Electronic speckle pattern interferometry: a novel non-invasive tool for studying drug transport rate through free films.

In this work, Electronic Speckle Pattern Interferometry (ESPI) is presented as a non-invasive tool to study drug transport in controlled release systems. ESPI is shown to be a feasible tool to measure drug film permeability via comparison with an ordinary diaphragm cell. A specially designed cuvette was used in the release study: the polymeric film separated the donor and the receiving chambers of the cuvette to create a diffusion cell with no mixing in the two chambers. Thus, the cuvette mimicked a coated system immersed in a stagnant bulk liquid. Concentration profile data were obtained for the two compartments. Using these data, it was possible to visually discriminate between a film subject only to diffusion and a film subject to diffusion as well as osmotic effects. Moreover, using the concentration profile data collected at different time intervals, it was possible to follow the film properties in terms of drug permeability, thus studying how drug permeability depended on drug concentration. Compared to other measuring techniques, ESPI offers the advantages that no invasive measurements are needed, and that no sampling and calibration are required. Furthermore, the permeability can be measured with no influence of mass transfer in the boundary layers.     

Phase I study of high-dose cytosine arabinoside and etoposide in patients with advanced malignancies

Summary Cytosine arabinsodie (ara-C) and etoposide (VP-16) display synergy in the laboratory. Twenty-six patients participated in a phase I study of high-dose ara-C in combination with VP-16. The dose of VP-16 was held constant at 50 mg/m² as an intermittent infusion over 33 h; escalating doses of ara-C were given as infusions during hours 9–12 and 21–24. Myelosuppression was the dose-limiting toxicity and occurred with doses considerably less than those expected from studies of the two drugs as single agents. The suggested initial doses for phase II trials with this schedule are 750 mg/m²×2 doses of ara-C and 50 mg/m² of VP-16. Nonhematologic toxicity was minimal; therefore, further dose escalation is feasible in patients in whom myelosuppression is acceptable.Supported in part by grants from the National Cancer Institute (CA-12197 and CA-09422) and the American Cancer Society CF-85-182     

1-beta-D-arabinofuranosylcytosine-diphosphate-choline is formed by the reversal of cholinephosphotransferase and not via cytidylyltransferase.

In an effort to identify the pathway leading to the formation of 1-beta-D-arabinofuranosylcytosine-diphosphate (ara-CDP)-choline from 1-beta-D-arabinofuranosylcytosine (ara-C) treatment of cultured cells, as well as of cells obtained from leukemia patients, we probed the enzymatic steps involved in the CDP-choline pathway for phosphatidylcholine biosynthesis. Ara-C-triphosphate was not a substrate for CTP:phosphocholine cytidylyltransferase activity under the conditions employed, whereas CTP and dCTP were utilized to form CDP-choline and dCDP-choline, respectively. When presented together, ara-C-triphosphate and CTP inhibited the enzymatic conversion of CTP to CDP-choline in the presence of phosphocholine, with a Ki of 6 mM. Since CTP:phosphocholine cytidylyltransferase did not appear to be responsible for the increased levels of ara-CDP-choline, we next studied the other enzyme in the pathway for phosphatidylcholine synthesis that could form ara-CDP-choline, CDP-choline:1,2-diacylglycerol cholinephosphotransferase. CDP-choline:1,2-diacylglycerol cholinephosphotransferase activity present in microsomes isolated from L5178Y murine leukemia cells exhibited a reversal of its normal catalytic activity, using CMP and 1-beta-D-arabinofuranosylcytosine-monophosphate (ara-CMP) along with phosphatidylcholine to produce either CDP-choline or ara-CDP-choline, plus diradylglycerol. The Vmax and Km values for CMP were 0.78 +/- 0.04 nmol/min/mg and 340 +/- 20 microM, respectively, whereas the Vmax and Km for ara-CMP were 0.22 +/- 0.06 nmol/min/mg and 1410 +/- 540 microM, respectively. A Ki value of 3 mM was obtained for ara-CMP under the cell-free assay conditions used. These results indicate that ara-CDP-choline most likely arises from a reversal of the CDP-choline:1,2-diacylglycerol cholinephosphotransferase utilizing ara-CMP, rather than from the catalysis of ara-C-triphosphate plus phosphocholine to ara-CDP-choline by CTP:phosphocholine cytidylyltransferase. It is speculated that this mechanism may explain, in part, the rapid cellular lysis observed with high dose ara-C therapy.     

Intermittent jaundice by tumor emboli from intrahepatic cholangiocarcinoma.

