Analysis of Memory B-Cell Responses Reveals Suboptimal Dosing Schedule of a Licensed Vaccine.

Current guidance recommends that adolescents receive a 2-dose human papillomavirus (HPV) vaccine, whereas young adults and immunocompromised persons receive 3 doses. We examined secondary responses of vaccine-elicited memory B cells (Bmem) in naive women receiving 3 doses of the quadrivalent HPV vaccine to understand the quality of B-cell memory generated by this highly effective vaccine. Unexpectedly, we observed a lower Bmem response rate and magnitude of Bmem responses to the third dose than to a booster dose administered at month 24. Moreover, high titers of antigen-specific serum antibody at vaccination inversely correlated with Bmem responses. As the purpose of additional doses/boosters is to stimulate Bmem to rapidly boost antibody levels, these results indicate the timing of the third dose is suboptimal and lend support to a 2-dose HPV vaccine for young adults. Our findings also indicate more broadly that multidose vaccine schedules should be rationally determined on the basis of Bmem responses.

Viral oncoprotein antibodies as a marker for recurrence of Merkel cell carcinoma: A prospective validation study.

BACKGROUND: Merkel cell carcinoma (MCC) is an aggressive skin cancer with a recurrence rate of >40%. Of the 2000 MCC cases per year in the United States, most are caused by the Merkel cell polyomavirus (MCPyV). Antibodies to MCPyV oncoprotein (T-antigens) have been correlated with MCC tumor burden. The present study assesses the clinical utility of MCPyV-oncoprotein antibody titers for MCC prognostication and surveillance. METHODS: MCPyV-oncoprotein antibody detection was optimized in a clinical laboratory. A cohort of 219 patients with newly diagnosed MCC were followed prospectively (median follow-up, 1.9 years). Among the seropositive patients, antibody titer and disease status were serially tracked. RESULTS: Antibodies to MCPyV oncoproteins were rare among healthy individuals (1%) but were present in most patients with MCC (114 of 219 patients [52%]; P < .01). Seropositivity at diagnosis independently predicted decreased recurrence risk (hazard ratio, 0.58; P = .04) in multivariate analyses adjusted for age, sex, stage, and immunosuppression. After initial treatment, seropositive patients whose disease did not recur had rapidly falling titers that became negative by a median of 8.4 months. Among seropositive patients who underwent serial evaluation (71 patients; 282 time points), an increasing oncoprotein titer had a positive predictive value of 66% for clinically evident recurrence, whereas a decreasing titer had a negative predictive value of 97%. CONCLUSIONS: Determination of oncoprotein antibody titer assists in the clinical management of patients with newly diagnosed MCC by stratifying them into a higher risk seronegative cohort, in which radiologic imaging may play a more prominent role, and into a lower risk seropositive cohort, in which disease status can be tracked in part by oncoprotein antibody titer. Cancer 2017;123:1464-1474. © 2016 American Cancer Society.

Re-detection vs. new acquisition of high-risk human papillomavirus in mid-adult women.

To understand high-risk (hr) human papillomavirus (HPV) epidemiology in mid-adulthood, we assessed whether associations between incident detection of hrHPV DNA and recent sexual behavior differed according to whether or not there was serologic evidence of prior infection. From 2011 to 2012, we enrolled 409 women aged 30-50 years into a 6-month longitudinal study. We collected health and sexual behavior histories, enrollment sera for HPV antibody testing, and monthly self-collected vaginal swabs for HPV DNA genotyping. Generalized estimating equations logistic regression identified risk factors for type-specific incident hrHPV DNA, stratified by type-specific hrHPV serostatus at enrollment. Population attributable risks of hrHPV due to prior and recent exposure were estimated. When type-specific hrHPV serology was negative, recent sexual risk behavior was positively associated with incident hrHPV DNA (odds ratio in women reporting ≥3 recent sexual risk behaviors [e.g., new or multiple partners] vs. no recent sexual activity = 9.8, 95% CI: 2.4-40.6). No associations with recent sexual behavior were observed with positive type-specific hrHPV serology. Thirty percent of incident hrHPV DNA detection was attributable to prior infection (with positive serology) and 40% was attributable to recent sexual risk behavior (with negative serology). The proportion of incident hrHPV DNA detection attributable to recent sexual risk behavior decreased with increasing age. Among women with serologic evidence of prior infection, re-detection of the same hrHPV type is likely due to reactivation or intermittent detection of persistent infection. Without serologic evidence of prior infection, new detection is likely due to new acquisition or to intermittent detection of persisting infection.

