produce almost any desired cell type from any patient's cells. Current developments in gene modification methods have opened the possibility for creating genetically corrected human iPSCs for certain genetic diseases that could be used later in autologous transplantation. Promising preclinical studies have demonstrated correction of disease-causing mutations in a number of hematological, neuronal, and muscular disorders. This review aims to summarize these recent advances with a focus on iPSC generation techniques, as well as gene modification methods. We will then further discuss some of the main obstacles remaining to be overcome before successful application of human pluripotent stem cell-based therapy arrives in the clinic and what the future of stem cell research may look like., (Copyright © 2013 Mosby, Inc. All rights reserved.)

Studies by Amabile et al reported in this issue of Blood use a novel strategy of teratoma formation from human induced pluripotent stem cells (iPSCs) to isolate hematopoietic stem/progenitor cells (HSPCs) capable of in vivo engraftment and producing functional lymphocytes.(1)

Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) provide an accessible, genetically tractable, and homogenous starting cell population to efficiently study human blood cell development. These cell populations provide platforms to develop new cell-based therapies to treat both malignant and nonmalignant hematological diseases. Our group previously demonstrated the ability of hESC-derived hematopoietic precursors to produce functional natural killer (NK) cells as well as an explanation of the underlying mechanism responsible for the inefficient development of T and B cells from hESCs. hESCs and iPSCs, which can be engineered reliably in vitro, provide an important new model system to study human lymphocyte development and produce enhanced cell-based therapies with the potential to serve as a "universal" source of antitumor lymphocytes. This review will focus on the application of hESC-derived NK cells with currently used and novel therapeutics for clinical trials, barriers to translation, and future applications through genetic engineering approaches., (Copyright 2010 Mosby, Inc. All rights reserved.)

Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) provide remarkable cellular platforms to better understand human hematopoiesis and to develop clinically applicable hematopoietic cell-based therapies. Over the past decade, hESCs have been used to characterize molecular and cellular mechanisms underpinning the differentiation of hematopoietic progenitors and mature, functional hematopoietic cells. These advances are now poised to lead to clinical translation of hESC- and iPSC-derived hematopoietic cells for novel therapies in the next few years. On the basis of areas of recent success, initial clinical use of hematopoietic cells derived from human pluripotent stem cells will probably be in the areas of transfusion therapies (erythrocytes and platelets) and immune therapies (natural killer cells). In contrast, efficient development and isolation of hematopoietic stem cells capable of long-term, multilineage engraftment still remains a significant challenge. Technical, safety, and regulatory concerns related to clinical applications of human PSCs must be appropriately addressed. However, proper consideration of these issues should facilitate and not inhibit clinical translation of new therapies. This review outlines the current status of hematopoietic cell development and what obstacles must be surmounted to bring hematopoietic cell therapies from human PSCs from "bench to bedside."

Bladder cancer is a heterogeneous disease, with 70% of patients presenting with superficial tumours, which tend to recur but are generally not life threatening, and 30% presenting as muscle-invasive disease associated with a high risk of death from distant metastases. The main presenting symptom of all bladder cancers is painless haematuria, and the diagnosis is established by urinary cytology and transurethral tumour resection. Intravesical treatment is used for carcinoma in situ and other high grade non-muscle-invasive tumours. The standard of care for muscle-invasive disease is radical cystoprostatectomy, and several types of urinary diversions are offered to patients, with quality of life as an important consideration. Bladder preservation with transurethral tumour resection, radiation, and chemotherapy can in some cases be equally curative. Several chemotherapeutic agents have proven to be useful as neoadjuvant or adjuvant treatment and in patients with metastatic disease. We discuss bladder preserving approaches, combination chemotherapy including new agents, targeted therapies, and advances in molecular biology.

Twenty-six patients with prostate cancer status post-radical prostatectomy who were candidates for salvage radiation therapy (SRT) underwent lymphotropic nanoparticle enhanced MRI (LNMRI) using superparamagnetic nanoparticle ferumoxtran-10. LNMRI was well tolerated, with only two adverse events, both Grade 2. Six (23%) of the 26 patients, previously believed to be node negative, tested lymph node positive by LNMRI. A total of nine positive lymph nodes were identified in these six patients, none of which were enlarged based on size criteria.

