Thyroid hormone may predict treatment failure in Kawasaki disease.

BACKGROUND: In systemic inflammatory conditions, inflammatory cytokines can cause low thyroid hormone levels. There are no reports discussing the relation between thyroid hormone levels and response to treatment for Kawasaki disease. METHODS: We investigated 67 patients who underwent treatment in the acute phase of Kawasaki disease. We divided patients into two groups based on their response to initial intravenous immunoglobulin (IVIG) treatment: the responder group (n = 40), and the non-responder group (n = 27). The serum levels of the thyroid hormones free triiodothyronine (FT3), free thyroxine (FT4), and thyroid-stimulating hormone (TSH) were compared before and after treatment in all patients, and between responder and non-responder groups. RESULTS: The FT3, FT4, and TSH levels were low before the initial treatment and increased significantly after treatment (p < 0.05). The FT3, FT4, and TSH levels before treatment were significantly lower in the non-responder group than in the responder group (p < 0.05). Logistic regression analysis suggested that the addition of pre-treatment FT4 values to Gunma score was useful in predicting treatment failure. CONCLUSIONS: Thyroid hormone and TSH levels were lower in the non-responder group than in the responder group in the initial IVIG treatment for Kawasaki disease. This study suggests that Kawasaki disease in the acute phase is associated with low thyroid hormone levels and TSH. It is possible that these hormone levels predict response to the initial IVIG.

M3 Muscarinic Receptor Signaling Stabilizes a Novel Mutant Human Ether-a-Go-Go-Related Gene Channel Protein via Phosphorylation of Heat Shock Factor 1 in Transfected Cells.

BACKGROUND: Long QT syndrome 2 (LQT2) is caused by mutations in the human ether-a-go-go-related gene (hERG). Most of its mutations give rise to unstable hERG proteins degraded by the proteasome. Recently, carbachol was reported to stabilize the wild-type hERG-FLAG via activation of the muscarinic type 3 receptor (M3-mAChR). Its action on mutant hERG-FLAG, however, remains uninvestigated.Methods and Results:A novel mutant hERG-FLAG carried 2 mutations: an amino acid substitution G572S and an in-frame insertion D1037_V1038insGD. When expressed in HEK293 cells, this mutant hERG-FLAG was degraded by the proteasome and failed to be transported to the cell surface. Carbachol restored stability of the mutant hERG-FLAG and facilitated cell-surface expression. Carbachol activated PKC, augmented phosphorylation of heat shock factor 1 (HSF1) and enhanced expression of heat shock proteins (hsps), hsp70 and hsp90. Both a M3-mAChR antagonist, 4-DAMP, and a PKC inhibitor, bisindolylmaleimide, abolished carbachol-induced stabilization of the mutant hERG-FLAG. CONCLUSIONS: M3-mAChR activation leads to enhancement of hsp expression via PKC-dependent phosphorylation of HSF1, thereby stabilizing the mutant hERG-FLAG protein. Thus, M3-mAChR activators may have a therapeutic value for patients with LQT2. (Circ J 2016; 80: 2443-2452).

Instability of KCNE1-D85N that causes long QT syndrome: stabilization by verapamil.

BACKGROUND: A KCNE1 polymorphism, D85N, causes long QT syndrome (LQTS) with a decrease in the slowly activating delayed-rectifier K(+) channel current (IKs ). We examined impacts of D85N polymorphism on KCNE1 protein stability and functions, and tested the ability of various drugs to modify them. METHODS: KCNE1-D85N or the wild-type protein was coexpressed in COS7 cells with KCNQ1 to form K(+) channels. Expression, degradation, and intracellular localization of KCNE1 proteins, as well as the currents conferred by KCNQ1/KCNE1 complexes, were determined using immunoblots, immunofluorescence, and patch-clamp techniques. RESULTS: The protein level of KCNE1-D85N was lower than that of the wild-type, in spite of the comparable levels of their mRNA. KCNE1-D85N was highly ubiquitinated and rapidly degraded as compared to the wild-type; a proteasome inhibitor, MG132, inhibited its degradation and increased its steady-state level. Both KCNE1-D85N and the wild-type proteins were co-immunoprecipitated with KCNQ1. Immunofluorescent signals of KCNE1-D85N accumulated in the endoplasmic reticulum and Golgi apparatus, with reduced levels on the cell membrane. Patch-clamp experiments demonstrated that the membrane current corresponding to IKs was much smaller in cells expressing KCNE1-D85N than in those expressing the wild-type. Verapamil (0.5-10 µM) increased the protein level of KCNE1-D85N, decreased its ubiquitination, slowed its degradation, and enhanced KCNQ1/KCNE1-D85N channel currents. Pretreatment with amiodarone abolished these effects of verapamil. CONCLUSION: KCNE1-D85N is less stable than the wild-type protein, and is rapidly degraded through the ubiquitin-proteasome system. Verapamil may be of a therapeutic value in LQTS patients via preventing degradation of KCNE1-D85N.

