A new case of sodium-dependent multivitamin transporter defect occurring as a life-threatening condition responsive to early vitamin supplementation and literature review.

BACKGROUND: Biallelic pathogenic variants in SLC5A6 resulting in sodium-dependent multivitamin transporter (SMVT) defect have recently been described as a vitamin-responsive inborn error of metabolism mimicking biotinidase deficiency. To our knowledge, only 16 patients have been reported so far with various clinical phenotypes such as neuropathy and other neurologic impairments, gastro-intestinal dysfunction and failure to thrive, osteopenia, immunodeficiency, metabolic acidosis, hypoglycemia, and recently severe cardiac symptoms. METHODS: We describe a case report of a 5-month-old girl presenting two recurrent episodes of metabolic decompensation and massive cardiac failure in the course of an infectious disease. We compare clinical, biological, and genetic findings of this patient to previous literature collected from Pubmed database (keywords: Sodium-dependent multivitamin transporter (SMVT), SMVT defect/disorder/deficiency, SLC5A6 gene/mutation). RESULTS: We highlight the life-threatening presentation of this disease, the stagnation of psychomotor development, the severe and persistent hypogammaglobulinemia, and additionally, the successful clinical response on early vitamin supplementation (biotin 15 mg a day and pantothenic acid 100 mg a day). Metabolic assessment showed a persistent increase of urinary 3-hydroxyisovaleric acid (3-HIA) as previously reported in this disease in literature. CONCLUSION: SMVT deficiency is a vitamin-responsive inborn error of metabolism that can lead to a wide range of symptoms. Increased and isolated excretion of urinary 3-hydroxyisovaleric acid may suggest, in the absence of markedly reduced biotinidase activity, a SMVT deficiency. Prompt supplementation with high doses of biotin and pantothenic acid should be initiated while awaiting results of SLC5A6 sequencing as this condition may be life-threatening.

Effects of biotin and coated cobalamin on lactation performance, nutrient digestion and rumen fermentation in Holstein dairy cows.

Biotin (BI) and cobalamin (CA) are essential for rumen propionate production and hepatic gluconeogenesis. The study evaluated the influence of BI or/and coated CA (CCA) on milk performance and nutrient digestion in cows. Sixty Holstein dairy cows were assigned in a 2 × 2 factorial arrangement and randomised block design to four groups. The factors were BI at 0 or 20 mg/day and CCA at 0 or 9 mg CA/day. Dry matter intake increased with BI addition but was unchanged with CCA supply. Addition of BI or CCA increased fat-corrected milk, milk fat and milk protein yields and feed efficiency. Moreover, lactose yield was increased by CCA addition. Dry matter, organic matter, crude protein and acid detergent fibre total-tract digestibility increased for BI or CCA supply. When CCA was supplemented, positive response of neutral detergent fibre digestibility to BI addition was enhanced. Supplementing BI did not affect pH, propionate content and acetate to propionate ratio, but increased total volatile fatty acids (VFA) and acetate contents. Supplementing CCA decreased pH and acetate to propionate ratio, but increased total VFA, acetate and propionate contents. Rumen protease and carboxymethyl-cellulase activities and fungi, bacteria and Butyrivibrio fibrisolvens numbers increased for BI or CCA supply. In addition, protozoa increased for BI addition, and protease activity and Prevotella ruminicola increased for CCA supply. When CCA was supplemented, positive responses of R. albus and Ruminobacter amylophilus numbers to BI addition were enhanced. Blood glucose concentration was unchanged with BI supply, but increased for CCA supply. Blood nonesterified fatty acids and ß-hydroxybutyrate contents reduced with BI or CCA supply. Supplementation with BI or CCA increased blood BI or CA content. The results showed that supplementing BI or/and CCA improved lactation performance and nutrient digestion, and CCA supply did not enhance the lactation performance response to BI supply.

Biotinylated Quinone as a Chemiluminescence Sensor for Biotin-Avidin Interaction and Biotin Detection Application.