Free-floating tumor debris or mucobilia as a cause of intermittent obstruction has been described infrequently. A patient with intermittent jaundice caused by tumor emboli from an intrahepatic polypoid mucinous cholangiocarcinoma is presented. Symptoms of intermittent jaundice and midepigastric pain persisted over 5 years despite an initial cholecystectomy and common bile duct exploration before definitive diagnosis and treatment of an hepatic trisegmentectomy (segments II, III, and IV). Intraductal mucin was confirmed intraoperatively and pathologically as the cause of the obstructive jaundice. The patient remains asymptomatic and without evidence of disease more than 5 years postoperatively. This report of a predominantly mucin-producing intrahepatic cholangiocarcinoma details a rare protracted clinical course of intermittent biliary obstruction from mucus emboli and highlights the possibility of long-term survival after complete resection.     

Protective levels of diphtheria-neutralizing antibody induced in healthy volunteers by unilateral priming-boosting intranasal immunization associated with restricted ipsilateral mucosal secretory immunoglobulin a

Subunit intranasal vaccines offer the prospect of inducing combined systemic-mucosal immunity against mucosally transmitted infections such as human immunodeficiency virus. However, although human studies have demonstrated the induction of active immunity, secretory immunoglobulin A (sIgA) responses are variable, and no study has demonstrated protection by accepted vaccine-licensing criteria as measured by direct toxin-neutralizing activity. Using the genetically inactivated mutant diphtheria toxoid CRM(197) in a bioadhesive polycationic polysaccharide chitosan delivery system, we found that a single nasal immunization was well tolerated and boosted antitoxin neutralizing activity in healthy volunteers, which could be further boosted by a second immunization. The neutralizing activity far exceeded accepted protective levels and was equivalent to that induced by standard intramuscular vaccine and significantly greater than intranasal immunization with CRM(197) in the absence of chitosan. A striking but unexpected observation was that although unilateral intranasal immunization induced circulating antitoxin antibody-secreting cells, a nasal antitoxin sIgA response was seen only after the second immunization and only in the vaccinated nostril. If these data are reproduced in larger studies, an intranasal diphtheria vaccine based on CRM(197)-chitosan could be rapidly licensed for human use. However, a restricted sIgA response suggests that care must be taken in the priming-boosting strategy and clinical sampling techniques when evaluating such vaccines for the induction of local mucosal immunity.     

CCL22-induced responses are powerfully enhanced by synergy inducing chemokines via CCR4: evidence for the involvement of first beta-strand of chemokine

In an attempt to clarify how cells integrate the signals provided by multiple chemokines expressed during inflammation, we have uncovered a novel mechanism regulating leukocyte trafficking. Our data indicate that the concomitant exposure to CCR4 agonists and CXCL10/IP-10 strongly enhances the chemotactic response of human T lymphocytes. This enhancement is synergistic rather than additive and occurs via CCR4 since it persists after CXCR3 blockade. Besides chemotaxis, other cellular responses are enhanced upon stimulation of CCR4-transfected cells with CCL22/MDC plus CXCL10. Several other chemokines in addition to CXCL10 were able to increase CCL22-mediated chemotaxis. The first beta-strand of the chemokine structure is highly and specifically implicated in this phenomenon, as established using synergy-inducing and non-synergy-inducing chimeric chemokines. As shown in situ for skin from atopic and allergic contact dermatitis patients, this organ becomes the ideal environment in which skin-homing CCR4(+) T lymphocytes can accumulate under the stimulus offered by CCR4 agonists, together with the synergistic chemokines that are concomitantly expressed. Overall, our results indicate that chemokine-induced synergism strengthens leukocyte recruitment towards tissues co-expressing several chemokines.     

Interrupted versus continuous chemotherapy in patients with metastatic breast cancer. The Piedmont Oncology Association

BACKGROUND. Chemotherapy for metastatic breast cancer is palliative, and the optimal duration of therapy is unknown. We designed a trial to determine whether continuous treatment is superior to stopping treatment after a brief induction period and resuming treatment when the disease progresses. METHODS. We treated 250 women with metastatic breast cancer with six courses of cyclophosphamide, doxorubicin, and fluorouracil given every three weeks. At the completion of this induction period, women whose disease either regressed or remained stable were randomly assigned to receive either continued treatment with cyclophosphamide, methotrexate, and fluorouracil (maintenance therapy) or no further treatment (observation) followed by treatment with cyclophosphamide, methotrexate, and fluorouracil when disease progression became evident (reinduction). RESULTS. The combined rate of complete and partial responses after initial therapy was 30 percent (71 of 233 patients who could be evaluated; 95 percent confidence interval, 25 percent to 37 percent). In another 42 percent (98 patients), the disease remained stable. A total of 145 patients were randomized. Seventy-one were randomly assigned to the maintenance-therapy group, and 74 to the observation group. The median time to progression was 9.4 months for patients in the maintenance-therapy group and 3.2 months for patients in the observation group (P less than 0.001). After reinduction therapy, the median time to progression was 3.5 months. The median length of survival from the time of initial therapy was 14.8 months for all 250 patients; it was 21.1 months for the 71 patients in the maintenance-therapy group and 19.6 months for the 74 patients in the observation group (P = 0.67). Maintenance therapy was the most important determinant of the time before progression (P less than 0.001), but it was not associated with prolonged survival. The changes in performance status were similar in the patients in both groups, but nausea, vomiting, and mucositis were significantly more frequent in the maintenance-therapy group. CONCLUSIONS. In patients with breast cancer who received induction chemotherapy for 18 weeks, subsequent continuous chemotherapy was associated with a significant prolongation of the time before progression as compared with those receiving no further therapy; overall survival, however, was not significantly different in the two groups.     

