Pharmacogenetic determinants of tenofovir diphosphate and lamivudine triphosphate concentrations in people with HIV/HBV coinfection.

The nucleos(t)ide analogs require phosphorylation to the pharmacologically active anabolites in cells. We investigated the hypothesis that single-nucleotide polymorphisms (SNPs) in genes that encode transporters and phosphodiesterase (PDE) enzymes involved in tenofovir (TFV), disoproxil fumarate (TDF), and lamivudine (3TC) disposition will be associated with concentrations of their phosphate anabolites and virologic response. Individuals with human immunodeficiency virus (HIV) and hepatitis B virus (HBV) coinfection receiving TDF/3TC-containing antiretroviral therapy were enrolled. Steady-state TFV diphosphate (TFV-DP) and 3TC triphosphate (3TC-TP) concentrations in peripheral blood mononuclear cells (PBMCs) and dried blood spot samples were quantified. The relationship between genetic variants and TFV-DP and 3TC-TP concentrations as well as with virologic response were examined using multivariable linear regression. Of the 136 participants (median age 43 years; 63% females), 6.6% had HBV non-suppression, and 7.4% had HIV non-suppression. The multidrug resistance protein 2 (encoded by ABCC2 rs2273697) SNP was associated with 3TC-TP concentrations in PBMCs. The human organic anion transporter-1 (encoded by SLC28A2) rs11854484 SNP was associated with HIV non-suppression, and when evaluated together with SNPs with marginal associations (ABCC2 rs717620 and PDE1C rs30561), participants with two or three variants compared to those with none of these variants had an adjusted odds ratio of 48.3 (confidence interval, 4.3-547.8) for HIV non-suppression. None of the SNPs were associated with HBV non-suppression. Our study identified ABCC2 SNP to be associated with 3TC-TP concentrations in PBMCs. Also, a combination of genetic variants of drug transporters and PDE was associated with HIV non-suppression.

Real-world effectiveness and safety of switching to dolutegravir/lamivudine among people living with HIV-1 aged over 50 years who are virologically suppressed.

OBJECTIVES: People living with HIV face several challenges as they age, including the potential for polypharmacy and increased susceptibility to drug-related adverse effects. Thus, effective and well-tolerated regimens with minimal or no drug interactions would be useful in this population. We present real-world effectiveness and safety data for individuals aged >50 years who achieved virological suppression (HIV-1 RNA <50 copies/mL) and switched to dolutegravir/lamivudine (DTG/3TC). METHODS: This retrospective, observational, single-centre study conducted in Portugal included individuals aged >50 years who switched to DTG/3TC while virologically suppressed and had ≥12 months of follow-up. Proportions of individuals maintaining virological suppression were described at 12 months; CD4+ cell counts were described at baseline and 12 months. Descriptive subgroup analyses were performed based on age, sex assigned at birth, and availability of historical genotypic resistance results. RESULTS: Overall, 538 individuals aged >50 years were included (74% male; mean age, 62 years; mean time on previous therapy, 160 months). High proportions (intention-to-treat population, 97%; on-treatment population, 98%) of individuals who switched to DTG/3TC maintained virological suppression through 12 months of follow-up. CD4+ cell counts remained stable (mean baseline: 727 cells/mm3 [range 94-2371]; mean month 12: 742 cells/mm3 [range 99-2659]). No individuals experienced virological failure. Nine (2%) individuals discontinued DTG/3TC for non-treatment-related reasons. Proportions with virological suppression at month 12 were similar between on-treatment subgroups by age, sex assigned at birth, and historical genotypic resistance results availability. CONCLUSIONS: DTG/3TC demonstrated robust effectiveness and a good safety profile in individuals aged >50 years with virological suppression in Portugal.

Efficacy and safety of dolutegravir/lamivudine in virologically suppressed female participants: week 48 data from the pooled TANGO and SALSA studies.

