The immune system prevents recurrence of transplanted but not autochthonous antigenic tumors after oncogene inactivation therapy

Targeted oncogene inactivation by small molecule inhibitors can be very effective but tumor recurrence is a frequent problem in the clinic. Therapy by inactivation of the cancer-driving oncogene in transplanted tumors was shown to be augmented in the presence of T cells. However, these experiments did not take into account the long-term, usually tolerogenic, interaction of de novo malignancies with the immune system. Here, we employed mice, in which SV40 large T (Tag) and firefly luciferase (Luc) as fusion protein (TagLuc) could be regulated with the Tet-on system and upon activation resulted in tumors after a long latency. TagLuc inactivation induced profound tumor regression, demonstrating sustained oncogene addiction. While tumor relapse after TagLuc inactivation was prevented in immunocompetent mice bearing transplanted tumors, autochthonous tumors relapsed or recurred after therapy discontinuation indicating that the immune system that coevolved with the malignancy over an extended period of time lost the potency to mount an efficient anti-tumor immune response. By contrast, adoptively transferred CD8(+) T cells targeting the cancer-driving oncogene eradicated recurrent autochthonous tumors, highlighting a suitable therapy option in a clinically relevant model

H3.3K27M mutation is not a suitable target for immunotherapy in HLA-A2(+) patients with diffuse midline glioma

Diffuse midline glioma is the leading cause of solid cancer-related deaths in children with very limited treatment options. A majority of the tumors carry a point mutation in the histone 3 variant (H3.3) creating a potential HLA-A*02:01 binding epitope (H3.3K27M(26-35)). Here, we isolated an H3.3K27M-specific T cell receptor (TCR) from transgenic mice expressing a diverse human TCR repertoire. Despite a high functional avidity of H3.3K27M-specific T cells, we were not able to achieve recognition of cells naturally expressing the H3.3K27M mutation, even when overexpressed as a transgene. Similar results were obtained with T cells expressing the published TCR 1H5 against the same epitope. CRISPR/Cas9 editing was used to exclude interference by endogenous TCRs in donor T cells. Overall, our data provide strong evidence that the H3.3K27M mutation is not a suitable target for cancer immunotherapy, most likely due to insufficient epitope processing and/or amount to be recognized by HLA-A*02:01 restricted CD8(+) T cells

Quality of compounded hydrocortisone capsules used in the treatment of children.

Objectives: Due to the lack of paediatric licensed formulations, children are often treated with individualized pharmacy-compounded adult medication. An international web-based survey about the types of medication in children with adrenal insufficiency (AI) revealed that the majority of paediatric physicians are using pharmacy-compounded medication to treat children with AI. Observations of loss of therapy control in children with congenital adrenal hyperplasia with compounded hydrocortisone capsules and regained control after prescribing a new hydrocortisone batch led to this "real world" evaluation of pharmacy-compounded paediatric hydrocortisone capsules. Methods: Capsule samples were collected randomly from volunteering parents of treated children suffering from congenital adrenal hyperplasia from all over Germany. Analysis of net mass and hydrocortisone content by HPLC-UV (high performance liquid chromatography) method were performed based on the European Pharmacopeia. Results: In total 61 batches were sent in. 5 batches could not be analyzed because of missing dose information, insufficient number of capsules or were not possible to be evaluated. 56 batches containing 1125 capsules were evaluated. 21.4% of the batches revealed insufficiency in uniformity of net mass or drug content and additional 3.6% failed because they did not contain the labelled drug. Conclusions: Compounded medication is a possible cause of variation of steroid doses in children with adrenal insufficiency or congenital adrenal hyperplasia, putting these vulnerable patients at risk of poor disease control and adrenal crisis. These data may apply to other individualized compounded oral medication as well, emphasizing the need for development of licensed paediatric formulations approved by regulatory authorities

Insights in the maturational processes influencing hydrocortisone pharmacokinetics in congenital adrenal hyperplasia patients using a middle-out approach

Introduction: Hydrocortisone is the standard of care in cortisol replacement therapy for congenital adrenal hyperplasia patients. Challenges in mimicking cortisol circadian rhythm and dosing individualization can be overcome by the support of mathematical modelling. Previously, a non-linear mixed-effects (NLME) model was developed based on clinical hydrocortisone pharmacokinetic (PK) pediatric and adult data. Additionally, a physiologically-based pharmacokinetic (PBPK) model was developed for adults and a pediatric model was obtained using maturation functions for relevant processes. In this work, a middle-out approach was applied. The aim was to investigate whether PBPK-derived maturation functions could provide a better description of hydrocortisone PK inter-individual variability when implemented in the NLME framework, with the goal of providing better individual predictions towards precision dosing at the patient level. Methods: Hydrocortisone PK data from 24 adrenal insufficiency pediatric patients and 30 adult healthy volunteers were used for NLME model development, while the PBPK model and maturation functions of clearance and cortisol binding globulin (CBG) were developed based on previous studies published in the literature. Results: Clearance (CL) estimates from both approaches were similar for children older than 1 year (CL/F increasing from around 150 L/h to 500 L/h), while CBG concentrations differed across the whole age range (CBGNLME stable around 0.5 μM vs. steady increase from 0.35 to 0.8 μM for CBG PBPK). PBPK-derived maturation functions were subsequently included in the NLME model. After inclusion of the maturation functions, none, a part of, or all parameters were re-estimated. However, the inclusion of CL and/or CBG maturation functions in the NLME model did not result in improved model performance for the CL maturation function (ΔOFV > −15.36) and the re-estimation of parameters using the CBG maturation function most often led to unstable models or individual CL prediction bias. Discussion: Three explanations for the observed discrepancies could be postulated, i) non-considered maturation of processes such as absorption or first-pass effect, ii) lack of patients between 1 and 12 months, iii) lack of correction of PBPK CL maturation functions derived from urinary concentration ratio data for the renal function relative to adults. These should be investigated in the future to determine how NLME and PBPK methods can work towards deriving insights into pediatric hydrocortisone PK

