Improvement in neoantigen prediction via integration of RNA sequencing data for variant calling

IntroductionNeoantigen-based immunotherapy has emerged as a promising strategy for improving the life expectancy of cancer patients. This therapeutic approach heavily relies on accurate identification of cancer mutations using DNA sequencing (DNAseq) data. However, current workflows tend to provide a large number of neoantigen candidates, of which only a limited number elicit efficient and immunogenic T-cell responses suitable for downstream clinical evaluation. To overcome this limitation and increase the number of high-quality immunogenic neoantigens, we propose integrating RNA sequencing (RNAseq) data into the mutation identification step in the neoantigen prediction workflow.MethodsIn this study, we characterize the mutation profiles identified from DNAseq and/or RNAseq data in tumor tissues of 25 patients with colorectal cancer (CRC). Immunogenicity was then validated by ELISpot assay using long synthesis peptides (sLP).ResultsWe detected only 22.4% of variants shared between the two methods. In contrast, RNAseq-derived variants displayed unique features of affinity and immunogenicity. We further established that neoantigen candidates identified by RNAseq data significantly increased the number of highly immunogenic neoantigens (confirmed by ELISpot) that would otherwise be overlooked if relying solely on DNAseq data.DiscussionThis integrative approach holds great potential for improving the selection of neoantigens for personalized cancer immunotherapy, ultimately leading to enhanced treatment outcomes and improved survival rates for cancer patients

Oseltamivir Is Adequately Absorbed Following Nasogastric Administration to Adult Patients with Severe H5N1 Influenza

In the absence of a parenteral drug, oral oseltamivir is currently recommended by the WHO for treating H5N1 influenza. Whether oseltamivir absorption is adequate in severe influenza is unknown. We measured the steady state, plasma concentrations of nasogastrically administered oseltamivir 150 mg bid and its active metabolite, oseltamivir carboxylate (OC), in three, mechanically ventilated patients with severe H5N1 (male, 30 yrs; pregnant female, 22 yrs) and severe H3N2 (female, 76 yrs). Treatments were started 6, 7 and 8 days after illness onset, respectively. Both females were sampled while on continuous venovenous haemofiltration. Admission and follow up specimens (trachea, nose, throat, rectum, blood) were tested for RNA viral load by reverse transcriptase PCR. In vitro virus susceptibility to OC was measured by a neuraminidase inhibition assay. Admission creatinine clearances were 66 (male, H5N1), 82 (female, H5N1) and 6 (H3N2) ml/min. Corresponding AUC0–12 values (5932, 10,951 and 34,670 ng.h/ml) and trough OC concentrations (376, 575 and 2730 ng/ml) were higher than previously reported in healthy volunteers; the latter exceeded 545 to 3956 fold the H5N1 IC50 (0.69 ng/ml) isolated from the H5N1 infected female. Two patients with follow-up respiratory specimens cleared their viruses after 5 (H5N1 male) and 5 (H3N2 female) days of oseltamivir. Both female patients died of respiratory failure; the male survived. 150 mg bid of oseltamivir was well absorbed and converted extensively to OC. Virus was cleared in two patients but two patients died, suggesting viral efficacy but poor clinical efficacy

The Outcomes of Three Surgical Approaches for Acromioclavicular Dislocation Treatment: Findings from Vietnam

Background: Acromioclavicular (AC) dislocation, one of the most common shoulder joint injuries, can be treated by several surgical methods. However, there are still few records about the treatment quality. This study aims to describe the outcomes of three surgical methods for acromioclavicular dislocation treatment at Viet Duc University Hospital, Vietnam. Methods: A cross-sectional study was conducted on 80 patients diagnosed with AC. We retrospectively collected data in the medical records and re-examined the patients. Results: There was no difference between the three groups of surgical approaches relating to the patient’s characteristics, except for the time from the accident to hospital admission. The median length of stay after surgery was highest in the Hook plate group (median (IQR) = 5(2) days), while it was lowest in the K-wire group (median (IQR) = 3(1) days) (p < 0.05). There is statistical significance in the difference of coracoclavicular distance between pre and post-operation in all three surgical method groups (p < 0.001). Conclusion: All of the methods—Hook plate, K-wire, and TightRope—were associated with optimistic outcomes and restored initial anatomy. While the three surgical methods are both safe and effective, the K-wire method is associated with a shorter length of stay and might be economical

