Drug hypersensitivity reactions are a serious concern in clinical practice because they can be severe and result in lifelong sequelae. An accurate diagnosis and identification of the culprit drug is essential to prevent future reactions as well as for the identification of safe treatment alternatives. Nonetheless, the diagnosis can be challenging. In vivo and in vitro tests can be helpful, although none are conclusive; therefore, the tests are not usually performed in isolation but as part of a diagnostic algorithm. In addition, some in vitro tests are only available in research laboratories, and standardization has not been fully accomplished. Collaborating research is needed to improve drug hypersensitivity reaction diagnosis. In this review, we update the current available in vivo and in vitro tools with their pros and cons and propose an algorithm to integrate them into clinical practice.

Frequent mislabelled causal relationship between drug hypersensitivity reactions and culprit drugs reinforces the need for an accurate diagnosis. The systematic reviews and meta-analyses of in vitro assays published so far focused on immediate reactions and the most severe delayed reactions, while the most frequent drug-induced delayed reactions-nonsevere exanthemas-have been underestimated. We aim to fill this gap. A systematic review of studies on in vitro assays used in the diagnosis of nonsevere drug-induced delayed reactions was conducted following the methodology of Preferred Reporting Items for a Systematic Review and Meta-analysis of Diagnostic Test Accuracy Studies Statement. The EMBASE and PubMed databases were searched. We have included 33 studies from which we extracted the data, then performed meta-analysis where possible, or synthesised the evidence narratively. The quality of the analysed studies was assessed with the QUADAS-2 tool. The tests identified the most frequently were lymphocyte transformation test (LTT), ELISpot, and ELISA. In the meta-analysis carried out for LTT in reactions induce by beta-lactams, the pool estimate of sensitivity and specificity amounted to 49.1% (95% CI: 14.0%, 85.0%) and 94.6% (95% CI: 81.7%, 98.6%), respectively. The studies showed heterogeneity in study design and laboratory settings, which resulted in a wide range of specificity and sensitivity of testing.

Drug hypersensitivity reactions (DHRs) occur in certain people and are often not predictable. DHRs can be classified as immediate and delayed reactions regarding to onset of clinical manifestations. Both reactions are considered to be an important public health problem because they can lead to life-threatening conditions; however, this review article will focus on delayed DHRs. The most important points for diagnosis of delayed DHRs are the recognition of drug hypersensitivity characteristics and culprit drug identification. While it is usually difficult to identify a culprit drug; clinical evaluation using the causality assessment method, a non-invasive process, can identify the culprit drug without the need for intensive investigation. Delayed DHRs can cause life-threatening conditions; therefore, in vivo skin tests, as well as drug provocation tests, have to be cautiously performed by a drug allergist and have not been recommended in uncontrolled conditions. ENDA/EAACI has recommended that in vitro tests (if available) be performed prior to any in vivo tests. Therefore, in vitro diagnostic tests can be alternative methods to identify a culprit drug for delayed DHR diagnosis as there is no or very low risk for patients under investigation. There are many testing approaches to identify causative agents for delayed DHRs such as: the lymphocyte transformation test (LTT), cytokine/mediator detection assays (i.e. ELISA and flow cytometry-based bead assays), multiplex bead-based immunoassay and ELISpot. The LTT is the most standardized method whereas it has been available in medical schools affiliated with university hospitals. Other in vitro tests, like cytokine detection assays, have also been used, even though they are still being evaluated. They could supplement LTT results that would provide drug allergist's with documentary evidence and prevent risk to patients by avoiding in vivo or drug provocation testing. Hence, the in vitro tests have been promising tests contributing to the management of the delayed DHR work-up process in clinical practice.

In the setting of an acute cutaneous adverse drug reaction there is increasing interest in selected phenotypes and hosts to continue drug therapy, especially in settings in which there are limited therapeutic options. This concept of "treating through," defined as the continued use of a drug in the setting of, in particular maculopapular exanthema, potentially avoids unnecessary drug discontinuation. A review of the recent literature, historical viewpoints, and expert opinion are provided within to form recommendations and algorithms for a "treating-through" approach.

Although the incidence of severe cutaneous adverse reactions (SCARs) is very low, if it is not diagnosed and treated in time, it can not only cause skin and mucous membrane involvement, but can also cause multiple organ failure and even death. The diagnostic criteria and treatment guidelines for severe drug eruptions have not been unified. Many medical centers have used human leukocyte antigen alleles to diagnose SCARs. Some prospective studies have shown that susceptibility gene testing can prevent SCARs as early as possible, but the widespread implementation of its technology is limited by being ethnically specific. With the unique advantages of cytokine detection technology, cytokines are increasingly important for the diagnosis and treatment of SCARs. Related cytokines/chemokines involved in the pathogenesis, adjuvant diagnosis, and treatment of SCARs are discussed.

