Tacrolimus Pharmacotherapy: Infectious Complications and Toxicity in Organ Transplant Recipients; An Updated Review.

BACKGROUND: After allogeneic organ transplantation, in order to reduce the risk of rejection, tacrolimus is given. In fact, infection is reported as one of the most common side effects of tacrolimus that might be associated with graft failure. OBJECTIVE: This study aims to review the association between the occurrence of infections due to toxicity following the administration of tacrolimus in organ transplant recipients. METHODS: Scientific literature on the pharmacotherapy of tacrolimus after organ transplantation, infections, and neurotoxicity were searched using PUBMED.Gov (https://pubmed.ncbi.nlm.nih.gov/), Web of Science, and Scopus (n=108). All articles were screened, and the data associated with the topic of interest was extracted. The primary outcome was infection and neurotoxicity. RESULTS: Total area under the curve exposure, the ratio of parent drug/metabolites of tacrolimus was reported to be correlated with aggressive events such as infection episodes. A trough/dose ratio may demonstrate the net state of immunosuppression and drug-related events. The most frequent infectious complication of tacrolimus after organ transplantation was reported as urinary tract infections (UTIs). Virulent strains of recombinant Listeria monocytogenes, in addition to an increase in bacterial burden in the liver and spleen tissues, were reported in experimental animal studies. Patient survival was significantly lower in recipients with UTIs in the first post-transplant month. A higher degree of immunosuppression was associated with recurrent UTIs and drug-resistant organisms. By inhibiting the cerebral immune system, tacrolimus could cause neurodegeneration. CONCLUSION: Transplant type, gut dysmotility, acute or chronic condition before transplant surgery, use of azole, antifungal, hematocrit, tacrolimus methods of detection, the total area under the curve, and duration of hospital stay could define the risk of infection through the first month of transplant surgery. In addition, neurological and infectious complications could be associated with the higher amounts of tacrolimus trough levels (C0). Polypharmacy based on tacrolimus, antiviral, and antifungal drugs, in addition to neurotoxicity, could increase the risk of opportunistic infections such as cytomegalovirus within the first year of organ transplantation.

Role of rs7903146 polymorphism and adropin serum level in patients with diabetes mellitus; a case-control study from Isfahan, Iran.

BACKGROUND: Type-2 diabetes mellitus (T2DM) is the common endocrinopathy which characterised by insulin resistance, insufficient expression or secretion of insulin and decrement of insulin effectiveness. Although T2DM has unknown aetiology, the strongest susceptible gene in this disease is TCF7L2. Adropin peptide may have roles in T2DM pathogenesis due to several roles in glucose tolerance, decrement of insulin resistance, lipid metabolism and energy homoeostasis. AIM: To evaluate the serum level of adropin in T2DM patients and comparing with healthy individuals as well as assessing frequency of rs7903146 genotypes/alleles in patients and control groups. METHODS: We analysed the frequency of rs7903146 genotypes/alleles in 93 patients with T2DM disease and 53 healthy individuals by the method of polymerase chain reaction-restriction fragment length polymorphism analysis. The serum level of adropin was measured by using enzyme-linked immunosorbent assay technique. RESULTS: The mean serum level of adropin was 12.32 ± 2.98 and 9.51 ± 2.73 in patients and control groups, respectively (p value < .001). Also, there were significant difference in frequency of genotypes and alleles of rs7903146 in patients and controls groups (p < .001). The rs7903146T/T and rs7903146C/T genotypes increased risk of T2DM disease (OR: 6.035 and OR: 3.082, respectively). Interestingly, the highest level of adropin was detected in T2DMpatients with rs7903146T/T genotype. CONCLUSION: Our analysis showed higher level of adropin in T2DM patients and increased risk of T2DM with rs7903146T/T and rs7903146C/T genotypes.

Novel Approach to Overcome Defects of Cell-SELEX in Developing Aptamers against Aspartate ß-Hydroxylase.

Cell-based aptamer selection (Cell-SELEX) against predefined protein targets that benefits using the native form of the targets is the most promising approach to achieve aptamer probes capable of recognizing targets under both in vitro and in vivo conditions. The major disadvantages in Cell-SELEX are the imperfectness of the negative selection step and the lengthy procedure of selection. Here, we introduced the Counter-SELEX as part of our modified Cell-SELEX and implemented deep sequencing to overcome these shortcomings in developing aptamers against aspartate ß-hydroxylase (ASPH) as a known tumor marker. In parallel with the conventional Cell-SELEX, five consecutive cycles of counter selection were accomplished using sequences bound to negative cells (the Counter-SELEX) to detect oligos that are not specific for ASPH. After high-throughput sequencing, the representative of each promising achieved family was subjected to further confirmatory analysis via flow cytometry, followed by the fluorescence immunostaining of histopathological sections. Implementing our innovative complementary method, annoying mis-selected sequences in Cell-SELEX enriched pools were effectively identified and removed. According to the affinity assay on the cells displaying ASPH, three aptamers, AP-Cell 1, AP-Cell 2, and AP-Cell 3, with K d values of 47.51, 39.38, and 65.23 nM, respectively, were obtained, while AP-Cell 1 and 3 could then successfully spot ASPH displayed on the tissues. Our study showed that the Counter-SELEX could be considered as a complementary method for Cell-SELEX to overcome the imperfectness of the negative selection step. Moreover, high-throughput nucleotide sequencing could help to shorten the overall process.

