Optimization of performance of Dutch newborn screening for cystic fibrosis.

BACKGROUND: Dutch newborn screening (NBS) for Cystic Fibrosis (CF) introduced in 2011 showed a sensitivity of 90% and a positive predictive value (PPV) of 63%. We describe a study including an optimization phase and evaluation of the modified protocol. METHODS: Dutch protocol consists of four steps: determination of immunoreactive trypsinogen (IRT) and pancreatitis-associated protein (PAP), DNA analysis by INNO-LiPA and extended gene analysis (EGA). For the optimization phase we used results of 556,952 newborns screened between April 2011 and June 2014 to calculate effects of 13 alternative protocols on sensitivity, specificity, PPV, ratios of CF to other diagnoses, and costs. One alternative protocol was selected based on calculated sensitivity, PPV and costs and was implemented on 1st July 2016. In this modified protocol DNA analysis is performed in samples with a combination of IRT ≥60 µg/l and PAP ≥3.0 µg/l, IRT ≥100 µg/l and PAP ≥1.2 µg/l or IRT ≥124 µg/l and PAP not relevant. Results of 599,137 newborns screened between 1st July 2016 and 31st December 2019 were similarly evaluated as in the optimization phase. RESULTS: The modified protocol showed a sensitivity of 95%, PPV of 76%, CF to CF transmembrane conductance regulator-related metabolic syndrome/CF screen positive, inconclusive diagnoses (CRMS/CFSPID) ratio 12/1, CF/CF carrier ratio 4/1. Costs per screened newborn were slightly higher. Eleven children, of whom five with classic CF, would not have been referred with the previous protocol. CONCLUSIONS: The modified protocol results in acceptable sensitivity (95%) and good PPV of 76% with minimal increase in costs.

FDA-approved drug screening in patient-derived organoids demonstrates potential of drug repurposing for rare cystic fibrosis genotypes.

BACKGROUND: Preclinical cell-based assays that recapitulate human disease play an important role in drug repurposing. We previously developed a functional forskolin induced swelling (FIS) assay using patient-derived intestinal organoids (PDIOs), allowing functional characterization of CFTR, the gene mutated in people with cystic fibrosis (pwCF). CFTR function-increasing pharmacotherapies have revolutionized treatment for approximately 85% of people with CF who carry the most prevalent F508del-CFTR mutation, but a large unmet need remains to identify new treatments for all pwCF. METHODS: We used 76 PDIOs not homozygous for F508del-CFTR to test the efficacy of 1400 FDA-approved drugs on improving CFTR function, as measured in FIS assays. The most promising hits were verified in a secondary FIS screen. Based on the results of this secondary screen, we further investigated CFTR elevating function of PDE4 inhibitors and currently existing CFTR modulators. RESULTS: In the primary screen, 30 hits were characterized that elevated CFTR function. In the secondary validation screen, 19 hits were confirmed and categorized in three main drug families: CFTR modulators, PDE4 inhibitors and tyrosine kinase inhibitors. We show that PDE4 inhibitors are potent CFTR function inducers in PDIOs where residual CFTR function is either present, or created by additional compound exposure. Additionally, upon CFTR modulator treatment we show rescue of CF genotypes that are currently not eligible for this therapy. CONCLUSION: This study exemplifies the feasibility of high-throughput compound screening using PDIOs. We show the potential of repurposing drugs for pwCF carrying non-F508del genotypes that are currently not eligible for therapies. ONE-SENTENCE SUMMARY: We screened 1400 FDA-approved drugs in CF patient-derived intestinal organoids using the previously established functional FIS assay, and show the potential of repurposing PDE4 inhibitors and CFTR modulators for rare CF genotypes.

Females with cystic fibrosis have a larger decrease in sweat chloride in response to lumacaftor/ivacaftor compared to males.

