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Homologous DNA recombination is a fundamental, regenerative process within living organisms. However, in most organisms, homologous recombination is a rare event, requiring a complex set of reactions and extensive homology. We demonstrate in this paper that Beta protein of phage λ generates recombinants in chromosomal DNA by using synthetic single-stranded DNAs (ssDNA) as short as 30 bases long. This ssDNA recombination can be used to mutagenize or repair the chromosome with efficiencies that generate up to 6% recombinants among treated cells. Mechanistically, it appears that Beta protein, a Rad52-like protein, binds and anneals the ssDNA donor to a complementary single-strand near the DNA replication fork to generate the recombinant. This type of homologous recombination with ssDNA provides new avenues for studying and modifying genomes ranging from bacterial pathogens to eukaryotes. Beta protein and ssDNA may prove generally applicable for repairing DNA in many organisms.

Moderate-to-severe ulcerative colitis (UC) and Crohn's disease (CD) are associated with substantial quality of life and economic burdens (Kawalec, 2016). The Communicating Needs and Features of IBD Experiences (CONFIDE) study aims to further the understanding of the experience and impact of symptoms on patients' lives and elucidate any gaps in communication between healthcare professionals (HCPs) and patients with moderate-to-severe UC and CD in the United States (US), Europe, and Japan. These data focus on US patients with UC and US HCPs. An online, quantitative, cross-sectional survey was conducted with HCPs (n=200) and patients with moderate-to-severe UC (n=200) in the US between May (HCPs) and July (patients) 2021. Moderate-to-severe UC was defined using criteria based on previous treatment experience, steroid use and/or hospitalization. The HCP survey included physicians (89%) and non-physician HCPs (11%) who are responsible for making prescribing decisions. Data collected included perspectives on the experience and impact of symptoms in patients with moderate-to-severe UC. The top three symptoms currently (past month) and ever suffered by patients (mean age: 40.4, 61.5% male) were diarrhea (62.5% and 74.0%, respectively), bowel urgency (47.0% and 61.5%) and increased stool frequency (38.5% and 57.5%). Blood in stool was reported by 27.0% of patients as currently suffering, and 51.0% ever. According to HCPs (78.0% male), the top three symptoms reported by patients were diarrhea (73.5% ranked in top 3), blood in stool (69.0%), and increased stool frequency (37.5%). Bowel urgency was recorded in the top 3 patient-reported symptoms by 24.0% of HCPs. Patients self-rated their disease-severity as 10.5% (n = 21) mild UC, 71.0% (n = 142) moderate UC, 17.5% (n = 35) severe UC, and 1.0% (n = 2) patients did not know. Bowel urgency was more frequently reported in patients with severe disease (62.9%, n = 22) when compared with those with mild-to-moderate disease (42.9%, n = 70). Among the overall patient population, 76.5% (n = 153) were receiving advanced therapies (biologic or novel oral therapy). Bowel urgency was currently experienced by 46.4% of these patients. Only 38.2% of patients felt completely comfortable reporting bowel urgency to their HCP. Of patients not comfortable reporting bowel urgency, 62.2% (n = 23) reported they felt embarrassed talking about it. Among HCPs, 75.5% (n = 151) reported they proactively discussed bowel urgency at routine appointments. Those HCPs who reported that they do not proactively discuss bowel urgency (24.5%, n = 49) cited the main reason as they expect the patient to bring it up (46.9%, n = 23). Bowel urgency is the second-most commonly reported symptom by patients with moderate-to-severe UC but is not among the HCP-perceived top three most reported symptoms. A substantial proportion of patients with moderate-to-severe UC receiving advanced therapies continue to report bowel urgency. A communication gap between patients and HCPs was identified and highlights the under appreciation of bowel urgency as an important symptom impacting patients' daily life.