OBJECTIVES: Women represent >50% of people with HIV globally but have historically been underrepresented in clinical trials. We evaluated the efficacy and safety of switching to dolutegravir/lamivudine (DTG/3TC) vs continuing their current antiretroviral regimen (CAR) by sex assigned at birth (female and male) in virologically suppressed adults with HIV-1 without prior virological failure in a pooled analysis of two randomized controlled trials. METHODS: This analysis included 48-week data from the phase 3 TANGO and SALSA studies. Primary and key secondary endpoints included proportions of participants with HIV-1 RNA ≥50 and <50 copies/mL at week 48, respectively. Safety was also assessed. RESULTS: Of 1234 participants, 250 (DTG/3TC, n = 133; CAR, n = 117) were female at birth. Week 48 proportions of participants with Snapshot HIV-1 RNA ≥50 copies/mL were similar regardless of sex at birth (DTG/3TC vs CAR: female, <1% [1/133] vs 2% [2/117]; male, <1% [1/482] vs <1% [3/502]). Proportions with HIV-1 RNA <50 copies/mL were high across sexes and treatment groups (DTG/3TC vs CAR: female, 91% [121/133] vs 89% [104/117]; male, 94% [455/482] vs 94% [471/502]). Immunological response with DTG/3TC was slightly higher in female participants. Incidences of adverse events leading to withdrawal and serious adverse events were low and comparable between treatment groups and across sexes. Weight gain was higher with DTG/3TC than with CAR among female participants aged ≥50 years (treatment difference 2.08 kg [95% confidence interval 0.40-3.75]). CONCLUSIONS: Results confirm the robustness of DTG/3TC as a switch option in virologically suppressed females with HIV-1, with outcomes similar to those in males.

Efficacy and safety of switching to dolutegravir/lamivudine in virologically suppressed people with HIV-1 aged ≥ 50 years: week 48 pooled results from the TANGO and SALSA studies.

BACKGROUND: As the population of people with HIV ages, concerns over managing age-related comorbidities, polypharmacy, immune recovery, and drug-drug interactions while maintaining viral suppression have arisen. We present pooled TANGO and SALSA efficacy and safety results dichotomized by age (< 50 and ≥ 50 years). METHODS: Week 48 data from the open-label phase 3 TANGO and SALSA trials evaluating switch to once-daily dolutegravir/lamivudine (DTG/3TC) fixed-dose combination vs continuing current antiretroviral regimen (CAR) were pooled. Proportions of participants with HIV-1 RNA ≥ 50 and < 50 copies/mL (Snapshot, intention-to-treat exposed) and safety were analyzed by age category. Adjusted mean change from baseline in CD4 + cell count was assessed using mixed-models repeated-measures analysis. RESULTS: Of 1234 participants, 80% of whom were male, 29% were aged ≥ 50 years. Among those aged ≥ 50 years, 1/177 (< 1%) DTG/3TC participant and 3/187 (2%) CAR participants had HIV-1 RNA ≥ 50 copies/mL at 48 weeks; proportions with HIV-1 RNA < 50 copies/mL were high in both treatment groups (≥ 92%), consistent with overall efficacy and similar to observations in participants aged < 50 years (≥ 93%). Regardless of age category, CD4 + cell count increased or was maintained from baseline with DTG/3TC. Change from baseline in CD4 + /CD8 + ratio was similar across age groups and between treatment groups. One CAR participant aged < 50 years had confirmed virologic withdrawal, but no resistance was detected. In the DTG/3TC group, incidence of adverse events (AEs) was similar across age groups. Proportions of AEs leading to withdrawal were low and comparable between age groups. Although drug-related AEs were generally low, across age groups, drug-related AEs were more frequent in participants who switched to DTG/3TC compared with those who continued CAR. While few serious AEs were observed in both treatment groups, more were reported in participants aged ≥ 50 years vs < 50 years. CONCLUSIONS: Among individuals with HIV-1, switching to DTG/3TC maintained high rates of virologic suppression and demonstrated a favorable safety profile, including in those aged ≥ 50 years despite higher prevalence of concomitant medication use and comorbidities. TRIAL REGISTRATION NUMBER: TANGO, NCT03446573 (February 27, 2018); SALSA, NCT04021290 (July 16, 2019).

Treatment persistence of bictegravir/emtricitabine/tenofovir alafenamide and efavirenz + lamivudine + tenofovir disoproxil among HIV-1 patients newly starting treatment in Hunan Province in China.

