FLT3L governs the development of partially overlapping hematopoietic lineages in humans and mice.

FMS-related tyrosine kinase 3 ligand (FLT3L), encoded by FLT3LG, is a hematopoietic factor essential for the development of natural killer (NK) cells, B cells, and dendritic cells (DCs) in mice. We describe three humans homozygous for a loss-of-function FLT3LG variant with a history of various recurrent infections, including severe cutaneous warts. The patients' bone marrow (BM) was hypoplastic, with low levels of hematopoietic progenitors, particularly myeloid and B cell precursors. Counts of B cells, monocytes, and DCs were low in the patients' blood, whereas the other blood subsets, including NK cells, were affected only moderately, if at all. The patients had normal counts of Langerhans cells (LCs) and dermal macrophages in the skin but lacked dermal DCs. Thus, FLT3L is required for B cell and DC development in mice and humans. However, unlike its murine counterpart, human FLT3L is required for the development of monocytes but not NK cells.

Urinary cytomegalovirus excretion: The unresolved issues.

Cytomegalovirus (CMV) excretion in urine is frequently observed in clinical practice. However, the specific circumstances and pathophysiological mechanisms underlying this shedding remain largely unknown. Here, we address some of the key questions regarding urinary CMV excretion, focusing on new hypotheses raised by recent advances in the field. Cellular origins of CMV shedding, clinical contexts of occurrence, systemic spread of the virus versus compartmentalization in the urinary tract, and clinical impact are successively discussed.

Human papillomavirus negative high grade cervical lesions and cancers: Suggested guidance for HPV testing quality assurance.

BACKGROUND: Some high-grade cervical lesions and cervical cancers (HSIL+) test negative for human papillomavirus (HPV). The HPV-negative fraction varies between 0.03 % and 15 % between different laboratories. Monitoring and extended re-analysis of HPV-negative HSIL+ could thus be helpful to monitor performance of HPV testing services. We aimed to a) provide a real-life example of a quality assurance (QA) program based on re-analysis of HPV-negative HSIL+ and b) develop international guidance for QA of HPV testing services based on standardized identification of apparently HPV-negative HSIL+ and extended re-analysis, either by the primary laboratory or by a national HPV reference laboratory (NRL). METHODS: There were 116 initially HPV-negative cervical specimens (31 histopathology specimens and 85 liquid-based cytology samples) sent to the Swedish HPV Reference Laboratory for re-testing. Based on the results, an international QA guidance was developed through an iterative consensus process. RESULT: Standard PCR testing detected HPV in 55.2 % (64/116) of initially "HPV-negative" samples. Whole genome sequencing of PCR-negative samples identified HPV in an additional 7 samples (overall 61.2 % HPV positivity). Reasons for failure to detect HPV in an HSIL+ lesion are listed and guidance to identify cases for extended re-testing, including which information should be included when referring samples to an NRL are presented. CONCLUSION: Monitoring the proportion of and reasons for failure to detect HPV in HSIL+ will help support high performance and quality improvement of HPV testing services. We encourage implementation of QA strategies based on re-analysis of "HPV negative" HSIL+ samples.

Prevalence and significance of HPV DNA detection below the clinical threshold of the commercial kit Alinity m HR-HPV assay (Abbott).

