Daniele Cammelli

Alkaptonuria, ochronosis, and ochronotic arthropathy.

OBJECTIVES To describe the clinical presentation and course of a relatively large group of Italian adult patients screened for mutation of the homogentisate dioxygenase gene causing alkaptonuria (AKU) and ochronosis, and to review typical and atypical facets of this condition. METHODS We reviewed the medical records of 9 patients affected by ochronotic arthropathy who were observed in our institutions between 1979 and 2001. All patients were diagnosed as having AKU through a rapid urine test with alkali. Mutation screening was performed by single-strand conformation analysis of all homogentisate dioxygenase exons, followed by sequencing of altered conformers. RESULTS Our 9 cases had similar clinical features and they reflected those described in the literature: a progressive degenerative arthropathy mainly affecting axial and weight-bearing joints associated with extraarticular manifestation. Musculoskeletal symptoms began in most of our patients around the age of 30 years with back pain and stiffness: involvement of the large peripheral joints usually occurred several years after spinal changes. Ochronotic peripheral arthropathy generally was degenerative, but joint inflammation was observed in some cases; this could be attributed to an inflammatory reaction of the ochronotic shard in the synovial membrane. CONCLUSIONS Ochronosis is a model of arthropathy with known etiologic factors. Over time, AKU, the genetically determined metabolic defect, leads to the accumulation of pigment and the development of this crippling condition. Most of the clinical findings may be explained by inhibition of collagen crosslinks, but some require additional interpretation. For example, inflammatory features of the ochronotic joint only occur in a minority of cases, and may be attributable to ochronotic shards. Further studies are needed to establish the genotype-phenotype correlation to identify mutations that are predictive of severe disease. For this purpose, the Italian Study Group on Alkaptonuria (www.dfc.unifi.it/aku) is enrolling affected patients in an on-line database to characterize the molecular defects and their relationship to clinical data.

Drug-Specific Th2 Cells and IgE Antibodies in a Patient with Anaphylaxis to Rituximab

Rituximab (RTX) is currently used in the treatment of lymphoproliferative diseases and of several rheumatologic disorders and is a frequent cause of acute infusion reactions, usually classified as cytokine release syndrome (CRS). Some infusion reactions to RTX raise concern for immediate type I hypersensitivity, even if to date RTX-specific IgE antibodies have not been reported. To improve knowledge of the mechanisms of reactions to RTX, we investigated humoral and cellular immune responses to this drug in a patient suffering from rheumatoid arthritis who displayed two immediate infusion-related reactions. RTX-exposed tolerant patients and healthy untreated subjects were used as controls. Non-isotype-specific and IgE anti-RTX antibodies were positive in the serum samples collected from the reactive patient but not in those from the control groups. Only the reactive patient also displayed skin testing positivity with RTX. More importantly, RTX-stimulated peripheral blood mononuclear cells from the reactive patient, but not from the controls, displayed a dose-dependent proliferative response associated with a Th2 cytokine production profile. Our results show the presence of RTX-specific Th2-type cells and IgE antibodies, thus suggesting that type I hypersensitivity may be an additional mechanism to CRS in the development of RTX reactions.

Aortic Connective Tissue in Ageing—A Biochemical Study

The biochemical analysis of samples of aortic connective tissue was carried out in 22 subjects from 9 to 84 years old. Aortic samples were taken at necropsy performed after sudden or, more often, traumatic death. The results suggest that aging of the aorta is accompanied by an increase both in collagen content and in total sugar content when expressed as mg/cm2 while the elastin content, when expressed in the same way, does not undergo any variation.

Connective Tissue in Varicose Veins

The collagen, elastin, total sugar, and nonscleroprotein content was eval uated in 32 samples of saphenous varicose vein and in 34 controls. A signifi cantly lower collagen and elastin content was found in the varicose samples without correlation with the degree of pathologic broadening. Otherwise the total sugars and the soluble nonscleroproteins were found to be increased in varicose samples. The results are more significant when expressed as milligrams per surface unit of endothelium. Our data support the hypothesis that the decrease in collagen and elastin content is a primary rather than secondary change.

Connective Tissue in Aging Lung

Collagen, elastin and structural glycoproteins were measured in 21 intact lungs taken at postmortem examination after sudden death from subjects aged from 15 to 83. The data were expressed as milligrams per cubic centimeter of lung peripheral parenchyma inflated and fixed at the standard pressure of 25 cm H2O to exclude the pitfall of referring to dried tissue weight. The volume/weight ratio of inflated dried lung parenchyma increased significantly with aging; likewise the collagen content decreased and so did the collagen/elastin ratio, while the elastin content did not show any significant correlation with age. The present findings indicate that biochemical, morphological and functional data on the senile lung agree well.