Biotin, or vitamin B7, is essential for metabolic reactions. It must be obtained from external sources such as food and biotin/vitamin supplements because it is not biosynthesized by mammals. Therefore, there is a need to monitor its levels in supplements. However, biotin detection methods, which include chromatographic, immune, enzymatic, and microbial assays, are tedious, time-consuming, and expensive. Thus, we synthesized a product called biotin-naphthoquinone, which produces chemiluminescence upon its redox cycle reaction with dithiothreitol and luminol; then it was used as a chemiluminescence sensor for biotin-avidin interaction. When a quinone biotinylated compound binds avidin, the chemiluminescence decreases noticeably due to the proximity between quinone and avidin, and when free biotin is added in a competitive assay, the chemiluminescence returns. The chemiluminescence is regained as the free biotin displaces biotinylated quinone in its complex with avidin, freeing biotin-naphthoquinone. Many experiments, including the use of a biotin-free quinone, proved the competitive nature of the assay. The competitive assay method used in this study was linear in the range of 1.0-100 µM with a detection limit of 0.58 µM. The competitive chemiluminescence assay could detect biotin in vitamin B7 tablets with good recovery of 91.3 to 110% and respectable precision (RSD < 8.7%).

An Analysis of the Biotin-(Strept)avidin System in Immunoassays: Interference and Mitigation Strategies.

An immunoassay is an analytical test method in which analyte quantitation is based on signal responses generated as a consequence of an antibody-antigen interaction. They are the method of choice for the measurement of a large panel of diagnostic markers. Not only are they fully automated, allowing for a short turnaround time and high throughput, but offer high sensitivity and specificity with low limits of detection for a wide range of analytes. Many immunoassay manufacturers exploit the extremely high affinity of biotin for streptavidin in their assay design architectures as a means to immobilize and detect analytes of interest. The biotin-(strept)avidin system is, however, vulnerable to interference with high levels of supplemental biotin that may cause elevated or suppressed test results. Since this system is heavily applied in clinical diagnostics, biotin interference has become a serious concern, prompting the FDA to issue a safety report alerting healthcare workers and the public about the potential harm of ingesting high levels of supplemental biotin contributing toward erroneous diagnostic test results. This review includes a general background and historical prospective of immunoassays with a focus on the biotin-streptavidin system, interferences within the system, and what mitigations are applied to minimize false diagnostic results.

High-dose pharmaceutical-grade biotin in patients with demyelinating neuropathies: a phase 2b open label, uncontrolled, pilot study.

BACKGROUND: We proposed to investigate high-dose pharmaceutical-grade biotin in a population of demyelinating neuropathies of different aetiologies, as a proof-of-concept. METHODS: Phase IIb open label, uncontrolled, single center, pilot study in 15 patients (three groups of five patients) with chronic demyelinating peripheral neuropathy, i.e. chronic inflammatory demyelinating polyradiculoneuropathy, anti-myelin-associated glycoprotein neuropathy and Charcot-Marie-Tooth 1a or 1b. The investigational product was high-dose pharmaceutical-grade biotin (100 mg taken orally three times a day over a maximum of 52 weeks. The primary endpoint was a 10% relative improvement in 2 of the following 4 electrophysiological variables: motor nerve conduction velocity, distal motor latency, F wave latency, duration of the compound muscle action potential. The secondary endpoints included Overall Neuropathy Limitations Scale (ONLS) score, Medical Research Council (MRC) sum score, Inflammatory Neuropathy Cause and Treatment (INCAT) sensory sum score, 10-m walk test, 6-min walk test, posturography parameters, and nerve excitability variables. RESULTS: The primary endpoint was reached in one patient. In the full population analysis, some secondary endpoints parameters improved: MRC score, INCAT sensory sum score, 6-min walk distance, strength-duration time constant, and rheobase. There was a positive correlation between the improvement in the 6-min walk distance and the strength-duration time constant. Regarding the safety results, 42 adverse events occurred, of which three were of severe intensity but none was considered as related to the investigational product. CONCLUSIONS: Even if the primary endpoint was not met, administration of high-dose pharmaceutical-grade biotin led to an improvement in various sensory and motor parameters, gait abilities, and nerve excitability parameters. The tolerance of the treatment was satisfactory. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02967679; date 2016/12/05.