A prospective study of children aged 0–8 years with CAH and adrenal insufficiency treated with hydrocortisone granules

Context Children with congenital adrenal hyperplasia (CAH) and adrenal insufficiency (AI) require daily hydrocortisone replacement with accurate dosing. Objective Prospective study of efficacy and safety of hydrocortisone granules in children with AI and CAH monitored by 17-OHP (17-hydroxyprogesterone) saliva profiles. Methods Seventeen children with CAH (9 male) and 1 with hypopituitarism (male), aged from birth to 6 years, had their hydrocortisone medication changed from pharmacy compounded capsules to hydrocortisone granules. Patients were followed prospectively for 2 years. In children with CAH, the therapy was adjusted by 17-OHP salivary profiles every 3 months. The following parameters were recorded: hydrocortisone dose, height, weight, pubertal status, adverse events, and incidence of adrenal crisis. Results The study medication was given thrice daily, and the median duration of treatment (range) was 795 (1–872) days, with 150 follow-up visits. Hydrocortisone doses were changed on 40/150 visits, with 32 based on salivary measurements and 8 on serum 17-OHP levels. The median daily mg/m2 hydrocortisone dose (range) at study entry for the different age groups 2–8 years, 1 month to 2 years, <28 days was 11.9 (7.2–15.5), 9.9 (8.6–12.2), and 12.0 (11.1–29.5), respectively, and at end of the study was 10.2 (7.0–14.4), 9.8 (8.9–13.1), and 8.6 (8.2–13.7), respectively. There were no trends for accelerated or reduced growth. No adrenal crises were observed despite 193 treatment-emergent adverse events, which were mainly common childhood illnesses. Interpretation This first prospective study of glucocorticoid treatment in children with AI and CAH demonstrates that accurate dosing and monitoring from birth results in hydrocortisone doses at the lower end of the recommended dose range and normal growth, without occurrence of adrenal crises

Model-Informed target morning 17α-hydroxyprogesterone concentrations in dried blood spots for pediatric congenital adrenal hyperplasia patients

Monitoring cortisol replacement therapy in congenital adrenal hyperplasia (CAH) patients is vital to avoid serious adverse events such as adrenal crises due to cortisol underexposure or metabolic consequences due to cortisol overexposure. The less invasive dried blood spot (DBS) sampling is an advantageous alternative to traditional plasma sampling, especially in pediatric patients. However, target concentrations for important disease biomarkers such as 17α-hydroxyprogesterone (17-OHP) are unknown using DBS. Therefore, a modeling and simulation framework, including a pharmacokinetic/pharmacodynamic model linking plasma cortisol concentrations to DBS 17-OHP concentrations, was used to derive a target morning DBS 17-OHP concentration range of 2–8 nmol/L in pediatric CAH patients. Since either capillary or venous DBS sampling is becoming more common in the clinics, the clinical applicability of this work was shown by demonstrating the comparability of capillary and venous cortisol and 17-OHP concentrations collected by DBS sampling, using a Bland-Altman and Passing-Bablok analysis. The derived target morning DBS 17-OHP concentration range is a first step towards providing improved therapy monitoring using DBS sampling and adjusting hydrocortisone (synthetic cortisol) dosing in children with CAH. In the future, this framework can be used to assess further research questions, e.g., target replacement ranges for the entire day

International practice of corticosteroid replacement therapy in congenital adrenal hyperplasia - data from the I-CAH registry.

OBJECTIVE: Despite published guidelines no unified approach to hormone replacement in congenital adrenal hyperplasia (CAH) exists. We aimed to explore geographical and temporal variations in the treatment with glucocorticoids and mineralocorticoids in CAH. DESIGN: This retrospective multi-center study, including 31 centers (16 countries), analyzed data from the International-CAH Registry. METHODS: Data was collected from 461 patients aged 0-18 years with classic 21-hydroxylase deficiency (54.9% females) under follow-up between 1982 - 2018. Type, dose and timing of glucocorticoid and mineralocorticoid replacement was analyzed from 4174 patient visits. RESULTS: The most frequently used glucocorticoid was hydrocortisone (87.6%). Overall, there were significant differences between age groups with regards to daily hydrocortisone-equivalent dose for body surface, with the lowest dose (median with interquartile range) of 12.0 (10.0 - 14.5) mg/ m2/ day at age 1 - 8 years and the highest dose of 14.0 (11.6 - 17.4) mg/ m2/ day at age 12-18 years. Glucocorticoid doses decreased after 2010 in patients 0-8 years (p<0.001) and remained unchanged in patients aged 8-18 years. Fludrocortisone was used in 92% of patients, with relative doses decreasing with age. A wide variation was observed among countries with regards to all aspects of steroid hormone replacement. CONCLUSIONS: Data from the I-CAH Registry suggests international variations in hormone replacement therapy, with a tendency to treatment with high doses in children