An innovative cell selection approach in developing human cells overexpressing aspartyl/asparaginyl ß-hydroxylase.

BACKGROUND AND PURPOSE: Aspartyl/asparaginyl ß-hydroxylase (ASPH) is abundantly expressed in malignant neoplastic cells. The establishment of a human cell line overexpressing ASPH could provide the native-like recombinant protein needed for developing theranostic probes. In the process of transfection, the obtained cells normally contain a range of cells expressing the different levels of the target of interest. In this paper, we report on our simple innovative approach in the selection of best-transfected cells with the highest expression of ASPH using subclone selection, quantitative real-time polymerase chain reaction, and gradual increment of hygromycin concentration. EXPERIMENTAL APPROACH: To achieve this goal, human embryonic kidney (HEK 293T) cells were transfected with an ASPH-bearing pcDNA3.1/Hygro(+) vector. During antibiotic selection, single accumulations of the resistant cells were separately cultured and the ASPH mRNA levels of each flask were evaluated. The best subclones were treated with a gradually increasing amount of hygromycin. The ASPH protein expression of the obtained cells was finally evaluated using flow cytometry and immunocytochemistry. FINDINGS / RESULTS: The results showed that different selected subclones expressed different levels of ASPH. Furthermore, the gradual increment of hygromycin (up to 400mg/mL) improved the expression of ASPH. The best relative fold change in mRNA levels was 57.59 ± 4.11. Approximately 90.2% of HEKASPH cells overexpressed ASPH on their surface. CONCLUSION AND IMPLICATIONS: The experiments indicated that we have successfully constructed and evaluated a recombinant human cell line overexpressing ASPH on the surface. Moreover, our innovative selection approach provided an effective procedure for enriching highly expressing recombinant cells.

Aptamer-based approaches for in vitro molecular detection of cancer.

Cancer is typically associated with abnormal production of various tumor-specific molecules known as tumor markers. Probing these markers by utilizing efficient approaches could be beneficial for cancer diagnosis. The current widely-used biorecognition probes, antibodies, suffer from some undeniable shortcomings. Fortunately, novel oligonucleotide-based molecular probes named aptamers are being emerged as alternative detection tools with distinctive advantages compared to antibodies. All of the existing strategies in cancer diagnostics, including those of in vitro detection, can potentially implement aptamers as the detecting moiety. Several studies have been performed in the field of in vitro cancer detection over the last decade. In order to direct future studies, it is necessary to comprehensively summarize and review the current status of the field. Most previous studies involve only a few cancer diagnostic strategies. Here, we thoroughly review recent significant advances on the applications of aptamer in various in vitro detection strategies. Furthermore, we will discuss the status of diagnostic aptamers in clinical trials.

In vivo/In vitro immune responses to L. major isolates from patients with no clinical response to Glucantime.

BACKGROUND: Leishmaniasis is a major health problem in some endemic areas of tropical and subtropical areas of the world. Interleukin-12 (IL-12) and interferon gamma (IFN-γ) are essential cytokines associated with initiation of Th1 response. The main objective of this study was to evaluate of the type of immune response to L. major isolates from patients with no clinical response to antimonite (Glucantime). MATERIALS AND METHODS: This experimental study was carried out during 2013-2014. In the current study Leishmania major were isolated from 10 CL patients with a history of at least one course of treatment with Meglumine antimonate (Sb5). The isolates were used to evaluate in vitro and in vivo response to Sb5. J774 murine macrophage cell line was used for in vitro tests and Balb/c mice was used for in vivo studies. IL-12 gene expression was evaluated using Real-time PCR and IFN-γ serum level was quantified using ELISA technique. SPSS (version: 20), analysis of Covariance (ANCOVA) was used for statistical analysis. RESULTS: PCR results confirmed that all 10 isolates were L. major. The mean of IL-12 gene expression in vitro, in vivo and IFN-γ serum levels (pg/ml) after 2 and 3 weeks treatment in vivo, increased significantly following the treatment with Glucantime in the two groups of Balb/c mice infected either with patients' isolates or standard L. major. No significant difference was seen between the patients' isolates and standard species. CONCLUSIONS: Although the L. major were isolated from patients with active lesion and no clinical response to Glucantime after at least one courses of Glucantime treatment but in vivo and in vitro immune response of L. major isolates showed no difference between the patients' isolates and standard L. major.

