MRI-guided optimisation of neoadjuvant chemotherapy duration in stage II-III HER2-positive breast cancer (TRAIN-3): a multicentre, single-arm, phase 2 study.

BACKGROUND: Patients with stage II-III HER2-positive breast cancer have good outcomes with the combination of neoadjuvant chemotherapy and HER2-targeted agents. Although increasing the number of chemotherapy cycles improves pathological complete response rates, early complete responses are common. We investigated whether the duration of chemotherapy could be tailored on the basis of radiological response. METHODS: TRAIN-3 is a single-arm, phase 2 study in 43 hospitals in the Netherlands. Patients with stage II-III HER2-positive breast cancer aged 18 years or older and a WHO performance status of 0 or 1 were enrolled. Patients received neoadjuvant chemotherapy consisting of paclitaxel (80 mg/m2 of body surface area on day 1 and 8 of each 21 day cycle), trastuzumab (loading dose on day 1 of cycle 1 of 8 mg/kg bodyweight, and then 6 mg/kg on day 1 on all subsequent cycles), and carboplatin (area under the concentration time curve 6 mg/mL per min on day 1 of each 3 week cycle) and pertuzumab (loading dose on day 1 of cycle 1 of 840 mg, and then 420 mg on day 1 of each subsequent cycle), all given intravenously. The response was monitored by breast MRI every three cycles and lymph node biopsy. Patients underwent surgery when a complete radiological response was observed or after a maximum of nine cycles of treatment. The primary endpoint was event-free survival at 3 years; however, follow-up for the primary endpoint is ongoing. Here, we present the radiological and pathological response rates (secondary endpoints) of all patients who underwent surgery and the toxicity data for all patients who received at least one cycle of treatment. Analyses were done in hormone receptor-positive and hormone receptor-negative patients separately. This trial is registered with ClinicalTrials.gov, number NCT03820063, recruitment is closed, and the follow-up for the primary endpoint is ongoing. FINDINGS: Between April 1, 2019, and May 12, 2021, 235 patients with hormone receptor-negative cancer and 232 with hormone receptor-positive cancer were enrolled. Median follow-up was 26·4 months (IQR 22·9-32·9) for patients who were hormone receptor-negative and 31·6 months (25·6-35·7) for patients who were hormone receptor-positive. Overall, the median age was 51 years (IQR 43-59). In 233 patients with hormone receptor-negative tumours, radiological complete response was seen in 84 (36%; 95% CI 30-43) patients after one to three cycles, 140 (60%; 53-66) patients after one to six cycles, and 169 (73%; 66-78) patients after one to nine cycles. In 232 patients with hormone receptor-positive tumours, radiological complete response was seen in 68 (29%; 24-36) patients after one to three cycles, 118 (51%; 44-57) patients after one to six cycles, and 138 (59%; 53-66) patients after one to nine cycles. Among patients with a radiological complete response after one to nine cycles, a pathological complete response was seen in 147 (87%; 95% CI 81-92) of 169 patients with hormone receptor-negative tumours and was seen in 73 (53%; 44-61) of 138 patients with hormone receptor-positive tumours. The most common grade 3-4 adverse events were neutropenia (175 [37%] of 467), anaemia (75 [16%]), and diarrhoea (57 [12%]). No treatment-related deaths were reported. INTERPRETATION: In our study, a third of patients with stage II-III hormone receptor-negative and HER2-positive breast cancer had a complete pathological response after only three cycles of neoadjuvant systemic therapy. A complete response on breast MRI could help identify early complete responders in patients who had hormone receptor negative tumours. An imaging-based strategy might limit the duration of chemotherapy in these patients, reduce side-effects, and maintain quality of life if confirmed by the analysis of the 3-year event-free survival primary endpoint. Better monitoring tools are needed for patients with hormone receptor-positive and HER2-positive breast cancer. FUNDING: Roche Netherlands.

Biotransformation of Trastuzumab and Pertuzumab in Breast Cancer Patients Assessed by Affinity Enrichment and Ion-Exchange Chromatography.