Characteristics of memory B cells elicited by a highly efficacious HPV vaccine in subjects with no pre-existing immunity.

Licensed human papillomavirus (HPV) vaccines provide near complete protection against the types of HPV that most commonly cause anogenital and oropharyngeal cancers (HPV 16 and 18) when administered to individuals naive to these types. These vaccines, like most other prophylactic vaccines, appear to protect by generating antibodies. However, almost nothing is known about the immunological memory that forms following HPV vaccination, which is required for long-term immunity. Here, we have identified and isolated HPV 16-specific memory B cells from female adolescents and young women who received the quadrivalent HPV vaccine in the absence of pre-existing immunity, using fluorescently conjugated HPV 16 pseudoviruses to label antigen receptors on the surface of memory B cells. Antibodies cloned and expressed from these singly sorted HPV 16-pseudovirus labeled memory B cells were predominantly IgG (>IgA>IgM), utilized diverse variable genes, and potently neutralized HPV 16 pseudoviruses in vitro despite possessing only average levels of somatic mutation. These findings suggest that the quadrivalent HPV vaccine provides an excellent model for studying the development of B cell memory; and, in the context of what is known about memory B cells elicited by influenza vaccination/infection, HIV-1 infection, or tetanus toxoid vaccination, indicates that extensive somatic hypermutation is not required to achieve potent vaccine-specific neutralizing antibody responses.

Determinants of High-Risk Human Papillomavirus Seroprevalence and DNA Prevalence in Mid-Adult Women.

BACKGROUND: The epidemiology of high-risk human papillomavirus (hrHPV) infections in mid-adult women is not well understood. METHODS: We conducted a cross-sectional analysis of 379 women 30 to 50 years of age. Vaginal samples were tested for type-specific HPV DNA by polymerase chain reaction. Sera were tested for type-specific HPV antibodies by Luminex-based assay. Assays included 13 hrHPV types (16/18/31/33/35/39/45/51/52/56/58/59/68). Self-reported health and sexual history were ascertained. Risk factors for seropositivity and DNA positivity to hrHPV were assessed in separate Poisson regression models. RESULTS: The mean (SD) age of participants was 38.7 (6.1) years, and the median lifetime number of male sex partners was 7. Approximately two-thirds (68.1%) were seropositive for any hrHPV, 15.0% were DNA positive, and 70.7% were seropositive or DNA positive. In multivariate analyses, women who were married/living with a partner were less likely to be seropositive than single/separated women (adjusted prevalence ratio [aPR], 0.86; 95% confidence interval [CI], 0.75-0.98). Compared with never hormonal contraceptive users, current (aPR, 1.53; 95% CI, 1.01-2.29) or former (aPR, 1.64; 95% CI, 1.10-2.45) users were more likely to be seropositive. Women with a lifetime number of sex partners of 12 or more were more likely to be seropositive compared with those with 0 to 4 partners (aPR, 1.29; 95% CI, 1.06-1.56). Similar associations were seen with DNA positivity. In addition, there was a positive association between current smoking and hrHPV DNA (aPR vs. never smokers, 2.51; 95% CI, 1.40-4.49). CONCLUSIONS: Seventy-one percent of mid-adult women had evidence of current or prior hrHPV infection. Measures of probable increased exposure to HPV infection were associated with both seropositivity and DNA positivity to hrHPV, whereas current smoking was positively associated with hrHPV DNA only.

Absence of an association of human polyomavirus and papillomavirus infection with lung cancer in China: a nested case-control study.