Screening of heat-killed lactic acid bacteria based on inhibitory activity against oral bacteria and effects of oral administration of heat-killed Ligilactobacillus salivarius CP3365 on periodontal health in healthy participants: a double-blinded, randomized, placebo-controlled trial.

Objectives: The aims of this study were to select heat-killed lactic acid bacteria (HKL) with antibiotic activity and investigate the efficacy of this bacteria in maintaining periodontal parameters in healthy participants. Materials and methods: An in vitro evaluation was conducted to assess the inhibitory efficacy of lactic acid bacteria against Porphyromonas gingivalis and Fusobacterium nucleatum subsp. nucleatum. The effects of HKL administration on various parameters (plaque control record, bleeding on probing, and probing pocket depth) were assessed in a randomized, placebo-controlled trial. Participants in the test and placebo groups (n = 32) consumed oral tablets containing placebo or HKL daily for 8 weeks. Oral bacteria in supra-plaque and saliva were identified using 16S rRNA gene community profiling analysis. Results: Heat-killed Ligilactobacillus salivarius CP3365 significantly (p < 0.05) decreased the viability of oral bacteria and was selected for clinical trials. Administration of HKL CP3365 significantly (p < 0.05) inhibited increases in each parameter. No changes in the relative abundance of P. gingivalis or F. nucleatum subsp. nucleatum were detected by HKL CP3365, but the relative abundance of oral bacteria (genera Porphyromonas, Fusobacterium, and Haemophilus) was significantly (p < 0.05) decreased. Conclusion: HKL CP3365 effectively inhibited oral bacteria growth and was useful for maintaining periodontal health. Clinical Trial Registration: [https://www.umin.ac.jp/ctr/index.htm], identifier [UMIN000045656].

Effects of cilnidipine on serum uric acid level and urinary nitrogen monoxide excretion in patients with hypertension.

The effects of cilnidipine on the serum uric acid level and urinary NO excretion in hypertensive patients were investigated. Blood and urine samples of 16 hypertensive outpatients were collected before and 2 months after cilnidipine therapy (10 mg). The serum uric acid level decreased significantly after cilnidipine treatment, while the uric acid-creatinine clearance ratio was unaffected. The cilnidipine medication produced a significant increase in urinary NO excretion, although amlodipine did not change it significantly. Therefore, cilnidipine has a profound antihypertensive effect and may reduce the serum uric acid level and increase NO production in the kidney.

A new microdeletion syndrome of 5q31.3 characterized by severe developmental delays, distinctive facial features, and delayed myelination.

Chromosomal deletion including 5q31 is rare and only a few patients have been reported to date. We report here the first two patients with a submicroscopic deletion of 5q31.3 identified by microarray-based comparative genomic hybridization. The common clinical features of both patients were marked hypotonia,feeding difficulty in infancy, severe developmental delay, and epileptic/nonepileptic encephalopathy associated with delayed myelination. Both patients also shared characteristic facial features,including narrow forehead, low-set and dysmorphic ears, bilateral ptosis, anteverted nares, long philtrum, tented upper vermilion,edematous cheeks, and high arched palate. The deleted region contains clustered PCDHs, including PCDHA [corrected]. and PCDHG, which are highly expressed in the brain where they function to guide neurons during brain development, neuronal differentiation, and synaptogenesis. The common deletion also contains neuregulin 2(NRG2), a major gene for neurodevelopment. We suggest that 5q31.3 deletion is responsible for severe brain developmental delay and distinctive facial features, and that the common findings in these two patients should be recognized as a new microdeletion syndrome. We need further investigations to determine which genes are really responsible for patients' characteristic features

Inhibitory Effect of the Glycerophosphate Moiety of Lipoteichoic Acid from Lactic Acid Bacteria on Dexamethasone-Induced Atrogin-1 Expression in C2C12 Myotubes.