Gene-modified cell vaccines are the best way to achieve the immunotherapy for all types of acute leukemia. In this study, the recombinant eukaryotic expression vector (pDisplay-HSP70) of heat shock protein 70 (HSP70) of Bacille Calmette-Guérin (BCG) was constructed by amplifying the whole BCG HSP70 gene using polymerase chain reaction (PCR) and sub-cloning into the polyclone endonuclease sites in pDisplay. Then the HL-60 cell vaccine expressing the protein onto the cell surface was prepared by lipofectamine transfection and its anti-tumor effect and mechanism were further studied. Results showed that the fragment of BCG HSP70 was consistent with Mycobacterium tuberculosis HSP70 gene published in GeneBank. DNA sequencing showed that the recombinant vector was correctly constructed and named pDisplay-HSP70. After BCG HSP70 gene transfection, the yellow-green fluorescence on the HL-60 cells surface was observed under a fluorescence microscope. The immunogenicity of HSP70-transfected HL-60 cells exhibited upregulated proliferation of lymphocytes, increased cytokine secretion (IFN-γ) and enhanced killing activity. These results suggested that gene transfection of BCG HSP70 could significantly enhance the immunogenicity of HL-60 cells. It may be used as a suitable candidate gene-modified cell vaccine for cancer immunotherapy.

Gene-modified cell vaccines are the best way to achieve the immunotherapy for all types of acute leukemia. In this study, heat shock protein 70 (HSP70) gene of BCG was transfected into the acute leukemia cells and its anti-leukemia effect was further studied. Results showed that short-term culture of the leukemia cells exhibited increased number and no change in antigen expression. After HSP70 gene transfection, the yellow-green fluorescence on the leukemia cell surface was observed under a fluorescence microscope. The immunogenicity of HSP70-transfected cells exhibited that autologous lymphocytes proliferated significantly and secreted higher amount of IFN-γ, and cytotoxic T lymphocytes induced more beneficial anti-leukemia effects. These results suggested that gene transfection of BCG HSP70 could significantly enhance the immunogenicity of leukemia cells. It may be used as a suitable candidate gene-modified cell vaccine for cancer immunotherapy.

Hypersensitivity drug reactions (HDR) consist of an individual abnormal response with the involvement of the immunological system. In addition to specific immunological mechanisms where specific antibodies or sensitised T cells participate, release of inflammatory mediators by non-specific immunological recognition may also occur. Within this category are one of the most common groups of drugs, the non-steroidal anti-inflammatory drugs. In addition to chemical drugs new emerging ones with an increasing protagonism are biological agents like humanised antibodies and others. For IgE dependent reactions both in vivo and in vitro tests can be used for the immunological evaluation. Sensitivity of these is not optimal and very often a drug provocation test must be considered for knowing the mechanism involved and/or establishing the diagnosis. For non-immediate reactions also both in vivo and in vitro tests can be used. Sensitivity for in vivo tests is generally low and in vitro tests may be needed for the immunological evaluation. Immunohistochemical studies of the affected tissue enable a more precise classification of non-immediate reactions. The monitorization of the acute response of the reactions has given clues for understanding these reactions and has promising results for the future of the immunological evaluation of HDR.

Drug-induced hypersensitivity syndrome (DIHS) is a severe reaction usually characterized by fever, rash, and multiorgan failure, occurring 2-6 wk after drug introduction. It is an immune-mediated reaction involving macrophage and T-lymphocyte activation and cytokine release. A 54-year-old woman was diagnosed with rheumatic arthritis and initiated salazosulfapyridine by mouth. About 10 d later, she had a high fever, skin rash and liver dysfunction. She was admitted to hospital and diagnosed with a drug eruption. She was treated with oral prednisolone 30 mg/d; however, she developed high fever again and her blood tests showed acute liver failure and cytopenia associated with hyperferritinemia. She was diagnosed with acute liver failure and hemophagocytosis caused by DIHS. She was transferred to the Department of Medicine and Bioregulatory Science, Kyushu University, where she was treated with arterial steroid injection therapy. Following this treatment, her liver function improved and serum ferritin immediately decreased. We hypothesized that an immune-mediated reaction in DIHS may have generated over-activation of macrophages and T-lymphocytes, followed by a cytokine storm that affected various organs. The measurement of serum ferritin might be a useful marker of the severity of DIHS.

Maculopapular exanthema (MPE) induced by drugs is a T-cell mediated reaction and effector cells may play an important role in its development. We assessed the effector and cutaneous homing phenotype in peripheral blood cells from allergic patients after drug stimulation. This study included 10 patients and 10 controls. The effector phenotype (CCR7(-)CD27(+/-)), chemokine receptors (CCR4 and CCR10), and activation (CD25(low)) and regulatory markers (CD25(high)) were measured by flow cytometry in both peripheral blood mononuclear cells (PBMCs) and CD4-T-lymphocytes. Proliferation was determined by 5-(-6)-carboxyfluorescein diacetate succinimidyl ester (CFSE) assay and the migratory capacity by a chemotaxis assay using CCL17 and CCL27. Compared to controls, CCR7(-)CD27(-) cells were increased in patients without (p=0.003) and with drug stimulation (p less than 0.001) and had significantly higher proliferation (p=0.010). CCR10 expression was increased in patients after drug stimulation in total and memory CD27(+) T-cells. Lymphocyte migration with CCL27 was higher in patients with drug stimulation (p=0.048), with a decrease in CCR7(-)CD27(-) (p less than 0.0001) and an increase in CCR7(-)CD27(+) (p=0.017). In patients, CD4-T-lymphocytes were significantly activated after drug stimulation (p less than 0.001). In conclusion, we show that effector memory CD4(+) T-cells (CCR7(-)CD27(+)) respond specifically to the drug responsible for MPE and confirm previous data about the involvement of CCR10 in cell trafficking to the skin.

