HPV integration hijacks and multimerizes a cellular enhancer to generate a viral-cellular super-enhancer that drives high viral oncogene expression.

Integration of human papillomavirus (HPV) genomes into cellular chromatin is common in HPV-associated cancers. Integration is random, and each site is unique depending on how and where the virus integrates. We recently showed that tandemly integrated HPV16 could result in the formation of a super-enhancer-like element that drives transcription of the viral oncogenes. Here, we characterize the chromatin landscape and genomic architecture of this integration locus to elucidate the mechanisms that promoted de novo super-enhancer formation. Using next-generation sequencing and molecular combing/fiber-FISH, we show that ~26 copies of HPV16 are integrated into an intergenic region of chromosome 2p23.2, interspersed with 25 kb of amplified, flanking cellular DNA. This interspersed, co-amplified viral-host pattern is frequent in HPV-associated cancers and here we designate it as Type III integration. An abundant viral-cellular fusion transcript encoding the viral E6/E7 oncogenes is expressed from the integration locus and the chromatin encompassing both the viral enhancer and a region in the adjacent amplified cellular sequences is strongly enriched in the super-enhancer markers H3K27ac and Brd4. Notably, the peak in the amplified cellular sequence corresponds to an epithelial-cell-type specific enhancer. Thus, HPV16 integration generated a super-enhancer-like element composed of tandem interspersed copies of the viral upstream regulatory region and a cellular enhancer, to drive high levels of oncogene expression.

PRESTO: A Phase III, Open-Label Study of Intensification of Androgen Blockade in Patients With High-Risk Biochemically Relapsed Castration-Sensitive Prostate Cancer (AFT-19).

PURPOSE: Patients with biochemically recurrent prostate cancer (BRPC) after radical prostatectomy and a short PSA doubling time are at risk for distant metastases. Apalutamide, an androgen receptor antagonist, and abiraterone acetate plus prednisone (AAP) prolong survival in the metastatic setting. We evaluated whether intensification of androgen-deprivation therapy (ADT) improves outcomes in BRPC. PATIENTS AND METHODS: PRESTO is a randomized phase III, open-label trial in patients with BRPC and PSA doubling time ≤9 months (ClinicalTrials.gov identifier: NCT03009981). Patients were randomly assigned 1:1:1 to receive a finite 52-week treatment course with ADT control, ADT + apalutamide, or ADT + apalutamide + AAP. The primary end point was PSA progression-free survival (PSA-PFS), defined as serum PSA >0.2 ng/mL after treatment completion. RESULTS: Five hundred three patients were enrolled. The median PSA was 1.8 ng/mL (IQR, 1.0-3.6). At the first planned interim analysis, both experimental arms significantly prolonged PSA-PFS compared with the control arm (median, 24.9 months for ADT + apalutamide v 20.3 months for ADT; hazard ratio [HR], 0.52 [95% CI, 0.35 to 0.77]; P = .00047; median, 26.0 months for ADT + apalutamide + AAP v 20.0 months for ADT; HR, 0.48 [95% CI, 0.32 to 0.71]; P = .00008). Median time to testosterone recovery did not differ across treatment arms. The most common grade ≥3 adverse event was hypertension (7.5%, 7.4%, and 18% in ADT, ADT + apalutamide, and ADT + apalutamide + AAP arms, respectively). CONCLUSION: Intensified AR blockade for a finite duration prolongs PSA-PFS with a manageable safety profile, without adversely affecting time to testosterone recovery. The addition of apalutamide to ADT should be considered in patients with high-risk BRPC.

Graded Organ Response and Progression Criteria for Kidney Immunoglobulin Light Chain Amyloidosis.

