Prospective, Multicenter, Head-to-Head Comparison Between Allergists Versus Nonallergists in Low-Risk Penicillin Allergy Delabeling: Effectiveness, Safety, and Quality of Life (HK-DADI2).

BACKGROUND: Penicillin "allergy" labels are prevalent but frequently misdiagnosed. Mislabelled allergies are associated with adverse outcomes and increased antimicrobial resistance. With an urgent need to delabel the overwhelming number of mislabeled allergies, nonallergist evaluations have been advocated for low-risk individuals. Despite growing interest in non-allergist-led initiatives, evidence on their effectiveness, safety, and impact by direct comparisons is lacking. OBJECTIVE: To assess the comparative outcomes of penicillin allergy evaluations conducted by allergists versus nonallergists. METHODS: A prospective, multicenter, pragmatic study was conducted at 4 tertiary hospitals (1 allergist- vs 3 non-allergist-led) for low-risk penicillin allergy patients in Hong Kong-the Hong Kong Drug Allergy Delabelling Initiative 2 (HK-DADI2). RESULTS: Among 228 low-risk patients who underwent testing (32.9% by allergists, 67.1% by nonallergists), only 14 (6.1%) had positive penicillin allergy testing results. Delabeling rates (94.1% vs 93.3%; P = .777), positive skin test results (2.6% vs 2.7%; P > .99), and drug provocation test results (3.3% vs 2.7%; P = 1.000) were similar between allergists and nonallergists. There were no systemic reactions in either cohort. All patients had significant improvements in health-related quality of life (Drug Hypersensitivity Quality of Life Questionnaire scores -5.00 vs -8.33; P = .072). Nonallergist evaluations had shorter waiting times (0.57 vs 15.7 months; P < .001), whereas allergists required fewer consultations with higher rate of completing evaluations within a single visit (odds ratio, 0.04; P < .001). CONCLUSIONS: With training and support, nonallergists can independently evaluate low-risk penicillin allergies. Compared with allergists, evaluation of low-risk penicillin allergy by nonallergists can be comparably effective, safe, and impactful on quality of life. More multidisciplinary partnerships to empower nonallergists to conduct allergy evaluations should be encouraged.

Effects of syphilis infection among HIV-1-positive individuals on suppressive antiretroviral therapy.

INTRODUCTION: Incident syphilis leads to changes in plasma HIV-1 RNA and CD4 + T-cell level in people with HIV (PWH) with viraemia. Its effect in PWH on suppressive antiretroviral therapy (ART) is less clear. METHODS: PWH on suppressive ART (plasma HIV-1 RNA < 50copies/mL) followed at the Queen Elizabeth Hospital, Hong Kong, China were regularly screened for syphilis. Their plasma HIV-1 RNA, CD4 + and CD8 + T-cell, and total lymphocyte levels before syphilis, during syphilis, and after successful treatment were compared. RESULTS: Between 2005 and 2020, 288 syphilis episodes from 180 individuals were identified; 287 episodes were related to male, with a median age of 41 at diagnosis; 221 (77%) were syphilis re-infection. The rates of plasma HIV-1 suppression were statistically unchanged across the time-points (97% pre-syphilis, 98% during syphilis, and 99% post-treatment). Total lymphocyte, CD4+ and CD8+ T-cell levels decreased during incident syphilis (p<0.01), and rebounded post-treatment (p<0.01). VDRL titre was associated with declines in CD4+ T-cell (p=0.045), CD8+ T-cell (p=0.004), and total lymphocyte levels (p=0.021). Pre-syphilis CD4/CD8 ratio was associated with increases in CD8+ T-cell (p=0.001) and total lymphocyte levels (p=0.046) during syphilis. Syphilis re-infection was associated with an increase in total lymphocyte level (p=0.037). In the multivariable analysis, only pre-syphilis CD4/CD8 ratio was independently associated with increases in CD8+ T-cell (p=0.014) and total lymphocyte levels (p=0.039) during syphilis. CONCLUSIONS: Among virally-suppressed PWH, total lymphocyte, CD4+, and CD8+ T-cell levels declined during incident syphilis but rebounded post-treatment. The status of plasma HIV suppression was unaffected by syphilis.

Hong Kong Drug Allergy Delabelling Initiative (HK-DADI) consensus statements for penicillin allergy testing by nonallergists.

