Quantifying nutrient throughput and DOM production by algae in continuous culture.

Freshwater and marine algae can balance nutrient demand and availability by regulating uptake, accumulation and exudation. To obtain insight into these processes under nitrogen (N) and phosphorus (P) limitation, we reanalyze published data from continuous cultures of the chlorophyte Selenastrum minutum. Based on mass budgets, we argue that much of the non-limiting N and P had passed through the organisms and was present as dissolved organic phosphorus or nitrogen (DOP or DON). We construct a model that describes the production of biomass and dissolved organic matter (DOM) as a function of the growth rate. A fit of this model against the chemostat data suggests a high turnover of the non-limiting N and P: at the highest growth rates, N and P atoms spent on average only about 3 h inside an organism, before they were exuded as DON and DOP, respectively. This DOM exudation can explain the observed trends in the algal stoichiometric ratios as a function of the dilution rate. We discuss independent evidence from isotope experiments for this apparently wasteful behavior and we suggest experiments to quantify and characterize DON and DOP exudation further.

Treatment of oropharyngeal squamous cell carcinoma with IMRT: patterns of failure after concurrent chemoradiotherapy and sequential therapy.

BACKGROUND: The optimal management of oropharyngeal squamous cell carcinoma (OPSCC) is controversial. Modern radiotherapy typically employs intensity-modulated radiation therapy (IMRT), and herein, we report the Dana-Farber Cancer Institute (DFCI) experience with IMRT-based treatment of OPSCC. DESIGN: Retrospective study of all patients treated at DFCI for OPSCC with definitive or adjuvant IMRT between 8/04 and 8/09. The primary end point was overall survival (OS); secondary end points were locoregional control (LRC) and freedom from distant metastases (FFDM). Propensity score matching was used to create concurrent chemoradiotherapy (CCRT) and sequential therapy (ST) cohorts equally balanced for patient and disease characteristics. RESULTS: One hundred and sixty-three patients were included with 75% presenting with stage IV disease. Fifty-six patients (34%) were treated with ST. The three-year actuarial OS, LRC, and FFDM rates for the entire cohort/ST subset were 86%/89%, 86%/87%, and 88%/93%, respectively. There were no differences in OS, LRC, or FFDM between CCRT and ST in the propensity-matched cohort. CONCLUSIONS: IMRT was associated with excellent OS, LRC, and FFDM. Although the results following ST were superb, there was no obvious benefit to ST after adjustment for selection bias. We recommend that ST be reserved for medically fit patients with a high risk of distant metastases.

TU-E-BRB-02: A Decision Support Tool for SBRT Planning Using a Searchable DVH Database.

PURPOSE: Develop a decision support tool that aids dosimetrists, physicians, and physicists in assessing and improving plan quality through comparison to plans previously used in similar clinical situations. METHODS: Software was developed to capture and store DVHs and other clinically relevant treatment plan characteristics in a database. In addition to the plan DVH, the database contains a total of 24 plan characteristics including fractionation, prescribed dose, treatment volume, prior surgery, tumor position, and smoking history. DVH and other plan data was captured from the treatment planning system via exported dicom RT files. Structures in the plan were automatically matched by name to a list of standard structures using a system of regular expressions. Additional fields were entered manually using a simple java interface. As a support tool, a plan under development can be quickly compared to similar plans in the database based on selected plan characteristics. A plot displaying the current and historical DVHs provides an easy visual comparison. Our interface also provides statistics for comparison for each dose/volume level such as average, minimum, maximum and standard deviation. RESULTS: DVHs from 111 lung SBRT plans treated from 2009-2011 were imported in accordance with an approved IRB protocol. As an example of data comparisons that can be easily performed to guide plan evaluation, we examined plans prescribing 5400cGy in 3 fractions and found that tumors >7.5cc (n=34) had an average PTV coverage of 94.2% (range: 73.5-95.0%), and tumors =7.5cc (n=35) had an average PTV coverage of 94.9% (range: 81.6-99.6%). CONCLUSION: A searchable DVH database was constructed to provide planners, physicists, and physicians with a straightforward means of comparing plans against historic distributions of DVHs. In the future, outcome data will be included in the database to strengthen its functionality as a decision support and research tool.

SU-E-T-401: Evaluation of Initial Setup Accuracy and Intra-Fraction Motion for Spine SBRT Using Stereotactic Body Frames.