BACKGROUND: Though bictegravir/emtricitabine/tenofovir (BIC/FTC/TAF) have been regulatory approved and included in the National Reimbursement Drug List in China, due to the affordability concern, generic version of efavirenz + lamivudine + tenofovir (EFV + 3TC + TDF) is still recommended as the first-line therapy in the clinical guideline and widely used in clinical practice. The aim of the study is to assess the persistence with first-line BIC/TAF/TAF and EFV + 3TC + TDF in newly treated HIV-1 patients in the real-world setting in Hunan Province in China. METHODS: A retrospective analysis of the medical records of HIV patients initiating first-line antiretroviral therapy in the First Hospital of Changsha in January 1st, 2021-July 31st, 2022 was conducted. Persistence was assessed as the number of days on the therapy from the index until treatment discontinuation or end of data availability. Kaplan-Meier Curves and Cox Proportional Hazard models were used to evaluate the discontinuation rates. Subgroup analysis was performed excluding BIC/FTC/TAF patients with treatment discontinuation due to economic reason, and EFV + 3TC + TDF patients with a viral load > 500,000 copies/mL. RESULTS: A total of 310 eligible patients were included in the study, with 244 and 66 patients in the BIC/FTC/TAF group and EFV + 3TC + TDF group, respectively. Compared with EFV + 3TC + TDF patients, BIC/FTC/TAF patients were older, more living in the capital city currently, and had significantly higher total cholesterol and low-density level (all p < 0.05). No significant difference was shown in the time to discontinuation between BIC/FTC/TAF patients and EFV + 3TC + TDF patients. After excluding BIC/FTC/TAF patients with treatment discontinuation due to economic reason, EFV + 3TC + TDF group were shown to have a significantly higher risk of discontinuation than BIC/FTC/TAF group (hazard ratio [HR] = 11.1, 95% confidence interval [CI] = 1.3-93.2). After further removing the EFV + 3TC + TDF patients with a viral load > 500,000 copies/mL, the analysis showed similar results (HR = 10.1, 95% CI = 1.2-84.1). 79.4% of the EFV + 3TC + TDF patients discontinued treatment due to clinical reasons, while 83.3% of the BIC/FTC/TAF patients discontinued treatment due to economic reasons. CONCLUSIONS: Compared with BIC/FTC/TAF, EFV + TDF + 3TC patients were significantly more likely to discontinue the first-line treatment in Hunan Province in China.

Evaluation of Pathogenicity and Causativity of Variants in the MPZ and SH3TC2 Genes in a Family Case of Hereditary Peripheral Neuropathy.

The implementation of NGS methods into clinical practice allowed researchers effectively to establish the molecular cause of a disorder in cases of a genetically heterogeneous pathology. In cases of several potentially causative variants, we need additional analysis that can help in choosing a proper causative variant. In the current study, we described a family case of hereditary motor and sensory neuropathy (HMSN) type 1 (Charcot-Marie-Tooth disease). DNA analysis revealed two variants in the SH3TC2 gene (c.279G>A and c.1177+5G>A), as well as a previously described variant c.449-9C>T in the MPZ gene, in a heterozygous state. This family segregation study was incomplete because of the proband's father's unavailability. To evaluate the variants' pathogenicity, minigene splicing assay was carried out. This study showed no effect of the MPZ variant on splicing, but the c.1177+5G>A variant in the SH3TC2 gene leads to the retention of 122 nucleotides from intron 10 in the RNA sequence, causing a frameshift and an occurrence of a premature stop codon (NP_078853.2:p.Ala393GlyfsTer2).

The effect of iron on Trichomonas vaginalis TvCP2: a cysteine proteinase found in vaginal secretions of trichomoniasis patients.

Trichomonas vaginalis (Tv) induces host cell damage through cysteine proteinases (CPs) modulated by iron. An immunoproteomic analysis showed that trichomoniasis patient sera recognize various CPs, also some of them are present in vaginal washes (VWs). Thus, the goal of this work was to determine whether TvCP2 is expressed during infection and to assess the effect of iron on TvCP2 expression, localization and contribution to in vitro cellular damage. Western-blotting (WB) assays using TvCP2r and vaginitis patient serum samples showed that 6/9 Tv (+) but none of the Tv (-) patient sera recognized TvCP2r. WB using an anti-TvCP2r antibody and VWs from the same patients showed that in all of the Tv (+) but none of the Tv (-) VWs, the anti-TvCP2r antibody detected a 27 kDa protein band that corresponded to the mature TvCP2, which was confirmed by mass spectrometry analysis. Iron decreased the amount of TvCP2 mRNA and the protein localized on the parasite surface and cytoplasmic vesicles concomitant with the cytotoxic effect of TvCP2 on HeLa cells. Parasites pretreated with the anti-TvCP2r antibody also showed reduced levels of cytotoxicity and apoptosis induction in HeLa cell monolayers. In conclusion, these results show that TvCP2 is expressed during trichomonal infection and plays an important role in the in vitro HeLa cell cytotoxic damage under iron-restricted conditions.