AIM: To explore which patient-related factors influence sweat test response to CFTR modulators, as well as examining the correlation between the sweat chloride response and ppFEV1 or BMI response, using systematically collected real-life clinical data. METHODS: 160 CF patients were identified who had used lumacaftor/ivacaftor for at least six months. Of these patients, age, sweat chloride levels, ppFEV1 weight and BMI at the start of treatment and after 6 months were collected retrospectively. Pearson and Spearman tests were performed to assess correlations. RESULTS: Females compared to males in this group showed a larger response in sweat chloride (mean difference 10.6 mmol/l, 95% CI: 5.7-15.4) and BMI (mean difference 0.27 kg/m2, 95% CI: 0.01-0.54). A modest but significant correlation was found between patient weight and sweat chloride response (Pearson R = 0.244, p = 0.001), which diminished upon correction for the other factors. The correlation between sex and sweat chloride response remained; R = 0.253, p = 0.001. Sweat chloride response did not correlate with ppFEV1 change or BMI change at 6 months after start of therapy. CONCLUSION: Sweat chloride response is larger in females compared to males, which also explains the negative correlation of weight with the response in sweat chloride concentration after start of lumacaftor/ivacaftor. Sweat chloride response does not correlate with the responses in ppFEV1 and BMI. This information may help the interpretation of sweat test results acquired for the follow up and evaluation of CFTR modulating treatments, and warrants further investigation into the underlying mechanisms of sex differences in response to CFTR modulators.

Clinical effect of lumacaftor/ivacaftor in F508del homozygous CF patients with FEV1 ≥ 90% predicted at baseline.

OBJECTIVE: The first available CFTR modulator combination for homozygous F508del patients, lumacaftor/ivacaftor, has not been tested in patients with percentage predicted (pp)FEV1 > 90 in the phase III trials. The objective of this study is to share real life experience about treatment results in this group. METHODS: In this retrospective observational study, patients aged 6 years or older starting on lumacaftor/ivacaftor in standard care were in strict follow up. For these patients, data were obtained about FEV1, BMI, CFQ-R and sweat chloride before start and after 6 months of treatment, and data about FEV1 and BMI were recorded every 3 months. Exacerbations were recorded continuously. RESULTS: We identified 40 patients with a ppFEV1 ≥ 90 at the start of lumacaftor/ivacaftor who had been in follow up for at least 12 months. After 12 months, ppFEV1 was unchanged, whereas mean absolute change in BMI was +0.88 (p = 0.001) with a mean change in SDS for BMI of +0.26 (p = 0.014). Mean CFQ-R overall score at 6 months improved by 2.6% (p = 0.004) and mean decrease in sweat chloride was -27.3 mEq/L (p = 0.000). Exacerbation rate declined from 1.03 to 0.53/person/year (p = 0.003). One patient discontinued treatment in the first 12 months because of progression of CFRLD, two paused treatment but resumed later. CONCLUSION: Homozygous F508del patients starting lumacaftor/ivacaftor at ppFEV1 ≥ 90 improved significantly in nutritional status, sweat chloride levels and exacerbation rate, but did not respond in ppFEV1. Treatment is well tolerated in this patient group. These effects make it worth considering to treat this group of patients with lumacaftor/ivacaftor.

Limited premature termination codon suppression by read-through agents in cystic fibrosis intestinal organoids.

Premature termination codon read-through drugs offer opportunities for treatment of multiple rare genetic diseases including cystic fibrosis. We here analyzed the read-through efficacy of PTC124 and G418 using human cystic fibrosis intestinal organoids (E60X/4015delATTT, E60X/F508del, G542X/F508del, R1162X/F508del, W1282X/F508del and F508del/F508del). G418-mediated read-through induced only limited CFTR function, but functional restoration of CFTR by PTC124 could not be confirmed. These studies suggest that better read-through agents are needed for robust treatment of nonsense mutations in cystic fibrosis.

A bioassay using intestinal organoids to measure CFTR modulators in human plasma.

Treatment efficacies of drugs depend on patient-specific pharmacokinetic and pharmacodynamic properties. Here, we developed an assay to measure functional levels of the CFTR potentiator VX-770 in human plasma and observed that VX-770 in plasma from different donors induced variable CFTR function in intestinal organoids. This assay can help to understand variability in treatment response to CFTR potentiators by functionally modeling individual pharmacokinetics.