Importance: Kidney light chain (AL) amyloidosis is associated with a risk of progression to kidney replacement therapy (KRT) and death. Several studies have shown that a greater reduction in proteinuria following successful anticlonal therapy is associated with improved outcomes. Objective: To validate graded kidney response criteria and their association with kidney and overall survival (OS). Design, Setting, and Participants: This retrospective, multicenter cohort was conducted at 10 referral centers for amyloidosis from 2010 to 2015 and included patients with kidney AL amyloidosis that was evaluable for kidney response and who achieved at least hematologic partial response within 12 months of diagnosis. The median follow-up was 69 (54-88) months. Data analysis was conducted in 2023. Exposure: Four kidney response categories based on the reduction in pretreatment 24-hour urine protein (24-hour UP) levels: complete response (kidCR, 24-hour UP ≤200 mg), very good partial response (kidVGPR, >60% reduction in 24-hour UP), partial response (kidPR, 31%-60% reduction), and no response (kidNR, ≤30% reduction). Kidney response was assessed at landmark points (6, 12, and 24 months) and best kidney response. Main Outcomes and Measures: Cumulative incidence of progression to KRT and OS. Results: Seven-hundred and thirty-two patients (335 women [45.8%]) were included, with a median (IQR) age of 63 (55-69) years. The median (IQR) baseline 24-hour proteinuria and estimated glomerular filtration rate was 5.3 (2.8-8.5) g per 24 hours and 72 (48-92) mL/min/1.73m2, respectively. In a competing-risk analysis, the 5-year cumulative incidence rates of progression to KRT decreased with deeper kidney responses as early as 6 months from therapy initiation (11%, 12%, 2.1%, and 0% for kidNR, kidPR, kidVGPR, and kidCR, respectively; P = .002) and were maintained at 12 months and 24 months and best kidney response. Patients able to achieve kidCR/kidVGPR by 24 months and at best response had significantly better OS compared with kidPR/kidNR. Kidney progression, defined as a 25% or greater decrease in estimated glomerular filtration rate, was associated with cumulative incidence of progression to KRT and OS. Conclusions and Relevance: The results of this cohort study suggest that graded kidney response criteria offers clinically and prognostically meaningful information for treating patients with kidney AL amyloidosis. The response criteria potentially inform kidney survival based on the depth of reduction in 24-hour proteinuria levels and demonstrate an OS advantage for those able to achieve kidCR/kidVGPR compared with kidPR/kidNR. Taken together, achievement of at least kidVGPR by 12 months is needed to ultimately improve kidney and patient survival.

Carcinoembryonic antigen-expressing oncolytic measles virus derivative in recurrent glioblastoma: a phase 1 trial.

Measles virus (MV) vaccine strains have shown significant preclinical antitumor activity against glioblastoma (GBM), the most lethal glioma histology. In this first in human trial (NCT00390299), a carcinoembryonic antigen-expressing oncolytic measles virus derivative (MV-CEA), was administered in recurrent GBM patients either at the resection cavity (Group A), or, intratumorally on day 1, followed by a second dose administered in the resection cavity after tumor resection on day 5 (Group B). A total of 22 patients received study treatment, 9 in Group A and 13 in Group B. Primary endpoint was safety and toxicity: treatment was well tolerated with no dose-limiting toxicity being observed up to the maximum feasible dose (2×107 TCID50). Median OS, a secondary endpoint, was 11.6 mo and one year survival was 45.5% comparing favorably with contemporary controls. Other secondary endpoints included assessment of viremia, MV replication and shedding, humoral and cellular immune response to the injected virus. A 22 interferon stimulated gene (ISG) diagonal linear discriminate analysis (DLDA) classification algorithm in a post-hoc analysis was found to be inversely (R = -0.6, p = 0.04) correlated with viral replication and tumor microenvironment remodeling including proinflammatory changes and CD8 + T cell infiltration in post treatment samples. This data supports that oncolytic MV derivatives warrant further clinical investigation and that an ISG-based DLDA algorithm can provide the basis for treatment personalization.

Association of circulating markers with cognitive decline after radiation therapy for brain metastasis.