PURPOSE: Quantify initial setup accuracy and intra-fraction motion using stereotactic body frames (SBF) for spine SBRT. METHODS: 10 patients (11 sites, 31 fractions) treated with spine SBRT using SBF immobilization were evaluated for initial setup accuracy and intra-fraction motion. Either the commercial Elekta SBF or an in-house developed SBF (BHS-SBF) were used. The BHS-SBF uses the same setup/immobilization principle as the Elekta but with increased interior space and couch indexing. Both frames include sidewalls to conform the vac-loc rigidly to the patient's sides. All patients were setup using the Brainlab ExacTrac system which includes IR and stereoscopic kV x-ray based positioning. Patients were initially positioned in the frame using skin tattoos then shifted to treatment isocenter based on IR markers affixed to the frame with known geometry relative to isocenter. kV imaging was acquired and automatic 6-D bony fusion performed. Resulting translations and rotations give the initial setup accuracy. Calculated shifts and rotations were performed using a robotic couch and verification imaging acquired. The imaging/fusion process was repeated multiple times during treatment providing intra-fraction motion data. RESULTS: Mean initial setup error in the VRT, LNG and LAT directions was 0.1+/-3.0 mm (0.1+/-0.6 deg), 0.5+/-5.2 mm (0.1+/-1.1 deg) and -0.3+/- 3.7 mm (0.4+/-0.8 deg) respectively. Mean 3-D error magnitude was 6.6 mm with a 95% certainty of 11.2 mm. Mean intra-fraction shifts observed in the VRT, LNG and LAT directions were -0.1+/-0.4 mm, -0.1+/-0.4 mm and 0.1+/-0.3 mm respectively. Mean 3-D intra-fraction shift magnitude was 0.6 mm with a 95% certainty of 1.4 mm. No significant difference was observed between the SBFs. CONCLUSIONS: Patient positioning is not sufficiently reproducible with the evaluated SBF to allow non-image guided treatment. However, provided image guidance is used for patient positioning, these frames provide excellent immobilization which is on par with mask based cranial radiosurgery.

Cost-effectiveness of CT and PET-CT for determining the need for adjuvant neck dissection in locally advanced head and neck cancer.

BACKGROUND: Patients with node-positive head and neck squamous cell carcinomas (HNC) have a significant risk of residual disease (RD) in the neck after treatment, despite optimal chemoradiotherapy (CRT). Adjuvant neck dissection (ND) after CRT has been considered standard treatment, but its morbidity has led investigators to consider using post-CRT imaging to determine the need for surgery. We analyzed the cost-effectiveness of computed tomography (CT) and positron emission tomography-computed tomography (PET-CT) as predictors of the need for ND compared with ND for all patients. MATERIALS AND METHODS: We developed a Markov model to describe health states in the 5 years after CRT for HNC in a 50-year-old man. We compared three strategies: dissect all patients, dissect patients with RD on CT, and dissect patients with RD on PET-CT. Probabilistic sensitivity analyses were carried out to model uncertainty in PET-CT performance, up-front and salvage dissection costs, and patient utilities. RESULTS: ND only for patients with RD on PET-CT was the dominant strategy over a wide range of realistic and exaggerated assumptions. Probabilistic sensitivity analyses confirmed that the PET-CT strategy was almost certainly cost-effective at a societal willingness-to-pay threshold of $500,000/quality-adjusted life year. CONCLUSION: Adjuvant ND reserved for patients with RD on PET-CT is the dominant and cost-effective strategy.

Prognostic significance of mid- and post-ABVD PET imaging in Hodgkin's lymphoma: the importance of involved-field radiotherapy.