BACKGROUND: Studies of human polyomavirus (HPyV) infection and lung cancer are limited and those regarding the association of human papillomavirus (HPV) infection and lung cancer have produced inconsistent results. METHODS: We conducted a nested case-control study to assess the association between incident lung cancer of various histologies and evidence of prior infection with HPyVs and HPVs. We selected serum from 183 cases and 217 frequency matched controls from the Yunnan Tin Miner's Cohort study, which was designed to identify biomarkers for early detection of lung cancer. Using multiplex liquid bead microarray (LBMA) antibody assays, we tested for antibodies to the VP1 structural protein and small T antigen (ST-Ag) of Merkel cell, KI, and WU HPyVs. We also tested for antibodies against HPV L1 structural proteins (high-risk types 16, 18, 31, 33, 52, and 58 and low-risk types 6 and 11) and E6 and E7 oncoproteins (high risk types 16 and 18). Measures of antibody reactivity were log transformed and analyzed using logistic regression. RESULTS: We found no association between KIV, WUV, and MCV antibody levels and incident lung cancer (P-corrected for multiple comparisons >0.10 for all trend tests). We also found no association with HPV-16, 18, 31, 33, 52, and 58 seropositivity (P-corrected for multiple comparisons >0.05 for all). CONCLUSIONS: Future studies of infectious etiologies of lung cancer should look beyond HPyVs and HPVs as candidate infectious agents.

Identification of an overprinting gene in Merkel cell polyomavirus provides evolutionary insight into the birth of viral genes.

Many viruses use overprinting (alternate reading frame utilization) as a means to increase protein diversity in genomes severely constrained by size. However, the evolutionary steps that facilitate the de novo generation of a novel protein within an ancestral ORF have remained poorly characterized. Here, we describe the identification of an overprinting gene, expressed from an Alternate frame of the Large T Open reading frame (ALTO) in the early region of Merkel cell polyomavirus (MCPyV), the causative agent of most Merkel cell carcinomas. ALTO is expressed during, but not required for, replication of the MCPyV genome. Phylogenetic analysis reveals that ALTO is evolutionarily related to the middle T antigen of murine polyomavirus despite almost no sequence similarity. ALTO/MT arose de novo by overprinting of the second exon of T antigen in the common ancestor of a large clade of mammalian polyomaviruses. Taking advantage of the low evolutionary divergence and diverse sampling of polyomaviruses, we propose evolutionary transitions that likely gave birth to this protein. We suggest that two highly constrained regions of the large T antigen ORF provided a start codon and C-terminal hydrophobic motif necessary for cellular localization of ALTO. These two key features, together with stochastic erasure of intervening stop codons, resulted in a unique protein-coding capacity that has been preserved ever since its birth. Our study not only reveals a previously undefined protein encoded by several polyomaviruses including MCPyV, but also provides insight into de novo protein evolution.

Natural Antibodies to Human Papillomavirus 16 and Recurrence of Vulvar High-Grade Intraepithelial Neoplasia (VIN3).

OBJECTIVES: Approximately 30% of women treated for squamous high-grade intraepithelial neoplasia (VIN3), often associated with human papillomavirus (HPV), have recurrent disease. In this study, we assess predictors of recurrence that may provide targets for early prevention or treatment. MATERIALS AND METHODS: Women with VIN3 who participated in a previous population-based case-control study with blood and tumor samples completed a follow-up telephone interview an average of 5 years after initial diagnosis. The risk of recurrence was determined by proportional hazards modeling. RESULTS: Women with VIN3 in the follow-up study (n = 65) were similar to women with VIN3 in the parent study (n = 215) with regard to age at primary diagnosis, level of current cigarette smoking (>60%), and lifetime number of partners. We found that 22 (33.8%) of 65 participants had a vulvar recurrence and that 73.4% recurred within 3 years of treatment. Recurrences occurred more often among women with common warts in the decade before diagnosis (hazard ratio [HR] = 2.5, 95% CI = 1.1-5.8) and among those with a previous anogenital cancer (HR = 2.7, 95% CI = 1.2-6.3). Interestingly, recurrence was less frequent among women who mounted a natural antibody response to HPV16 (HR = 0.4, 95% CI = 0.2-0.9). CONCLUSIONS: These data provide strong preliminary evidence that VIN3 recurrence was less frequent among those with HPV16 antibodies. Vaccination with the currently licensed HPV vaccine as part of adjunctive therapy for VIN3 would increase antibody response and may decrease risk of recurrence. Randomized controlled trials are needed to determine whether HPV vaccination is effective against VIN3 recurrence.

Prior human polyomavirus and papillomavirus infection and incident lung cancer: a nested case-control study.

