Mitoquinone Mesylate and Mitochondrial DNA in End Organs in Humanized Mouse Model of Chronic Treated Human Immunodeficiency Virus Infection.

No treatment exists for mitochondrial dysfunction, a contributor to end-organ disease in human immunodeficiency virus (HIV). The mitochondrial antioxidant mitoquinone mesylate (MitoQ) attenuates mitochondrial dysfunction in preclinical mouse models of various diseases but has not been used in HIV. We used a humanized murine model of chronic HIV infection and polymerase chain reaction to show that HIV-1-infected mice treated with antiretroviral therapy and MitoQ for 90 days had higher ratios of human and murine mitochondrial to nuclear DNA in end organs compared with HIV-1-infected mice on antiretroviral therapy. We offer translational evidence of MitoQ as treatment for mitochondrial dysfunction in HIV.

Healthcare disparity and its associations with cytomegalovirus disease in pediatric liver transplant recipients in South Africa.

BACKGROUND: Cytomegalovirus (CMV) infection and disease are preventable complications following pediatric liver transplantation (PLT), despite the use of prophylaxis to minimize the risk of CMV disease. We evaluated the incidence and complications of CMV disease in PLT recipients in South Africa (SA), with particular reference to potential differences in outcome between state and private sector patients. METHODS: Medical records of patients younger than 16 years of age who received liver transplants between January 1, 2012, and August 31, 2018 were analyzed. RESULTS: Records of all 150 PLT patients were retrieved. The median age at transplant was 29.2 months (95% confidence interval 15.6-58.4) and follow-up was 46.3 months (interquartile range 27.6-63.1). Sixty-six (44%) patients were high risk, 79 (52.7%) were intermediate risk, and five (3.3%) were low risk for CMV infection. Forty-three (28.9%) patients had CMV DNAemia following transplantation, and 30 (20.1%) developed CMV disease. Receipt of care in the private sector was consistently associated with a lower hazard of CMV disease (adjusted hazard ratio [aHR] ranging from 0.36 to 0.43) and a consistently lower hazard of death among recipients at high risk for CMV disease and/or those who developed CMV disease (aHR ranging from 0.28 to 0.33). CONCLUSION: Receipt of care in the private health sector was associated with a consistently lower hazard of CMV disease and death in individuals with CMV disease and/or at high risk for CMV disease. Policies aimed at creating a more equitable healthcare system in SA may mitigate the differential burden of illness associated with CMV in PLT recipients.

Cytomegalovirus DNAemia and disease: current-era epidemiology, clinical characteristics and outcomes in cancer patients other than allogeneic haemopoietic transplantation.

BACKGROUND: High-intensity chemotherapy and advances in novel immunotherapies have seen the emergence of cytomegalovirus (CMV) infections in cancer patients other than allogeneic haemopoietic cell transplantation (HCT). Aim To evaluate the epidemiology, clinical characteristics and outcomes of CMV infection in this population. METHODS: A retrospective review of cancer patients other than allogeneic HCT who had CMV DNAemia and/or disease from July 2013 till May 2020 at a quaternary cancer centre was performed. RESULTS: Of 11 485 cancer patients who underwent treatment during this period, 953 patients had CMV DNA testing performed and 238 of them had CMV DNAemia. After excluding patients with allogeneic HCT, 62 patients with CMV DNAemia were identified, of whom 10 had concurrent CMV disease. The most frequent underlying malignancies were B-cell lymphoproliferative disease (LPD) (31%; 19/62), T-cell LPD (21%; 13/62), chronic lymphocytic leukaemia (11%; 7/62) and multiple myeloma (10%; 6/62). Most patients had lymphopenia (77%; 48/62), multiple cancer therapies (63%; 39/62 received ≥2 previous therapies), co-infection (56%; 35/62 had ≥1 co-infection) and corticosteroid therapy (48%; 30/62) within 1 month before CMV diagnosis. CMV DNAemia and disease were observed in patients receiving novel immunotherapies, including bispecific antibody therapy, chimeric-antigen receptor T-cell therapy and immune checkpoint inhibitors. CONCLUSION: Patients with haematological malignancy, particularly B-cell LPD, T-cell LPD, chronic lymphocytic leukaemia and multiple myeloma, were frequently identified to have CMV DNAemia and disease. Lymphopenia, multiple cancer therapies, co-infection and recent receipt of systemic corticosteroids were also commonly observed. Future studies are necessary to determine optimal identification and management of CMV in these patients.

Cytomegalovirus Viral Load Kinetics Predict Cytomegalovirus End-Organ Disease and Mortality After Hematopoietic Cell Transplant.