A long-acting 3TC ProTide nanoformulation suppresses HBV replication in humanized mice.

Nowadays, there is a strong request for the treatment of chronic HBV-infection with direct acting antivirals. Furthermore, prevalent human immunodeficiency virus (HIV-1) and hepatitis B (HBV) co-infections highlight an immediate need for dual long-acting and easily administered antivirals. To this end, we modified lamivudine (3TC), a nucleoside analog inhibitor of both viruses, into a lipophilic monophosphorylated prodrug (M23TC). Prior work demonstrated that nanoformulation of M23TC (NM23TC) enhanced drug stability, controlled dissolution and improved access to sites of viral replication. The present study evaluated the efficacy of a NM23TC in HBV-infected chimeric liver humanized mice. Levels of HBV DNA and HBsAg in plasma were monitored up to 8 weeks posttreatment. A single intramuscular dose of 75 mg/kg 3TC equivalents of nanoformulated NM23TC provided sustained drug levels and suppressed HBV replication in humanized mice for 4 weeks. The results support further development of this long-acting 3TC nanoformulation for HBV treatment and prevention.

Assessment of complex genomic alterations induced by AZT, 3TC, and the combination AZT +3TC.

Highly active antiretroviral therapy (HAART) regimens are based on the use of nucleoside reverse transcriptase inhibitors (NRTIs), which are the main drugs used by patients infected with the human immunodeficiency virus (HIV). The use of NRTIs combinations has afforded clear clinical benefits to patients undergoing HAART. However, the combination of two NRTIs may increase the risk of genomic instability in comparison with the drugs administered individually. We analyzed the ability of zidovudine (AZT) and lamivudine (3TC), and the combination AZT +3TC to induce complex genomic alterations using the cytokinesis-block micronucleus (CBMN) assay in Chinese hamster ovary (CHO)-K1 cells. The 24-h cell treatment with individual NRTIs showed that AZT increased micronucleus frequencies and nucleoplasmic bridges (NPBs). No significant differences were observed for any parameters investigated after exposure of CHO-K1 cells to 3TC. The combination AZT +3TC significantly increased micronucleus frequencies. Analysis of interaction between these drugs suggested that antagonism occurs in all AZT +3TC concentrations. These results highlight the importance to investigate the genotoxic profile of NRTIs to develop safer intervention strategies in antiretroviral treatment protocols.

Mutational screening of the SH3TC2 gene in Greek patients with suspected demyelinating recessive Charcot-Marie-Tooth disease reveals a varied and unusual phenotypic spectrum.

Charcot-Marie-Tooth disease type 4 C (CMT4C) is an autosomal recessive form of demyelinating peripheral neuropathy caused by mutations in SH3TC2, characterized by early onset, spine deformities, and cranial nerve involvement. We screened SH3TC2 in 50 unrelated Greek patients with suspected demyelinating Charcot-Marie-Tooth disease and pedigree compatible with recessive inheritance. All patients had been previously screened for PMP22, GJB1, and MPZ mutations. We found five previously identified pathogenic mutations in SH3TC2 distributed among 13 patients in homozygosity or compound heterozygosity (p. Arg954Stop, Arg1109Stop, Gln892Stop, Ala878Asp, and Arg648Trp). Although most cases had early onset and spine deformities were almost omnipresent, a wide phenotypic spectrum was observed. Particularly notable were two siblings with Roussy-Lévy syndrome and one patient with young-onset trigeminal neuralgia. In conclusion, mutations in SH3TC2 are responsible for 26% of Greek patients with suspected CMT4, identifying CMT4C as the most common recessive demyelinating neuropathy in the Greek population, in accordance with other Mediterranean cohorts.

Attenuating Diabetic Vascular and Neuronal Defects by Targeting P2rx7.