PURPOSE: To test whether infection with select human polyomaviruses (HPyV) and human papillomaviruses (HPV) is associated with incident lung cancer. METHODS: We performed a nested case-control study, testing serum from the carotene and retinol efficacy trial, conducted 1985-2005, for antibodies to Merkel cell (MCV), KI (KIV), and WU (WUV) HPyVs as well as to six high-risk and two low-risk HPV types. Incident lung cancer cases (n = 200) were frequency-matched with controls (n = 200) on age, enrollment and blood draw dates, intervention arm assignment, and the number of serum freeze/thaw cycles. Sera were tested using multiplex liquid bead microarray antibody assays. We used logistic regression to assess the association between HPyV and HPV antibodies and lung cancer. RESULTS: There was no evidence of a positive association between levels of MCV, KIV, or WUV antibodies and incident lung cancer (p corrected >0.10 for all trend tests; odds ratio (OR) range 0.72-1.09, p corrected >0.10 for all). There was also no evidence for a positive association between HPV 16 or 18 infection and incident lung cancer (p corrected ≥0.10 for all trend tests; OR range 0.25-2.54, p > 0.05 for all OR > 1), but the number of persons with serologic evidence of these infections was small. CONCLUSIONS: Prior infection with any of several types of HPyV or HPV was not associated with subsequent diagnosis of lung cancer. Infection with these viruses likely does not influence a person's risk of lung cancer in Western smoking populations.

Polyomavirus ALTOs, but not MTs, downregulate viral early gene expression by activating the NF-κB pathway.

Polyomaviruses are small, circular dsDNA viruses that can cause cancer. Alternative splicing of polyomavirus early transcripts generates large and small tumor antigens (LT, ST) that play essential roles in viral replication and tumorigenesis. Some polyomaviruses also express middle tumor antigens (MTs) or Alternate LT ORFs (ALTOs), which are evolutionarily related but have distinct gene structures. MTs are a splice variant of the early transcript whereas ALTOs are overprinted on the second exon of the LT transcript in an alternate reading frame and are translated via an alternative start codon. Merkel cell polyomavirus (MCPyV), the only human polyomavirus that causes cancer, encodes an ALTO but its role in the viral lifecycle and tumorigenesis has remained elusive. Here, we show MCPyV ALTO acts as a tumor suppressor and is silenced in Merkel cell carcinoma (MCC). Rescuing ALTO in MCC cells induces growth arrest and activates NF-κB signaling. ALTO activates NF-κB by binding SQSTM1 and TRAF2&3 via two N-Terminal Activating Regions (NTAR1+2), resembling Epstein-Barr virus (EBV) Latent Membrane Protein 1 (LMP1).. Following activation, NF-κB dimers bind the MCPyV non-coding control region (NCCR) and downregulate early transcription. Beyond MCPyV, NTAR motifs are conserved in other polyomavirus ALTOs, which activate NF-κB signaling, but are lacking in MTs that do not. Furthermore, polyomavirus ALTOs downregulate their respective viral early transcription in an NF-κB and NTAR dependent manner. Our findings suggest that ALTOs evolved to suppress viral replication and promote viral latency and that MCPyV ALTO must be silenced for MCC to develop.

LAMP1 targeting of the large T antigen of Merkel cell polyomavirus results in potent CD4 T cell responses and tumor inhibition.

Introduction: Most cases of Merkel cell carcinoma (MCC), a rare and highly aggressive type of neuroendocrine skin cancer, are associated with Merkel cell polyomavirus (MCPyV) infection. MCPyV integrates into the host genome, resulting in expression of oncoproteins including a truncated form of the viral large T antigen (LT) in infected cells. These oncoproteins are an attractive target for a therapeutic cancer vaccine. Methods: We designed a cancer vaccine that promotes potent, antigen-specific CD4 T cell responses to MCPyV-LT. To activate antigen-specific CD4 T cells in vivo, we utilized our nucleic acid platform, UNITE™ (UNiversal Intracellular Targeted Expression), which fuses a tumor-associated antigen with lysosomal-associated membrane protein 1 (LAMP1). This lysosomal targeting technology results in enhanced antigen presentation and potent antigen-specific T cell responses. LTS220A, encoding a mutated form of MCPyV-LT that diminishes its pro-oncogenic properties, was introduced into the UNITE™ platform. Results: Vaccination with LTS220A-UNITE™ DNA vaccine (ITI-3000) induced antigen-specific CD4 T cell responses and a strong humoral response that were sufficient to delay tumor growth of a B16F10 melanoma line expressing LTS220A. This effect was dependent on the CD4 T cells' ability to produce IFNγ. Moreover, ITI-3000 induced a favorable tumor microenvironment (TME), including Th1-type cytokines and significantly enhanced numbers of CD4 and CD8 T cells as well as NK and NKT cells. Additionally, ITI-3000 synergized with an α-PD-1 immune checkpoint inhibitor to further slow tumor growth and enhance survival. Conclusions: These findings strongly suggest that in pre-clinical studies, DNA vaccination with ITI-3000, using the UNITE™ platform, enhances CD4 T cell responses to MCPyV-LT that result in significant anti-tumor immune responses. These data support the initiation of a first-in-human (FIH) Phase 1 open-label study to evaluate the safety, tolerability, and immunogenicity of ITI-3000 in patients with polyomavirus-positive MCC (NCT05422781).