Atrogin-1, which is an important regulator of ubiquitin-mediated protein degradation in skeletal muscle, is a major marker of muscle loss and disuse muscle atrophy. To investigate which components of lactic acid bacteria (LAB) suppress dexamethasone (DEX)-induced atrogin-1 expression, mouse skeletal muscle C2C12 myotubes were treated with DEX in the presence or absence of components of LAB. Heat-killed cells and lipoteichoic acid (LTA) derived from five LAB strains significantly suppressed DEX-induced atrogin-1 expression. The glycerophosphate (GroP) fraction prepared from chemically-degraded LTA and sn-glycerol-1-phosphate suppressed DEX-induced atrogin-1 expression, whereas the glycolipid anchor fraction of LTA did not. Heat-killed cells obtained by culturing under low-Mn2+ conditions, which generated fewer poly-GroP polymers in LTA, displayed significantly lower inhibitory activity compared to heat-killed cells grown under normal conditions. These results suggested that LTA of LAB contributed to suppressing atrogin-1 expression and that the GroP moiety of LTA was responsible for its inhibitory activity.

Synthesis and antiviral evaluation of α-D-2',3'-didehydro-2',3'-dideoxy-3'-C-hydroxymethyl nucleosides.

We have identified a selective S(N)2' reaction triggered by iodide ion that leads to the ring-opening of 2,2'-anhydro-α-nucleosides. By applying the method, we have synthesized α-D-2',3'-didehydro-2',3'-dideoxy-3'-C-hydroxymethyl nucleosides, designed as potential antiviral agents.

Fluctuating liver functions in siblings with MPV17 mutations and possible improvement associated with dietary and pharmaceutical treatments targeting respiratory chain complex II.

BACKGROUND/AIMS: To describe the clinical and biological findings of two Japanese siblings with novel MPV17 gene mutations (c.451insC/c.509C > T) manifesting hepatic mitochondrial DNA depletion syndrome. METHODS: We observed these brothers and sought to determine the efficacy of treatment targeting respiratory chain complex II for the younger brother. RESULTS: A 3-month-old boy had presented with profound liver dysfunction, failure to thrive, and watery diarrhea. Although he was then placed on a carbohydrate-rich diet, his liver function thereafter fluctuated greatly in association with viral infections, and rapidly deteriorated to liver failure. He underwent liver transplantation at 17 months of age but died at 22 months of age. The younger brother, aged 47 months at the time of this writing, presented with liver dysfunction from 8 months of age. His transaminase levels also fluctuated considerably fluctuations in association with viral infections. At 31 months of age, treatment with succinate and ubiquinone was initiated together with a lipid-rich diet using ketone milk. Thereafter, his transaminase levels normalized and never fluctuated, and the liver histology improved. CONCLUSIONS: These cases suggested that the clinical courses of patients with MPV17 mutations are greatly influenced by viral infections and that dietary and pharmaceutical treatments targeting the mitochondrial respiratory chain complex II may be beneficial in the clinical management of MPV17 mutant patients.

Diversity of mucosa-associated microbiota in active and inactive ulcerative colitis.

OBJECTIVE: Recent findings indicate that bacteria play an important role in the pathogenesis of inflammatory bowel disease (IBD). However, the exact role of bacteria in ulcerative colitis (UC) has still to be elucidated. The objective of the study was to investigate the potential differences in the intestinal microbiota between patients with UC and control subjects, using terminal restriction fragment length polymorphism (T-RFLP) analysis of the mucosa-associated microbiota from UC patients and non-IBD controls. MATERIAL AND METHODS: Nine active UC patients and 11 non-IBD controls were included in the study. Seven patients with active UC who entered into the inactive phase after antibiotic combination treatment were also classified as patients with inactive UC. Mucosa-associated microbiota was compared between non-IBD controls and UC patients using T-RFLP analysis. Microbiota in both the active and inactive phase was also analyzed in UC patients receiving antibiotic treatment. RESULTS: T-RFLP patterns of mucosa-associated microbiota differed between active UC patients and non-IBD controls. Microbial compositions of active UC patients were significantly less diverse. The difference resulted from loss of commensals. From the viewpoint of disease activity before and after antibiotic combination treatment, T-RFLP patterns were also different between the active and inactive phases in the identical patients. Inactive UC patients possessed more diverse microbial compositions. No specific terminal restriction fragments were observed in UC patients. CONCLUSIONS: T-RFLP analysis showed that the mucosa-associated microbiota of patients with active UC differed from that of non-IBD controls. Active UC patients possessed significantly fewer diverse microbial compositions.