Therapeutic proteins (TPs) are known to be heterogeneous due to modifications that occur during the production process and storage. Modifications may also occur in TPs after their administration to patients due to in vivo biotransformation. Ligand binding assays, which are widely used in the bioanalysis of TPs in body fluids, are typically unable to distinguish such modifications. Liquid chromatography coupled to mass spectrometry is being increasingly used to study modifications in TPs, but its use to study in vivo biotransformation has been limited until now. We present a novel approach that combines affinity enrichment using Affimer reagents with ion-exchange chromatography (IEX) to analyze charge variants of the TPs trastuzumab and pertuzumab in plasma of patients undergoing therapy for HER2-positive breast cancer. Affimer reagents were immobilized via engineered Cys tags to maleimide beads, and the TPs were eluted under acidic conditions followed by rapid neutralization. The enriched TPs were analyzed by cation-exchange chromatography (IEX) using pH-gradient elution, resulting in the separation of about 20 charge variants for trastuzumab and about five charge variants for pertuzumab. A comparison between in vitro stressed TPs spiked into plasma, and TPs enriched from patient plasma showed that the observed profiles were highly similar. This indicates that in vitro stress testing in plasma can mimic the situation in patient plasma, as far as the generation of charge variants is concerned. SIGNIFICANCE STATEMENT: This research attempts to elucidate the modifications that occur in therapeutic proteins (TPs) after they have been administered to patients. This is important because there is little knowledge about the fate of TPs in this regard, and certain modifications could affect their efficiency. Our results show that the modifications discovered are most likely due to a chemical process and are not patient specific.

Analytical and pharmacological consequences of the in vivo deamidation of trastuzumab and pertuzumab.

A liquid chromatography-tandem mass spectrometry method is presented for the quantitative determination of the in vivo deamidation of the biopharmaceutical proteins trastuzumab and pertuzumab at an asparagine in their complementarity determining regions (CDRs). For each analyte, two surrogate peptides are quantified after tryptic digestion of the entire plasma protein content: one from a stable part of the molecule, representing the total concentration, and one containing the deamidation-sensitive asparagine, corresponding to the remaining non-deamidated concentration. Using a plasma volume of 10 µL and a 2-h digestion at pH 7, concentrations between 2 and 1000 µg/mL can be determined for the various protein forms with values for bias and CV below 15% and without unacceptable in vitro deamidation taking place. A considerable difference between the total and non-deamidated concentrations, and thus a substantial degree of deamidation, was observed in plasma for both trastuzumab and pertuzumab. After a 56-day forced deamidation test 40% of trastuzumab and 68% of pertuzumab was deamidated, while trastuzumab and pertuzumab showed up to 47% and 35% of deamidation, respectively, in samples collected from breast cancer patients during treatment with a combination of both drugs. A good correlation between the non-deamidated concentration results and those of a receptor binding assay indicate a loss of receptor binding for both trastuzumab and pertuzumab along with the deamidation in their CDRs. Deamidated trastuzumab also lost its capability to inhibit the growth of breast cancer cells in a cell-based viability assay, suggesting a relation between the degree of deamidation and pharmacological activity.

Effects, costs and implementation of monitoring kidney transplant patients' tacrolimus levels with dried blood spot sampling: A randomized controlled hybrid implementation trial.