The positive clinical threshold of human papillomavirus (HPV) tests validated for primary cervical cancer screening (CCS) is designed to offer an optimal balance between clinical sensitivity and specificity. However, there may be a gap between the analytical sensitivity of the test and the positive clinical threshold, referred to here as the "gray-zone." This study aims to determine the prevalence and significance of HPV results obtained in the gray-zone in routine practice. Cervical samples obtained in our institution for CCS over a 22-month-period were tested with the Alinity m HR-HPV Assay (Abbott). Clinical and biological data, including cytological results and patients' HPV history were collected. Of the 6101 samples collected, 1.7% had an HPV result in the gray-zone (102 patients). The proportion of gray-zone results varied according to HPV genotype, reaching 11.8% of samples with detectable HPV DNA in the case of HPV31/33/52/58 genotypes. Reflex cytologies showed no abnormalities or Atypical Squamous Cells of Undetermined Significance results in 74.6% and 17.9% of cases, respectively. A previous or subsequent HPV-positive result with a (possibly) identical genotype was observed in 58% and 38% of cases, respectively. Two women with a history of persistent HPV detection had a CIN2+ lesion 1 year after the gray-zone result. In conclusion, the proportion of HPV results in the gray-zone varies according to genotype. No cytological abnormality is observed in the majority of cases, but a few rare patients with a history of persistent HPV infection should be closely monitored even if the HPV result is transiently located in the gray-zone.

Comparison of high-risk HPV detection by the AmpFire® HPV Screening 16/18/HR technique (Atila Biosystems) and the hybrid capture 2 test (Qiagen).

BACKGROUND: Detection of high-risk human papillomaviruses (hrHPV) is widely used at the first line of cervical cancer screening, requiring rigorous validation of the clinical performance of commercial kits designed for this indication. METHODS: Performance of the AmpFire HPV Screening 16/18/HR test (AF, Atila Biosystems) and the Hybrid Capture 2 test (HC2, Qiagen) for detecting hrHPV was cross-compared in 200 cervical samples in our institution. RESULTS: The global percentage of agreement between the 2 techniques was 95.0% (95%CI 92-98%) with a Cohen's kappa coefficient of 0.85 (95%CI 0.75-0.94). Ten samples showed discordant results between the 2 techniques in both directions (5 HC2+/AF- and 5 HC2-/AF+). Among possible explanations for these discrepancies was the detection of HPV66 and HPV53 genotypes in two samples, since these genotypes are targeted by the Ampfire test but not by the HC2 test, as well as intrinsic differences in analytical performance to target specific genotypes. CONCLUSIONS: A high level of agreement was observed between the two techniques, which encourages further testing in order to definitively validate the use of the Ampfire kit for primary cervical cancer screening.

Anogenital distribution of mucosal HPV in males and females before and after renal transplantation.

INTRODUCTION: Immunosuppressive drugs taken by transplant recipients may favor HPV infection at anogenital sites. HPV-type prevalence was studied in males and females before and after renal transplantation. PATIENTS AND METHODS: Anal, cervical and penile samples were taken from 62 patients before transplantation and from 41 patients after transplantation. HPV DNA was investigated using the INNO-LiPA HPV genotyping extra test and HPV-type distribution determined. RESULTS: Before transplantation, up to 30% of analyzed samples harbored HPV DNA, with the highest prevalence found in cervical specimens (60%). After transplantation, a trend toward HPV clearance was observed in females. By contrast, a trend toward incident infections by a wide variety of HPV genotypes at the penis and anal level was documented in men. CONCLUSION: High prevalence of HPV at anogenital sites was documented before and after renal transplantation. Immunosuppressive drugs taken after transplantation may impact HPV acquisition or reactivation, especially in males. Special attention should be paid in view of preventing HPV-associated diseases in this vulnerable population.

Anal and oropharyngeal HPV distribution in HIV-negative multipartner MSM using self-sampling kits for HIV and sexually transmitted infection screening.

Men who have sex with men (MSM) are at high risk of sexually transmitted infections, among which HPV infections are particularly prominent. We took advantage of the MémoDépistages study to evaluate HPV distribution at anal and oropharyngeal sites in HIV-negative multipartner MSM. HPV DNA was detected in 82% (n = 344) of anal and 11% (n = 45) of oropharyngeal self-collected samples taken from 421 participants. Multiple HPV types were detected in 70% of anal samples, and single HPV types in 91% of oropharyngeal samples. HPV16 was the most frequent type detected in the anus, followed by HPV6, HPV51, and HPV52. HPV6, HPV16, and HPV11 were the most prevalent types in the oropharynx. HPV targeted by the nonavalent vaccine was detected in 71% and 50% of HPV-positive anal and oropharyngeal samples, respectively. The main risk factor associated with HPV detection was frequenting gay meeting places, living in large cities, and having an anal Chlamydia trachomatis/Neisseria gonorrhoeae infection. In this cohort of highly sexually active MSM, HPV detection was highly frequent and rendered them at high risk of precancerous and cancerous lesions. Universal vaccination against HPV before sexual debut is an important public health strategy to prevent HPV-associated cancers in this highly vulnerable population of HIV-negative MSM.