BACKGROUND: Although positron emission tomography (PET) response to chemotherapy (CT) has prognostic significance in Hodgkin's lymphoma (HL), it is unclear whether patients with 2-[fluorine-18]fluoro-2-deoxy-D-glucose (FDG)-PET positivity during and/or after CT can be rendered disease free with consolidative involved-field radiotherapy (IFRT). METHODS: Patients with HL treated with adriamycin, bleomycin, vinblastine and dacarbazine (ABVD)-based CT and radiotherapy (RT) at our institution from January 2000 to March 2007 were eligible. All patients had either a post-treatment PET or PET-CT before initiation of RT or a negative midtreatment PET or PET-CT. The primary end point was failure-free survival (FFS) for patients with and without residual FDG avidity after ABVD. The treatment outcome of patients with interim PET positivity during CT was also reported. RESULTS: Seventy-three patients were included in this study. Twenty patients (out of 46) were PET positive on interim PET, and 13 patients (out of 73) were PET positive at the conclusion of CT. At a median follow-up of 3.4 years for surviving patients, the 2-year FFSs for patients PET-negative versus PET-positive disease after ABVD were 95% and 69%, respectively (P < 0.01). On bivariable Cox regression, post-ABVD positivity (hazard ratio 4.8, P = 0.05) was predictive of disease recurrence after controlling for bulky disease. Of the 20 patients with interim PET positivity, three recurred, with a 2-year FFS of 85%. Among the 13 patients with interim PET positivity, but became PET negative at the completion of CT, the 2-year FFS was 92%. CONCLUSION: Sixty-nine per cent of patients with residual FDG avidity after ABVD were free of disease after consolidative RT, indicating a majority of patients with persistent lymphoma can be cured by sterilizing this PET-positive disease.

Brusatol-mediated induction of leukemic cell differentiation and G(1) arrest is associated with down-regulation of c-myc.

Employing the natural product quassinoid brusatol, we currently report cellular and molecular events leading to cell death or terminal differentiation in a panel of leukemic cells. Brusatol and bruceantin exerted significant cytotoxic effects with several leukemic cell lines, but not with K562 or normal lymphocytic cells. Cell lines that were less sensitive to the cytotoxic effects of brusatol responded primarily through induction of terminal differentiation. The differentiated phenotype in cell lines derived from acute or chronic myeloid leukemias (HL-60, K562, Kasumi-1, NB4, U937, BV173) was characterized for producing superoxide and non-specific esterase, and some with up-regulation of CD13 (cluster of differentiation) and down-regulation of CD15. Chronic myeloid leukemic cell lines, K562 and BV173, and acute lymphoblastic cell lines, SUPB13 and RS4;11, were induced to differentiate along the erythrocytic pathway. Withdrawal studies showed that brusatol treatment for 48 h was sufficient to induce commitment towards terminal differentiation in HL-60, K562 and SUPB13. Reh cells did not undergo maturation. Analysis of c-MYC protein expression revealed that brusatol or bruceantin down-regulated expression to undetectable levels in cell lines that were most sensitive, based on cell death or terminal differentiation. Generally, c-myc RNA was reduced, but to a lower extent than c-MYC protein levels, indicating c-myc expression was regulated by quassinoids at the post-transcriptional level. Thus, regulation of c-myc expression may represent a critical event that leads to terminal differentiation. Since these responses are facilitated at clinically achievable concentrations, quassinoids may be of value for the management of hematological malignancies.

Stem cell transplantation eliminates alloantibody in a highly sensitized patient.

Highly sensitized patients are forced to stay on transplant waiting lists for many years and ultimately may never find a donor. Peripheral blood stem cell (PBSC) transplantation may provide a strategy to decrease host alloreactivity through the production of a chimeric state. We investigated alloreactivity and chimerism in a highly sensitized 40-year-old patient with sickle cell disease who underwent a nonradiation based conditioning regimen consisting of fludarabine, ATG, and high dose melphalan, for allogeneic stem cell transplant. Host monocytes and lymphocytes became donor in origin by day 14. PRA, initially 100% pretransplant, fell to 0 by day 263. Anti-red blood cells antibody became undetectable by day 152. The use of a new nonradiation-based conditioning regimen enabled successful engraftment of allogeneic donor PBSCs and the elimination of alloantibody. As new less toxic conditioning regimens are developed, PBSC transplantation might provide a new solution to allosensitization.

Mesenchymal stem cells are capable of homing to the bone marrow of non-human primates following systemic infusion.