Introduction: Penicillin allergy testing has been traditionally performed by allergists, but there remains a huge deficit of specialists. A multidisciplinary effort with nonallergists would be invaluable to overcome the magnitude of penicillin allergy labels via the Hong Kong Drug Allergy Delabelling Initiative (HK-DADI). These consensus statements (CSs) offer recommendations and guidance to enable nonallergists to screen for low-risk (LR) patients and perform penicillin allergy testing. Methods: CSs were formulated by the HK-DADI Group using the Delphi method. An agreement was defined as greater than or equal to 80% consensus. Results: A total of 26 CSs reached consensus after multiple rounds of Delphi. CSs were categorized into risk assessment, skin testing, drug provocation testing (DPT), and post-testing management. For risk assessment, the essentials of allergy history and exclusion criteria were detailed. Patients with only LR features can proceed with testing by nonallergists. Skin tests should be performed prior to DPT. Details regarding the timing, preparation, and interpretation of skin tests were elaborated. DPT remains the gold standard to diagnose genuine allergy or tolerance and should be performed when there is a low pretest probability following negative skin testing. Details of DPT preparations, dosing protocols, and interpretation were elaborated. For post-testing management, inaccurate allergy labels should be delabeled following negative DPT with proper patient counseling. Conclusion: CSs support penicillin allergy testing by nonallergists in Hong Kong. LR cases can be managed by nonallergists at Spoke Clinics, with training and support of an allergist-led Hub.

Metabolic active peritoneal sites affect tumor debulking in ovarian and peritoneal cancers.

RATIONALE AND OBJECTIVES: To evaluate the impact of metabolic parameters in the peritoneal cavity on the likelihood of achieving complete tumor debulking in patients with ovarian and peritoneal cancers. MATERIALS AND METHODS: Forty-nine patients with ovarian and peritoneal cancers were included, who underwent pre-operative 18F-fluorodeoxyglucose Positron Emission Tomography/Computed Tomography (18F-FDG PET/CT). The immediate surgical outcome was dichotomized into complete and incomplete tumor debulking. 18F-FDG PET/CT was qualitatively and quantitatively assessed by scrutinizing 15 anatomical sites for the presence of peritoneal carcinomatosis (PC). Patient-based and site-based diagnostic characteristics were described. Metabolic parameters (SUVmax, metabolic tumor volume and total lesion glycolysis) and the number of 18F-FDG avid peritoneal sites were evaluated between the two groups. Receiver operating curve (ROC) analysis was performed to determine the optimal cut-off value in predicting incomplete tumor debulking. RESULTS: Twenty-seven out of the 49 patients had PC and 11 had incomplete debulking. Patient-based and site-based accuracies for detection of PC were 87.8 and 97.6%, respectively. The number of 18F-FDG avid peritoneal sites was significantly different between complete and incomplete debulking groups (0.6 ± 0.8 versus 2.3 ± 1.7 sites respectively, p = 0.001), and the only independent significant risk factor among other metabolic parameters tested (odd ratio = 2.983, 95% CI 1.104-8.062) for incomplete tumor debulking with an optimal cut-off value of ≥4 (AUC = 0.816). CONCLUSION: The number of 18F-FDG avid peritoneal sites increased the risk of incomplete tumor debulking after surgery and potentially useful in assisting treatment stratification in patients with ovarian and peritoneal cancers.

Risk of nosocomial transmission of coronavirus disease 2019: an experience in a general ward setting in Hong Kong.

BACKGROUND: Coronavirus disease 2019 (COVID-19) was first reported in Wuhan in December 2019 and has rapidly spread across different cities within and outside China. Hong Kong started to prepare for COVID-19 on 31st December 2019 and infection control measures in public hospitals were tightened to limit nosocomial transmission within healthcare facilities. However, the recommendations on the transmission-based precautions required for COVID-19 in hospital settings vary from droplet and contact precautions, to contact and airborne precautions with placement of patients in airborne infection isolation rooms. AIM: To describe an outbreak investigation of a patient with COVID-19 who was nursed in an open cubicle of a general ward before the diagnosis was made. METHOD: Contacts were identified and risk categorized as 'close' or 'casual' for decisions on quarantine and/or medical surveillance. Respiratory specimens were collected from contacts who developed fever, and/or respiratory symptoms during the surveillance period and were tested for SARS-CoV-2. FINDINGS: A total of 71 staff and 49 patients were identified from contact tracing, seven staff and 10 patients fulfilled the criteria of 'close contact'. At the end of 28-day surveillance, 76 tests were performed on 52 contacts and all were negative, including all patient close contacts and six of the seven staff close contacts. The remaining contacts were asymptomatic throughout the surveillance period. CONCLUSION: Our findings suggest that SARS-CoV-2 is not spread by an airborne route, and nosocomial transmissions can be prevented through vigilant basic infection control measures, including wearing of surgical masks, hand and environmental hygiene.