Prevention and control of HPV and HPV-related cancers in France: the evolving landscape and the way forward - a meeting report.

Misinformation regarding HPV vaccine safety and benefits has resulted in low coverage within the eligible French population. HPV vaccination is safe and efficacious in preventing HPV infections in adolescents. However, reaching optimal coverage in countries such as France is challenging due to misinformation, among other factors. Moreover, disparities exist in cervical cancer screening programs. To support the government health promotion policy aimed at improving prevention and control of HPV-related cancers in France, the Human Papillomavirus Prevention and Control Board (HPV-PCB), in collaboration with local experts, held a meeting in Annecy, France (December 2021).HPV-PCB is an independent, multidisciplinary board of international experts that disseminates relevant information on HPV to a broad array of stakeholders and provides guidance on strategic, technical and policy issues in the implementation of HPV control programs.After a one-and-a-half-day meeting, participants concluded that multi-pronged strategies are required to expand vaccination coverage and screening. Vaccine acceptance could be improved by: 1) strenghtening existing trust in clinicians by continuous training of current and upcoming/pre-service healthcare professionals (HCPs), 2) improving health literacy among adolescents and the public through school and social media platforms, and 3) providing full reimbursement of the gender-neutral HPV vaccine, as a strong signal that this vaccination is essential.The discussions on HPV infections control focused on the need to: 1) encourage HCPs to facilitate patient data collection to support performance assessment of the national cervical cancer screening program, 2) advance the transition from cytology to HPV-based screening, 3) improve cancer prevention training and awareness for all HCPs involved in screening, including midwives, 4) identifying patient barriers to invitation acceptance, and 5) promoting urine or vaginal self-sampling screening techniques to improve acceptability, while establishing appropriate follow-up strategies for HPV-positive women. This report covers some critical findings, key challenges, and future steps to improve the status of HPV prevention and control measures in the country.

Human papilloma virus-16-specific CD8+ T-cell expansions characterize different clinical forms of lichen planus and not lichen sclerosus et atrophicus.

Lichen planus (LP) is a cutaneomucosal chronic inflammatory disease characterized by a CD8+ cytotoxic T-lymphocytes (CTL) infiltrate. In erosive oral LP, we found HPV16-specific activated CTL in lesions, supporting a pathogenic contribution of HPV16. Here, we investigated whether a similar scenario occurs in other clinical forms of LP and in lichen sclerosus et atrophicus (LSA), another chronic disease also affecting the mucosa and/or the skin. Blood CTL from LP and LSA patients expressed significant higher levels of granzyme B, perforin and CD107a proteins than healthy donors. Expansions of TCRVß3+ CTL, with presence of TCR clonotypes identical to those previously detected in erosive oral LP, were found both in blood and mucosal/skin lesions of LP, and not of LSA patients. These expansions were enriched with HPV16-specific CD8+ T-cells as shown by their recognition of the E711-20 immunodominant epitope. In LSA patients, the peripheral repertoire of CTL was oligoclonal for TCRVß6+ CTL. Finally, although patients with LP and LSA have developed antibodies against HPV16 capsid L1, antibodies against HPV16 E6 were only observed in patients with LP. Overall, our data collectively suggest an involvement of HPV16-specific CTL in different clinical forms of LP, not only in erosive oral LP, while a different scenario operates in LSA.