BACKGROUND: A recent phase III trial (NCT01372774) comparing use of stereotactic radiosurgery [SRS] versus whole-brain radiation therapy [WBRT] after surgical resection of a single brain metastasis revealed that declines in cognitive function were more common with WBRT than with SRS. A secondary endpoint in that trial, and the primary objective in this secondary analysis, was to identify baseline biomarkers associated with cognitive impairment after either form of radiotherapy for brain metastasis. Here we report our findings on APOE genotype and serum levels of associated proteins and their association with radiation-induced neurocognitive decline. METHODS: In this retrospective analysis of prospectively collected samples from a completed randomized clinical trial, patients provided blood samples every 3 months that were tested by genotyping and enzyme-linked immunosorbent assay, and results were analyzed in association with cognitive impairment. RESULTS: The APOE genotype was not associated with neurocognitive impairment at 3 months. However, low serum levels of ApoJ, ApoE, or ApoA protein (all P < .01) and higher amyloid beta (Aß 1-42) levels (P = .048) at baseline indicated a greater likelihood of neurocognitive decline at 3 months after SRS, whereas lower ApoJ levels were associated with decline after WBRT (P = .014). CONCLUSIONS: Patients with these pretreatment serum markers should be counseled about radiation-related neurocognitive decline.

Biomedical contraceptives and their association with condom use consistency among female sex workers in China: Results from a national cross-sectional study.

PURPOSE: To characterize contraceptive needs and patterns of use among Chinese FSW and investigate the effect of contraceptive choice on condom use practices with clients. METHODS: Data come from a cross-sectional study conducted in eight cities (Beijing, Tianjin, Shenzhen, Kunming, Jiaozhou, Yunfu, Xiangyang and Longnan) throughout seven provinces in China between August 17 and October 17, 2019. We used logistic regression to compare odds of consistent condom use with clients between FSW who do and do not use biomedical contraceptives (defined as oral pills, injectables/implants/patches/rings, intrauterine devices, and sterilization) and assessed subgroup effects of interest. RESULTS: Of the 1229 participants, 62.4% reported ever having an unintended pregnancy. Biomedical contraceptive use was non-significantly associated with consistent condom use with clients generally (aOR=0.95, 95% CI:0.75-1.21). This association between biomedical contraceptive use and consistent condom use was lower among FSW with at least a high school education (aOR=0.33, 95% CI: 0.18-0.62) and those who had never married (aOR=0.60, 95% CI: 0.38-0.93); it was higher among FSW employed at middle tier commercial sex venues (aOR=1.67, 95% CI: 1.13-2.45). CONCLUSIONS: Taken together these findings highlight the pressing unmet need for contraceptives in Chinese FSW and the complex interplay between reproductive and sexual health needs in this key population.

Tandemly Integrated HPV16 Can Form a Brd4-Dependent Super-Enhancer-Like Element That Drives Transcription of Viral Oncogenes.

UNLABELLED: In cancer cells associated with human papillomavirus (HPV) infections, the viral genome is very often found integrated into the cellular genome. The viral oncogenes E6 and E7 are transcribed from the viral promoter, and integration events that alter transcriptional regulation of this promoter contribute to carcinogenic progression. In this study, we detected highly enriched binding of the super-enhancer markers Brd4, MED1, and H3K27ac, visible as a prominent nuclear focus by immunofluorescence, at the tandemly integrated copies of HPV16 in cells of the cervical neoplasia cell line W12 subclone 20861. Tumor cells are often addicted to super-enhancer-driven oncogenes and are particularly sensitive to disruption of transcription factor binding to the enhancers. Treatment of 20861 cells with bromodomain inhibitors displaced Brd4 from the HPV integration site, greatly decreased E6/E7 transcription, and inhibited cellular proliferation. Thus, Brd4 activates viral transcription at this integration site, and strong selection for E6/E7 expression can drive the formation of a super-enhancer-like element to promote oncogenesis. IMPORTANCE: Oncogenic human papillomaviruses play an essential role in the development of cervical cancer, and growth of these cancer cells requires continued expression of the viral E6 and E7 oncogenes. Integration of the virus into the host genome often results in deregulation of E6 and E7 expression, which provides a selective growth advantage and increases genetic instability of infected cells. We show here that tandemly integrated copies of the viral genome can form a super-enhancer-like element that drives E6/E7 transcription. Targeted disruption of factors binding to this element decreases viral transcription and causes cell death. Thus, cancer cells that harbor integrated HPV could be targeted by therapeutics that disrupt super-enhancer function.