Effect of RAGE polymorphisms on susceptibility to and severity of osteoarthritis in a Han Chinese population: a case-control study.

Recent studies have revealed that the inflammatory process plays a role in the pathogenesis of osteoarthritis (OA). The S100 family and receptor for advanced glycation end products (RAGE) participate in regulating inflammation, even in the production of matrix metalloproteinases (MMPs). MMP-1 degrades cartilage, which may result in OA development. Moreover, polymorphisms in RAGE, S100A8, and MMP-1 have a marked effect on ligand binding and transcription regulating. In this study, we investigated the potential genetic contribution of the RAGE, S100A8, and MMP-1 genes to OA. We performed a matched case-control association study and genotyped OA patients and healthy controls, who were analyzed by polymerase chain reaction-restriction fragment length polymorphism assays. A total of 207 patients were diagnosed with knee OA and underwent total knee replacement. The control group included 207 individuals who had standard X-rays of the knee joints to confirm K/L < 2 and were matched by age and gender. Single-nucleotide polymorphisms in RAGE (-429T/C, -374T/A, and 557G/A), S100A8 (rs3795391A/G), and MMP-1 (-1607 1G/2G, -755G/T, and -519A/G) were evaluated. RAGE -374T/A, S100A8 rs3795391A/G, MMP-1 -1607 1G/2G, -755G/T, and -519A/G showed no significant difference between OA patients and healthy controls. RAGE -429T/C and 557G/A showed a significant association between OA patients and healthy controls (P = 0.016 and 0.047, respectively). In haplotype analyses, no RAGE and MMP-1 haplotypes showed associations with OA. Our results suggest that the investigated polymorphism in the RAGE gene play a role in OA in the Han Chinese population.

Denaturing HPLC for high-throughput screening of rifampicin-resistant Mycobacterium tuberculosis isolates.

OBJECTIVE: To evaluate the use of denaturation high-performance liquid chromatography (dHPLC) as a rapid method to detect rifampicin (RMP) resistance based on mutations in the rpoB gene in a high-volume laboratory setting. METHODS: A total of 132 RMP-resistant Mycobacterium tuberculosis strains with different rpoB mutation were used to optimise the running condition of dHPLC as a pilot study. A blind correlation study was subsequently done between dHPLC and in vitro RMP susceptibility tests on 3167 M. tuberculosis strains in a high-throughput clinical setting. RESULTS: In the pilot study, rpoB mutation could be detected on 116/132 (87.9%) RMP-resistant strains by dHPLC. In the second phase of the study, 84/3107 (2.7%) clinical M. tuberculosis isolates were RMP-resistant. The sensitivity and specificity of dHPLC in the prediction of RMP resistance were 70/84 (83.3%) and 70/77 (91.0%), respectively. The specificity became 100% when 511 Leu to Pro mutation was excluded from the RMP resistance-related genetic changes. CONCLUSION: In the detection of RMP resistance in a high-throughput laboratory setting, dHPLC has been demonstrated to be rapid, simple, workable, automatable and inexpensive in terms of running costs and the labour involved.

Use of developmental language scales in Chinese children.

The Reynell Developmental Language Scale (RDLS) and Symbolic Play Test (SPT) have been useful language tests for assessing the language age of children. Both tests have been validated in English-speaking children. However, there have been no studies conducted for Chinese children, whether Mandarin (Northerners) or Cantonese (Southerners) is used as the main dialect. As the Chinese population is the largest ethnic group in the world, and Chinese emigration occurred to nearly all parts of the world, it is essential to test whether these language tools can be applied for this ethnic group. The objective of this research was to study whether RDLS and SPT are useful in assessing the language age of Chinese children. Both RDLS (Chinese version) and SPT are conducted for 116 Chinese (Cantonese-speaking) children, aged 13-59 months, in Hong Kong. There is a significant positive correlation of the language age using RDLS and SPT with the chronological age of Chinese children. Both RDLS and SPT can be adopted in determining the language/mental age of Chinese (Cantonese-speaking) children.