Change of intestinal microbiota with elemental diet and its impact on therapeutic effects in a murine model of chronic colitis.

Elemental diet (ED) has been used as an enteral nutritional therapy for Crohn's disease. However, the precise mechanisms of ED remain unclear. In interleukin-10 (IL-10)-deficient cell-transferred mice, we investigated the change of intestinal microbiota with ED using molecular terminal-restriction fragment length polymorphism (T-RFLP) analysis and culture method, and evaluated its influence on therapeutic effects of ED. ED significantly suppressed intestinal inflammation. The total amount of bacteria in colitis mice fed the regular diet was higher than in normal mice but decreased in colitis mice fed ED. T-RFLP profiles of the ED group markedly differed from those of the regular diet groups. The diversity of bacterial species in the ED group decreased to 60% of that found in the regular diet groups. Among the cultivated bacteria, the change in lactic acid bacteria composition was remarkable. Lactobacillus reuteri and L. johnsonii decreased and Enterococcus faecalis and E. durans increased in the ED group. The culture supernatant of L. reuteri isolates induced significant tumor necrosis factor-alpha (TNF-alpha) and IL-6 activity in RAW 264 cells, while the culture supernatant of E. faecalis and E. durans barely induced their activity. These data suggested that reduction in amount and diversity of intestinal microbiota and decrease of proinflammatory cytokines via a change in composition of lactic acid bacteria by ED seem to contribute to reduction of bowel inflammation in this model.

Lactobacillus curvatus CP2998 Prevents Dexamethasone-Induced Muscle Atrophy in C2C12 Myotubes.

To investigate whether heat-killed Lactobacillus curvatus CP2998 (CP2998) inhibits glucocorticoid-induced myotube atrophy which is associated with the ubiquitin-proteasome system, mouse skeletal muscle C2C12 myotubes were treated with dexamethasone (DEX) in the presence or absence of CP2998. DEX exposure significantly decreased myotube diameters and increased mRNA expression levels of MuRF1 and MAFbx, E3 ubiquitin ligases. CP2998 treatment restored myotube diameters and dose dependently decreased mRNA expression levels of these E3 ubiquitin ligases. CP2998 treatment also inhibited DEX-induced glucocorticoid dependent transcription. Our results suggest that CP2998 prevents DEX-induced muscle atrophy by suppressing glucocorticoid receptor activation.

Terminal restriction fragment length polymorphism analysis of the diversity of fecal microbiota in patients with ulcerative colitis.

BACKGROUND: Terminal restriction fragment length polymorphism (T-RFLP) analysis is a powerful tool to assess the diversity of complexed microbiota. This permits rapid comparison of microbiota from many samples. In this study, we performed T-RFLP analysis of the fecal microbiota from patients with ulcerative colitis (UC). METHODS: Forty-four patients with UC (23 women and 21 men, median age 25 years) and 46 healthy individuals (25 women and 21 men, median age 34 years) were enrolled in this study. DNA was extracted from their stool samples, and the 16S rRNA genes were amplified by PCR. The PCR products were then digested with HhaI and/or MspI restriction enzymes, and the length of the T-RF was determined. RESULTS: The fecal microbial communities were classified in 8 clusters. Almost all the healthy individuals (39 of 46) were included in cluster I, and most of the UC patients could be divided into the other 7 clusters, indicating that fecal bacterial communities are different between healthy individuals and active UC patients. Some T-RFs, derived from the unclassified bacteria, Ruminococcus, Eubacterium, Fusobacterium, gammaproteobacteria, unclassified Bacteroides, and unclassified Lactobacillus, were detected in the UC patients, but not in the healthy individuals. The T-RFLP patterns were also different between the active patients and inactive (remission) patients. The T-RF derived from the unclassified bacteria, Ruminococcus and Eubacterium, and the T-RFs derived from the unclassified bacteria, Eubacterium, and Fusobacterium were predominantly detected in the active patients not the inactive patients. In contrast, the T-RFs derived from Lactobacillus and unclassified Lactobacillus were more predominant in the inactive (remission) patients. In 4 patients with proctitis, the pattern of fecal microbial diversity was very similar. CONCLUSIONS: T-RFLP analyses showed that the diversity of fecal microbiota in patients with UC was different from that in healthy individuals. Unclassified bacteria, as well as known bacteria, can contribute to alterations in the bacterial diversity of UC patients.

