Uffe S. Løve

BACKGROUND: Increasingly, circulating tumor DNA (ctDNA) is proposed as a tool for minimal residual disease (MRD) assessment. Digital PCR (dPCR) offers low analysis costs and turnaround times of less than a day, making it ripe for clinical implementation. Here, we used tumor-informed dPCR for ctDNA detection in a large colorectal cancer (CRC) cohort to evaluate the potential for post-operative risk assessment and serial monitoring, and how the metastatic site may impact ctDNA detection. Additionally, we assessed how altering the ctDNA-calling algorithm could customize performance for different clinical settings. PATIENTS AND METHODS: Stage II-III CRC patients (N = 851) treated with a curative intent were recruited. Based on whole-exome sequencing on matched tumor and germline DNA, a mutational target was selected for dPCR analysis. Plasma samples (8 ml) were collected within 60 days after operation and-for a patient subset (n = 246)-every 3-4 months for up to 36 months. Single-target dPCR was used for ctDNA detection. RESULTS: Both post-operative and serial ctDNA detection were prognostic of recurrence [hazard ratio (HR) = 11.3, 95% confidence interval (CI) 7.8-16.4, P < 0.001; HR = 30.7, 95% CI 20.2-46.7, P < 0.001], with a cumulative ctDNA detection rate of 87% at the end of sample collection in recurrence patients. The ctDNA growth rate was prognostic of survival (HR = 2.6, 95% CI 1.5-4.4, P = 0.001). In recurrence patients, post-operative ctDNA detection was challenging for lung metastases (4/21 detected) and peritoneal metastases (2/10 detected). By modifying the cut-off for calling a sample ctDNA positive, we were able to adjust the sensitivity and specificity of our test for different clinical contexts. CONCLUSIONS: The presented results from 851 stage II-III CRC patients demonstrate that our personalized dPCR approach effectively detects MRD after operation and shows promise for serial ctDNA detection for recurrence surveillance. The ability to adjust sensitivity and specificity shows exciting potential to customize the ctDNA caller for specific clinical settings.

11 Background: Timely detection of recurrence, as well as identification of patients at high risk of recurrence after surgery and after completion of adjuvant therapy, are major challenges in the treatment of colorectal cancer (CRC). Postsurgical circulating tumor DNA (ctDNA) analysis is a promising tool for the identification of patients with minimal residual disease (MRD) and a high risk of recurrence. The objective of this prospective, multicenter study was to determine whether serial postsurgical ctDNA analysis could identify the patients at high risk of recurrence, provide an assessment of adjuvant therapy efficacy and detect relapse earlier than standard-of-care radiological imaging. Methods: The cohort comprises 265 stage I-III CRC patients, the to-date largest cohort assessed for ctDNA. All patients had the tumor resected and a subset of 62.6% (166 /265) was additionally treated with ACT. Plasma samples (n = 1503) were collected at various time points for a median follow-up of 28.4 months (range: 1.2-51.0 months). Individual tumors and matched germline DNA were whole-exome sequenced and somatic single nucleotide variants (SNVs) were identified. Personalized multiplex PCR assays were designed to track tumor-specific SNVs (Signatera, bespoke mPCR NGS assay) in each patient’s plasma sample. Results: Postoperative ctDNA status prior to ACT was assessed in 218 patients, of which 9.17% (20/218) were identified to be MRD-positive and 75% (15/20) eventually relapsed. The remaining 25% (5/20) of MRD-positive patients that did not relapse, received ACT. In contrast, only 13.6% (27/198) of MRD-negative cases relapsed (HR: 11: 95% CI: 5.7-20; p &lt; 0.001). Longitudinal ctDNA-positive status, post-definitive therapy (n = 202) was associated with a HR of 36 (95% CI: 16-81; p &lt; 0.001). For a subset of 155 patients postoperative CEA and ctDNA measurements were compared, wherein, ctDNA-positive status was found to be significantly associated with RFS (HR, 7.1; 95% CI, 3.4-15; P &lt; 0.001) compared to CEA (HR, 1.2; 95% CI, 0.46-3.1; P = 0.73). Serial ctDNA analysis detected MRD up to a median of 8 months (0.56 - 21.6 months) ahead of radiologic relapse. Conclusion: Postoperative ctDNA positive status was associated with markedly reduced RFS compared to CEA. The study also shows that effective therapy can be curative in a portion of MRD-positive patients. In a longitudinal setting, ctDNA analysis predicted the risk of recurrence and is a more reliable biomarker for treatment response monitoring.