Retinal vascular and neuronal degeneration are established pathological features of diabetic retinopathy. Data suggest that defects in the neuroglial network precede the clinically recognisable vascular lesions in the retina. Therefore, new treatments that target early-onset neurodegeneration would be expected to have great value in preventing the early stages of diabetic retinopathy. Here, we show that the nucleoside reverse transcriptase inhibitor lamivudine (3TC), a newly discovered P2rx7 inhibitor, can attenuate progression of both neuronal and vascular pathology in diabetic retinopathy. We found that the expression of P2rx7 was increased in the murine retina as early as one month following diabetes induction. Compared to non-diabetic controls, diabetic mice treated with 3TC were protected against the formation of acellular capillaries in the retina. This occurred concomitantly with a maintenance in neuroglial function, as shown by improved a- and b-wave amplitude, as well as oscillatory potentials. An improvement in the number of GABAergic amacrine cells and the synaptophysin-positive area was also observed in the inner retina of 3TC-treated diabetic mice. Our data suggest that 3TC has therapeutic potential since it can target both neuronal and vascular defects caused by diabetes.

Charcot-Marie-Tooth Disease type 4C: Novel mutations, clinical presentations, and diagnostic challenges.

INTRODUCTION: This study analyzes and describes atypical presentations of Charcot-Marie-Tooth disease type 4C (CMT4C). METHODS: We present clinical and physiologic features of 5 patients with CMT4C caused by biallelic private mutations of SH3TC2. RESULTS: All patients manifested scoliosis, and nerve conduction study indicated results in the demyelinating range. All patients exhibited signs of motor impairment within the first years of life. We describe 2 or more different genetic diseases in the same patient, atypical presentations of CMT, and 3 new mutations in CMT4C patients. DISCUSSION: A new era of unbiased genetic testing has led to this small case series of individuals with CMT4C and highlights the recognition of different genetic diseases in CMT4C patients for accurate diagnosis, genetic risk identification, and therapeutic intervention. The phenotype of CMT4C, in addition, appears to be enriched by a number of features unusual for the broad CMT category. Muscle Nerve 57: 749-755, 2018.

Puerarin inhibits TRPM3/miR-204 to promote MC3T3-E1 cells proliferation, differentiation and mineralization.

Puerarin is an isoflavonoid phytoestrogen extracted from the root of Radix Pueraria, has attracted increasing attention because of its beneficial effects on anti-osteoporosis, but the molecular mechanisms underlying its actions on osteoblasts are not fully understood. The current study aimed to investigate the effect of puerarin on MC3T3-E1 osteoblastic cells proliferation, differentiation and mineralization, in vitro and its underlying mechanisms. The results indicated that puerarin significantly promoted the osteoblasts proliferation, enhanced alkaline phosphatase activity and increased the formation of mineralized nodules. Following treatment with puerarin, the expression levels of transient receptor potential Melastatin 3 (TRPM3) and microRNA-204 (miR-204) were decreased, whereas that of Runt-related transcription Factor 2 (Runx2) increased. TRPM3-small interfering RNA and 2-aminoethoxydiphenyl borate (2-APB, inhibitor of TRPM3) promoted the expression of Runx2 and thus improved the development of osteoblasts, but pregnenolone sulfate, which is the agonist of TRPM3, inhibited the effects. In addition, puerarin induced the changes of intracellular Ca2+ concentration ([Ca2+ ]i ) and extracellular Ca2+ concentration ([Ca2+ ]0 ) through TRPM3 might be involved in the biological process of MC3T3-E1 cells. These results suggested that puerarin may promote MC3T3-E1 cell proliferation, differentiation and mineralization, which may be related to the downregulation of TRPM3/miR-204 and following regulating [Ca2+ ]i and [Ca2+ ]0 , and activation of Runx2.

Neuromuscular Junction Changes in a Mouse Model of Charcot-Marie-Tooth Disease Type 4C.