OBJECTIVE: The human bone marrow contains mesenchymal stem cells capable of differentiating along multiple mesenchymal cell lineages. Using a non-human primate model, we sought to determine whether the systemic infusion of baboon-derived mesenchymal stem cells was associated with toxicity and whether these cells were capable of homing to and persisting within the bone marrow. MATERIALS AND METHODS: Five baboons (Papio anubis) were administered lethal irradiation followed by intravenous autologous hematopoietic progenitor cells combined with either autologous (n = 3) or allogeneic (n = 2) mesenchymal stem cells that had been expanded in culture. In four of these baboons, the mesenchymal stem cells were genetically modified with a retroviral vector encoding either the enhanced green fluorescent protein gene (n = 3) or the human placental alkaline phosphatase gene (n = 1) for tracking purposes. A sixth animal received only intravenous gene marked autologous mesenchymal stem cells but no hematopoietic stem cells or conditioning irradiation. RESULTS: Following culture, baboon mesenchymal stem cells appeared morphologically as a homogeneous population of spindle-shaped cells that were identified by the monoclonal antibodies SH-3 and SH-4. These cells did not express the hematopoietic markers CD34 or CD45. Baboon mesenchymal stem cells isolated from primary culture were capable of differentiating along both adipogenic and osteogenic lineages. There was no acute or chronic toxicity associated with the intravenous infusion of mesenchymal stem cells. In all five recipients of gene marked mesenchymal stem cells, transgene was detected in post-transplant bone marrow biopsies. In two animals receiving autologous mesenchymal stem cells, including the one non-conditioned recipient, transgene could be detected over 1 year following infusion. In one recipient of allogeneic gene marked mesenchymal stem cells, transgene was detected in the bone marrow at 76 days following infusion. CONCLUSION: These data demonstrate that baboon mesenchymal stem cells: 1) are not associated with significant toxicity when administered intravenously, 2) are capable of homing to the bone marrow following intravenous infusion, and 3) have the capacity to establish residence within the bone marrow for an extended duration following systemic administration.

Fludarabine-based conditioning for allogeneic transplantation in adults with sickle cell disease.

Although allogeneic transplantation can be curative for patients with sickle cell disease, the toxicity of conditioning regimens has precluded its use in adults with significant end-organ damage. Newer conditioning regimens have been developed that are less toxic and that may broaden the applicability of allogeneic transplantation in this disorder. We report two adults with end-stage sickle cell disease, who underwent allogeneic transplantation from an HLA-identical sibling donor after conditioning with fludarabine/melphalan and ATG. Both patients had been extensively transfused and one had multiple RBC antibodies. One of the patients also had end-stage renal disease, and was dialysis dependent. Engraftment occurred promptly in both patients. Both achieved 100% donor chimerism and both were free of pain crises after transplant. The first patient died of a respiratory failure related to chronic graft-versus-host disease (GVHD) on day 335 after transplantation. The second patient developed severe gastro-intestinal GVHD and TTP and died on day 147 after transplantation. Conditioning with fludarabine/melphalan and ATG followed by allogeneic stem cell transplantation resulted in prompt and reliable engraftment in adults with end-stage sickle cell disease. The incidence of severe GVHD was unacceptably high and may be related to the ethnicity of the patients or to the inflammatory state associated with pre-existing sickle cell disease.

Cell cycle regulatory gene abnormalities are important determinants of leukemogenesis and disease biology in adult acute lymphoblastic leukemia.

To test the hypothesis that cell cycle regulatory gene abnormalities are determinants of clinical outcome in adult acute lymphoblastic leukemia (ALL), we screened lymphoblasts from patients on a Southwest Oncology Group protocol for abnormalities of the genes, retinoblastoma (Rb), p53, p15(INK4B), and p16(INK4A). Aberrant expression occurred in 33 (85%) patients in the following frequencies: Rb, 51%; p16(INK4A), 41%; p53, 26%. Thirteen patients (33%) had abnormalities in 2 or more genes. Outcomes were compared in patients with 0 to 1 abnormality versus patients with multiple abnormalities. The 2 groups did not differ in a large number of clinical and laboratory characteristics. The CR rates for patients with 0 to 1 and multiple abnormalities were similar (69% and 54%, respectively). Patients with 0 to 1 abnormality had a median survival time of 25 months (n = 26; 95% CI, 13-46 months) versus 8 months (n = 13; 95% CI, 4-12 months) for those with multiple abnormalities (P <.01). Stem cells (CD34+lin-) were isolated from adult ALL bone marrows and tested for p16(INK4A) expression by immunocytochemistry. In 3 of 5 patients lymphoblasts and sorted stem cells lacked p16(INK4A) expression. In 2 other patients only 50% of sorted stem cells expressed p16(INK4A). By contrast, p16 expression was present in the CD34+ lin- compartment in 95% (median) of 9 patients whose lymphoblasts expressed p16(INK4A). Therefore, cell cycle regulatory gene abnormalities are frequently present in adult ALL lymphoblasts, and they may be important determinants of disease outcome. The presence of these abnormalities in the stem compartment suggests that they contribute to leukemogenesis. Eradication of the stem cell subset harboring these abnormalities may be important to achieve cure.