Measurement of nasal nitric oxide: evaluation of six different sampling methods.

BACKGROUND: Specific guidelines are developed for the measurement of bronchial FE(NO), however, nasal nitric oxide (nNO) measurement is not standardised yet, resulting in divergent nNO values. This study compares six different sampling methods for nNO as described in the literature, to analyse their outcome and short term and long term reproducibility. DESIGN: nNO concentrations were measured in 38 healthy subjects. Each subject performed nNO measurements during tidal breathing (nNO-TB), single breath quiet exhalations (nNO-QE), QE with oral exhalation against a resistance (nNO-QE + R), breath holding (nNO-BH) and during single-breath humming exhalations at 128 and 440 Hz (nNO-HE(128) and nNO-HE(440), respectively). To assess short term and long term reproducibility all manoeuvres were repeated after one and 24 h. RESULTS: Lowest values were found during quiet exhalation (mean nNO-QE was 364 p.p.b., SEM 27). Methods in which there is turbulence of nasal flow (as in TB, HE(128) and HE(440)) result in higher nNO levels. Highest values were found in methods with decreased nasal flow [when there is no nasal flow as in BH or when the velum is closed as in QE + R: mean nNO 763 p.p.b. (SEM 61)]. NNO during humming at 440 Hz was significantly higher than at 128 Hz (P < 0.01). The within-subject coefficient of variation of repeated measurements was lowest during humming and breath holding, 3.4 and 3.8%, respectively. Concerning short term and long term reproducibility, best agreement is reached with humming and second best with breath holding. CONCLUSIONS: Different methods result in different levels and reproducibility of nNO. In regard to this, methods of humming and breath holding are recommended for standardised measurement of nasal NO.

Excision of a giant hydatid cyst of the lung under thoracic epidural anaesthesia.

We present a patient with a large pulmonary hydatid cyst compressing underlying lung, with previous pulmonary tuberculosis, who presented in respiratory failure. After institution of thoracic epidural anaesthesia employing 0.25% bupivacaine, 1% lignocaine and fentanyl, the patient was placed in the sitting position and the hydatid cyst excised and drained after a limited rib resection. An air leak persisted until the 16th postoperative day. A marked improvement in symptoms as well as in spirometly and arterial blood gases occurred, and the patient was discharged on the 20th day. Thoracic epidural anaesthesia may be a safer method than general anaesthesia for removal of a hydatid cyst in a patient with severe respiratory compromise.

Blunt cardiac rupture.

Cardiac rupture as a result of blunt trauma is not commonly encountered. Seven patients with this injury have been treated at Groote Schuur Hospital over the past 14 years. All presented with cardiovascular collapse and 4 developed signs of cardiac tamponade. A clinical diagnosis was made in 4 patients and echocardiography was done in 3. Pericardiocentesis was used in 1 patient to confirm the diagnosis. Significant diagnostic delay occurred in 1 patient with associated liver rupture. Two patients required emergency room thoracotomy. All other patients were approached using a median sternotomy. Five patients survived, giving an overall survival rate of 71%. Five patients had right atrial ruptures and 2 right ventricular ruptures. One patient with right ventricular rupture died in the operating room, while another patient with multiple right atrial ruptures died from multiple organ failure after 11 days. We also briefly review the history, mechanisms and pathology.

Penetrating injuries involving the intrathoracic great vessels.

Forty-four consecutive patients with injuries to the intrathoracic great vessels admitted to our department from January 1982 to June 1994 were reviewed retrospectively. Forty-two patients (95%) sustained stabwounds and 2 (5%) patients had gunshot wounds. The most frequent radiological abnormality was mediastinal widening in 26 patients (59%). Eighteen patients (41%) were haemodynamically stable on admission with the remainder being unstable (46%), agonal (11%) or lifeless (2%). Twenty-two patients (50%) underwent angiography with 1 false-negative study. A total of 48 arterial and 16 venous injuries were identified with the innominate artery (N = 17, 39% of patients) and left innominate vein (N = 8, 18% of patients) the most frequently injured structures. Associated injuries to thoracic viscera occurred in 13 patients (30%). Two patients required cardiopulmonary bypass to repair their injuries. Arterial shunts were not used in any case. Overall mortality was 5% (2/44) and complications occurred in 7 patients (16%).