Drug-induced hypersensitivity syndrome (DIHS), also called drug rash with eosinophilia and systemic symptoms (DRESS), is a severe reaction usually characterized by fever, rash, and multiorgan failure, occurring 1-8 weeks after drug introduction. It is an immune-mediated reaction involving macrophage and T-lymphocyte activation and cytokine release, although no consensus has been reached as to its etiology. The skin, hematopoietic system, and liver are frequently involved. DIHS can mimic severe sepsis, viral infection, adult-onset Still disease (AOSD), or lymphoproliferation.We describe 24 consecutive patients with DIHS who were hospitalized between September 2004 and March 2008. Criteria for inclusion in this observational study were suspected drug reaction, eosinophilia >or=500/microL and/or atypical lymphocytes, involvement of at least 2 organs (skin being 1 of them), with suggestive chronology and exclusion of other diagnoses. Our cohort of 12 women and 12 men had a median age of 49 years (range, 22-82 yr), and 11 had skin phototype V or VI. Patients with mild or no rash were immunocompromised (7/24)- defined as treatment with prednisone (>or=10 mg/d) and another immunosuppressant drug, or human immunodeficiency virus infection. All patients were febrile (>38 degrees C), 14 had localized or generalized edema, 7 had pharyngitis, 8 had lymphadenopathy, 22 had hepatitis, 4 had nephritis, 2 had noninfectious and nonlithiasic angiocholitis or cholecystitis. Ten patients were hypotensive, 5 of whom had associated laboratory signs and/or imaging findings suggestive of acute myocardial dysfunction. Half of the patients had hemogram abnormalities, including eosinophilia. Nine DIHS patients fulfilled the Fautrel criteria for AOSD diagnosis, including glycosylated ferritin <20% in 4/11, with or without laboratory characteristics of hemophagocytosis. Twenty DIHS episodes occurred during the less sunny months of October to March.We determined 25-hydroxyvitamin D3 (25[OH]D3) levels in 18 patients and found that 9 patients had vitamin D deficiency (<25 nmol/L or <10 microg/L) and 5 had vitamin D insufficiency (25-50 nmol/L). Moreover, 25(OH)D3 levels were inversely correlated with ferritin values. After culprit-drug withdrawal, outcomes were favorable for all patients, including those with cardiac abnormalities under slow tapering of glucocorticoids.We recommend looking for the frequent but underdiagnosed hypersensitivity myocarditis with noninvasive diagnostic tools, such as N-terminal probrain natriuretic peptide, and promptly withdrawing the culprit drug and starting glucocorticoids. Vitamin D deficiency might be a DIHS risk or severity factor, especially for patients with high skin phototype and during the winter. Because DIHS clinical and laboratory patterns share similarities with AOSD and hemophagocytosis, DIHS should be included in their differential diagnoses.

Maculopapular exanthema (MPE) is the most frequent clinical manifestation of nonimmediate allergic reactions to drugs and T helper 1 (Th1) cytokines and CD4(+) T cells have been shown to play an important role in its pathogenesis. We assessed the role of cytokines and chemokines and their receptors in the pathogenesis of MPE.

We evaluated skin biopsies and peripheral CD4(+) and CD8(+) T cells from 27 patients during the acute phase of the reaction and 26 exposed controls. Semiquantitative real-time PCR was performed to determine the expression of cytokines and chemokines and their receptors and immunohistochemistry was used to determine the same chemokines and their receptor proteins in skin.

There was a high expression of the Th1 cytokines interferon-gamma (P = 0.006) and tumor necrosis factor-alpha (P = 0.022) in skin and CD4(+) T cells (P = 0.007 and P = 0.005, respectively); and of the Th1 chemokines CXCL9 (P = 0.005) and CXCL10 (P = 0.028) in the skin, while their receptor CXCR3 was increased in skin (P = 0.006) and CD4(+) T cells (P = 0.03). Homing chemokine receptors were also increased: CCR6 in skin (P = 0.026) and CD4(+) T cells (P = 0.016), and CCR10 only in CD4(+) T cells (P = 0.016), as well as their ligands, CCL20 and CCL27, in skin alone. Immunohistochemistry confirmed these results.

These data show significant differences in the expression of chemokines and chemokine receptors, related with a Th1 profile, in both skin biopsies and peripheral CD4(+) T cells in patients with drug-induced MPE.