Natural killer (NK) cells serve as important effectors for antitumor immunity, and CD56+CD45+ NK cells can be routinely derived from human embryonic stem cells (hESCs). However, little is know about the ability of hESC-derived NK cells to mediate an effective in vivo antitumor response. Using bioluminescent imaging, we now demonstrate that H9 line hESC-derived NK cells mediate effective clearance of human tumor cells in vivo. In addition to increased in vitro killing of diverse tumor targets, the in vivo tumor clearance by H9 hESC-derived NK cells was more effective compared with NK cells derived from umbilical cord blood (UCB). Phenotypic analysis demonstrates the hESC-derived NK cells are uniformly CD94+CD117(low/-), an NK-cell population characterized by potent cytolytic activity and thus more competent to mediate tumor clearance. These studies demonstrate that hESCs provide an important model to study human lymphocyte development and may serve as a novel source for antitumor immunotherapy.

Background: This study explored the efficacy and tolerability of sunitinib, an inhibitor of tyrosine kinase receptors, in men with castration-resistant prostate cancer (CRPC)., Methods: Men with no prior chemotherapy (group A) and men with docetaxel (Taxotere)-resistant prostate cancer (group B) were treated with sunitinib. The primary end point was confirmed 50% prostate-specific antigen (PSA) decline. Secondary end points included objective response rate and safety. Serum-soluble biomarkers were measured., Results: Seventeen men were enrolled in each group. One confirmed PSA response was observed in each group, and an additional eight men and seven men had stable PSA at week 12 in groups A and B, respectively. Improvements in imaging were observed in the absence of post-treatment PSA declines. Common adverse effects included fatigue, nausea, diarrhea, myelosuppression and transaminase elevation. Significant changes following sunitinib treatment were observed in serum-soluble biomarkers including soluble vascular endothelial growth factor receptor-2, platelet-derived growth factor aa, placental growth factor and leptin., Conclusions: Sunitinib monotherapy resulted in few confirmed 50% post-treatment declines in PSA in men with CRPC. Serum markers of angiogenesis confirmed on-target effects of sunitinib. Assessments of radiographic disease status were often discordant with changes in PSA, indicating that alternate end points are important in future trials.

Hematopoietic progenitor cells derived from human embryonic stem cells (hESCs) develop into diverse mature hematopoietic lineages, including lymphocytes. Whereas functional natural killer (NK) cells can be efficiently generated in vitro from hESC-derived CD34(+) cells, studies of T- and B-cell development from hESCs have been much more limited. Here, we demonstrate that despite expressing functional Notch-1, CD34(+) cells from hESCs did not derive T cells when cocultured with OP9 cells expressing Delta-like 1, or in fetal thymus organ culture. hESC-derived CD34(+) cells also did not produce B cells in vitro. In contrast, CD34(+) cells isolated from UCB routinely generated T and B cells when cultured in the same conditions. Notably, both undifferentiated hESCs, and sorted hESC-derived populations with hematopoietic developmental potential exhibited constitutive expression of ID family genes and of transcriptional targets of stem cell factor-induced signaling. These pathways both inhibit T-cell development and promote NK-cell development. Together, these results demonstrate fundamental differences between hESC-derived hematopoietic progenitors and analogous primary human cells. Therefore, hESCs can be more readily supported to differentiate into certain cell types than others, findings that have important implications for derivation of defined lineage-committed populations from hESCs.

When mollusc shells are analysed conventionally for amino acid geochronology, the entire population of amino acids is included, both inter- and intra-crystalline. This study investigates the utility of removing the amino acids that are most susceptible to environmental effects by isolating the fraction of amino acids encapsulated within mineral crystals of mollusc shells (intra-crystalline fraction). Bleaching, heating and leaching (diffusive loss) experiments were undertaken on modern and fossil Corbicula fluminalis, Margaritifera falcata, Bithynia tentaculata and Valvata piscinalis shells. Exposure of powdered mollusc shells to concentrated NaOCl for 48 h effectively reduced the amino acid content of the four taxa to a residual level, assumed to represent the intra-crystalline fraction. When heated in water at 140 degrees C for 24 h, only 1% of amino acids were leached from the intra-crystalline fraction of modern shells compared with 40% from whole shell. Free amino acids were more effectively retained in the intra-crystalline fraction, comprising 55% (compared with 18%) of the whole shell after 24 h at 140 degrees C. For fossil gastropods, the inter-shell variability in D/L values for the intra-crystalline fraction of a single-age population was reduced by 50% compared with conventionally analysed shells. In contrast, analysis of the intra-crystalline fraction of C. fluminalis does not appear to improve the results for this taxon, possibly due to variability in shell ultrastructure. Nonetheless, the intra-crystalline fraction in gastropods approximates a closed system of amino acids and appears to provide a superior subset of amino acids for geochronological applications.