The IRS-pathway operates distinctively from the Stat-pathway in hematopoietic cells and transduces common and distinct signals during engagement of the insulin or interferon-alpha receptors.

Binding of interferon-alpha (IFN-alpha) to its receptor on hematopoietic cells activates the signal transducers and activators of transcription (Stat)- and insulin receptor substrate (IRS)-pathways, and regulates expression of antiproliferative and antiviral activities. However, it remains unknown whether these two pathways cooperate in the generation of IFN-alpha responses or function independently, and whether IRS-proteins transduce distinct downstream signals in response to IFNs or insulin/insulin-like growth factor (IGF)-1-mediated activation. Our data show that in response to IFN-alpha treatment, IRS-1 functions selectively as a docking protein for the SH2 domains of the p85 subunit of the PI 3'-kinase, but not the SH2 domain of Grb-2 which is engaged during insulin/IGF-1 signaling. In studies with THP-1 human myelomonocytic cells and 32D mouse myeloid cells, which are IRS-defective, we found that the IFN-alpha-regulated activation of Stat-1, Stat-2, and Stat-3 does not require the function of the IRS-system. Furthermore, THP-1 cells are responsive to the protective effect of IFN-alpha against vesicular stomatitis virus. Both 32D and THP-1 cells were resistant to the growth inhibitory effect of IFN-alpha, but this effect was not reversible by expression of IRS-1 or IRS-2 alone in 32D cells. Taken altogether these data show that: (1) The IRS-system transduces common and distinct signals in response to IFN-alpha or insulin/lGF-1 stimulation of hematopoietic cells. (2) The IRS-pathway operates separately from the Stat-pathway, and its function is not essential for the generation of the antiviral effect of IFN-alpha. (3) Neither the IRS- nor the Stat-pathways alone are sufficient to mediate the antiproliferative effects of IFN-alpha in hematopoietic cells, and additional signaling elements are required.

Anterior cruciate ligament graft fixation. Initial comparison of patellar tendon and semitendinosus autografts in young fresh cadavers.

The initial biomechanical properties of semitendinosus and patellar tendon autografts and their fixation strengths were investigated. Twenty fresh cadaveric knees from donors under 42 years of age were used in the study. After removing all soft tissues other than the anterior cruciate ligament, we determined the ultimate tensile strength (2195 +/- 427 N) and stiffness (306 +/- 80 N/mm) of the anterior cruciate ligament in nine knees. In six knees, anterior cruciate ligaments were reconstructed using an autologous patellar tendon graft with proximal and distal interference fit screws; this resulted in an ultimate tensile strength of 416 +/- 66 N. Five knees were reconstructed with quadruple-stranded (double-looped) semitendinosus tendons fixed proximally by a titanium button and braided tape and distally by tibial post screw. This resulted in an ultimate tensile strength of 612 +/- 73 N, which was significantly higher than the strength in the patellar tendon group. Graft stiffness did not differ between the groups and was 47 +/- 19 N/mm (N = 11). This study demonstrates that the reconstructed knees had only 20% to 30% of the ultimate tensile strength of the normal anterior cruciate ligament. In summary, the semitendinosus reconstruction using a button for proximal fixation is, at the time of surgery, approximately 50% stronger than patellar tendon reconstructions with similar stiffness.

Interaction of p59fyn with interferon-activated Jak kinases.

During IFN alpha stimulation, p59(fyn) associates with the Type I IFNR-associated Tyk-2 kinase in several human hematopoietic cell lines in vivo. This interaction is direct, and is mediated by the SH2 domain in p59(fyn), as shown by binding studies using glutathione-S-transferase fusion proteins and far western blots. Furthermore, in response to IFN alpha-treatment of cells, the SH2 domain of Fyn interacts with the Tyk-2-associated c-cbl proto-oncogene product. In a similar manner, during IFN gamma stimulation, p59(fyn) associates via its SH2 domain with the activated form of the IFN gamma-dependent Jak-2 kinase. These data suggest that p59(fyn) is a common element in IFN alpha and IFN gamma signaling, and is selectively engaged by the Type I or II IFN receptors via specific interactions with distinct Jak kinases.