Protein-Based Anchoring Methods for Nucleic Acid Detection in Lateral Flow Format Assays.

The use of lateral flow assays to detect nucleic acid targets has many applications including point-of-care diagnostics, environmental monitoring, and food safety. A sandwich format, similar to that in protein immunoassays, is often used to capture the target nucleic acid sequence with an immobilized complementary strand anchored to a substrate, and then to visualize this event using a complementary label nucleic acid bound to a nanoparticle label. A critical component of high-sensitivity nucleic acid detection is to utilize high-density capture surfaces for the effective capture of target nucleic acid. Multiple methods have been reported, including the use of streptavidin-based protein anchors that can be adsorbed to the lateral flow substrate and that can utilize the high-affinity streptavidin-biotin linkage to bind biotinylated nucleic acid capture sequences for subsequent target nucleic acid binding. However, these protein anchors have not been systematically characterized for use in the context of nucleic acid detection. In this work, we characterize several protein-based anchors on nitrocellulose for (i) capturing the robustness of the attachment of the protein anchor, (ii) capturing nucleic acid density, and (iii) targeting nucleic acid capture. Further, we demonstrate the signal gains in target nucleic acid hybridization made by increasing the density of capture nucleic acid on a nitrocellulose substrate using multiple applications of protein loading onto nitrocellulose. Finally, we use our high-density capture surfaces to demonstrate high-sensitivity nucleic acid detection in a lateral flow assay (in the context of a SARS-CoV-2 sequence), achieving a LOD of approximately 0.2 nM.

Targeted co-delivery of curcumin and erlotinib by MoS2 nanosheets for the combination of synergetic chemotherapy and photothermal therapy of lung cancer.

BACKGROUND: Curcumin (Cur), a bioactive component of Chinese traditional medicine, has demonstrated inhibitory properties against cancer cell proliferation while synergistically enhancing the anticancer efficacy of erlotinib (Er). However, the individual limitations of both drugs, including poor aqueous solubility, lack of targeting ability, short half-life, etc., and their distinct pharmacokinetic profiles mitigate or eliminate their combined antitumor potential. RESULTS: In this study, we developed a molybdenum disulfide (MoS2)-based delivery system, functionalized with polyethylene glycol (PEG) and biotin, and co-loaded with Cur and Er, to achieve efficient cancer therapy. The MoS2-PEG-Biotin-Cur/Er system effectively converted near-infrared (NIR) light into heat, thereby inducing direct photothermal ablation of cancer cells and promoting controlled release of Cur and Er. Biotin-mediated tumor targeting facilitated the selective accumulation of MoS2-PEG-Biotin-Cur/Er at the tumor site, thus enhancing the synergistic antitumor effects of Cur and Er. Remarkably, MoS2-PEG-Biotin-Cur/Er achieved the combination of synergistic chemotherapy and photothermal therapy (PTT) upon NIR irradiation, effectively suppressing lung cancer cell proliferation and inhabiting tumor growth in vivo. CONCLUSIONS: The as-synthesized MoS2-PEG-Biotin-Cur/Er, featuring high targeting ability, NIR light-responsive drug release, and the integration of synergistic chemotherapy and PTT, may provide a promising strategy for the treatment of lung cancer in clinical practice.

Biotinidase Deficiency in the Second Decade with Atypical Neuroimaging Findings.

Biotinidase deficiency is a rare autosomal recessive neurometabolic disorder resulting in biotin deficiency. Our patient presented with seizures and developmental delay since infancy and was started on megavitamin supplements. At 14 years, she presented with motor regression with encephalopathy after discontinuation of vitamins. There were no skin and hair changes. Magnetic resonance imaging (MRI) of the brain showed bilateral symmetrical posterior putamen signal changes. Tandem mass spectroscopy showed increased methyl malonyl carnitine and 3-OH isovaleryl carnitine. There was a low biotinidase level, and a pathogenic variant in the BTD gene in the next-generation sequencing was identified. Special importance is placed on the unusual symmetric posterior putamen involvement seen in MRI of the brain.