Inactivation of SSI-1, a JAK/STAT inhibitor, in human hepatocellular carcinomas, as revealed by two-dimensional electrophoresis.

BACKGROUND/AIMS: Hepatocellular carcinoma (HCC) is one of the most common human cancers, and many efforts have been paid to discover aberrant expression control in HCC, however the specific molecular mechanisms involved in hepatocarcinogenesis remain to be determined. METHODS: To investigate genomic changes that occur in human primary hepatocellular carcinomas (HCC), we carried out restriction landmark genomic scanning. This two-dimensional electrophoretic system displays 2000-3000 NotI-landmark sites in a single gel. RESULTS: We detected one landmark spot that showed diminished signal intensities in a majority of the HCCs we examined. Cloning revealed that this spot represented a NotI-cluster sequence that was enriched with CpG dinucleotides in the promoter region of a gene encoding Janus kinase (JAK)-binding protein, SSI-1 (also known as JAB1 or SOCS-1). Expression of the SSI-1 gene was markedly reduced in half of eight HCCs analyzed. CONCLUSIONS: This protein regulates the Janus kinase signal transducers and activators of transcription signal transduction pathway, which transmits signals from cytokines to the intracellular apparatus. These data suggest that dysregulation of the pathway relate with progression of HCC.

Apoptosis in human hepatoma cell line SMMC-7721 induced by water-soluble macromolecular components of Artemisia capillaris Thunberg.

The aim of this study was to investigate the effect of water-soluble macromolecular components of Artemisia capillaris Thunberg (ACT) on human hepatoma cell line SMMC-7721 (SMMC-7721). The morphological changes of SMMC-7721 were observed under a light microscope and an electron microscope. Inhibition of proliferation was measured with a colorimetric MTT assay. It was discovered that ACT extract-treated cells exhibit morphological changes typical of apoptosis, including condensed chromatin and a reduction in volume. ACT extract at 25-200 microg/ml dose-dependently inhibited the proliferation of SMMC-7721. The 50% effective dose, evaluated on day 3 of exposure to the extract, was 64.52+/-3.53 microg/ml. Upon gel electrophoresis, the fragmented DNA showed a characteristic ladder pattern. Cell cycle analyses revealed that ACT induced cell cycle arrest at the G0/G1 phase.

Predictive sensitivity of human cancer cells in vivo using semipermeable polysulfone fibers.

An in vivo experimental model was developed to predict efficiently and accurately chemosensitivity of human tumors. Human cancer cells either from cultured cell lines or from patients' tumors were injected directly into semipermeable polysulfone fibers subsequently implanted into immunocompetent rats. Results suggest utility of this novel model system for predicting tumor sensitivity to a wide range of anticancer agents and for potentially guiding the treatment of cancer patients in the clinical setting.

Phase I clinical and pharmacological studies of benzylacyclouridine, a uridine phosphorylase inhibitor.

Benzylacyclouridine (BAU, IND 039655) is a potent and specific inhibitor of uridine phosphorylase (UrdPase; EC 2.4.2.3). This enzyme plays a major role in regulating uridine homeostasis and also catalyzes the conversion of fluoropyrimidine nucleosides to their respective bases. Inhibition of UrdPase enzyme activity 18-24 h after 5-fluorouracil (5-FU) administration increased plasma levels of uridine and enhanced the therapeutic index of 5-FU by rescuing normal tissues. Moreover, in vitro preclinical studies have also shown that inhibiting UrdPase enzyme activity by BAU prior to administration of 5-FU increased cytotoxicity in a number of human cancer cell lines. A series of preclinical studies was performed in dogs and pigs to evaluate the pharmacological and pharmacodynamic properties of BAU. These data showed a sustained elevation in plasma uridine concentration in both animal models. The rapid degradation of a tracer dose of uridine into uracil was virtually arrested by BAU administered both p.o. or i.v. The t1/2 of BAU was 1.8-3.6 h in dogs, with bioavailability levels of 85% (30 mg/kg) and 42.5% (120 mg/kg). In pigs, the half-life varied from 1.6 to 2.3 h, with a bioavailability of 40% at 120 mg/kg. The drug was distributed into most tissues with a tissue: plasma ratio of approximately 0.7. On the basis of these preclinical studies, we performed a Phase I clinical trial of BAU in patients with advanced cancer. Patients received 200, 400, 800, and 1600 mg/m2 BAU as a single oral dose. Toxicities included grade 2 anemia, grade 1 fever, grade 1 fatigue, grade 1 constipation, and grade 1 elevation in alkaline phosphatase; none of these toxicities were observed to be dose dependent. The maximum tolerated dose and dose-limiting toxicity were not reached at the doses given. BAU plasma concentrations and area under the curve correlated linearly with the oral dose level. The pharmacokinetics of BAU were consistent with a first-order clearance, with average peak concentrations ranging from 19 microM (200 mg/m2) to 99 microM (1600 mg/m2) and tbeta1/2 ranging from 3.0 to 3.9 h at the four dose levels. Compared with baseline plasma uridine, treatment of patients with 200, 400, 800, and 1600 mg/m2 BAU increased peak uridine concentrations by 120, 150, 250, and 175%, respectively. On the basis of this clinical study, the suggested Phase II starting dose of BAU in combination with 5-FU is 800 mg/m2. Studies combining BAU with 5-FU and incorporating appropriate molecular and biochemical end points to assess the effects of this drug combination on tumor and/or surrogate tumor tissue are under way.