High levels of HPV16-L1 antibody but not HPV16 DNA load or integration predict oropharyngeal patient outcome: The Papillophar study.

The incidence of oropharyngeal cancers (OPC) is increasing in the world. Among OPC, those induced by human papillomaviruses have a better prognosis than non-HPV-associated OPC. The objective of this study was to highlight the relevance of HPV16 load, HPV16 DNA integration and HPV16-L1 serology on progression-free survival and overall survival of OPC patients. The PAPILLOPHAR cohort consists of 362 patients with oropharyngeal squamous cell carcinomas prospectively followed up for 5 years after treatment. Tumor biopsies and sera were collected at inclusion to investigate tumor HPV DNA/RNA characteristics and HPV16 L1 serology, respectively. Twenty-seven percent of tumor biopsies were HPV DNA- and RNA-positive and HPV16 represented 93% of HPV-positive cases. Among them, neither HPV16 viral load nor HPV16 DNA integration was associated with overall survival (OS) or progression-free survival (PFS). In contrast, high anti-HPV16 L1 antibody titers were significantly associated with a better OS and PFS. This study reveals that HPV16 load and integration are not relevant prognosis biomarkers in OPC patients.Clinical Relevance: High levels of HPV16 L1 antibodies may be useful to predict OPC patient outcome following treatment.ClinicalTrials.gov Identifier: NCT00918710, May 2017.

Prognostic role of HPV integration status and molecular profile in advanced anal carcinoma: An ancillary study to the epitopes-HPV02 trial.

Squamous Cell Carcinoma of the Anal canal (SCCA) is a rare disease associated with a Human Papillomavirus (HPV) infection in most cases, predominantly the HPV16 genotype. About 15% of SCCA are diagnosed in metastatic stage and some will relapse after initial chemoradiotherapy (CRT). Treatment of patients by Docetaxel, Cisplatin and 5-fluorouracil (DCF) has been recently shown to improve their complete remission and progression-free survival. The aim of this retrospective study was to explore the impact of HPV infection, HPV DNA integration, TERT promoter mutational status and somatic mutations of oncogenes on both progression-free (PFS) and overall survivals (OS) of patients treated by DCF. Samples obtained from 49 patients included in the Epitopes-HPV02 clinical trial, diagnosed with metastatic or non-resectable local recurrent SCCA treated by DCF, were used for analyses. Median PFS and OS were not associated with HPV status. Patients with episomal HPV had an improved PFS compared with SCCA patients with integrated HPV genome (p=0.07). TERT promoter mutations were rarely observed and did not specifically distribute in a subset of SCCA and did not impact DCF efficacy. Among the 42 genes investigated, few gene alterations were observed, and were in majority amplifications (68.4%), but none were significantly correlated to PFS. As no biomarker is significantly associated with patients' survival, it prompts us to include every patient failing CRT or with metastatic disease in DCF strategy.

Longitudinal follow-up of HPV16 sequence after cervical infection: Low intrahost variation and no correlation with clinical evolution.

Human papillomavirus (HPV) 16 exhibits different variants that may differ in their carcinogenic risk. To identify some high-risk variants, we sequenced and compared HPV16 whole genomes obtained from a longitudinal cohort of 34 HPV16-infected women who had either spontaneously cleared their infection (clearance group or "C"), or developed cervical high-grade lesions following a viral persistence (group persistence or "P"). Phylogenetic analysis of paired samples obtained at the beginning (C0 or P0) and at the end (C2 or P2) of the follow-up (median intervals between C0-C2 and between P0-P2 were 16 and 36.5 months, respectively) revealed a low genetic variability within the host compared to the genetic interhost diversity. By comparing our HPV16 sequences to a reference sequence, we observed 301 different substitutions, more often transitions (60.9%) than transversions (39.1%), that occurred throughout the viral genome, but with a low frequency in E6 and E7 oncogenes (10 and 9 substitutions), suggesting a high conservation of these genes. Deletions and insertions were mostly observed in intergenic regions of the virus. The only significant substitution found between the subgroups C2 and P2 was observed in the L2 gene (L330F), with an unclear biological relevance. Our results suggest a low longitudinal intrahost evolution of HPV16 sequences and no correlation between genetic variations and clinical evolution.