A recombination system has been developed for efficient chromosome engineering in Escherichia coli by using electroporated linear DNA. A defective λ prophage supplies functions that protect and recombine an electroporated linear DNA substrate in the bacterial cell. The use of recombination eliminates the requirement for standard cloning as all novel joints are engineered by chemical synthesis in vitro and the linear DNA is efficiently recombined into place in vivo. The technology and manipulations required are simple and straightforward. A temperature-dependent repressor tightly controls prophage expression, and, thus, recombination functions can be transiently supplied by shifting cultures to 42 degrees C for 15 min. The efficient prophage recombination system does not require host RecA function and depends primarily on Exo, Beta, and Gam functions expressed from the defective λ prophage. The defective prophage can be moved to other strains and can be easily removed from any strain. Gene disruptions and modifications of both the bacterial chromosome and bacterial plasmids are possible. This system will be especially useful for the engineering of large bacterial plasmids such as those from bacterial artificial chromosome libraries.

A phage λ-based recombination system, Red, can be used for high-efficiency mutagenesis, repair, and engineering of chromosomal or episomal DNA in vivo in Escherichia coli. When long linear double-stranded DNA with short flanking homologies to their targets are used for the recombination, the X Exo, Beta, and Gam proteins are required. The current model is: (i) Gam inhibits the host RecBCD activity, thereby protecting the DNA substrate for recombination; (ii) Exo degrades from each DNA end in a 5′ → 3′ direction, creating double-stranded DNA with 3′ single-stranded DNA tails; and (iii) Beta binds these 3′ overhangs to protect and anneal them to complementary sequences. We have tested this model for Red recombination by using electroporation to introduce overlapping, complementary oligonucleotides that when annealed in vivo approximate the recombination intermediate that Exo should create. Using this technique we found Exo-independent recombination. Surprisingly, a similarly constructed substrate with 5′ overhangs recombined more efficiently. This 5′ overhang recombination required both Exo and Beta for high levels of recombination and the two oligonucleotides need to overlap by only 6 bp on their 3′ ends. Results indicate that Exo may load Beta onto the 3′ overhang it produces. In addition, multiple overlapping oligonucleotides were successfully used to generate recombinants in vivo, a technique that could prove useful for many genetic engineering procedures.

Introduction/BackgroundPatient preference on treatment options following frontline platinum-based chemotherapy for epithelial ovarian cancer (EOC) remains unstudied. Multiple treatment options are available, including PARP inhibitors, so understanding patient preference is critical.MethodologyA cross-sectional survey was completed by US patients with newly-diagnosed EOC eligible for frontline maintenance therapy. Maintenance preference was assessed via time trade-off simulation. Patients selected their preferred post-chemotherapy treatment approach: surveillance, oral daily (QD), oral twice daily (BID), intravenous every 3 weeks (IV-Q3W), or combination IV-Q3W/BID, assuming equivalent efficacy (for all scenarios) and safety (medication scenarios only). Patients were asked to select between a series of maintenance scenarios comparing decreased time to progression (TTP) on their preferred option with constant TTP with alternative options. Relative disutility of each scenario was calculated.Abstract 2022-RA-949-ESGO Table 1Patient (N=153) preferences for formulation and dosing frequency of frontline maintenance for EOCResults153 patients completed the survey, median age was 52.3 years; 30% were non-White, and 83% had health insurance covering full EOC treatment. Of all medication strategies, QD treatment was preferred (38%, table 1); patients were willing to trade the least amount of time (2.3 months) without progression on this scenario versus other choices. For patients who preferred to take a medication even when surveillance offered the same amount of time without progression (n=86), the most common reason was a feeling of taking an active approach to treatment (66%), having a reason to regularly visit a doctor/hospital (30%), being cared for/monitored more regularly and carefully (28%), and because taking medication is reassuring (24%).ConclusionPatients preferred QD treatment more than other medication strategies for EOC maintenance following frontline platinum-based chemotherapy; patients who preferred medication felt they were taking an active approach to treatment. Patient preferences should be considered in treatment decisions and further studied.Previously submitted to the IGCS Global Meeting (29 Sept–1 Oct 2022; New York City, USA)Funding statementGSK (214511/NCT02655016).