AIMS: Dried blood spot (DBS) home sampling allows monitoring creatinine levels and tacrolimus trough levels as an alternative for blood sampling in the hospital, which is important in kidney transplant patient follow-up. This study aims to assess whether DBS home sampling results in decreased patient travel burden and lower societal costs. METHODS: In this single-centre randomized controlled hybrid implementation trial, adult kidney transplant patients were enrolled. The intervention group (n = 25) used DBS home sampling on top of usual care in the first 6 months after transplantation. The control group (n = 23) received usual care only. The primary endpoint was the number of outpatient visits. Other endpoints were costs per patient, patient satisfaction and implementation. RESULTS: There was no statistically significant difference in the average number of outpatient visits between the DBS group (11.2, standard deviation: 1.7) and the control group (10.9, standard deviation: 1.4; P = .48). Average costs per visit in the DBS group were not significantly different (€542, 95% confidence interval €316-990) compared to the control group (€533, 95% confidence interval €278-1093; P = .66). Most patients (n = 19/23, 82.6%) were willing to perform DBS home-sampling if this would reduce the number of hospital visits. Only 55.9% (n = 143/256) of the expected DBS samples were received and 1/5 analysed on time (n = 52/256). CONCLUSION: Adult kidney transplant patients are willing to perform DBS home sampling. However, to decrease patient travel burden and costs in post-transplant care, optimization of the logistical process concerning mailing and analysis of DBS samples is crucial.

Neoadjuvant chemotherapy with or without anthracyclines in the presence of dual HER2 blockade for HER2-positive breast cancer (TRAIN-2): a multicentre, open-label, randomised, phase 3 trial.

BACKGROUND: The optimal chemotherapy backbone for dual HER2 blockade in the neoadjuvant setting for early breast cancer is unknown. We investigated whether the addition of anthracyclines would improve pathological complete response compared with a carboplatin-taxane regimen, when given in combination with the HER2-targeted agents trastuzumab and pertuzumab. METHODS: The TRAIN-2 study is an open-label, randomised, controlled, phase 3 trial being done in 37 hospitals in the Netherlands. We recruited patients aged 18 years or older with previously untreated, histologically confirmed stage II-III HER2-positive breast cancer. Patients were randomly allocated using central randomisation software (1:1 ratio) with minimisation without a random component, stratified by tumour stage, nodal stage, oestrogen receptor status, and age, to receive 5-fluorouracil (500 mg/m2), epirubicin (90 mg/m2), and cyclophosphamide (500 mg/m2) every 3 weeks for three cycles followed by paclitaxel (80 mg/m2 on days 1 and 8) and carboplatin (area under the concentration-time curve [AUC] 6 mg/mL per min on day 1 or optionally, as per hospital preference, AUC 3 mg/mL per min on days 1 and 8) every 3 weeks for six cycles, or to receive nine cycles of paclitaxel and carboplatin at the same dose and schedule as in the anthracycline group. Patients in both study groups received trastuzumab (6 mg/kg, loading dose 8 mg/kg) and pertuzumab (420 mg, loading dose 840 mg) concurrently with all chemotherapy cycles. The primary endpoint was the proportion of patients who achieved a pathological complete response in breast and axilla (ypT0/is ypN0) in the intention-to-treat population. Safety was analysed in patients who received at least one treatment cycle according to actual treatment received. This trial is registered with ClinicalTrials.gov, number NCT01996267, and follow-up for long-term outcome is ongoing. FINDINGS: Between Dec 9, 2013, and Jan 14, 2016, 438 patients were enrolled and randomly assigned to the two treatment groups (219 patients to each group), of whom 418 were evaluable for the primary endpoint (212 in the anthracycline group and 206 in the non-anthracycline group). The median follow-up for all patients was 19 months (IQR 16-23 months). A pathological complete response was recorded in 141 (67%, 95% CI 60-73) of 212 patients in the anthracycline group and in 140 (68%, 61-74) of 206 in the non-anthracycline group (p=0·95). One patient randomly allocated to the non-anthracycline group did receive anthracyclines and was thus included in the anthracycline group for safety analyses; therefore, for the safety analyses there were 220 patients in the anthracycline group and 218 in the non-anthracycline group. Serious adverse events were reported in 61 (28%) of 220 patients in the anthracycline group and in 49 (22%) of 218 in the non-anthracycline group. The most common adverse events of any cause were grade 3 or worse neutropenia (in 131 [60%] of 220 patients in the anthracycline group vs 118 [54%] of 218 in the non-anthracycline group), grade 3 or worse diarrhoea (26 [12%] vs 37 [18%]), and grade 2 or worse peripheral neuropathy (66 [30%] vs 68 [31%]), with no substantial differences between the groups. Grade 3 or worse febrile neutropenia was more common in the anthracycline group than in the non-anthracycline group (23 [10%] vs three [1%], p<0·0001). Symptomatic left ventricular systolic dysfunction was rare in both groups (two [1%] of 220 vs 0 of 218). One patient in the anthracycline group died because of a pulmonary embolism, which was possibly treatment related. INTERPRETATION: In view of the high proportion of pathological complete responses recorded in both groups and the fact that febrile neutropenia was more frequent in the anthracycline group, omitting anthracyclines from neoadjuvant treatment regimens might be a preferred approach in the presence of dual HER2 blockade in patients with early HER2-positive breast cancer. Long-term follow-up is required to confirm these results. FUNDING: Roche Netherlands.