A Snapshot on the On-Label and Off-Label Use of the Interleukin-1 Inhibitors in Italy among Rheumatologists and Pediatric Rheumatologists: A Nationwide Multi-Center Retrospective Observational Study

Background: Interleukin (IL)-1 inhibitors have been suggested as possible therapeutic options in a large number of old and new clinical entities characterized by an IL-1 driven pathogenesis. Objectives: To perform a nationwide snapshot of the on-label and off-label use of anakinra (ANA) and canakinumab (CAN) for different conditions both in children and adults. Methods: We retrospectively collected demographic, clinical, and therapeutic data from both adult and pediatric patients treated with IL-1 inhibitors from January 2008 to July 2016. Results: Five hundred and twenty-six treatment courses given to 475 patients (195 males, 280 females; 111 children and 364 adults) were evaluated. ANA was administered in 421 (80.04%) courses, CAN in 105 (19.96%). Sixty-two (32.1%) patients had been treated with both agents. IL-1 inhibitors were employed in 38 different indications (37 with ANA, 16 with CAN). Off-label use was more frequent for ANA than CAN (p < 0.0001). ANA was employed as first-line biologic approach in 323 (76.7%) cases, while CAN in 37 cases (35.2%). IL-1 inhibitors were associated with corticosteroids in 285 (54.18%) courses and disease modifying anti-rheumatic drugs (DMARDs) in 156 (29.65%). ANA dosage ranged from 30 to 200 mg/day (or 1.0–2.0 mg/kg/day) among adults and 2–4 mg/kg/day among children; regarding CAN, the most frequently used posologies were 150mg every 8 weeks, 150mg every 4 weeks and 150mg every 6 weeks. The frequency of failure was higher among patients treated with ANA at a dosage of 100 mg/day than those treated with 2 mg/kg/day (p = 0.03). Seventy-six patients (14.4%) reported an adverse event (AE) and 10 (1.9%) a severe AE. AEs occurred more frequently after the age of 65 compared to both children and patients aged between 16 and 65 (p = 0.003 and p = 0.03, respectively). Conclusions: IL-1 inhibitors are mostly used off-label, especially ANA, during adulthood. The high frequency of good clinical responses suggests that IL-1 inhibitors are used with awareness of pathogenetic mechanisms; adult healthcare physicians generally employ standard dosages, while pediatricians are more prone in using a weight-based posology. Dose adjustments and switching between different agents showed to be effective treatment strategies. Our data confirm the good safety profile of IL-1 inhibitors.

Response to interleukin-1 inhibitors in 140 Italian patients with adult-onset still's disease: A multicentre retrospective observational study

Background: Interleukin (IL)-1 plays a crucial role in the pathogenesis of Adult onset Still’s disease (AOSD). Objectives: To evaluate the efficacy and safety of anakinra (ANA) and canakinumab (CAN) in a large group of AOSD patients. Methods: Data on clinical, serological features, and concomitant treatments were retrospectively collected at baseline and after 3, 6, and 12 months from AOSD patients (Yamaguchi criteria) referred by 18 Italian centers. Pouchot’s score was used to evaluate disease severity. Results: One hundred forty patients were treated with ANA; 4 were subsequently switched to CAN after ANA failure. The systemic pattern of AOSD was identified in 104 (74.2%) of the ANA-treated and in 3 (75%) of the CAN-treated groups; the chronic-articular type of AOSD was identified in 48 (25.8%) of the ANA-treated and in 1 (25%) of the CAN-treated groups. Methotrexate (MTX) was the most frequent disease modifying anti-rheumatic drug (DMARD) used before beginning ANA or CAN [91/140 (75.8%), 2/4 (50%), respectively]. As a second-line biologic DMARD therapy in 29/140 (20.7%) of the patients, ANA was found effective in improving all clinical and serological manifestations (p < 0.0001), and Pouchot’s score was found to be significantly reduced at all time points (p < 0.0001). No differences in treatment response were identified in the ANA-group when the patients were stratified according to age, sex, disease pattern or mono/combination therapy profile. ANA primary and secondary inefficacy at the 12-month time point was 15/140 (10.7%) and 11/140 (7.8%), respectively. Adverse events (AEs) [mainly represented by in situ (28/47, 59.5%) or diffuse (12/47, 25.5%) skin reactions and infections (7/47, 14.8%)] were the main causes for discontinuation. Pouchot’s score and clinical and serological features were significantly ameliorated at all time points (p < 0.0001) in the CAN-group, and no AEs were registered during CAN therapy. Treatment was suspended for loss of efficacy only in one case (1/4, 25%). Conclusion: This is the largest retrospective observational study evaluating the efficacy and safety of IL-1 inhibitors in AOSD patients. A good response was noted at 3 months after therapy onset in both the ANA- and CAN-groups. Skin reaction may nevertheless represent a non-negligible AE during ANA treatment.