A Retrospective Evaluation of the Laboratory Findings of Dermatology Patients Whose Biotin Levels Were Checked.

Objective Biotin is widely known to be beneficial for the hair, nails, and skin, but there are only a few studies on biotin. We evaluated whether there is a relationship between biotin levels and age, gender, and frequently observed laboratory findings. We also evaluated biotin levels according to the reason for checking biotin levels. Methods One hundred five patients applied to the dermatology outpatient clinic and had their biotin levels checked. Patient files were retrospectively analyzed. Results There were a weak positive (r=0.207) relationship between biotin levels and basophil count, a weak positive (r=0.201) relationship between biotin levels and creatinine, and a weak positive (r=0.314) relationship between biotin levels and cholesterol/triglyceride ratio. There were a weak negative (r=-0.216) relationship between biotin levels and mean platelet volume (MPV) and a moderately negative (r=-0.315) relationship between biotin levels and triglyceride levels. Conclusion Biotin levels do not significantly differ with gender but increase with age. Although a weak correlation was detected between hemogram parameters and biotin levels with basophil percentage and mean platelet volume values, biotin did not significantly change hemogram parameters. The relationship between biotin levels and triglyceride levels was the most critical finding of our study. We recommend examining biotin levels in the patients with high triglyceride levels. When we encounter dermatological side effects related to the use of epidermal growth factor receptor tyrosine kinase inhibitors, we recommend evaluating biotin levels. We recommend that biotin supplementation be made only in the patients with deficiencies and that biotin levels be measured in the follow-up.

Effects of rumen-protected biotin on the growth performance, nitrogen utilization and blood parameters of yearling Liaoning cashmere doelings.

OBJECTIVE: This study was conducted to investigate the effects of rumen-protected biotin (RPB) on growth performance, nutrient digestibility, nitrogen utilization and plasma biochemical parameters of Liaoning cashmere goats during the cashmere fiber growing period. METHODS: Sixteen 6-month-old Liaoning cashmere twin-doelings (24.8±1.20 kg) were allocated to 2 diet groups that were individually ad libitum fed 30% concentrate and 70% forage diet (dry matter [DM]) by a paired experimental design. Goats of the control group were fed the basal diet, while goats belonging to the RPB group were fed the basal diet with 10 mg RPB/d per animal. The duration of the experiment was 16 weeks with two 8-week periods. Digestibility was determined at weeks 7 and 15, and other measures were taken every four weeks. RESULTS: Compared with the control group, the average daily gain of the RPB group increased by 10.94% (p<0.05), and the intake of neutral detergent fiber was increased (p = 0.045). There were some increasing tendencies for the intake of DM, acid detergent fiber and ether extract (p = 0.070, 0.088, and 0.070, respectively). The intake and digestibility of N tended to increase (p = 0.062 and 0.093, respectively), while the N fecal excretion percentage of N intake was decreased (p = 0.093) in the RPB compared with the control group. N retention tended to increase (p = 0.084) with the addition of adding RPB to the diet. Plasma total protein was increased (p = 0.037), whereas the urea-N concentration was decreased (p = 0.049) in the RPB diet group compared with the control diet group. The levels of propionyl-CoA carboxylase (p<0.001) and methylmalonyl-CoA (p = 0.013) were increased in the RPB group. CONCLUSION: Supplementation of rumen-protected biotin in the diet of cashmere goats can enhance the utilization of N and improve daily weight gain during cashmere fiber growing period.

Technical Note: Approach to Identify and Eliminate Biotin Interference in Thyroid Function Tests Using Beckman DXI 800 Analyzer by Taking Advantage of Assay Harmonization with Alinity i Analyzer.