Serum antibody response following genital {alpha}9 human papillomavirus infection in young men.

Background. Although the prevalence of human papillomavirus (HPV) genital infection is similarly high in males and females, seroprevalence is lower in males. This study assessed rates and determinants of seroconversion after detection of genital HPV infection in young men. Methods. We investigated HPV type-specific seroconversion in a cohort of heterosexual male university students who had an α9 HPV type (HPV-16, -31, -33, -35, -52, -58, or -67) detected in the genital tract (n = 156). HPV DNA and antibodies were detected and typed using liquid bead-based multiplex assays. We calculated seroconversion using Kaplan-Meier survival analysis. Cox proportional hazards models with generalized estimating equations were used to examine associations with seroconversion. Results. Within 24 months of detecting genital HPV infection, type-specific seroconversion ranged from 4% for HPV-52 to 36% for HPV-31. HPV-16 seroconversion at 24 months was 13% (95% confidence interval [CI], 7%-25%). Among incident HPV infections, ever cigarette smoking and infection site(s) (shaft/scrotum and glans/urine vs shaft/scrotum or glans/urine only) were positively associated with type-specific seroconversion. Conclusions. For each of the α9 HPV types, type-specific seroconversion within 24 months was observed in 36% or less of infected men. Seroconversion might be related to cigarette smoking and genital site(s) infected.

B Cell Responses upon Human Papillomavirus (HPV) Infection and Vaccination.

Infection with human papillomavirus (HPV) is the necessary cause of cervical cancer. Availability of vaccines against HPV makes it a highly preventable disease. HPV vaccines act through type-specific neutralizing antibodies produced by antigen-specific plasma cells known as long-lived plasma cells (LLPC). However, just as any other vaccine, success of HPV vaccine is attributed to the immunologic memory that it builds, which is largely attained through generation and maintenance of a class of B cells named memory B cells (Bmem). Both LLPCs and Bmems are important in inducing and maintaining immune memory and it is therefore necessary to understand their role after HPV vaccination to better predict outcomes. This review summarizes current knowledge of B-cell responses following HPV vaccination and natural infection, including molecular signatures associated with these responses.

Generation of a cost-effective cell line for support of high-throughput isolation of primary human B cells and monoclonal neutralizing antibodies.

The isolation of human monoclonal antibodies (mAbs) arising from natural infection with human pathogens has proven to be a powerful technology, facilitating the understanding of the host response to infection at a molecular level. mAbs can reveal sites of vulnerability on pathogens and illuminate the biological function of the antigenic targets. Moreover, mAbs have the potential to be used directly for therapeutic applications such as passive delivery to prevent infection in susceptible target populations, and as treatment of established infection. The isolation of antigen-specific B cells from vaccine trials can also assist in deciphering whether the desired B cells are being targeted by a given vaccine. Several different processes have been developed to isolate mAbs, but all are generally labor-intensive and result in varying degrees of efficiency. Here, we describe the development of a cost-effective feeder cell line that stably expresses CD40-ligand, interleukin-2 and interleukin-21. Sorting of single B cells onto a layer of irradiated feeder cells sustained antibody production that permits functional screening of secreted antibodies in a manner that enables subsequent recovery of B cells for recombinant antibody cloning. As a proof of concept, we show that this approach can be used to isolate B cells that secrete antibodies that neutralize human papilloma virus (HPV) from participants of an HPV vaccine study.

Exertional compartment syndrome of the thigh: a rare diagnosis and literature review.