Viral load testing to monitor the HIV epidemic among PWID in Vietnam

Objective: To share Vietnam’s experiences piloting the integration of viral load (VL) testing into the national HIV sentinel surveillance (HSS) system to better understand the level of HIV viral transmission among people who inject drugs (PWID).Introduction: Vietnam initiated the HSS system in 1994 in selected provinces with high HIV burden. The surveillance has two components: monitor HIV sero-prevalence and risk behaviors among key population including PWID. However, no VL data were collected among HIV infected people. In 2016, Vietnam piloted an added component of VL testing to the existing HSS system. The purpose was to test the feasibility of adding VL testing to the HSS so that VL data among PWID would be available. The pilot was conducted in two provinces in southern Vietnam-Ho Chi Minh City and Long An. It was expected that adding the VL testing to the existing HSS would also save resources and help monitor HIV viral transmission among PWID in the community regardless if they are currently on anti-retroviral therapy (ART).Methods: Male PWIDs were enrolled into 2016 HSS+ following the standard operating procedure (SOP)[1]. Community-based sampling was based on random selection of wards/communes listed in the sampling frame. In each selected ward/commune, all eligible PWID were invited to voluntarily participate in the survey. Eligibility criteria were males 16 years of age or older, reporting injecting drug in the past month, and residing in the selected area. . The survey included an interview using a standardized questionnaire and 7ml blood drawn for HIV testing. Blood specimens were transferred from districts to provincial labs for plasma separation in the same day. Each plasma specimen was divided into three aliquots of 1ml each. One aliquot was used to test for HIV diagnosis at provincial labs, using the national HIV testing strategy III[2]. The remaining 2 aliquots were stored at provincial labs at 2-80C and within 5 days, were shipped to Pasteur Institute in Ho Chi Minh City (PIHCM) where the plasma specimens were stored at -800C. Processing of samples for VL testing was conducted at the end of the survey where all plasma specimen were transferred to PIHCM lab, which was 2 months since the collection of the first blood specimen. VL was undertaken on COBAS AMPLYPREP/COBAS TAQMAN 48, with identification threshold 20 cps/ml and specificity of 100% using Kit CAP-G/CTM HIV-1 V 2.0. The VL testing results were sent back to relevant Provicial AIDS Centers to return to respective participants, within 3 months.Results: Five hundred male PWID (HCMC: 300; LA: 200) were enrolled into 2016 HSS/HSS+ and agreed to provide blood specimen without any refusal. 84 tested positive for HIV (16.8%. HCMC: 15.0%; LA: 19.5%), 43 (51.2%) specimens had unsuppressed VL (>1000 copies/ml) (HCMC: 66.7%; LA: 33.3%), 35 (41.7%) specimens had undetected level (<50 copies/ml or undetected) (HCMC: 31.1%; LA: 53.9%), and 7.1% had VL that ranged from 50-1000 copies/ml (HCMC: 2.2%; LA: 12.8%). Among those who had VL < 1000 copies/ml, 22 (53.7%) had ever been on ART.Conclusions: The pilot survey has measured VL among male PWID, including those who were aware of their HIV status and those who did not know their status before. Findings indicate that a significant proportion of PWID do not have their VL suppressed leading to high-risk of HIV transmission from PWID to their sexual partners[3] in the community although level of unsuppressed viral load is not a direct measure of HIV viral transmission in itself. This pilot indicated that it was feasible to add VL testing into HSS and Vietnam government can add it as a routine practice in HSS and can be expanded in the coming years

Viral load suppression and acquired HIV drug resistance in adults receiving antiretroviral therapy in Viet Nam: results from a nationally representative survey

Objective: The purpose of this survey was to estimate the prevalence of viral load (VL) suppression and emergence of HIV drug resistance (HIVDR) among individuals receiving antiretroviral therapy (ART) for 36 months or longer in Viet Nam using a nationally representative sampling method. Methods: The survey was conducted between May and August 2014 using a two-stage cluster design. Sixteen ART clinics were selected using probability proportional to proxy size sampling, and patients receiving ART for at least 36 months were consecutively enrolled. Epidemiological information and blood specimens were collected for HIV-1 VL and HIVDR testing; HIVDR was defined by the Stanford University HIVDR algorithm. Results: Overall, 365 eligible individuals were recruited with a mean age of 38.2 years; 68.4% were men. The mean time on ART was 75.5 months (95% confidence interval [CI]: 69.0–81.9 months), and 93.7% of the patients were receiving non-nucleoside reverse transcriptase inhibitor-based regimens. Of the 365 individuals, 345 (94.7%, 95% CI: 64.1–99.4%) had VL below 1000 copies/mL and 19 (4.6%, 95% CI: 2.8-–7.5) had HIVDR mutations. Discussion: Our nationally representative survey found a high level of VL suppression and a low prevalence of HIVDR among individuals who received ART for at least 36 months in Viet Nam. Continued surveillance for HIVDR is important for evaluating and improving HIV programs

Evaluation of a liquid biopsy protocol using ultra-deep massive parallel sequencing for detecting and quantifying circulation tumor DNA in colorectal cancer patients

The identification and quantification of actionable mutations are critical for guiding targeted therapy and monitoring drug response in colorectal cancer. Liquid biopsy (LB) based on plasma cell-free DNA analysis has emerged as a non-invasive approach with many clinical advantages over conventional tissue sampling. Here, we developed a LB protocol using ultra-deep massive parallel sequencing and validated its clinical performance for detection and quantification of actionable mutations in three major driver genes ( and ). The assay showed a 92% concordance for mutation detection between plasma and paired tissues and great reliability in quantification of variant allele frequency

Viral clearance in patients A.

<p>Day 0 is the day of illness onset.</p

Patients' clinical, pharmacokinetic and virological features.

*<p>Cockroft Gault formula.</p>¶<p>Arterial blood gas prior to ventilation, mmHg: PO₂, PCO₂, mmol/L:HCO₃⁻.</p>#<p>OP = oseltamivir phosphate, OC = oseltamivir carboxylate.</p

Viral clearance in patient C.

<p>Day 0 is the day of illness onset.</p