OBJECTIVE: We reported significant interference of biotin in FT3 and FT4 assays using Beckman DXI 800 analyzer. Recently we acquired Alinity i analyzer where TSH, FT3 and FT4 assays are not biotin based. We hypothesized that if thyroid function tests on DXI 800 and Alinity i are harmonized, then it is possible to eliminate biotin interference. METHODS: We investigated assay harmonization by analyzing 35 specimens for TSH, FT4 and FT3 using both analyzers. We prepared one serum pool using left-over specimens where thyroid tests were ordered. Then aliquots of the pool were supplemented with various amounts of biotin followed by measuring thyroid function tests again. RESULTS: We observed assay harmonization between both analyzers for TSH, FT3 and FT4 Tests. TSH assay was not affected in the presence of biotin, but FT3 and FT4 values were significantly elevated using DXI 800 analyzer. In contrast, TSH, FT3 and FT4 assays were not affected by biotin using Alinity i analyzer. CONCLUSIONS: Elevated FT3 and FT4 using DXI 800 analyzer may be due to biotin interference which can be eliminated by observing normal values using Alinity i analyzer. However, normal or slightly elevated TSH with elevated FT3 and FT4 using both analyzers may indicate rare type of TSH producing tumor of pituitary, not biotin interference.

Pharmacotechnical, Physico-Chemical, and Antioxidant Evaluation of Newly Developed Capsule Formulations.

The excess of free radicals causes numerous imbalances in the body that lead to premature aging, the degradation of internal structures, and the appearance of numerous pathologies responsible for the increased risk of premature death. The present work aims to evaluate the physical, chemical, pharmacotechnical, and antioxidant activity of newly achieved capsule formulations. These two formulations were F1a.i., which contains melatonin:biotin:coenzyme Q10 (weight ratio of 1:2:60), and F2a.i., which contains quercetin:resveratrol:biotin:coenzyme Q10 (weight ratio of 10:10:1:10). The adequate selection of the excipient types and amounts for final capsule formulations (F1c.c., F2c.c.) was based on preformulation studies performed on the powders containing active ingredients. The antioxidant activity assessed using three methods (ABTS, DPPH, and FRAP) compared with acid ascorbic as a positive control demonstrated that the F2c.c. formulation possesses the strongest antioxidant capacity. The results confirmed the suitable formulation and the accurate selection of the types and amounts of active ingredients, as well as the auxiliary excipients used in newly developed capsule formulations as supplements with an excellent antioxidant effect on the human body.

Biotin induced biochemical hyperthyroidism: a case report and review of the literature.

BACKGROUND: Biotin is a commonly used supplement for hair, nail, and skin. Recent literature suggests that high-dose biotin therapy for neurological diseases like Multiple sclerosis can interfere with lab results that use biotin/streptavidin immunoassay, called biotin interference. Biotin interference can affect thyroid lab results, giving biochemical hyperthyroidism. CASE PRESENTATION: Our patient, a 64-year-old white man with a known history of multiple sclerosis, presented with elevated free T3, free T4, and low TSH that resembled hyperthyroidism. He had no symptoms of hyperthyroidism except some fatigue and tachycardia on the first encounter. He was started on anti-thyroid medications. He was then re-evaluated since his lab results remained the same after two months of anti-thyroid medications. It was found that he was on biotin, 10000mcg/day, for his multiple sclerosis. Biotin was discontinued, and five days later his lab results returned to normal values. CONCLUSION: The lack of knowledge of biotin use by patients can lead to misdiagnosis of patients' thyroid lab results and improper management. Awareness about biotin interference and abnormal thyroid lab values should be a priority among clinicians and the public. If the biotin is discontinued on time, such misdiagnosis can be avoided.

Elevated free thyroxine and free triiodothyronine probably caused by high-dose biotin intake in a patient with Graves' disease: a case report.