The neuromuscular junction (NMJ) appears to be a site of pathology in a number of peripheral nerve diseases. Charcot-Marie-Tooth (CMT) 4C is an autosomal recessive, early onset, demyelinating neuropathy. Numerous mutations in the SH3TC2 gene have been shown to underlie the condition often associated with scoliosis, foot deformities, and reduced nerve conduction velocities. Mice with exon 1 of the Sh3tc2 gene knocked out demonstrate many of the features seen in patients. To determine if NMJ pathology is contributory to the pathomechanisms of CMT4C we examined NMJs in the gastrocnemius muscle of SH3TC2-deficient mice. In addition, we performed proteomic assessment of the sciatic nerve to identify protein factors contributing to the NMJ alterations and the survival of demyelinated axons. Morphological and gene expression analysis of NMJs revealed a lack of continuity between the pre- and post-synaptic apparatus, increases in post-synaptic fragmentation and dispersal, and an increase in expression of the gamma subunit of the acetylcholine receptor. There were no changes in axonal width or the number of axonal inputs to the NMJ. Proteome investigations of the sciatic nerve revealed altered expression of extracellular matrix proteins important for NMJ integrity. Together these observations suggest that CMT4C pathology includes a compromised NMJ even in the absence of changes to the innervating axon.

Multiple Lipid Nanoparticles (MLN), a New Generation of Lipid Nanoparticles for Drug Delivery Systems: Lamivudine-MLN Experimental Design.

PURPOSE: An optimized methodology for the development of a new generation of lipid nanoparticles, the multiple lipid nanoparticles (MLN) is described. MLN have characteristics between nanostructured lipid carriers (NLC) and multiple emulsions (W/O/W), but without the outer aqueous phase. METHODS: The production is based on a hot homogenization method combined with high shear and ultrasonication. The antiretroviral agent lamivudine (3TC), was loaded in the MLN. For comparison purposes, NLC-3TC formulation was also developed and physico-chemically characterized by the same parameters as MLN-3TC. The development and optimization of MLN and NLC formulations were supported by a Quality by Design (QbD) approach. RESULTS: The MLN-3TC formulation exhibited a size of about 450 nm, polydispersity <0.3 and negative zeta potential > -20 mV. Furthermore, the morphology assessed by TEM showed a structure with multiples aqueous vacuoles. MLN-3TC was physically stable for at least 45 days, had low cytotoxicity and drug release studies showed a sustained and controlled release of 3TC under gastric and plasma-simulated conditions (at pH 7.4 for about 45 h). CONCLUSIONS: The optimized formulations present suitable profiles for oral administration. Overall, the results reveal that MLN present higher loading capacity and storage stability than NLC.

Screening for SH3TC2 gene mutations in a series of demyelinating recessive Charcot-Marie-Tooth disease (CMT4).

Charcot-Marie-Tooth disease type 4C (CMT4C) is an autosomal recessive (AR) demyelinating neuropathy associated to SH3TC2 mutations, characterized by early onset, spine deformities, and cranial nerve involvement. We screened 43 CMT4 patients (36 index cases) with AR inheritance, demyelinating nerve conductions, and negative testing for PMP22 duplication, GJB1 and MPZ mutations, for SH3TC2 mutations. Twelve patients (11 index cases) had CMT4C as they carried homozygous or compound heterozygous mutations in SH3TC2. We found six mutations: three nonsense (p.R1109*, p.R954*, p.Q892*), one splice site (c.805+2T>C), one synonymous variant (p.K93K) predicting altered splicing, and one frameshift (p.F491Lfs*32) mutation. The splice site and the frameshift mutations are novel. Mean onset age was 7 years (range: 1-14). Neuropathy was moderate-to-severe. Scoliosis was present in 11 patients (severe in 4), and cranial nerve deficits in 9 (hearing loss in 7). Scoliosis and cranial nerve involvement are frequent features of this CMT4 subtype, and their presence should prompt the clinician to look for SH3TC2 gene mutations. In our series of undiagnosed CMT4 patients, SH3TC2 mutation frequency is 30%, confirming that CMT4C may be the most common AR-CMT type.

Fluorescence Based Turn-on Probe for the Determination of Caffeine Using Europium-Tetracycline as Energy Transfer Complex.