Exercise-induced acute compartment syndrome of the thigh is an uncommon entity. We present a rare case of bilateral exercise-induced three-compartment syndrome of the thighs that required fasciotomies. The objective of this study was to understand the history, physical examination, signs, symptoms, pathophysiology, diagnosis, and treatment of compartment syndrome and rhabdomyolysis. A 42-year-old man presented to the Emergency Department (ED) complaining of worsening pain and swelling in both thighs 45 h after performing a lower extremity exercise regimen. The patient's thighs were tender and swollen, but there was no ecchymosis or evidence of trauma. Admitting serum creatinine kinase (CK) was 106,289 U/L. Treatment for rhabdomyolysis was initiated. The next day, he complained of escalating bilateral thigh pain. Repeat serum CK was 346,580 U/L. The patient was diagnosed with bilateral thigh compartment syndrome and immediately taken to the operating room for fasciotomies. Postoperatively, the patient's symptoms improved rapidly and his serum CK quickly returned to normal. His incisions were closed and he returned to normal activities of daily living. Because exercise-induced compartment syndrome is an extremely rare diagnosis with a high risk of poor outcome, this article serves to emphasize the importance of considering this diagnosis during the work-up of patients presenting to the ED with rhabdomyolysis.

Antibody responses in oral fluid after administration of prophylactic human papillomavirus vaccines.

We sought to determine whether oral fluid can be used to assess serum human papillomavirus (HPV) antibody status by enrolling women who had received a prophylactic HPV-16 vaccine in a new follow-up study. After the prophylactic HPV-6/11/16/18 vaccine was licensed in the United States, we administered it to consenting participants. With serologic findings used as the reference standard, The sensitivity of oral fluid was 49.6% (95% confidence interval [CI], 42.0%-57.3%) before and 100% (95% CI, 92.0%-100%) after administration of the quadrivalent vaccine. Oral fluid may have the potential to be used for monitoring of prophylactic HPV vaccines in the future.

Age of diagnosis of squamous cell cervical carcinoma and early sexual experience.

BACKGROUND: Given the established links among young age at first intercourse (AFI), number of sex partners, high-risk human papillomavirus infection, and squamous cell cervical cancer (SCC), we hypothesized that women diagnosed with SCC at younger ages would be more likely to report young AFI than women diagnosed later in life. METHODS: We performed a population-based investigation among invasive SCC cases who were diagnosed between 1986 and 2004, were ages 22 to 53 years, and lived in the metropolitan Seattle-Puget Sound region (n = 333). Using multivariate linear regression, we estimated coefficients and 95% confidence intervals (95% CI) to assess the association between age at SCC diagnosis and AFI (<15, 15-18, > or =19 years) and number of sex partners at age <20 years (0, 1, 2-4, 5-14, > or =15), accounting for birth year and other factors. Interactions were assessed using the likelihood ratio test. RESULTS: The interval between AFI and SCC diagnosis ranged from 4 to 35 years. In a multivariate model, compared with SCC cases reporting AFI > or =19, the mean age of diagnosis was 3.1 years younger for SCC cases reporting AFI <15 (95% CI, -5.8 to -0.5) and 2.6 years younger for SCC cases reporting AFI 15 to 18 (95% CI, -4.6 to -0.6). Although number of sex partners at age <20 years was associated with age at SCC diagnosis in a crude analysis, the association was not independent of AFI. However, in the AFI > or =19 and <15 groups, differences in effect were seen by number of sex partners at age <20 years (P(interaction) = 0.08), with the association remaining strong and significant only in the AFI <15 group that had > or =2 partners at age <20 years (coefficient, -4.2; 95% CI, -6.3 to -2.1). CONCLUSION: Among younger and middle-aged women with SCC, early age of diagnosis was associated with early AFI, although the effect appeared to be modified by number of sex partners at age <20 years.

Quantitative human papillomavirus 16 and 18 levels in incident infections and cervical lesion development.