BACKGROUND: The use of the vertical rectus abdominis myocutaneous flap in reconstruction after abdominoperineal resection or pelvic exenteration for neoplasia is well documented. However, functional outcomes after vaginal reconstruction, including sexual function, are poorly described. OBJECTIVE: This study aimed to examine sexual function in women following extensive pelvic surgery with colpectomy and vaginal reconstruction with the use of a vertical rectus abdominis myocutaneous flap. DESIGN: This study is a retrospective review of medical records in combination with patient questionnaires. Nonresponders were followed up with a second contact. SETTINGS: This study was performed at a tertiary care university medical center (Colorectal Section, Department of Surgery P, Aarhus University Hospital, Denmark) PATIENTS: All women undergoing pelvic surgery and simultaneous vaginal reconstruction with the use of a vertical rectus abdominis myocutaneous flap between 2004 and 2010 at our department were identified from a patient database. Thirty women who were alive at the time of identification were included in the study. MAIN OUTCOME MEASURES: Sexual function before and after surgery was evaluated by the use of the Sexual function Vaginal changes Questionnaire. The main outcome end point was whether the patient was sexually active after vaginal reconstruction. RESULTS: Twenty-six participants (87%) answered the questionnaire. Fifty percent of patients reported an active sex life before surgery. In general, patients reported an unchanged desire for both physical and sexual contact after surgery. However, only 2 patients (14%) reported being sexually active after surgery. LIMITATIONS: This was a retrospective study with a heterogeneous cohort involving several types of cancers and surgical procedures. Factors other than vertical rectus abdominis myocutaneous flap reconstruction itself may interfere with the sexual function. CONCLUSION: Extensive pelvic surgery with colpectomy leads to sexual dysfunction even when the vagina is reconstructed with a vertical rectus abdominis myocutaneous flap. This knowledge may improve the quality of information given to this group of patients before surgery.

BACKGROUND: Secondary peritonitis is a severe condition with a 20-32% reported mortality. The accepted treatment modalities are vacuum-assisted closure (VAC) or primary closure with relaparotomy on-demand (ROD). However, no randomised controlled trial has been completed to compare the two methods potential benefits and disadvantages. METHODS: This study will be a randomised controlled multicentre trial, including patients aged 18 years or older with purulent or faecal peritonitis confined to at least two of the four abdominal quadrants originating from the small intestine, colon, or rectum. Randomisation will be web-based to either primary closure with ROD or VAC in blocks of 2, 4, and 6. The primary endpoint is peritonitis-related complications within 30 or 90 days and one year after index operation. Secondary outcomes are comprehensive complication index (CCI) and mortality after 30 or 90 days and one year; quality of life assessment by (SF-36) after three and 12 months, the development of incisional hernia after 12 months assessed by clinical examination and CT-scanning and healthcare resource utilisation. With an estimated superiority of 15% in the primary outcome for VAC, 340 patients must be included. Hospitals in Denmark and Europe will be invited to participate. DISCUSSION: There is no robust evidence for choosing either open abdomen with VAC treatment or primary closure with relaparotomy on-demand in patients with secondary peritonitis. The present study has the potential to answer this important clinical question. TRIAL REGISTRATION: The study protocol has been registered at clinicaltrials.gov (NCT03932461). Protocol version 1.0, 9 January 2022.