Benzylacyclouridine enhances 5-fluorouracil cytotoxicity against human prostate cancer cell lines.

At a nontoxic growth inhibitory concentration benzyloxyacyclouridine (BAU), a potent and specific inhibitor of uridine phosphorylase (UrdPase), enhanced 5-fluorouracil (5-FU) cytotoxic activity against human prostate cancer PC-3 and DU-145 cell lines. The BAU/5-FU combination exhibited greater antitumor activity in vivo using PC-3 human xenografts compared to 5-FU alone, with no associated increase in animal host toxicity. The mechanism(s) responsible for the enhanced in vitro and in vivo activity of this combination may involve enhanced formation of the 5-FU nucleotide metabolites FdUMP, FdUTP, and FUTP resulting in enhanced inhibition of thymidylate synthase (TS) and increased incorporation of fluoropyrimidine metabolites into tumoral RNA and DNA.

Inhibition of growth factor mitogenicity and growth of tumor cell xenografts by a sulfonated distamycin A derivative.

Interference with growth factor-receptor interactions may have particular relevance in efforts to intervene clinically in both autocrine and paracrine aspects of malignancy. Suramin is a synthetic anticancer agent that works, in part, by blocking the binding of growth factors to their receptors. While initial clinical trials have been encouraging, its use in clinical applications is associated with significant toxicities. Suradista is a novel sulfonated distamycin derivative that is also effective at complexing and inactivating growth factors and cytokines while remaining relatively nontoxic. The goal of this study was to compare the antineoplastic properties of suramin and Suradista. To achieve this, the effects of these compounds on growth factor induced mitogenesis in normal mouse fibroblasts and human umbilical vein endothelial cells were examined, as well as their ability to inhibit the growth of NIH/3T3 cells that had been transformed by the introduction of a fibroblast growth factor (FGF) 1 coding region (residues 1-154) fused to the signal peptide of the hst/KS3 gene (sp-hst/KS3:FGF1-154). In each case, Suradista was more effective than suramin in inhibiting mitogenesis in normal cells, as well as the growth of the transformed cells. Furthermore, Suradista was also shown to be as effective as suramin at inhibiting the growth of sp-hst/KS3:FGF1-154-transformed NIH/3T3 xenografts grown in athymic nude mice when given at only 50% the dosage used for suramin (50 mg/kg for Suradista versus 100 mg/kg for suramin). In summary, these results indicate that novel compounds acting like suramin may be developed as effective antineoplastic agents and may also prove to be of clinical benefit.

Expression of mRNA for brain-derived neurotrophic factor in the dorsal root ganglion following peripheral inflammation.

It is well known that the nerve growth factor (NGF) may serve as a link between inflammation and hyperalgesia. Recent experiments showed that systemic injection of NGF dramatically stimulated the expression of brain-derived neurotrophic factor (BDNF) mRNA in the dorsal root ganglion (DRG). In the present study, we evaluated the change of BDNF mRNA in the DRG following peripheral inflammation and also observed colocalization of BDNF and trkA mRNAs by means of in situ hybridization histochemistry in rats. Peripheral tissue inflammation produced by an intraplantar injection of Freund's adjuvant into the paws significantly increased BDNF mRNA levels in the DRG and many neurons expressing trkA mRNA showed increased expession of BDNF mRNA. Intraplantar injection of antibody to NGF together with Freund's adjuvant prevented the increase in BDNF mRNA. These findings suggest that peripheral inflammation induces an increased expression of BDNF mRNA which is mediated by NGF in DRG.