The study describes a simple and sensitive fluorometric sensor based on the enhancement of fluorescence intensity of Europium ion (Eu(3+)) - tetracycline (TC) charge transfer complex on addition of caffeine. The Eu(3+)-TC ternary complex has a characteristic emission peak at 615 nm (λex = 375 nm), the intensity of which increases with increase in concentration of caffeine. The caffeine sensor assay was found to be linear in the range of 0.0515 mM to 51.5 mM. The limit of detection and quantification were found to be 0.0515 mM and 0.382 mM, respectively. A caffeine recovery of 90 to 110 % in biological samples (serum and urine) indicated minimal interference by commonly present excipients in the samples. Rosenthal plots to calculate the binding capacity of caffeine with the Eu(3+)- TC complex revealed an association constant (K) of 238 x 10(3) L/mol and binding number (N) of 1.9. Bland-Altman plot comparing the developed assay and HPLC showed good agreement between values obtained by both the methods. The proposed fluorescent chemical sensor is a rapid and convenient method to determine caffeine with excellent recovery and low detection limit. The probable reaction mechanism for the formation of the turn on fluorescent probe enhancer is discussed.

Novel mutations in SH3TC2 in a young Japanese girl with Charcot-Marie-Tooth disease type 4C.

Charcot-Marie-Tooth disease type 4C (CMT4C) is an autosomal recessive demyelinating form of CMT characterized clinically by early onset and severe spinal deformities, and is caused by mutations in SH3TC2. We describe the case of a 10-year-old Japanese girl diagnosed with CMT4C. The patient developed progressive foot deformities such as marked pes cavus and ankle contracture, with mild muscle weakness in both legs, and generalized areflexia. On electrophysiological studies, motor nerve conduction velocity ranged from 22.3 m/s in the tibial nerve to 48.2 m/s in the median nerve. Sensory nerve conduction velocity ranged from 30.3 m/s in the sural nerve to 52.8 m/s in the median nerve. Sequence analysis of candidate genes identified two novel heterozygous mutations, c.229C>T and c.2775G>A, in SH3TC2. The patient was diagnosed as having CMT4C with novel mutations, making this the first documented Japanese pediatric case.

Phenotypic variability of CMT4C in a French-Canadian kindred.

INTRODUCTION: Charcot-Marie-Tooth type 4C (CMT4C) is an autosomal recessive dysmyelinating neuropathy characterized by precocious and rapidly progressive scoliosis. METHODS: Patients in a French-Canadian kindred were evaluated with clinical examination, electrophysiologic study, and genomic DNA extraction. RESULTS: Six of 10 siblings were clinically symptomatic with supportive electrophysiologic features. The proband presented with regional side-to-side sensorimotor asymmetry, typical pes cavus without obvious scoliosis, and unremarkable plain films of the spine. Affected siblings all share symptoms of foot deformity but have variable onset of neuropathic symptoms, degree of extremity weakness, progression of symptoms, and, most notably, evidence of scoliosis. DNA sequence analysis revealed a novel combination of 2 known recessive mutations, p.R904X and p.R954X, in the SH3TC2 gene. CONCLUSIONS: A broad spectrum of phenotypes should be considered in the possible diagnosis of CMT4C. The absence of scoliosis or late-onset symptoms should not exclude SH3TC2 from the list of candidate genes under consideration. Age of onset and clinical features were variable and suggest that polygenic factors contribute to the final phenotype.

The reverse transcriptase inhibitor 3TC modulates hippocampal transcriptome signatures of inflammation in tauopathy model mice.

Reducing neuroinflammation, a key contributor to brain aging and neurodegenerative diseases, is a promising strategy for improving cognitive function in these settings. The FDA-approved nucleoside reverse transcriptase inhibitor 3TC (Lamivudine) has been reported to improve cognitive function in old wild-type mice and multiple mouse models of neurodegenerative disease, but its effects on the brain have not been comprehensively investigated. In the current study, we used transcriptomics to broadly characterize the effects of long-term supplementation with a human-equivalent therapeutic dose of 3TC on the hippocampal transcriptome in male and female rTg4510 mice (a commonly studied model of tauopathy-associated neurodegeneration). We found that tauopathy increased hippocampal transcriptomic signatures of neuroinflammation/immune activation, but 3TC treatment reversed some of these effects. We also found that 3TC mitigated tauopathy-associated activation of key transcription factors that contribute to neuroinflammation and immune activation, and these changes were related to improved recognition memory performance. Collectively, our findings suggest that 3TC exerts protective effects against tauopathy in the hippocampus by modulating inflammation and immune activation, and they may provide helpful insight for ongoing clinical efforts to determine if 3TC and/or related therapeutics hold promise for treating neurodegeneration.