Efficacy of neoadjuvant treatment with or without pertuzumab in patients with stage II and III HER2-positive breast cancer: a nationwide cohort analysis of pathologic response and 5-year survival.

BACKGROUND: Pathologic complete response (pCR) rates in early stage HER2-positive breast cancer improved after pertuzumab was added to neoadjuvant treatment. However, survival benefit is less-well established and seems mostly limited to node-positive patients. We used national cancer registry data to compare outcomes of patients treated with and without pertuzumab. METHODS: We identified stage II-III HER2-positive breast cancer patients treated with neoadjuvant trastuzumab-based chemotherapy between November 2013 until January 2016 from the Netherlands Cancer Registry. During that period pertuzumab was only available in the 37 hospitals that participated in the TRAIN-2 study. Missing grade and pCR-status were obtained from the Dutch Pathology Registry (PALGA) and cause of death from Statistics Netherlands. We used multiple imputation to impute missing data, multivariable logistic regression to evaluate the association between pertuzumab and pCR (ypT0/is, ypN0) and multivariable Cox regression models for overall survival and breast cancer specific survival (BCSS). RESULTS: We identified 1124 patients of whom 453 received pertuzumab. Baseline characteristics were comparable, although tumor grade was missing more often in patients treated without pertuzumab (12% vs. 2%). Pertuzumab improved pCR rates (41% vs 65%, adjusted odds ratio [aOR] 2.91; 95% CI:2.20-3.94). After a median follow-up of 6.0 years, 5-year BCSS rates were 95% and 98% respectively (adjusted hazard ratio [aHR]: 0.58; 95% CI:0.36-0.95). Younger patients derived more benefit from pertuzumab, but no other significant interactions were found. CONCLUSION: These results support earlier data of a small survival benefit with the addition of pertuzumab to trastuzumab-based neoadjuvant chemotherapy which is most meaningful in younger patients.

Three-Year Follow-up of Neoadjuvant Chemotherapy With or Without Anthracyclines in the Presence of Dual ERBB2 Blockade in Patients With ERBB2-Positive Breast Cancer: A Secondary Analysis of the TRAIN-2 Randomized, Phase 3 Trial.