Biotin is a water-soluble vitamin that acts as a cofactor for carboxylase, and is often used as a component in several immunoassays. We present a case of a 46-year-old male with Graves' disease (GD) who revealed elevated free thyroxine (FT4) and free triiodothyronine (FT3) levels after high-dose biotin intake. Levels of these hormones had been within the reference range when he was on thiamazole 5 mg/day for 7 years; however, the levels increased from 1.04 to 2.20 ng/dL and from 3.05 to 9.84 pg/mL for FT4 and FT3, respectively, after he started taking biotin 72 mg/day. Despite these high levels, his symptoms and the other laboratory results, including the thyroid-stimulating hormone level, did not suggest GD relapse. His thyroid hormone data was decreased and returned within the reference range immediately after the laboratory assays for FT3 and FT4 had been coincidentally changed from those containing streptavidin-biotin complexes to biotin-free ones. Biotin interference, which is caused by high-dose biotin intake and immunoassays using some form of streptavidin-biotin complex, is sometimes clinically problematic, giving high or low results. To our knowledge, this is the first case report of a patient with GD on high-dose biotin receiving high thyroid hormone level results that were initially misunderstood as an aggravation of the disease; there are some reports of misdiagnosis of hyperthyroidism due to biotin administration. Unexpected fluctuations in thyroid function test results in patients with GD should be checked for biotin intake, immunoassays and the limiting concentration of biotin to avoid misdiagnosis of relapse.

A novel SLC5A6 homozygous variant in a family with multivitamin-dependent neurometabolic disorder: Phenotype expansion and long-term follow-up.

The sodium-dependent multivitamin transporter (hSMVT) encoded by the SLC5A6 gene is required for the intestinal absorption of biotin, pantothenic acid and lipoate, three micronutrients essential for normal growth and development. Systemic deficiency of these elements, either occurring from nutritional causes or genetic defects, is associated with neurological disorders, growth delay, skin and hair changes, metabolic and immunological abnormalities. A few patients with biallelic variants of SLC5A6 have been reported, exhibiting a spectrum of neurological and systemic clinical features with variable severity. We describe three patients from a single family carrying a homozygous p.(Leu566Valfs*33) variant of SLC5A6 disrupting the frame of the C-terminal portion of the hSMVT. In these patients, we documented a severe disorder featuring developmental delay, sensory polyneuropathy, optic atrophy, recurrent infections, and repeated episodes of intestinal pseudo-obstruction. Two patients who did not receive multivitamin supplementation therapy died in early infancy. In a third patient, early supplementation of biotin and pantothenic acid stabilized the clinical picture changing the course of the disease. These findings extend genotype-phenotype correlations and show how a timely and lifelong multivitamin treatment may be crucial to reduce the risk of life-threatening events in patients with pathogenic variants of the SLC5A6 gene.

Twin Premature Infants With Riboflavin and Biotin Deficiency Presenting With Refractory Lactic Acidosis, Rash, and Multiorgan Failure During Prolonged Parenteral Nutrition.

We are reporting monochorionic, diamniotic twin premature infants born at 25 weeks and 6 days gestation with riboflavin (vitamin B2) and biotin (vitamin B7) deficiency, while on prolonged total parenteral nutrition (TPN) during vitamin shortage. They presented initially with skin rash, lactic acidosis, and thrombocytopenia. Both twins progressed to severe respiratory failure, severe lactic acidosis, with refractory vasodilatory shock, pancytopenia, ischemic bowel injury, acute kidney injury, liver injury, and capillary leak syndrome leading to death of twin A. The surviving twin B was diagnosed with riboflavin and biotin deficiency that presented with abnormal metabolic work up suggestive of maple syrup urine disease, glutaric acidemia type 2, and X-linked adrenoleukodystrophy. Twin B was started on riboflavin and biotin supplementation at 41 days of life, with rapid improvement in clinical findings and laboratory abnormalities within days of starting biotin and riboflavin supplementation. He was discharged home in stable condition at 49 weeks of postmenstrual age.

Functionalization of a Fully Integrated Electrophotonic Silicon Circuit for Biotin Sensing.