Low correlation between Ki67 assessed by qRT-PCR in Oncotype Dx score and Ki67 assessed by Immunohistochemistry.

Breast cancers expressing high levels of Ki67 are associated with poor outcomes. Oncotype DX test was designed for ER+/HER2- early-stage breast cancers to help adjuvant chemotherapy decision by providing a Recurrent Score (RS). RS measures the expression of 21 specific genes from tumor tissue, including Ki67. The primary aim of this study was to assess the agreement between Ki67RNA obtained with Oncotype DX RS and Ki67IHC. Other objectives were to analyze the association between the event free survival (EFS) and the expression level of Ki67RNA; and association between RS and Ki67RNA. Herein, we report a low agreement of 0.288 by Pearson correlation coefficient test between Ki67IHC and Ki67RNA in a cohort of 98 patients with early ER+/HER2- breast cancers. Moreover, Ki67RNAhigh tumors were significantly associated with the occurrence of events (p = 0.03). On the other hand, we did not find any association between Ki67IHC and EFS (p = 0.26). We observed a low agreement between expression level of Ki67RNA and Ki67 protein labelling by IHC. Unlike Ki67IHC and independently of the RS, Ki67RNA could have a prognostic value. It would be interesting to better assess the prognosis and predictive value of Ki67RNA measured by qRT-PCR. The Ki67RNA in medical routine could be a good support in countries where Oncotype DX is not accessible.

HPV status and therapeutic initial strategy impact on survival and oncologic outcomes: 5-year results from the multicentric prospective cohort of oropharyngeal cancers Papillophar.

PURPOSE: After the 2 years of follow-up, we aimed to evaluate at 5 years the impact of human papillomavirus (HPV) status, tobacco, and initial treatment approach on progression-free survival (PFS) and overall survival (OS) of patients with oropharyngeal cancer (OPC) in France. METHODS: Papillophar study was designed as a prospective cohort of 340 OPC patients in 14 French hospitals. HPV-positive status (21.7%) was defined with PCR (positivity for HPV DNA and E6/E7 mRNA). Cox proportional hazard models were used to assess the relationship between PFS, OS, HPV, and other prognostic factors. The combined effect of HPV status with smoking, stage, or initial treatment on PFS was also evaluated. RESULTS: HPV-pos patients had better PFS than HPV-neg patients (HR = 0.46; 95% CI: 0.29-0.74), and worse for older patients (HR for 5-year age increase = 1.14), UICC stage 4 from the 7th TNM classification compared to stage 1-2 (HR = 2.58; CI: 1.33-5.00), and those having had radiotherapy (HR = 2.07; 95% CI: 1.36-3.16) or induction chemotherapy (HR = 2.11; 95% CI: 1.32-3.38) instead of upfront surgery. HPV-neg patients encountered a larger incidence of loco-regional disease than HPV-pos patients (31.5% and 14.0%, respectively, p = 0.0001). Distant metastases proportion was similar. HPV-neg patients developed more second primary tumor than HPV-pos patients (11.7% vs. 3.3%, p = 0.02). CONCLUSIONS: 5-year follow-up confirms the specifically improved prognosis in HPV-positive patients. The prognosis is nevertheless significantly modified through clinical and therapeutical variations.

The detection of specific hypermethylated WIF1 and NPY genes in circulating DNA by crystal digital PCR™ is a powerful new tool for colorectal cancer diagnosis and screening.