Human embryonic stem cells (hESCs) provide an important means to effectively study soluble and cell-bound mediators that regulate development of early blood and endothelial cells in a human model system. Here, several complementary methods are used to demonstrate canonical Wnt signaling is important for development of hESC-derived cells with both hematopoietic and endothelial potential. Analyses using both standard flow cy-tometry, as well the more detailed high-throughput image scanning flow cytometry, characterizes sequential development of distinct early developing CD34(bright)CD31(+)Flk1(+) cells and a later population of CD34(dim)CD45(+) cells. While the CD34(bright)CD31(+)Flk1(+) have a more complex morphology and can develop into both endothelial cells and hematopoietic cells, the CD34(dim)CD45(+) cells have a simpler morphology and give rise to only hematopoietic cells. Treatment with dickkopf1 to inhibit Wnt signaling results in a dramatic decrease in development of cells with hematoendothelial potential. In addition, activation of the canonical Wnt signaling pathway in hESCs by coculture with stromal cells that express Wnt1, but not use of noncanonical Wnt5-expressing stromal cells, results in an accelerated differentiation and higher percentage of CD34(bright)CD31(+)Flk1(+) cells at earlier stages of differentiation. These studies effectively demonstrate the importance of canonical Wnt signaling to mediate development of early hematoendothelial progenitors during human development.

Donor cell leukemia (DCL) is a rare complication of hematopoietic cell transplantation (HCT). Its incidence has been reported between 0.12% and 5%, although the majority of cases are anecdotal. The mechanisms of leukemogenesis in DCL may be distinct from other types of leukemia. Possible causes of DCL include oncogenic alteration or premature aging of transplanted donor cells in an immunosuppressed person. Although many studies have recently better characterized leukemic stem cells, it is important to also consider that both intrinsic cell factors and external signals from the hematopoietic microenvironment govern the developmental fate of hematopoietic stem cells (HSCs). Therefore, in cases of DCL, alteration of the microenvironment after HCT may increase the likelihood that some progeny of normal HSCs become leukemic. This complex intercommunication between cells, growth factors, and cytokines in the hematopoietic microenvironment are critical to balance HSC self-renewal, proliferation, and differentiation. However, this homeostasis is likely perturbed in the development of DCL, allowing unique insight into the stimuli that regulate normal and potentially abnormal hematopoietic development. In this article, we discuss the possible pathogenesis of DCL, its association with stem cells, and its likely dependence on a less-supportive stem cell niche.

Radiation therapy has a multifaceted role in the treatment of muscle-invasive bladder cancer, from being a component of bladder sparing regimens to adjuvant therapy for patients after partial cystectomy, to palliative treatment in patients with metastatic disease. Here, we review the techniques currently used and the settings in which these techniques are applied. Advances in imaging and radiation delivery have allowed for definition of more precise treatment volumes, permitting the delivery of higher tumor doses and lesser doses to critical targets. Better tumor control, fewer therapeutic complications, and better quality of life outcomes are anticipated. In the United States, the most rapidly growing use of radiation in the treatment of bladder cancer is as a component of selective bladder conservation. It uses trimodality therapy, consisting of a maximal transurethral resection followed by concurrent chemotherapy and radiation. Careful cystoscopic surveillance by an experienced urologist ensures a prompt cystectomy at the fist sign of treatment failure. The majority of patients retain a well-functioning bladder with no survival decrement. Radiation therapy is also used as adjuvant therapy after partial cystectomy in select patients. In this setting, it decreases the risk of local or incisional recurrence. It is also used in patients with pelvic recurrences after cystectomy, often combined with concurrent chemotherapy. Radiation is a very effective palliative agent for patients with locally advanced or metastatic disease. It can palliate bleeding and pain for patients with local progression or alleviate pain from bony metastases.