BACKGROUND:Moderate-to-severe ulcerative colitis (UC) and Crohn's disease (CD) are associated with substantial quality of life and economic burdens (Kawalec, 2016). The Communicating Needs and Features of IBD Experiences (CONFIDE) study aims to further the understanding of the experience and impact of symptoms on patients' lives and elucidate any gaps in communication between healthcare professionals (HCPs) and patients with moderate-to-severe UC and CD in the United States (US), Europe, and Japan. These data focus on US patients with UC and US HCPs.METHODS:An online, quantitative, cross-sectional survey was conducted with HCPs (n=200) and patients with moderate-to-severe UC (n=200) in the US between May (HCPs) and July (patients) 2021. Moderate-to-severe UC was defined using criteria based on previous treatment experience, steroid use and/or hospitalization. The HCP survey included physicians (89%) and non-physician HCPs (11%) who are responsible for making prescribing decisions. Data collected included perspectives on the experience and impact of symptoms in patients with moderate-to-severe UC.RESULTS:The top three symptoms currently (past month) and ever suffered by patients (mean age: 40.4, 61.5% male) were diarrhea (62.5% and 74.0%, respectively), bowel urgency (47.0% and 61.5%) and increased stool frequency (38.5% and 57.5%). Blood in stool was reported by 27.0% of patients as currently suffering, and 51.0% ever. According to HCPs (78.0% male), the top three symptoms reported by patients were diarrhea (73.5% ranked in top 3), blood in stool (69.0%), and increased stool frequency (37.5%). Bowel urgency was recorded in the top 3 patient-reported symptoms by 24.0% of HCPs. Patients self-rated their disease-severity as 10.5% (n = 21) mild UC, 71.0% (n = 142) moderate UC, 17.5% (n = 35) severe UC, and 1.0% (n = 2) patients did not know. Bowel urgency was more frequently reported in patients with severe disease (62.9%, n = 22) when compared with those with mild-to-moderate disease (42.9%, n = 70). Among the overall patient population, 76.5% (n = 153) were receiving advanced therapies (biologic or novel oral therapy). Bowel urgency was currently experienced by 46.4% of these patients. Only 38.2% of patients felt completely comfortable reporting bowel urgency to their HCP. Of patients not comfortable reporting bowel urgency, 62.2% (n = 23) reported they felt embarrassed talking about it. Among HCPs, 75.5% (n = 151) reported they proactively discussed bowel urgency at routine appointments. Those HCPs who reported that they do not proactively discuss bowel urgency (24.5%, n = 49) cited the main reason as they expect the patient to bring it up (46.9%, n = 23).CONCLUSION:Bowel urgency is the second-most commonly reported symptom by patients with moderate-to-severe UC but is not among the HCP-perceived top three most reported symptoms. A substantial proportion of patients with moderate-to-severe UC receiving advanced therapies continue to report bowel urgency. A communication gap between patients and HCPs was identified and highlights the under appreciation of bowel urgency as an important symptom impacting patients' daily life.