Activation of the phosphatidylinositol 3-kinase serine kinase by IFN-alpha.

During engagement of the type I IFN receptor, IRS-1 is phosphorylated on tyrosine and associates with the p85 regulatory subunit of the phosphatidylinositol (PI) 3'-kinase, which is a dual-specificity enzyme possessing both lipid and serine kinase activities. We sought to determine whether treatment of cells with IFN-alpha activates the PI 3'-kinase serine kinase. 32P-labeling experiments and phosphoaminoacid analysis of immunoprecipitated IRS-1 protein demonstrated that, in addition to tyrosine phosphorylation, IFN-alpha induces its phosphorylation on serine residues. In vitro kinase assays on alphaIRS-1 immunoprecipitates also demonstrated IFN-alpha-dependent serine phosphorylation of IRS-1, suggesting that the protein associates with an IFN-alpha-regulated serine kinase. Furthermore, IFN-alpha-dependent phosphorylation of IRS-1 was detected in in vitro kinase assays on alpha p85 immunoprecipitates, and was inhibited by pretreatment of cells with the specific PI 3'-kinase inhibitor wortmannin, consistent with a regulatory role of the PI 3'-kinase serine kinase on the phosphorylation of the protein. Treatment of cells with wortmannin also inhibited the phosphorylation of the p85 subunit of PI 3'-kinase and the type I IFN-regulated activation of the Map kinase, but had no inhibitory effect on the IFN-alpha-induced activation of Tyk-2 and Jak-1 kinases nor on the activation of Stat-1, Stat-2, and Stat-3. Taken all together, these data establish that the PI 3'-kinase serine kinase is activated by IFN-alpha and may play an important role in the transmission of type I IFN receptor-generated signals.

Regional differences in gastric acidity and antacid distribution: is a single pH electrode sufficient?

UNLABELLED: Accurate measurement of intragastric acidity has both clinical and investigational importance in studying gastric pathophysiology. OBJECTIVES: The aims of this study were fourfold: (1) to investigate whether regional differences in intragastric acidity exist; (2) to determine intragastric acidity after a standard antacid was administered in both the fasting and fed states; (3) to monitor gastric emptying of and anatomic distribution of radiolabeled antacid during fasting and postprandial periods; and (4) to determine whether the regional effects of ingested antacid correlated with the anatomic distribution of the antacid. METHODS: Eight normal male volunteers were studied after fluoroscopically guided nasogastric placement of a tube assembly containing four pH electrodes, with one electrode in each quartile of the stomach. Simultaneous pH readings were made from the four electrodes while fasting, after administration of fasting antacid (30 ml, 79 mEq buffering capacity), postprandially, and after postprandial antacid ingestion. All subjects repeated the protocol on a separate day, five of them using radiolabeled antacid. Gastric emptying and gastric distribution over time of radiolabeled antacid were determined for comparison to regional intragastric acidity. RESULTS: Intragastric acidity varied regionally over time in response to meals and to fasting and postprandial antacid. In the fasting state, intragastric acidity returned to baseline after antacid ingestion in a proximal to distal (cardia to antrum) sequence, while postprandial antacid resulted in a return to baseline acidity in a distal to proximal (antrum to cardia) sequence. Radiolabeled antacid distribution paralleled intragastric pH and hydrogen ion concentration in the fasting state, with 82% of the antacid localizing in the distal half of the stomach within the first minute after antacid ingestion. Postprandially, the greatest initial and most prolonged antacid buffering effect occurred proximally, correlating with the presence of radiolabeled antacid. Postprandial antacid remained in the stomach for a longer time (T1/2 = 93.1 +/- 23.4 min) compared with fasting antacid (T1/2 = 23.6 +/- 11.1 min). CONCLUSIONS: Measurement of acidity in the four quartiles of the stomach demonstrated regional variation in response to both food and a standard antacid. A single pH electrode does not detect regional intragastric pH differences.

Value of molecular monitoring during the treatment of chronic myeloid leukemia: a Cancer and Leukemia Group B study.