Seventy to eighty percent of patients with newly-diagnosed bladder cancer will present with superficial tumors (Ta, Tis or T(1)). There is, however, a continuum between superficial and muscle-invasive cancer, with the advanced cases usually associated with less-differentiated histology and aneuploidy. Common sites of metastasis include regional lymph nodes, bone, lung, skin and liver. From the low cure rates achieved with radical cystectomy, there is strong evidence that bladder cancer, from the outset, is a systemic disease. The limitations of local treatment are well-documented: a local control rate of 30% with radiation treatment, and 50-70% with radical cystectomy; and no improvement in surgical cure was seen with the use of preoperative radiation. Over the past 30 years, since the initial reports of the effectiveness of cisplatin in the treatment of advanced bladder cancer, there has been a steady flow of chemotherapeutic agents, singly and in combination, shown to be effective in the treatment of this tumor. While response rates and CR rates have increased with the use of combination chemotherapy, this has not translated into survival in advanced disease of greater than 16 months. While the search for more effective agents and combinations continues, attention has also been given to the roles of neoadjuvant and adjuvant chemotherapy in an effort to improve the cure rate achieved with surgery alone. Although radical cystectomy, with continent diversion or neobladder construction in selected cases remains the standard of care in the United States for patients with muscle-invasive bladder cancer, several groups have explored therapeutic strategies that aim at bladder preservation. Early approaches with the goal of bladder preservation consisted of radiation treatment as monotherapy (largely abandoned) or aggressive TURBT for smaller tumors. Over the past 20 years, the Massachusetts General Hospital (MGH) and the Radiation Therapy Oncology Group (RTOG) have studied patients with muscle-invading bladder cancer utilizing tri-modality treatment: a visibly complete transurethral resection followed by radiation with concurrent radiosensitizing chemotherapy and, subsequently, adjuvant chemotherapy. Thus, chemotherapy has been used in two phases of treatment (1) as radiosensitizers, given concurrently with radiation treatment and (2) as adjuvant treatment, recognizing that survival will only be improved by the successful treatment of micrometastases. Based on preliminary information from reports of the effectiveness of gemcitabine/cisplatin in advanced disease, that combination was chosen as the adjuvant regimen in one of our earlier protocols, recently completed and reported. Our current protocol utilizes the Bellmunt regimen as our adjuvant program with the highest RR in advanced disease. This study is ongoing, with early reports of tolerance of the three-drug regimen encouraging. The treatment options for muscularis propria-invasive bladder tumors can broadly be divided into those that spare the bladder and those that involve removing it. In the United States, radical cystectomy with pelvic lymph node dissection is the standard method used to treat patients with this tumor.

The human/sheep xenograft model has proven valuable in assessing the in vivo hematopoietic activity of stem cells from a variety of fetal and postnatal human sources. CD34+/lineage- or CD34+/CD38- cells isolated from human embryonic stem cells (hESCs) differentiated on S17 feeder layer were transplanted by intraperitoneal injections into fetal sheep. Chimerism in primary transplants was established with polymerase chain reaction (PCR) and flow cytometry of bone marrow and peripheral blood samples. Whole bone marrow cells harvested from a primary recipient were transplanted into a secondary recipient. Chimerism was established as described before. This animal was stimulated with human GM-CSF, and an increase in human hematopoietic activity was noted by flow cytometry. Bone marrow aspirations cultured in methylcellulose generated colonies identified by PCR to be of human origin. We therefore conclude that hESCs are capable of generating hematopoietic cells that engraft primary recipients. These cells also fulfill the criteria for long-term engrafting hematopoietic stem cells as demonstrated by engraftment and differentiation in the secondary recipient.

Embryonic stem cells (ESCs) and adult stem cells both provide important resources to define the mechanisms of hematopoietic cell development. To date, studies that utilize hematopoietic stem cells (HSCs) isolated from sites such as bone marrow or umbilical cord blood have been the primary means to identify molecular and phenotypic characteristics of blood cell populations able to mediate long-term hematopoietic engraftment. Although these HSCs are very useful clinically, they are difficult to expand in culture. Now, basic research on human ESCs provides opportunities for novel investigations into the mechanisms of HSC self-renewal. Eventually, the long history of basic and clinical research with adult hematopoietic cell transplantation could translate to establish human ESCs as a suitable alternative starting cell source for clinical hematopoietic reconstitution.