Abstract BACKGROUND Moderate-to-severe ulcerative colitis (UC) is associated with burden on quality of life (Kawalec, 2016). The Communicating Needs and Features of IBD Experiences (CONFIDE) study aims to increase understanding of patients’ experiences and the impact of IBD on their lives in the United States (US), Europe, and Japan. These data focus on US patients with moderate-to-severe UC. METHODS An online, quantitative, cross-sectional survey was conducted (recruitment via panel) with patients with moderate-to-severe UC in the US in July 2021. Data collected included patient perspectives on their experiences with UC. Moderate-to-severe UC was defined using criteria based on previous treatment, steroid use, and/or hospitalization. Patients who reported ever experiencing symptoms were asked to report the frequency of those symptoms over the past three months. RESULTS - 200 (of 756 total contacted) patients (62% male, mean age 40 years) completed the survey. - Most patients who have ever experienced bowel urgency and/or urge incontinence (bowel urgency-related accidents) report doing so at least once-weekly (79%, n=97; 58%, n=52, respectively) over the past three months (Table 1). - Wearing a diaper/pad/protection at least once in the past three months due to fear of urge incontinence was reported by 76% (n=152) of patients with UC (Table 2). Of those patients, 47% (n=72) reported that they currently (past month) suffer from bowel urgency. - Almost half of all patients (45%, n=90) report wearing a diaper/pad/protection due to fear of urge incontinence at least once a week (Table 2). - A total of 77% (n=153) of patients were receiving advanced therapies (biologic or novel oral therapy). Many of these patients reported ever suffering bowel urgency, n=91, 59%, and/or urge incontinence, n=65, 42% (Table 1). Wearing a diaper/pad/protection at least once in the past three months due to fear of urge incontinence was reported by 80% (n=122) of these patients (Table 2). CONCLUSIONS - Bowel urgency markedly impacts the majority of patients with moderate-to-severe UC. - Despite the young patient population, and regardless of whether they were receiving advanced therapies or not, over three quarters of the surveyed patients reported wearing diapers/pads/protection in the past three months due to fear of urge incontinence. - Many patients who did not report bowel urgency as a current symptom still used diapers/pads/protection out of fear they will experience urge incontinence. - Over half of patients receiving advanced therapies who have ever experienced bowel urgency and/or urge incontinence reported having these symptoms at least once a week in the last three months. - Even if patients are not currently experiencing bowel urgency, the persistent impact of moderate-to-severe UC is demonstrated by the use of diaper/pads/other protection due to fear of urge incontinence.

IntroductionCommunicating Needs and Features of IBD Experiences (CONFIDE) aims to further understand the experience and impact of symptoms on patients’ (pts’) lives and elucidate any gaps in communication between healthcare professionals (HCPs) and pts with moderate-severe ulcerative colitis (UC) and Crohn’s disease (CD) in the US, Europe, and Japan. These data focus on US UC pts and HCPs.MethodsAn online, quantitative, cross-sectional survey was conducted on HCPs (n=200) and pts with moderate-severe UC (using criteria from previous treatment experience, steroid use and/or hospitalization) (n=200) in the US between May (HCPs) and July (pts) 2021. The HCP survey included physicians (89%) and non-physician HCPs (11%) responsible for making prescribing decisions.ResultsThe top 3 symptoms currently and ever suffered by pts (mean age: 40.4, 61.5% male) were diarrhea (62.5% and 74.0%), bowel urgency (47.0% and 61.5%), and increased stool frequency (38.5% and 57.5%). Blood in stool was reported by 27.0% pts as currently suffering, and 51.0% ever. Per HCPs (78.0% male), the top 3 pt-reported symptoms were diarrhea (73.5% ranked in top 3), blood in stool (69.0%), and increased stool frequency (37.5%). 24.0% HCPs reported bowel urgency in the top 3 pt-reported symptoms. Pts self-rated their disease severity as 10.5% (n=21) mild UC, 71.0% (n=142) moderate UC, 17.5% (n=35) severe UC, and not known 1.0% (n=2). Bowel urgency was more frequent with severe UC (62.9%, n=22) than mild-moderate UC (42.9%, n=70). 76.5% (n=153) pts were receiving advanced therapies; 46.4% currently experiencing bowel urgency. Only 38.2% pts felt comfortable reporting bowel urgency to their HCP, while 62.2% (n=23) reported feeling embarrassed talking about it. 75.5% (n=151) HCPs reported they proactively discussed bowel urgency at appointments. HCPs not proactively discussing bowel urgency (24.5%, n=49) expect the pt to bring it up (46.9%, n=23).ConclusionsBowel urgency is the second-most commonly reported symptom by moderate-severe UC pts, but is not among the HCP-perceived top 3. A substantial proportion of moderate-severe UC pts receiving advanced therapies continue to report bowel urgency. Communication gap between pts and HCPs was identified and highlights the under-appreciation of bowel urgency as an important symptom impacting pts’ daily life.