Manufacturing classification system in the real world: factors influencing manufacturing process choices for filed commercial oral solid dosage formulations, case studies from industry and considerations for continuous processing.

Following the first Manufacturing Classification System (MCS) paper, the team conducted surveys to establish which active pharmaceutical ingredient (API) properties were important when selecting or modifying materials to enable an efficient and robust pharmaceutical manufacturing process. The most commonly identified factors were (1) API particle size: small particle sizes are known to increase risk of processing issues; (2) Drug loading in the formulation: high drug loadings allow less opportunity to mitigate poor API properties through the use of excipients. The next step was to establish linkages with process decisions by identifying publicly-available proxies for these important parameters: dose (in place of drug loading) and BCS class (in place of particle size). Poorly-soluble API were seen as more likely to have controlled (smaller) particle size than more highly soluble API. Analysis of 435 regulatory filings revealed that higher doses and more poorly-soluble API was associated with more complex processing routes. Replacing the proxy factors with the original parameters should give the opportunity to demonstrate stronger trends. This assumption was tested by accessing a dataset relating to commercial tablet products. This showed that, for dry processes, a larger particle size was associated with higher achievable drug loading as determined by percolation threshold.

Lower serum extracellular superoxide dismutase levels are associated with polyneuropathy in recent-onset diabetes.

Increased oxidative stress is implicated in the pathogenesis of experimental diabetic neuropathy, but translational evidence in recent-onset diabetes is scarce. We aimed to determine whether markers of systemic oxidative stress are associated with diabetic sensorimotor polyneuropathy (DSPN) in recent-onset diabetes. In this cross-sectional study, we measured serum concentrations of extracellular superoxide dismutase (SOD3), thiobarbituric acid reactive substances (TBARS), and reduced glutathione (GSH) in 107 type 1 and 215 type 2 diabetes patients from the German Diabetes Study baseline cohort and 37 glucose-tolerant individuals (controls). DSPN was defined by electrophysiological and clinical criteria (Toronto Consensus, 2011). SOD3 and GSH concentrations were lower in individuals with type 1 and type 2 diabetes compared with concentrations in controls (P<0.0001). In contrast, the TBARS concentration was higher in participants with type 1 diabetes and type 2 diabetes compared with levels in controls (P<0.0001). In addition, the SOD3 concentration was higher in participants with type 1 diabetes compared to concentrations in those with type 2 diabetes (P<0.0001). A low SOD3 concentration was associated with DSPN in individuals with type 1 diabetes (ß=-0.306, P=0.002), type 2 diabetes (ß=-0.164, P=0.017), and in both groups combined (ß=-0.206, P=0.0003). Lower SOD3 concentrations were associated with decreased motor nerve conduction velocity (NCV) in men and, to a lesser degree, with reduced sensory NCV in women with diabetes. In conclusion, several biomarkers of oxidative stress are altered in recent-onset diabetes, with only a lower SOD3 concentration being linked to the presence of DSPN, suggesting a role for reduced extracellular antioxidative defense against superoxide in the early development of DSPN.

Spatial analysis improves the detection of early corneal nerve fiber loss in patients with recently diagnosed type 2 diabetes.

Corneal confocal microscopy (CCM) has revealed reduced corneal nerve fiber (CNF) length and density (CNFL, CNFD) in patients with diabetes, but the spatial pattern of CNF loss has not been studied. We aimed to determine whether spatial analysis of the distribution of corneal nerve branching points (CNBPs) may contribute to improving the detection of early CNF loss. We hypothesized that early CNF decline follows a clustered rather than random distribution pattern of CNBPs. CCM, nerve conduction studies (NCS), and quantitative sensory testing (QST) were performed in a cross-sectional study including 86 patients recently diagnosed with type 2 diabetes and 47 control subjects. In addition to CNFL, CNFD, and branch density (CNBD), CNBPs were analyzed using spatial point pattern analysis (SPPA) including 10 indices and functional statistics. Compared to controls, patients with diabetes showed lower CNBP density and higher nearest neighbor distances, and all SPPA parameters indicated increased clustering of CNBPs (all P<0.05). SPPA parameters were abnormally increased >97.5th percentile of controls in up to 23.5% of patients. When combining an individual SPPA parameter with CNFL, ≥1 of 2 indices were >99th or <1st percentile of controls in 28.6% of patients compared to 2.1% of controls, while for the conventional CNFL/CNFD/CNBD combination the corresponding rates were 16.3% vs 2.1%. SPPA parameters correlated with CNFL and several NCS and QST indices in the controls (all P<0.001), whereas in patients with diabetes these correlations were markedly weaker or lost. In conclusion, SPPA reveals increased clustering of early CNF loss and substantially improves its detection when combined with a conventional CCM measure in patients with recently diagnosed type 2 diabetes.