BACKGROUND: In oncology, liquid biopsy is of major relevance from theranostic point of view. The searching for mutations in circulating tumor DNA (ctDNA) in case of colorectal cancers (CRCs) allows the optimization of patient care. In this context, independent of mutation status biomarkers are required for its detection to confirm the presence of ctDNA in liquid biopsies. Indeed, the hypermethylation of NPY and WIF1 genes appear to be an ideal biomarker for the specific detection of ctDNA in CRCs. The objective of this work is to develop the research of hypermethylation of NPY and WIF1 by Crystal Digital PCR™ for the detection of ctDNA in CRCs. METHODS: Detection of hypermethylated NPY and WIF1 was developed on Cristal digital PCR™. Biological validation was performed from a local cohort of 22 liquid biopsies and 23 tissue samples from patients with CRC. These patients were treated at the University Hospital of Besancon (France). RESULTS: The local cohort study confirmed that NPY and WIF1 were significantly hypermethylated in tumor tissues compared to adjacent non-tumor tissues (WIF1 p < 0.001; NPY p < 0.001; non-parametric Wilcoxon paired-series test). Histological characteristics, tumor stages or mutation status were not correlated to the methylation profiles. On the other hand, hypermethylation of NPY or WIF1 in liquid biopsy had a 95.5% [95%CI 77-100%] sensitivity and 100% [95%CI 69-100%] specificity. CONCLUSION: Using Crystal digital PCR™, this study shows that hypermethylation of NPY and WIF1 are constant specific biomarkers of CRCs regardless of a potential role in carcinogenesis.

HPV16 Load Is a Potential Biomarker to Predict Risk of High-Grade Cervical Lesions in High-Risk HPV-Infected Women: A Large Longitudinal French Hospital-Based Cohort Study.

High-risk HPV (hrHPV) testing has been implemented as a primary screening tool for cervical cancer in numerous countries. However, there is still a need for relevant triage strategies to manage hrHPV positive women to avoid excessive referral to colposcopy. The objective of this study was to assess, in women infected by hrHPV and presenting no or mild cytological abnormalities, HPV16 and HPV18 viral loads to predict the development of cervical high-grade lesion. Among 2102 women positive for hrHPV, 885 had no lesion or mild cytological abnormalities at baseline and had at least one follow-up (FU) visit. HPV16 and HPV18 prevalence was 25.9% and 8.4%, respectively. Of those women, 15% developed a high-grade lesion during the FU. An HPV16 viral load cut-off set at 3.2 log10GE/103 cells permitted to identify a subgroup of women at high risk of developing high-grade cervical lesion (HR = 2.67; 95% CI 1.80-3.97; p ≤ 0.0001). No specific HPV18 viral load threshold could have been defined in regard to the present study. In multivariate analysis, HPV16 load (absence/log10GE/103 cells < 3.2 vs. ≥3.2), RLU/PC 239 (1-100 pg/mL vs. >100 pg/mL) and cytology (normal vs abnormal) were independently associated with a significant increased risk of high-grade lesion development and were used to construct the prognostic score. In conclusion, HPV16 load is a relevant biomarker to identify women at high risk for developing cervical precancerous lesions.

Human papillomavirus DNA and p16 expression in head and neck squamous cell carcinoma in young French patients.

OBJECTIVES: Human papillomavirus (HPV) is a risk factor for head and neck squamous cell carcinoma (HNSCC), which is currently increasing worldwide. We evaluated the prevalence of HPV DNA and p16 expression in HNSCC patients age <45 years compared with patients aged ≥45 years. METHODS: Thirty-nine patients aged <45 years who presented at Besançon University Hospital with HNSCC since 2005 were included in this retrospective study. HPV DNA was detected by HPV genotyping and p16 expression was determined by immunohistochemistry using paraffin-embedded tissues. A matched-group of 38 patients aged ≥45 years from Besançon University Hospital was included. RESULTS: The overall prevalence of HPV infection was 11.7%. HPV16 was the only genotype detected in 4/39 and 5/38 patients, and p16 was expressed in 6/39 and 4/38 patients aged <45 years and ≥45 years, respectively. CONCLUSIONS: HPV-positivity and p16 expression were similar in both age groups. The results suggest that p16 immunohistochemistry may provide a prognosis biomarker for all HNSCCs, not only oropharyngeal cancers, and this should be addressed in large clinical trials.