Electrophotonic (EPh) circuits are novel systems where photons and electrons can be controlled simultaneously in the same integrated circuit, attaining the development of innovative sensors for different applications. In this work, we present a complementary metal-oxide-semiconductor (CMOS)-compatible EPh circuit for biotin sensing, in which a silicon-based light source is monolithically integrated. The device is composed of an integrated light source, a waveguide, and a p-n photodiode, which are all fabricated in the same chip. The functionalization of the waveguide's surface was investigated to biotinylate the EPh system for potential biosensing applications. The modified surfaces were characterized by AFM, optical microscopy, and Raman spectroscopy, as well as by photoluminescence measurements. The changes on the waveguide's surface due to functionalization and biotinylation translated into different photocurrent intensities detected in the photodiode, demonstrating the potential uses of the EPh circuit as a biosensor.

Significant Interference of Biotin in Thyroid Function Tests Using Beckman Analyzer: How to Identify such Interferences?

OBJECTIVE: Biotin in elevated concentration interferes with biotin-based immunoassays. We studied biotin interferences in TSH, FT4, FT3, total T4, total T3 and thyroglobulin assays both in vitro and in vivo using Beckman DXI800 analyzer. METHODS: Two serum pools were prepared from left-over specimens. Then aliquots of each pool (and serum control) were supplemented with various amounts of biotin followed by measuring thyroid function tests again. Three volunteers each took 10 mg biotin supplement. We compared thyroid function tests before and 2 h after taking biotin. RESULTS: We observed significant biotin interferences in biotin-based assays (positive interference with FT4, FT3, and total T3 assay but negative interference with thyroglobulin) both in vitro and in vivo but non-biotin-based assays (TSH and total T4) were not affected. CONCLUSIONS: Elevated FT3 and FT4 in the presence of normal TSH is inconsistent with hyperthyroidism and should be followed up with total T3 and T4 test. Significant discrepancy between total T3 (falsely elevated value due to biotin) and total T4 (not affected as the assay is not biotin based) maybe an indication of biotin interference.

Biotinylated selenocyanates: Potent and selective cytostatic agents.

Most of the currently available cytotoxic agents for tackling cancer are devoid of selectivity, thus causing severe side-effects. This situation stimulated us to develop new antiproliferative agents with enhanced affinity towards tumour cells. We focused our attention on novel chalcogen-containing compounds (thiosemicarbazones, disulfides, selenoureas, thio- and selenocyanates), and particularly on selenium derivatives, as it has been documented that this kind of compounds might act as prodrugs releasing selenium-based reactive species on tumour cells. Particularly interesting in terms of potency and selectivity was a pharmacophore comprised by a selenocyanato-alkyl fragment connected to a p-phenylenediamine residue, where the nature of the second amino moiety (free, Boc-protected, enamine-protected) provided a wide variety of antiproliferative activities, ranging from the low micromolar to the nanomolar values. The optimized structure was in turn conjugated through a peptide linkage with biotin (vitamin B7), a cellular growth promoter, whose receptor is overexpressed in numerous cancer cells; the purpose was to develop a selective vector towards malignant cells. Such biotinylated derivative behaved as a very strong antiproliferative agent, achieving GI50 values in the low nM range for most of the tested cancer cells; moreover, it was featured with an outstanding selectivity, with GI50 > 100 µM against human fibroblasts. Mechanistic studies on the mode of inhibition of the biotinylated selenocyanate revealed (Annexin-V assay) a remarkable increase in the number of apoptotic cells compared to the control experiment; moreover, depolarization of the mitochondrial membrane was detected by flow cytometry analysis, and with fluorescent microscopy, what supports the apoptotic cell death. Prior to the apoptotic events, cytostatic effects were observed against SW1573 cells using label-free cell-living imaging; therefore, tumour cell division was prevented. Multidrug resistant cell lines exhibited a reduced sensitivity towards the biotinylated selenocyanate, probably due to its P-gp-mediated efflux. Remarkably, antiproliferative levels could be restored by co-administration with tariquidar, a P-gp inhibitor; this approach can, therefore, overcome multidrug resistance mediated by the P-gp efflux system.