Human papillomavirus (HPV) RNA levels may be a more sensitive early indicator of predisposition to carcinogenesis than DNA levels. We evaluated whether levels of HPV-16 and HPV-18 DNA and messenger RNA (mRNA) in newly detected infections are associated with cervical lesion development. Female university students were recruited from 1990 to 2004. Cervical samples for HPV DNA, HPV mRNA, and Papanicolaou testing were collected tri-annually, and women were referred for colposcopically directed biopsy when indicated. Quantitative real-time polymerase chain reaction of L1 and E7 DNA and E7 mRNA was performed on samples from women with HPV-16 and HPV-18 infections that were incidently detected by consensus PCR. Adjusting for other HPV types, increasing E7 cervical HPV-16 mRNA levels at the time of incident HPV-16 DNA detection were associated with an increased risk of cervical intraepithelial neoplasia grade 2-3 (HR per 1 log(10) increase in mRNA = 6.36, 95% CI = 2.00-20.23). Increasing HPV-16 mRNA levels were also associated with an increased risk of cervical squamous intraepithelial lesions; the risk was highest at the incident positive visit and decreased over time. Neither HPV-16 E7 DNA levels nor HPV-18 E7 DNA nor mRNA levels were significantly associated with cervical lesion development. Report of >1 new partner in the past 8 months (relative to no new partners) was associated with increased HPV mRNA (viral level ratio [VLR] = 10.05, 95% CI = 1.09-92.56) and increased HPV DNA (VLR = 16.80, 95% CI = 1.46-193.01). In newly detected HPV-16 infections, increasing levels of E7 mRNA appear to be associated with an increased risk of developing cervical pre-cancer.

The effect of drug concentration expression on epinephrine dosing errors: a randomized trial.

BACKGROUND: The expression of drug concentration as a ratio may cause dosing errors. OBJECTIVE: To examine the effect of ratio expressions on drug administration. DESIGN: Randomized, blinded, controlled study. SETTING: Simulation center in an urban hospital. PARTICIPANTS: 28 physicians. INTERVENTION: Participants managed a simulated pediatric acute anaphylaxis scenario by using epinephrine ampules labeled with mass concentration (1 mg in 1 mL) or a ratio (1 mL of a 1:1000 solution). MEASUREMENTS: The amount of epinephrine given and the time taken to administer it. RESULTS: Compared with providers using ampules with mass concentration labels, those using ratio labels gave more epinephrine (adjusted mean dose, 213 microg above target [95% CI, 76.4 to 350.1 microg]; P = 0.003), and took longer to do so (adjusted mean delay, 91 seconds, [CI, 61.0 to 122.1 seconds]; P < or = 0.0001). LIMITATIONS: Performance in simulated scenarios may not reflect clinical practice. In reality, ampule labels provide both expressions of concentration. CONCLUSION: The use of ratios to express drug concentration may be a source of drug administration error. Patient safety might be improved by expressing drug concentrations exclusively as mass concentration.

Cervical and vulvar cancer risk in relation to the joint effects of cigarette smoking and genetic variation in interleukin 2.

Cigarette smoking is an established cofactor to human papillomavirus (HPV) in the development of cervical and vulvar squamous cell carcinoma (SCC), and may influence risk through an immunosuppressive pathway. Genetic variation in interleukin 2 (IL2), associated in some studies with the inhibition of HPV-targeted immunity, may modify the effect of smoking on the risk of HPV-related anogenital cancers. We conducted a population-based case-only study to measure the departure from a multiplicative joint effect of cigarette smoking and IL2 variation on cervical and vulvar SCC. Genotyping of the four IL2 tagSNPs (rs2069762, rs2069763, rs2069777, and rs2069778) was done in 399 cervical and 486 vulvar SCC cases who had been interviewed regarding their smoking history. Compared with cases carrying the rs2069762 TT genotype, we observed significant departures from multiplicativity for smoking and carriership of the TG or GG genotypes in vulvar SCC risk [interaction odds ratio (IOR), 1.67; 95% confidence interval (CI), 1.16-2.41]. Carriership of one of three diplotypes, together with cigarette smoking, was associated with either a supramultiplicative (TGCT/GGCC; IOR, 2.09; 95% CI, 0.98-4.46) or submultiplicative (TTCC/TGTC; IOR, 0.37; 95% CI, 0.16-0.85 or TGCT/TGCC; IOR, 0.37; 95% CI, 0.15-0.87) joint effect in vulvar cancer risk. For cervical SCC, departure from multiplicativity was observed for smokers homozygous for the rs2069763 variant allele (TT versus GG or GT genotypes; IOR, 1.87; 95% CI, 1.00-3.48), and for carriership of the TTCC/TTCC diplotype (IOR, 2.08; 95% CI, 1.01-4.30). These results suggest that cervical and vulvar SCC risk among cigarette smokers is modified by genetic variation in IL2.