Successful retrieval of human papillomavirus DNA after a 4.5 year storage on FTA elute cards.

Efficient primary (vaccination) and secondary (screening) prevention strategies have the potential to eliminate cervical cancer worldwide. In this context, surveillance of HPV infections remains mandatory to assess the efficacy and the impact of screening and vaccination policies. Therefore there is a need to safely store cervical samples to conduct long-term studies in vaccinated and non-vaccinated subjects. Up-dated data on cervical specimen preservation on FTA® cards indicate that HPV DNA can be safely retrieved after 54 months of storage. A concordance of 97 % was achieved between HPV genotypes detected in initial cervical samples and on FTA® cards 4.5 years later. Even if a drop in HPV viral loads was observed in some cases at 4.5 years, using FTA® cards for safe and long-term storage of cervical samples represents an interesting option.

The exon junction complex core factor eIF4A3 is a key regulator of HPV16 gene expression.

High-risk human papillomavirus (hrHPVs), particularly HPV16 and HPV18, are the etiologic factors of ano-genital cancers and some head and neck squamous cell carcinomas (HNSCCs). Viral E6 and E7 oncoproteins, controlled at both transcriptional and post-transcriptional levels, drive hrHPVs-induced carcinogenesis. In the present study, we investigated the implication of the DEAD-box helicase eukaryotic translation initiation factor 4A3 (eIF4A3,) an Exon Junction Complex factor, in the regulation of HPV16 gene expression. Our data revealed that the depletion of the factor eIF4A3 up-regulated E7 oncoprotein levels. We also showed that the inhibition of the nonsense-mediated RNA decay (NMD) pathway, resulted in the up-regulation of E7 at both RNA and protein levels. We therefore proposed that HPV16 transcripts might present different susceptibilities to NMD and that this pathway could play a key role in the levels of expression of these viral oncoproteins during the development of HPV-related cancers.

Treatment algorithm and prognostic factors for patients with stage I-III carcinoma of the anal canal: a 20-year multicenter study.

Despite a growing incidence in developed countries and a recent improved understanding of its pathogenesis, anal cancer management has not evolved over the past decades and drug combination used as first-line regimen still largely depends on clinician preferences. Aiming at paving the way for precision medicine, a large cohort of 372 HIV-negative patients diagnosed over a 20-year time period with locally advanced anal carcinoma was collected and carefully characterized at the clinical, demographic, histopathologic, immunologic, and virologic levels. Both the prognostic relevance of each clinicopathological parameter and the efficacy of different concurrent chemoradiation strategies were determined. Overall, the incidence of anal cancer peaked during the sixth decade (mean: 63.4) and females outnumbered males (ratio: 2.51). After completion of treatment, 95 (25.5%) patients experienced progression of persistent disease or local/distant recurrence and 102 (27.4%) died during the follow-up period (median: 53.8 months). Importantly, uni-multivariate analyses indicated that both negative HPV/p16ink4a status and aberrant p53 expression were far better predictors for reduced progression-free survival than traditional risk factors such as tumor size and nodal status. As for overall survival, the significant influences of age at diagnosis, p16ink4a status, cTNM classification as well as both CD3+ and CD4+ T-cell infiltrations within tumor microenvironment were highlighted. Cisplatin-based chemoradiotherapy was superior to both radiotherapy alone and other concurrent chemoradiation therapies in the treatment of HPV-positive tumors. Regarding their HPV-uninfected counterparts, frequent relapses were observed, whatever the treatment regimen administered. Taken together, our findings reveal that current anal cancer management and treatment have reached their limits. A dualistic classification according to HPV/p53 status should be considered with implications for therapy personalization and optimization.