Intramuscular haemangioma of the chest wall. A case report.

Intramuscular haemangiomas of thoracic skeletal muscle are uncommon tumours. They are locally invasive and tend to recur if not completely and widely excised. This report illustrates the diagnosis and management of a 25-year-old man with an intramuscular haemangioma of the chest wall. A literature report on the aetiology and management of these tumours is also given.

Syphilitic aortic aneurysm eroding through the sternum.

Syphilitic aortic aneurysms are uncommon today. A syphilitic aneurysm eroding through the anterior chest wall with successful surgical treatment is reported. The large size these aneurysms reach, in conjunction with the overlying pressure-induced skin necrosis necrosis can represent a technical challenge to the surgeon, both in the method of repairing the aneurysm as well as reconstructing the chest wall. Syphilitic aortic disease is also briefly reviewed.

Spinal cord protection in the absence of collateral circulation: meta-analysis of mortality and paraplegia.

A meta-analysis of paraplegia complicating aortic surgery on patients having neither intercostal nor spinal collaterals, epitomized by patients with acute traumatic aortic rupture, was done. Index Medicus and Medline were searched for all suitable English publications between 1972 and 1992. New paraplegia occurred in 9.9% of 1492 patients who underwent surgery. However, 19.2% of patients undergoing surgery with only simple aortic cross-clamping developed paraplegia, in contrast to 6.1% if distal aortic perfusion was augmented by either "passive" or "active" methods (p < 0.00001). The risk of paraplegia increased progressively as cross-clamp times lengthened if simple aortic cross-clamping was used (p < 0.00001), but only once did the cross-clamp time exceed 30 minutes (p < 0.05). Paraplegia occurred in 8.2% of patients with "passive" shunts from the ascending aorta (p < 0.001 vs simple cross-clamping). Shunts from the left ventricular apex, however, had an incidence of paraplegia of 26.1% and, therefore, did not decrease the risk of paraplegia. "Active" augmentation of distal perfusion had the lowest risk of paraplegia: 2.3% (p < 0.00001 vs simple cross-clamping or "passive" shunts). Mortality, however, was higher in these potentially polytraumatized patients when they were perfused distally using methods requiring full systemic heparinization (18.2%), compared to mortality with methods not requiring heparin (11.9%; p < 0.01). In conclusion, simple aortic cross-clamping has a high risk of paraplegia if the cross-clamp time extends beyond 30 minutes. "Active" modalities of augmenting distal perfusion provide optimal spinal protection.

p53 gene mutants expression, cellular proliferation and differentiation in oesophageal carcinoma and non-cancerous epithelium.

An immunohistochemical study of 155 squamous cell carcinomas of the oesophagus, squamous epithelium adjacent to the tumour (n = 80), dysplastic epithelium (n = 16) and controls (n = 23) indicates that Ki67 and p53 expression is frequently present in premalignant oesophageal lesions and their related squamous cell carcinomas. Positive expression of both antibodies in apparently normal epithelium can signify early steps of malignant transformation of oesophageal epithelium and can serve in the detection of early precancerous lesions. The expression of growth factors EGF and TGF-alpha was higher in carcinomas and dysplastic lesions than in apparently normal squamous epithelium. EGF expression was unevenly distributed according to histological grade, indicating a lack of EGF immunoreactivity in poorly differentiated oesophageal carcinomas.

Intrinsic obstruction of the Procter-Livingstone tube.

Four instances of intrinsic obstruction of a Procter-Livingstone tube inserted for oesophageal carcinoma are described. The exact cause of the blistering is unknown, but it is thought to occur by absorption through the latex. It is suggested that if this complication is found, relief of obstruction may be obtained by perforation of the blister.