Characterization of the novel mutant A78T-HERG from a long QT syndrome type 2 patient: Instability of the mutant protein and stabilization by heat shock factor 1.

BACKGROUND: The human ether-a-go-go-related gene (HERG) encodes the α-subunit of rapidly activating delayed-rectifier potassium channels. Mutations in this gene cause long QT syndrome type 2 (LQT2). In most cases, mutations reduce the stability of the channel protein, which can be restored by heat shock (HS). METHODS: We identified the novel mutant A78T-HERG in a patient with LQT2. The purpose of the current study was to characterize this mutant protein and test whether HS and heat shock factors (HSFs) could stabilize the mutant protein. A78T-HERG and wild-type HERG (WT-HERG) were expressed in HEK293 cells and analyzed by immunoblotting, immunoprecipitation, immunofluorescence, and whole-cell patch clamping. RESULTS: When expressed in HEK293 cells, WT-HERG gave rise to immature and mature forms of the protein at 135 and 155 kDa, respectively. A78T-HERG gave rise only to the immature form, which was heavily ubiquitinated. The proteasome inhibitor MG132 increased the expression of immature A78T-HERG and increased both the immature and mature forms of WT-HERG. WT-HERG, but not A78T-HERG, was expressed on the plasma membrane. In whole-cell patch clamping experiments, depolarizing pulses evoked E4031-sensitive HERG channel currents in cells transfected with WT-HERG, but not in cells transfected with A78T-HERG. The A78V mutant, but not A78G mutant, remained in the immature form similarly to A78T. Maturation of the A78T-HERG protein was facilitated by HS, expression of HSF-1, or exposure to geranyl geranyl acetone. CONCLUSIONS: A78T-HERG was characterized by protein instability and reduced expression on the plasma membrane. The stability of the mutant was partially restored by HSF-1, indicating that HSF-1 is a target for the treatment for LQT2 caused by the A78T mutation in HERG.

Culture-independent analysis of fecal microbiota in infants, with special reference to Bifidobacterium species.

Fecal microbiota of 31 breast-fed, 26 mix-fed, and 11 bottle-fed infants were analyzed by using terminal restriction fragment length polymorphism (T-RFLP), and culture method. We first determined the total and cultivated bacterial counts in infant fecal microbiota. Only approximately 30% of bacteria present in fecal microbiota were cultivable while the remainder was yet-to-be cultured bacteria. Sixty-eight fecal samples were divided into two clusters (I and II) by T-RFLP analysis, and then subdivided into five subclusters (Ia, Ib, IIa, IIb and IIc). There was no clear relationship between clusters and feeding method. A proportion of bifidobacteria was detected in the fecal material by PCR method using species-specific primers. The predominant Bifidobacterium spp. was Bifidobacterium longum longum type (43 samples (63.2%)), followed by B. longum infantis type (23 samples (33.8%)) and B. breve (16 samples (23.5%)). The distribution of Bifidobacterium spp. was similar in the three feeding groups. In contrast, the high incidence of B. breve in cluster I, especially subcluster Ia and B. longum longum type in cluster II, especially subcluster IIa and IIc were characterized by T-RFLP method. Our results showed that the colonization of Bifidobacterium spp. in infant feces correlated with the T-RFLP clusters.

Attempt to reduce cytotoxicity by synthesizing the L-enantiomer of 4'-C-ethynyl-2'-deoxypurine nucleosides as antiviral agents against HIV and HBV.

We investigated the potential of 4'-C-substituted nucleosides for the treatment of HIV-1 and HBV. Of the nucleosides we prepared, several 4'-C-ethynyl-2'-deoxypurine nucleosides showed the most potent anti-HIV activity. However, two candidates, 4'-C-ethynyl-2'-deoxyguanosine and 9-(2-deoxy-4-C-ethynyl-beta-D-ribo-pentofuranosyl)-2,6-diaminopurine, were very toxic during in vivo study. On the other hand, lamivudine (3TC) is known to show remarkable activity against HIV and HBV with lower cytotoxicity. Therefore, we attempted to synthesize the L-enantiomer of 4'-C-ethynyl-2'-deoxypurine nucleosides in 20-21 steps. These methods consisted of preparing 4-C-ethynyl-L-sugar, starting from D-arabinose and then condensing the L-sugar derivative with 2,6-diaminopurine. 4'-C-Ethynyl-2'-deoxyguanosine was also prepared by enzymatic deamination from the 2,6-diaminopurine derivative. The compounds' antiviral activity against HIV and HBV was then evaluated. Unfortunately, they demonstrated no activity and no cytotoxicity.