Background: Effective care of each patient throughout a hospital admission involves executing a specific set of tasks to produce a favorable outcome within an appropriate time frame. The ProjectRounds methodology, which can be implemented using widely available software, incorporates the principles of project management in planning and control hospital inpatient care. It consists of four stages--clinical assessment, planning, scheduling, and tracking. OVERVIEW OF PROJECTROUNDS AND EXAMPLE: As an example, a 68-year-old-man is admitted with pneumonia. In clinical assessment, the admitting physician uses an assessment tool that prompts her to list all the patient's clinical issues, define the conditions that need to be met to discharge the patient, highlight special problems, and list any consultations, diagnostic tests, and procedures that are planned. In planning, the work breakdown structure--a tabulation of all the tasks in the "project" (the admission)--is created. In scheduling, a project schedule is generated, and in tracking, the clinical team evaluates and monitors the project's course. During interdisciplinary clinical rounds, the progress of the patient's hospital care can be tracked and quantified by employing the percent complete method. Tracking can be used as a "dashboard," providing a concise summary of the care that needs to be and has been rendered to the patient., Summary and Next Steps: Applying the tenets of project management can optimize the process of providing health care to hospital inpatients.

The purpose of the study was to evaluate response and survival in patients with metastatic urothelial cancer treated with combination gemcitabine and paclitaxel administered on a biweekly schedule at doses of 3000 mg/m2 and 150 mg/m2, respectively. Patients with adequate organ function and performance status were accrued through 7 institutions, stratified by prior therapy status, and treated as noted. Response was evaluated by 1979 bi-dimensional World Health Organization (WHO) criteria. Of 55 eligible patients, 17 had a partial and 5 had a complete response rate for an overall response rate of 40% (27-54%). One complete response and one partial response were observed in the 6 previously treated patients. Overall median survival was 11.8 months (11.9 months in the chemonaive cohort). Grade 3 or 4 myelosuppression occurred in 56%, but only 4 serious infections were observed. We conclude that because of a lower than expected complete response rate, even when corrected for prognostic groupings, this regimen is not recommended for routine use in patients with metastatic urothelial cancer. Insufficient patients with poor renal function or prior therapy were accrued to reach conclusions regarding its utility in these subgroups.

Background: Medical or surgical castration is effective in advanced prostate cancer but with profound side-effects, particularly on sexual function. Effective, less toxic therapies are needed. This study examined whether the addition of finasteride to high-dose bicalutamide enhanced disease control, as measured by additional decreases in serum prostate-specific antigen (PSA)., Patients and Methods: Forty-one patients with advanced prostate cancer received bicalutamide (150 mg/day). Finasteride (5 mg/day) was added at first PSA nadir. Serum PSA was measured every 2 weeks until disease progression. Questionnaires were administered to assess sexual function., Results: Median follow-up is 3.9 years. At the first PSA nadir, median decrease in PSA from baseline was 96.5%. Thirty of 41 patients (73%) achieved a second PSA nadir and median decrease of 98.5% from baseline. Median time to each nadir was 3.7 and 5.8 weeks, respectively. Median time to treatment failure was 21.3 months. Toxicities were minor, including gynecomastia. Seventeen of 29 (59%) and 12 of 24 (50%) men had normal sex drive at baseline and at second PSA nadir, respectively. One-third of men had spontaneous erection at both time points., Conclusion: Finasteride provides additional intracellular androgen blockade when added to bicalutamide. Duration of control is comparable to castration, with preserved sexual function in some patients.

We have used rhesus monkey embryonic stem (ES) cells to study endothelial cell development. Rhesus ES cells (R366.4 cell line) exposed to medium containing vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), insulin-like growth factor (IGF), and epidermal growth factor (EGF) assumed a relatively uniform endothelial cell morphology and could be propagated and expanded with a consistent phenotype and normal karyotype. When placed in Matrigel, these rhesus ES cell-derived endothelial cells (RESDECs) formed capillary-like structures characteristic of endothelial cells. Immunohistochemical and flow cytometric analysis of RESDECs showed that they take up acetylated low-density lipoprotein (LDL), express CD146, von Willebrand factor, and the integrin alpha v beta 3, and bind the lectin ulex europaeus agglutinin-1. These cells also express the VEGF receptor Flk-1 and secrete VEGF. When introduced in a Matrigel plug implanted subcutaneously in mice, RESDECs formed intact vessels and recruited new endothelial cell growth. In vivo function was demonstrated by coinjection of RESDECs with murine tumor cells subcutaneously into immunocompromised adult mice. RESDECs injected alone did not form measurable tumors. Tumor cells grew more rapidly and had increased vascularization when coinjected with the RESDECs. Immunohistochemical staining demonstrated that the RESDECs participated in forming the tumor neovasculature. RESDECs provide a novel means to examine the mechanisms of endothelial cell development, and may open up new therapeutic strategies.