Overexpression of cutaneous mitochondrial superoxide dismutase in recent-onset type 2 diabetes.

AIMS/HYPOTHESIS: Oxidative stress and microvascular damage have been implicated in the pathogenesis of diabetic neuropathy, with manganese superoxide dismutase 2 (SOD2) responsible for superoxide detoxification in mitochondria. We hypothesised that patients with recently diagnosed type 2 diabetes would show an altered cutaneous expression of SOD2 and endothelial cell area. METHODS: In this cross-sectional study, we assessed skin biopsies using immunohistochemistry, peripheral nerve function and heart rate variability in 69 participants of the German Diabetes Study with recently diagnosed type 2 diabetes and 51 control individuals. RESULTS: Subepidermal SOD2 area in the distal leg was increased by ~60% in the diabetic group vs the controls (0.24 ± 0.02% vs 0.15 ± 0.02%; p = 0.0005) and was correlated with an increasing duration of diabetes (r = 0.271; p = 0.024) and with the low frequency/high frequency ratio (ß = 0.381; p = 0.002) as an indicator of sympathovagal balance. The area of the subepidermal endothelial cells (measured by CD31 staining) did not differ between the groups. CONCLUSIONS/INTERPRETATION: Cutaneous antioxidative defence is enhanced in relation to the duration of diabetes and is linked to a cardiac sympathovagal imbalance towards a sympathetic predominance in individuals with recently diagnosed type 2 diabetes without evidence of endothelial cell damage. Whether cutaneous SOD2 levels can predict the development of diabetic neuropathy remains to be determined in prospective studies.

Early detection of nerve fiber loss by corneal confocal microscopy and skin biopsy in recently diagnosed type 2 diabetes.

We sought to determine whether early nerve damage may be detected by corneal confocal microscopy (CCM), skin biopsy, and neurophysiological tests in 86 recently diagnosed type 2 diabetic patients compared with 48 control subjects. CCM analysis using novel algorithms to reconstruct nerve fiber images was performed for all fibers and major nerve fibers (MNF) only. Intraepidermal nerve fiber density (IENFD) was assessed in skin specimens. Neurophysiological measures included nerve conduction studies (NCS), quantitative sensory testing (QST), and cardiovascular autonomic function tests (AFTs). Compared with control subjects, diabetic patients exhibited significantly reduced corneal nerve fiber length (CNFL-MNF), fiber density (CNFD-MNF), branch density (CNBD-MNF), connecting points (CNCP), IENFD, NCS, QST, and AFTs. CNFD-MNF and IENFD were reduced below the 2.5th percentile in 21% and 14% of the diabetic patients, respectively. However, the vast majority of patients with abnormal CNFD showed concomitantly normal IENFD and vice versa. In conclusion, CCM and skin biopsy both detect nerve fiber loss in recently diagnosed type 2 diabetes, but largely in different patients, suggesting a patchy manifestation pattern of small fiber neuropathy. Concomitant NCS impairment points to an early parallel involvement of small and large fibers, but the precise temporal sequence should be clarified in prospective studies.

Pronounced reduction of cutaneous Langerhans cell density in recently diagnosed type 2 diabetes.

Immune-mediated processes have been implicated in the pathogenesis of diabetic polyneuropathy. Langerhans cells (LCs) are the sole dendritic cell type located in the healthy epidermis and exert tolerogenic immune functions. We aimed to determine whether alterations in cutaneous LC density and intraepidermal nerve fiber density (IENFD) are present in patients with recently diagnosed type 2 diabetes. Skin biopsy specimens from the distal leg from 96 type 2 diabetic patients and 75 healthy control subjects were used for quantification of LC density and IENFD. LCs and IENFs were labeled using immunohistochemistry. Nerve conduction studies, quantitative sensory testing, and neurological examination were used to assess peripheral nerve function. LC density was markedly reduced in the diabetic group compared with the control group, but did not correlate with reduced IENFD or peripheral nerve function. Multivariate linear regression analysis revealed a strong association between LC density and whole-body insulin sensitivity in women but not in men with diabetes. Prospective studies should establish whether the pronounced reduction of cutaneous LCs detected in recently diagnosed type 2 diabetes could promote a cutaneous immunogenic imbalance toward inflammation predisposing to polyneuropathy and foot ulcers.