PURPOSE: Disappearance of the Philadelphia chromosome during treatment for chronic myeloid leukemia (CML) has become an important therapeutic end point. To determine the additional value of molecular monitoring during treatment for CML, we performed a prospective, sequential analysis using quantitative Southern blot monitoring of BCR gene rearrangements of blood and marrow samples from Cancer and Leukemia Group B (CALGB) study 8761. PATIENTS AND METHODS: Sixty-four previously untreated adults with chronic-phase CML who were enrolled onto CALGB 8761, a molecular-monitoring companion study to a treatment study for adults with chronic-phase CML (CALGB 9013). Treatment consisted of repetitive cycles of interferon alfa and low-dose subcutaneous cytarabine. Blood and marrow Southern blot quantitation of BCR gene rearrangements was compared with marrow cytogenetic analysis before the initiation of treatment and of specified points during therapy. Reverse-transcriptase polymerase chain reaction (RT-PCR) analysis was performed to detect residual disease in patients who achieved a complete response by Southern blot or cytogenetic analysis. RESULTS: Quantitative molecular monitoring by Southern blot analysis of blood samples was found to be equivalent to marrow monitoring at all time points. Twelve of 62 (19%) follow-up samples studied by Southern blot analysis had a complete loss of BCR gene rearrangement in matched marrow and blood specimens. Southern blot monitoring of blood samples was also found to be highly correlated to marrow cytogenetic evaluation at all points, although there were four discordant cases in which Southern blot analysis of blood showed no BCR gene rearrangement, yet demonstrated from 12% to 20% Philadelphia chromosome-positive metaphase cells in the marrow. RT-PCR analysis detected residual disease in five of six patients in whom no malignant cells were detected using Southern blot or cytogenetic analyses. CONCLUSION: Quantitative Southern blot analysis of blood samples may be substituted for bone marrow to monitor the response to therapy in CML and results in the need for fewer bone marrow examinations. To avoid overestimating the degree of response, marrow cytogenetic analysis should be performed when patients achieve a complete response by Southern blot monitoring. This approach provides a rational, cost-effective strategy to monitor the effect of treatment of individual patients, as well as to analyze large clinical trials in CML.

A single intragastric pH electrode does not accurately measure intragastric acidity.

OBJECTIVES: Recent studies have raised concerns about the validity of using a single intragastric pH electrode to measure gastric acidity accurately and reproducibly. The aim of this study was to compare simultaneous intragastric pH measurements obtained from an indwelling glass pH electrode to those determined by aspirations from the gastric pool and from ex vivo measurement. METHODS: Twenty two normal volunteers were studied after fluoroscopically guided placement of a combined nasogastric tube-pH probe assembly. Simultaneous intragastric pH electrode and aspirate pH determinations were made basally for 120 min after administration of 15 ml of antacid (40 mEq buffering capacity) and for another 120 min an hour postprandially after administration of a second 15-ml dose of antacid. Gastric acid concentration (pH) measurements were recorded every 15 min during the following study protocols: 1) fasting baseline (30 min); 2) fasting antacid (120 min); 3) test meal (60 min); and 4) postprandial antacid (120 min). RESULTS: Intragastric pH was consistently and significantly lower as measured by intragastric pH electrode than by aspiration. Baseline hydrogen ion concentration ([H+]) was 4.3 times higher by direct electrode measurement than by aspirate. Antacid-administered fasting decreased [H+] maximally at 15 min to 48% and 82% of baseline by electrode and aspiration, respectively. The minimal residual intragastric [H+] after fasting antacid was 12.4 times higher by electrode than by aspiration. Postprandial antacid maximally reduced [H+] by 46% at 15 min when recorded using an electrode compared with 60% at 30 min by aspiration. Correlation coefficients for intragastric electrode [H+] versus aspiration [H+] were 0.26 (p = 0.253), 0.61 (p < 0.001), 0.56 (p < 0.01), and 0.31 (p < 0.001), for baseline, fasting antacid, meal, and postprandial antacid, respectively. CONCLUSIONS: Quantitative evaluations of intragastric acidity (pH) using an intragastric pH electrode and aspiration of gastric juice may yield remarkably different results. Studies that rely on a single intragastric electrode to quantitate intragastric acidity may be highly inaccurate.