Enhancement of 5-fluoro-2'-deoxyuridine antineoplastic activity by 5-benzyloxybenzyloxybenzylacyclouridine in a human colon carcinoma cell line.

The pyrimidine acyclonucleoside benzyloxybenzyloxybenzylacyclouridine (BBBAU) showed growth inhibitory activity against the human colon cancer HCT-8 cell line with a 50% inhibitory concentration of 55 microM. Unlike its parent compounds, BBBAU was an extremely weak inhibitor of uridine phosphorylase. This acyclonucleoside analogue is an inhibitor of thymidylate synthase (TS) as determined by inhibition of [6-3H]-2'-deoxyuridine incorporation into DNA, inhibition of 3H release from [5-3H]-2'-deoxyuridine, and decrease in both the free and total TS 5'-fluoro-2'-deoxyuridine 5'-monophosphate binding sites. Kinetic analysis revealed that BBBAUMP, the monophosphate analogue of BBBAU, is a competitive inhibitor of purified human recombinant TS with a Ki of 8.0 microM. Nucleoside transport and uptake studies revealed that BBBAU (30 microM) inhibited the initial rate of transport and the total uptake of thymidine (25 microM). In contrast, while BBBAU (30 microM) inhibited the initial rate of transport of 5-fluoro-2'-deoxyuridine (FdUrd, 25 microM), its intracellular accumulation was increased. BBBAU (10 and 50 microM, respectively) potentiated FdUrd growth inhibition of HCT-8 cells and significantly enhanced the cytotoxic effects of FdUrd (0.3 and 1 microM, respectively) against HCT-8 cells using a clonogenic assay system. This combination resulted in additive inhibitory effects on TS activity resulting in greater depletion of dTTP pools. Moreover, the incorporation of radiolabeled FdUrd into the DNA fraction of HCT-8 cells was enhanced. The potential importance of this novel combination for human colon cancer chemotherapy is discussed.

Cell culture of the mucinous variant of human colorectal carcinoma.

Two cell lines, RW-2982 and RW-7213, have been established for the first time from the mucinous variant of human colorectal carcinoma, which is a distinctive and important subtype that has a worse prognosis than the more common nonmucogenic large bowel carcinoma. Methods of establishment and observations made during 7 and 3 years, respectively, of continuous culture are described. These cell lines required 4-9 months of adaptation to tissue culture conditions before noticeable growth occurred. Both cell lines have the following unique properties: (a) growth in vitro as delicate branching three-dimensional tumor particles within a wide gel of insoluble, often translucent mucus (proteoglycan); (b) production of large quantities of carcinoembryonic antigen; (c) ability to survive or adapt to growth in media free of serum, hormones, growth factors, and all protein; and (d) tumorigenicity in multiple sites in nude mice, including liver, with especially rapid growth in the peritoneal cavity as gelatinous material that is nonadherent and noninvasive and thus resembles pseudomyxoma peritonei. Unlike other reported colorectal cell lines, these mucus-coated particulate cell lines will not readily grow as monolayers and grow much more slowly with a doubling time of 2 weeks or more. A serially transplantable tumor from the RW-7213 surgical specimen has also been maintained in nude mice since August 8, 1984. This tumor retains properties of the original specimen. Observations made on the tumor biology of mucogenic colorectal carcinoma using these cell lines are discussed.

Inhibitors of purine nucleoside phosphorylase: effects of 9-deazapurine ribonucleosides and synthesis of 5'-deoxy-5'-iodo-9-deazainosine.