Synthesis of Novel Iso-4'-thionucleosides Using the Mitsunobu Reaction.

A novel class of isomeric 4'-thionucleosides with the base moiety at the 2'-position was synthesized from D-glucose. The coupling of 1,4-anhydro-4-thioarabitol (13) with various nucleobases using the Mitsunobu reaction was investigated. With both purines and N(3)-benzoyluracils, the reaction predominantly gave beta-isomers, suggesting that these were produced via an episulfonium intermediate. The beta-anomers produced by the reaction of N(3)-benzoyluracils included both N- and O-alkylated derivatives. Interestingly, only the reaction of N(3)-benzoyluracil gave a mixture of N-alkylated adduct (20d) and O-alkylated bipyrimidinyl adduct (22), the structure of which was unambiguously determined by NMR spectroscopic data including HMBC and NOE. Deprotection of the Mitsunobu reaction products gave the desired iso-4'-thionucleosides.

A Novel Synthesis of 2'-Modified 2'-Deoxy-4'-thiocytidines from D-Glucose(1).

Novel 2'-deoxycytidine antimetabolites, specifically several 2'-modified 2'-deoxy-4'-thiocytidines, were synthesized as potential new antineoplastic agents. Methyl 3-O-benzylxylofuranoside was converted to a 1,4-anhydro-4-thioarabitol 24. Protection of the primary alcohol of 24 gave a common intermediate (15) which was useful for the synthesis of various 2'-modified 2'-deoxy-4'-thionucleosides. Oxidation of the secondary hydroxyl group of 15, followed by the Wittig reaction or treatment with (diethylamido)sulfur trifluoride (DAST) produced 2-deoxy-2-methylene (26) and 2-deoxy-2,2-difluoro (34) derivatives, respectively. Unique Pummerer-type glycosylation between the corresponding sulfoxides and trimethylsilylated N(4)-acetylcytosine produced 2'-deoxy-2'-methylene- (10) and 2'-deoxy-2',2'-difluoro-4'-thiocytidines (11). On the other hand, treatment of 15 with DAST introduced a fluorine atom with retention of the 2'-stereochemistry, yielding 40. In contrast, the Mitsunobu reaction of 3-O-benzoyl derivative 53 which was obtained from 15 in five steps, using diphenylphosphoryl azide gave azide derivative 54 with inverted stereochemistry. These derivatives were converted to the corresponding 1-O-acetyl derivatives via the usual Pummerer rearrangement, which were in turn used to synthesize 4'-thiocytidines 12 and 58. Among the 2'-modified 4'-thiocytidines obtained, 2'-methylene (10) and 2'-fluoro (12) derivatives were found to have potent antineoplastic properties in vitro.

Synthesis of 4'-C-ethynyl and 4'-C-cyano purine nucleosides from natural nucleosides and their anti-HIV activity.

Purine 2'-deoxynucleosides bearing an ethynyl or a cyano group at C-4' of the sugar moiety were synthesized from the corresponding 2'-deoxynucleosides. These compounds exhibited very potent anti-HIV activity, and remained active against drug resistant HIV strains.

Design, efficient synthesis, and anti-HIV activity of 4'-C-cyano- and 4'-C-ethynyl-2'-deoxy purine nucleosides.

Some 4'-C-ethynyl-2'-deoxy purine nucleosides showed the most potent anti-HIV activity among the series of 4'-C-substituted 2'-deoxynucleosides whose 4'-C-substituents were methyl, ethyl, ethynyl and so on. Our hypothesis is that the smaller the substituent at the C-4' position they have, the more acceptable biological activity they show. Thus, 4'-C-cyano-2'-deoxy purine nucleosides, whose substituent is smaller than the ethynyl group, will have more potent antiviral activity. To prove our hypothesis, we planned to develop an efficient synthesis of 4'-C-cyano-2'-deoxy purine nucleosides (4'-CNdNs) and 4'-C-ethynyl-2'-deoxy purine nucleosides (4'-EdNs). Consequently, we succeeded in developing an efficient synthesis of six 2'-deoxy purine nucleosides bearing either a cyano or an ethynyl group at the C-4' position of the sugar moiety from 2'-deoxyadenosine and 2,6-diaminopurine 2'-deoxyriboside. Unfortunately, 4'-C-cyano derivatives showed lower activity against HIV-1, and two 4'-C-ethynyl derivatives suggested high toxicity in vivo.