The purpose of these studies was to define the effect of oxidant stress on intracellular pH in renal tubular epithelial cells. Cell pH was therefore quantitated from the fluorescence ratio of 2',7'-bis-(2-carboxyethyl)-5(6)-carboxyfluorescein in the BSC-1 and opossum kidney (OK) established cell lines, both of which are known to be susceptible to oxidant injury. Exposure to 1 mM H2O2 acidified cytosolic pH in both cell types, and this acidification reversed on withdrawal of the H2O2. Half-maximal acidification was seen at 10 microM H2O2. Effects on Na/H antiport were first measured as the recovery from an NH4Cl-pulse acid load: 1 mM H2O2 inhibited Na/H antiport-mediated pH recovery by approximately 40% in both cell types. H2O2 substantially increased the rate of passive Na-independent H+ flux in the OK cell but under the conditions used did not effect this flux in the BSC-1 cell. When both the Na/H antiport and Na-independent transport systems were blocked by perfusion with a 0-Na depolarizing buffer, the H2O2-induced acidification was totally abolished. Measured from the rate of H+ appearance in the medium, H2O2 decreased the rates of endogenous H+ production in both cell types. As measured from the fluorescence of endocytosed fluorescein isothiocyanate-dextran, H2O2 alkalinized lysosomes/late-endosomes, but at a rate much slower than the rate of cytosolic acidification. The results indicate that H2O2 acidifies cell pH in these cell lines by inhibiting Na/H antiport, by accelerating the passive influx of H+, and to some extent by releasing H+ from subcellular compartments. The acidification may have important effects in modulating the toxicity of H2O2.

Monocyte/macrophages are important components of cell-mediated immune responses in presentation of antigen, as regulators of lymphocyte function, and as sources of cytokines that modulate functions of cells other than those of the immune system. Their role in the pathogenesis of alopecia areata (AA) and universalis (AU) has not been explored. This study is an investigation of the function of peripheral blood monocytes from normal subjects and patients with AA, AU, and alopecia totalis (AT), with respect to the principal macrophage-derived angiogenic factor, tumor necrosis factor alpha (TNF alpha). Because neovascularization is a necessary component in the anagen phase of hair growth and may play a role in the pathology of these disorders, we asked whether monocyte/macrophage angiogenic activity was compromised in these alopecias. Purified preparations of monocytes were activated in culture. Conditioned media were assessed for angiogenic activity on the chick chorioallantoic membrane and for concentration of TNF alpha by enzyme-linked immunosorbent assay (ELISA). Both angiogenic and the TNF concentration were significantly diminished in conditioned media from AU monocytes when compared to those from normal subjects and patients with AA. These results show that the function of AU monocytes may be abnormal and that the abnormality may distinguish AU from AA. Defective monocyte/macrophage function could also play a pathogenic role via effects on neovascularization and/or modulation of the immune response.

The effect of human immunodeficiency virus (HIV) recombinant envelope glycoprotein 120 (rgp 120) on the functions of peripheral blood mononuclear cells (PBMC) in vitro was investigated. The results demonstrate that rgp 120 used at concentrations less than 1 microgram/ml has no significant effects on PBMC function in vitro. However, the addition of 1-20 micrograms/ml of rgp 120 significantly inhibits the tetanus toxoid-induced PBMC proliferative response in a dose-related manner as determined by [3H]thymidine incorporation. The data also show that rgp 120 (5 micrograms/ml) causes up to 70% reduction in the number of immunoglobulin G-secreting cells in pokeweed mitogen-stimulated PBMC cultures. Further, rgp 120 can selectively interact with the CD4a epitope of the CD4 helper cell membrane receptor. These results indicate that microgram per milliliter levels of rgp 120 can depress certain immune functions in vitro. The significance of these findings to the pathogenesis of immunodeficiency in HIV infection remains to be determined.