IMPORTANCE: Primary analysis of the TRAIN-2 study showed high pathologic complete response rates after neoadjuvant chemotherapy with or without anthracyclines plus dual ERBB2 (formerly HER2) blockade. OBJECTIVE: To evaluate 3-year event-free survival (EFS) and overall survival (OS) of an anthracycline-free and anthracycline-containing regimen with dual ERBB2 blockade in patients with stage II and III ERBB2-positive breast cancer. DESIGN, SETTING, AND PARTICIPANTS: A total of 438 patients with stage II and III ERBB2-positive breast cancer were enrolled in this randomized, clinical, open-label phase 3 trial across 37 hospitals in the Netherlands from December 9, 2013, until January 14, 2016. Follow-up analyses were performed after a median follow-up of 48.8 months (interquartile range, 44.1-55.2 months). Analysis was performed on an intention-to-treat basis. INTERVENTIONS: Participants were randomly assigned on a 1:1 basis, stratified by age, tumor stage, nodal stage, and estrogen receptor status, to receive 3 cycles of fluorouracil (500 mg/m2), epirubicin (90 mg/m2), and cyclophosphamide (500 mg/m2), followed by 6 cycles of paclitaxel and carboplatin or 9 cycles of paclitaxel (80 mg/m2 days 1 and 8) and carboplatin (area under the concentration-time curve, 6 mg/mL/min). Both groups received trastuzumab (6 mg/kg; loading dose 8 mg/kg) and pertuzumab (420 mg intravenously; loading dose 840 mg) every 3 weeks. MAIN OUTCOMES AND MEASURES: Three-year EFS, OS, and safety. RESULTS: A total of 438 women were randomized, with 219 per group (anthracycline group, median age, 49 years [interquartile range, 43-55 years]; and nonanthracycline group, median age, 48 years [interquartile range, 43-56 years]). A total of 23 EFS events (10.5%) occurred in the anthracycline group and 21 EFS events (9.6%) occurred in the nonanthracycline group (hazard ratio, 0.90; 95% CI, 0.50-1.63; favoring nonanthracyclines). Three-year EFS estimates were 92.7% (95% CI, 89.3%-96.2%) in the anthracycline group and 93.6% (95% CI, 90.4%-96.9%) in the nonanthracycline group and 3-year OS estimates were 97.7% (95% CI, 95.7%-99.7%) in the anthracycline group and 98.2% (95% CI, 96.4%-100%) in the nonanthracycline group. The results were irrespective of hormone receptor and nodal status. A decline in left ventricular ejection fraction of 10% or more from baseline to less than 50% was more common in patients who received anthracyclines than those who did not (17 of 220 [7.7%] vs 7 of 218 [3.2%]; P = .04). Two patients treated with anthracyclines developed acute leukemia. CONCLUSIONS AND RELEVANCE: This follow-up analysis of the TRAIN-2 study shows similar 3-year EFS and OS estimates with or without anthracyclines in patients with stage II and III ERBB2-positive breast cancer. Anthracycline use is associated with increased risk of febrile neutropenia, cardiotoxic effects, and secondary malignant neoplasms. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT01996267.

Possible missed opportunities for diagnosing colorectal cancer in Dutch primary care: a multimethods approach.

BACKGROUND: Early detection of colorectal cancer (CRC) is important to achieve better survival. Discriminating symptoms suggestive of CRC from benign conditions is a challenge for GPs because most known 'alarm symptoms' have low predictive values. AIM: To further understand the diagnostic process of CRC in general practice in terms of healthcare use and by analysing factors related to diagnostic intervals. DESIGN AND SETTING: A multimethod approach comprising a historical, prospective registry study and qualitative content analysis. METHOD: Healthcare use in the year before referral for colonoscopy was compared between patients diagnosed with CRC and an age-, sex,- and GP-matched control population. Qualitative content analysis was performed on free texts in electronic patient records from a purposive sample of patients with CRC. RESULTS: Patients with CRC (n = 287) had 41% (25-59%) more face-to-face contacts and 21% (7-37%) more medication prescriptions than controls (n = 828). Forty-six per cent of patients with CRC had two or more contacts for digestive reasons, compared with 12.2% of controls, more often for symptoms than diagnoses. From qualitative analysis two themes emerged: 'possible missed diagnostic opportunities' and 'improvements in diagnostic process unlikely'. Possible missed diagnostic opportunities were related to patients waiting before presenting symptoms, doctors attributing symptoms to comorbid conditions or medication use, or doctors sticking to an initial diagnosis. CONCLUSION: Fewer missed diagnostic opportunities might occur if GPs are aware of pitfalls in diagnosing CRC: the assumption that symptoms are caused by comorbid conditions or medication, or relating complaints to pre-existing medical conditions. GPs also need to be aware that repeated digestive complaints warrant rethinking an earlier diagnosis.