Preliminary study on patients located at the Kashani/Isfahan Hospital with multiple sclerosis between the years 2011 and 2013.

BACKGROUND: Multiple sclerosis (MS) is a multifactorial disease that could result from demyelination of the myelin sheath. The aim of this study is to investigate the demographic features and rank the immunomodulating drugs in patients with MS. MATERIALS AND METHODS: This study was conducted in the MS clinic of the Isfahan Kashani Hospital, from 22 May, 2011 to 18 March, 2013. The data analyses (n = 1067) were divided into two periods: (1) 2011/05/22 to 2012/03/18 denoted as P1 and (2) 2012/04/02 to 2013/03/18 denoted as P2. RESULTS: Most of drugs prescribed within the population studied were: Avonex, Betaferone, and Rebif. There was an increase in the number of female (n = 811) and male subjects (n = 256). During P1/P2 there was an increase from 460 to 607 in the total number of patients, respectively. The number of patients who attended the MS clinic once was 250 (P1) versus 430 (P2), and those more than four times was 71 (P1) versus 59 (P2) correspondingly. CONCLUSION: The number of females increased from 2011 to 2013. Because of dissimilar ingredients additive of different pharmaceutical companies, it could be suggested that pharmacotherapy strategies, especially in Iranian population of MS with first-line treatment using Avonex, Betaferone and Rebif, more spotlighted on inter- and intra-individual variability based on clinical pharmacokinetics parameters.

Preliminary analysis of month of birth in Iranian/Isfahan patients with multiple sclerosis.

BACKGROUND: Previous publications reported that an individual's month of birth (MOB) might have an important correlation to that consequent risk of multiple sclerosis (MS). AIM: The aim of this preliminary study was to investigate the distribution of different MOBs inpatients with MS in Isfahan, Iran. MATERIALS AND METHODS: This investigation was conducted to the Isfahan Neurosciences Research Centre. 1283 patients with MS were studied. Clinical data were recorded in d-Base and analyzed using SPSS (version 18) for Windows. RESULTS: Of the total population studied there were 979 females and 304 males. The mean age of all the patients was 34.6 years (range 10-87 years). Within the total population, the MOBs in the 62% of patients were in the season's spring and summer, and in 38% of patients they were in the season's autumn and winter. As the MOB might be recognized to have a bearing on an individual's risk of contracting MS, the highest and lowest correlations seem to be linked with April, September, May (↑), and November (↓), respectively. CONCLUSIONS: The seasonal relationship between MOBs and MS risk might be pointed toward a potential function for vitamin D throughout pregnancy or the early life of the newborn. Further studies are needed to confirm these correlations.

The investigation of correlation between Iminoral concentration and neurotoxic levels after kidney transplantation.

BACKGROUND: Neurotoxicity side effects related to cyclosporine kinetics could lead to dysfunction of kidney graft and patient outcome after transplantation. The aim of this study was evidence-based pharmacotherapy of kidney transplant recipients and to investigate neurotoxic levels of Iminoral. MATERIALS AND METHODS: The results of 2239 cyclosporine trough levels obtained from 743 patients were studied. Seventy-five adult kidney recipients who received Iminoral were studied for neurotoxicity symptoms. Demographic, clinical, hematology and biochemical data were recorded in d-base and analyzed using SPSS application for windows. RESULTS: The mean value related to cyclosporine C0 was 246.3 µg/l. In the 48% the signs of neurotoxicity such as tremor and headache were noted, but only in 9% the levels of cyclosporine C0 were >400 µg/l. Further studies on 75 patients showed that the incidence of neurotoxic side effects were as follows: Tremor in 35, headache in 24 and anxiety in 34 recipients of kidney. The prescribed drug regimens from the day of transplant in most patients were based on mycophenolic acid or cellcept, pulse therapy using methylprednisolone (daily from kidney transplant up to 3 days after transplant), cyclosporine or Iminoral plus other drugs related to each individual. Administrations of ganciclovir, thymoglobulin, clotrimazol and prednisolone were also distinguished with immunosuppressant-based therapy simultaneously. CONCLUSION: Evidence-based study related to pharmacotherapy of Iminoral showed that clinical presentation related to neurotoxic side effects such as tremor, headache and anxiety might be due to many factors such as polypharmacy. Planning immunosuppression to individual patients based on programmed therapeutic Iminoral monitoring, avoiding polypharmacy in terms of removal or drug minimization and focusing on first week after transplant seem to be a realistic option.