9-Deazapurine ribonucleosides constitute a new class of noncleavable purine nucleoside phosphorylase inhibitors that have at least 30-fold greater affinity for the enzyme than the corresponding C-nucleosides of the formycin B series. 9-Deazaguanosine, 9-deazainosine, and 5'-deoxy-5'-iodo-9-deazainosine competitively inhibited human erythrocytic purine nucleoside phosphorylase with Ki values of 29, 20, and 1.8 X 10(-7) M. The last compound is the most potent nucleoside inhibitor of the enzyme presently available and its synthesis is described. In contrast, 7,9-dideaza-7-thiainosine is a very weak inhibitor of the enzyme. When tested as an inhibitor of 2'-deoxyguanosine phosphorolysis in intact human erythrocytes and MOLT-3 human T-cell lymphoblastic leukemia cells, 5'-deoxy-5'-iodo-9-deazainosine was equipotent with 8-aminoguanosine (which is a precursor for 8-aminoguanine, Ki = 2 X 10(-7) M). Similarly, 5'-deoxy-5'-iodo-9-deazainosine and 8-aminoguanosine both potentiated the growth inhibition of human T-lymphocytic MOLT-3 cells by 2'-deoxyguanosine, reducing the 50% inhibitory concentration from approximately 2 X 10(-5) to approximately 2 X 10(-6) M.

Differential characteristics of two newly established human breast carcinoma cell lines.

Two human breast carcinoma cell lines, EP and MW, were established in culture from malignant pleural effusions. In addition to producing tumors in antithymocyte serum-immunosuppressed mice, both cell lines showed epithelial characteristics and anchorage-independent growth in soft agar. EP and MW differed in morphology (spindle-shaped versus round), chromosomal mode (hyperdiploid versus near triploid), estrogen receptor content (43.8 versus 5.1 fmol/mg protein), cloning efficiency (0.24 versus 15%), and activities (milliunits/10(6) cells) of creatine phosphokinase (25.7 versus 62.6) and lactate dehydrogenase (346.7 versus 778.5). Electron microscopy revealed that MW cells had more perinuclear filamentous material and more frequent intracytoplasmic vacuole formation than did EP cells. While having no effect on MW cells at the concentrations studied (10(-5) to 10(-11) M), beta-estradiol (10(-7) M) stimulated the growth of EP cells by 106% over the hormone-depleted control. In a variety of systems, EP was consistently the more drug sensitive of the two lines. In vitro, EP was significantly (p less than 0.001) more sensitive to methotrexate, vincristine, and 5-fluorouracil, respectively. In antithymocyte serum-mouse xenografts, EP displayed a greater response to three different dosages of a combination of cyclophosphamide, methotrexate, and 5-fluorouracil. One such dosage (cyclophosphamide, 32.0 mg/kg/day; methotrexate, 13.0 mg/kg/day; 5-fluorouracil, 190.0 mg/kg/day; for 1 day) reduced EP and MW tumor weights to 5.9 and 41% of controls, respectively. These results correlated well with the clinical responses.

5'-Deoxy-5'-methylthioadenosine phosphorylase--III. Role of the enzyme in the metabolism and action of 5'-halogenated adenosine analogs.

5'-Deoxy-5'-halogenated adenosines are alternative substrates for 5'-deoxy-5'-methylthioadenosine phosphorylase (MTAPase), an enzyme responsible for the metabolism of 5'-deoxy-5'-methylthioadenosine (MTA), a by-product of polyamine biosynthesis. The relative reactivity of these nucleosides with MTAPase from HL-60 human promyelocytic leukemia cells is MTA greater than 5'-deoxy-5'-fluoroadenosine (5'-FlAdo) greater than 5'-chloro-5'-deoxyadenosine (5'-ClAdo) greter than 5'-bromo-5'-deoxyadenosine (5'-BrAdo) greater than 5'-deoxy-5'-iodoadenosine (5'-IAdo). In MTAPase-containing cells, the adenine released from the 5'-halogenated adenosine was incorporated into adenine nucleotide pools; cleavage by (MTAPase appeared to be the rate-limiting step in this process. 5'-BrAdo and 5'-IAdo were growth inhibitors (EC50 values less than 10 microM) of MTAPase-containing cell lines (HL-60 human promyelocytic leukemia and the L5178Y murine lymphoblastic leukemia) but were much less active (EC50 values greater than 65 microM) against MTAPase-deficient cell lines (the CCRF-CEM human T cell leukemia and the L1210 murine leukemia). The full cytotoxicity of these compounds, therefore, appeared to be related to their phosphorolysis by MTAPase. Indirect evidence suggests that 5-halogenated ribose-1-phosphate derivatives of 5'-BrAdo or 5'-IAdo produced by the MTAPase reaction were the active metabolites of these 5'-halogenated adenosines.