Impact of COVID-19 Among Immigrant and Communities of Color Living with HIV in Oregon, 2020: Two Pandemics Rooted in Racism

Journal of Immigrant and Minority Health - Over 8100 people living with HIV (PLWH) in Oregon are at risk of acquiring COVID-19, and communities of color are disproportionately impacted by both...     

The effect of ara-C-induced inhibition of DNA synthesis on its cellular pharmacology

Summary The cytotoxicity of ara-C is believed to result from incorporation of ara-CTP into DNA and inhibition of DNA synthesis. Since complete inhibition of DNA synthesis would prevent further incorporation of ara-CTP, ara-C may have a self-limiting effect on its own cytotoxicity, particularly at the high concentrations typical of highdose ara-C clinical protocols. In this study, the incorporation of [³H]-dThd and [³H]-ara-C into DNA were compared. Within 1 h of exposure of L5178Y cells to ara-C, the rate of [³H]-dThd incorporation into the acid-insoluble fraction was reduced by 98%. Despite this nearly complete block in [³H]-dThd incorpration, DNA synthesis was not completely inhibited since [³H]-ara-C continued to be incorporated for up to 6 h, although a plateau in ara-CDNA synthesis was observed between 2 and 3 h exposure when ara-CTP levels were maximal. The effect of ara-C on [³H]-dThd incorporation into DNA was due in part to an indirect effect of ara-C on the metabolism of intracellular [³H]-dThd to [³H]-dTTP. Within 30 min exposure to 10 M ara-C, the rate of cellular [³H]-dTTP synthesis was slowed to only 15% of the control rate. This was not due to inhibition of [³H]-dThd transport, since the intracellular and extracellular concentrations of the nucleoside were equal. The effect of ara-C on [³H]-dTTP synthesis resulted from significant changes in deoxynucleoside 5-triphosphate (dNTP) pools. dTTP, dATP, and dGTP levels were increased, whereas the dCTP concentration was decreased. When dThd kinase from L5178Y cells was assayed with increased dTTP levels induced by ara-C vs the dTTP level in control cells, its activity was reduced by 72%. Thus, the [³H]-dThd incorporation experiment overestimated the extent of inhibition of DNA synthesis by ara-C due to increased feedback inhibition of dThd kinase and increased competition for DNA polymerase between the elevated unlabeled dTTP pool and the decreased levels of [³H]-dTTP. In vitro assay of DNA polymerase in the presence of the ara-CTP concentration achieved after 0.5 or 3 h exposure to 10 M ara-C (60 M and 200 M, respectively), plus the mixture of dNTPs found intracellularly at these times, resulted in 57% and 80% inhibition of the polymerase, respectively. This inhibition may account for the plateau in the accumulation of ara-CDNA that was observed at 3 h and suggests that ara-C incorporation may be self-limiting at high cellular concentrations of ara-CTP. The ara-C-induced decline in dCTP noted above was apparently a secondary effect resulting from the inhibition of ribonucleotide reductase by the elevated dTTP and dATP. CDP reductase activity in the presence of dATP and dTTP at the concentrations found in ara-C-treated cells was 58% of the activity observed in the presence of nucleotide levels found in control cells. The decrease in dCTP levels was associated with a reciprocal increase in the rate of [³H]-ara-C phosphorylation following subsequent exposure to unlabeled ara-C. Thus, ara-C self-potentiated its own uptake in these cells. These observations of the self-limiting and self-potentiating effects of high concentrations of ara-C may be relevant to the selection of the optimal dose and the duration of exposure in the clinical use of high-dose ara-C infusions.Abbreviations ara-C I--d-arabinofuranosyl, cytosine (cytosine arabinoside) - ara-CTP ara-C triphosphate - NTP unspecified nucleoside 5-triphosphate - dNTP deoxynucleoside 5-triphosphate - PBS phosphate-buffered saline This research was supported by grants from the American Cancer Society (CH35J), the National Institutes of Health (CA 12197), the Gaston County Cancer Society, and Dr. George Royer of the Upjohn Company