Liver stiffness correlates with methotrexate cumulative dose in patients with rheumatoid arthritis

BACKGROUND Liver stiffness values were recently proposed to identify patients with methotrexate-induced liver fibrosis. Aim of this study was to assess the clinical and laboratory determinants of the association between liver stiffness, measured by transient elastography, and methotrexate treatment in patients with rheumatoid arthritis in the absence of other factors contributing to liver damage and fibrosis. METHODS 100 patients with rheumatoid arthritis, with a cumulative methotrexate dose ranging from 1530 to 13,000 mg over a mean period of 7.07±3.89 yrs, were retrospectively evaluated. RESULTS The average liver stiffness value in the whole population was 4.93±1.8 kPa, excluding the presence of significant fibrosis. At univariate analysis, a significant correlation was found between liver stiffness and methotrexate cumulative dose, duration of treatment, alanine transaminases levels, body mass index, gamma glutamyl-transpeptidase and the presence of steatosis. At multivariate analysis, a significant association was detected only between liver stiffness and methotrexate cumulative dose. Out of 11 patients with liver stiffness >7.0 kPa, five were subjected to liver biopsy and mild or moderate perisinusoidal fibrosis was detected in two patients with a cumulative dose >4000 mg and liver stiffness >9 kPa. CONCLUSIONS Chronic methotrexate treatment induces a progressive increase in liver stiffness corresponding to mild or moderate perisinusoidal fibrosis for values >9 kPa.

Biochemical analysis of dermal connective tissue in subjects affected by primary uncomplicated varicose veins

Biochemical analysis of dermal connective tissue was carried out in 14 sub jects affected by primary uncomplicated varicose veins and 14 controls. Skin samples were taken, according to fixed criteria, from operation pieces of total mastectomy for breast cancer. The results suggest that the dermal tissue in these subjects is just thinner than that of controls, confirming previous similar clinical findings. The elective reduction of the collagen content observed, unassociated with changes of other components of the dermal connective tissue, brings evidence for a systemic biochemical defect of the extracellular matrix i.e. a collagen de fect affecting the entire body structure and not only the varicose or pre-varicose veins of the lower limbs.

Circulating T cells to infliximab are detectable mainly in treated patients developing anti-drug antibodies and hypersensitivity reactions

Antibodies recognizing infliximab (IFX) may develop in a proportion of treated patients, leading to loss of response or hypersensitivity reactions (HRs). T cell response to IFX has been poorly investigated. This paper was addressed to detect IFX‐specific T cells in treated patients with inflammatory diseases developing, or not, anti‐drug antibodies (ADA) and to correlate the presence of specific T cells with the clinical outcomes of the treatment. A co‐culture system of IFX‐loaded dendritic cells and purified autologous CD4+ T cells was used to detect memory T cells in 32 ADA+ and 39 ADA– IFX‐treated patients and control groups. The cytokine profile of IFX‐specific T cells was also studied in culture supernatants. IFX‐specific cell proliferation was detected mainly in cells from ADA+ patients, irrespective of their different diseases. HR patients displayed higher T cell proliferation than non‐responder and tolerant patients. A mixed [interferon (IFN)‐γ, interleukin (IL)‐13, IL‐10] cytokine profile was shown in cells from ADA+ patients, while IL‐10 was the most frequently detected cytokine in the supernatants of cultures from ADA‐ patients. Immunoglobulin (Ig)E+ADA+ patients with previous HRs exhibited a more pronounced type 2 profile than IgE–ADA+ patients. This work provides evidence that IFX‐specific circulating T cells are detectable mainly in ADA+ patients with HRs, regardless of their disease. The IFX‐induced cytokine pattern partially correlates with the ADA isotype.