BACKGROUND: We investigatedthe association between time-averaged area under the curve (AAUC) of cytomegalovirus (CMV) viral load (VL) by day 100 and overall survival (OS) at 1-year after hematopoietic cell transplantation (HCT). METHODS: In a retrospective cohort study, including patients receiving HCT between June 2010 and December 2017 from Memorial Sloan Kettering Cancer Center, AAUC was calculated for patients with detected VL. Patients were categorized into non-controllers (Q4) and controllers (Q1-Q3) using the highest AAUC quartile as cutoff. Cox models were used to estimate the association between AAUC and OS. Patients with non-detected CMV VL were categorized into elite-controllers (recipient+ [R+] or R-/donor+ [D+]) and R-/D-. RESULTS: The study (N = 952) included 282 controllers, 93 non-controllers, 275 elite-controllers, and 302 R-/D-. OS was 80.1% and 58.1% for controllers and non-controllers, respectively. In multivariable models, non-controllers had worse OS versus controllers (adjusted hazard ratio [HR] = 2.65; 95% confidence interval [CI], 1.71-4.12). In landmark analyses, controllers had similar OS as elite-controllers (HR = 1.26; 95% CI, .83-1.91) or R-/D- (HR = 0.98; 95% CI, .64-1.5). CONCLUSIONS: Non-controllers had worse OS 1-year post-HCT. Controllers had similar OS as elite-controllers or R-/D-. Future studies are needed to validate our AAUC cutoff across different cohorts and CMV management strategies.

Interleukin-18 and tumor necrosis factor-α are elevated in solid organ transplant recipients with possible cytomegalovirus end-organ disease.

End-organ cytomegalovirus (CMV) disease can be life threatening to solid organ transplant recipients. Diagnosis is often complicated by variation in amount of CMV DNA in plasma and the need for an invasive procedure to obtain a biopsy of the suspected affected organ, which can delay recognition and treatment. Several inflammatory cytokines are elevated in CMV disease, and the purpose of this study was to determine if they could be used to distinguish solid organ transplant recipients with CMV DNAemia alone from those with possible end-organ CMV disease. We found that regardless of pre-transplant CMV serostatus, plasma interleukin (IL)-18, tumor necrosis factor-α (TNF-α), and amount of CMV DNA in plasma were increased in possible end-organ CMV disease, with elevated IL-18 associated with increased odds of possible end-organ CMV disease even after adjusting for amount of CMV DNA. These findings highlight IL-18 and TNF-α as potential non-invasive markers of possible end-organ CMV disease regardless of transplanted organ or serostatus in solid organ transplant recipients.

High human herpesvirus 6 viral load in pediatric allogeneic hematopoietic stem cell transplant patients is associated with detection in end organs and high mortality.

Human Herpes Virus 6 (HHV-6) reactivation occurs in approximately half of patients following allogeneic hematopoietic stem cell transplant (HSCT). While encephalitis and delayed engraftment are well-documented complications of HHV-6 following HSCT, the extent to which HHV-6 viremia causes disease in children is controversial. We performed a retrospective review of HHV-6 reactivation and possible manifestations in pediatric allogeneic HSCT patients at a single institution. Of 89 children and young adults who underwent allogeneic HSCT over a three-and-a-half-year period, 34 patients reactivated HHV-6 early post-transplant. Unrelated donor stem cell source and lack of antiviral prophylaxis were risk factors for the development of HHV-6 viremia. Viremia correlated with the presence of acute graft-versus-host disease, but not chronic graft-versus-host disease. We identified two subgroups within the viremic patients-a high-risk viremic and tissue-positive group that reactivated HHV-6 and had suspected end-organ disease and a low-risk viremic but asymptomatic group that reactivated HHV-6 but did not exhibit symptoms or signs of end-organ disease. Peak viral load was found to be strongly associated with mortality. Prospective studies in larger numbers of patients are needed to further investigate the role of HHV-6 in causing symptomatic end-organ disease as well as the association of viral load with mortality.

Cytomegalovirus and HIV: A Dangerous Pas de Deux.

Human immunodeficiency virus (HIV)-infected adults who take stable antiretroviral therapy (ART) are at risk for early onset of age-related diseases. This is likely due to a complex interaction between traditional risk factors, HIV infection itself, and other factors, such as underlying immune dysfunction and persistent inflammation. HIV disrupts the balance between the host and coinfecting microbes, worsening control of these potential pathogens. For example, HIV-infected adults are more likely than the general population to have subclinical bursts of cytomegalovirus (CMV) replication at mucosal sites. Production of antigens can activate the immune system and stimulate HIV replication, and it could contribute to the pathogenesis of adverse outcomes of aging, like cardiovascular disease and neurocognitive impairment. Further investigation of the relationships between CMV, immune dysfunction, and unsuccessful aging during chronic HIV infection is warranted.

An investigation on the relationship between the occurrence of CMV DNAemia and the development of invasive aspergillosis in the allogeneic stem cell transplantation setting.

Cytomegalovirus (CMV) infection might increase the risk of fungal superinfection in allogeneic stem cell transplant patients. The potential association between the occurrence of CMV DNAemia and the development of invasive aspergillosis in this clinical setting was investigated. The current retrospective observational study included 167 patients undergoing T cell-replete allogeneic stem cell transplantation. Virological monitoring of active CMV infection was performed by the pp65 antigenemia assay and/or by a plasma real-time PCR assay. A total of 109 out of 167 patients developed CMV DNAemia. Twenty-three patients had proven (n = 4) or probable (n = 19) invasive aspergillosis. The occurrence of CMV DNAemia was not significantly associated with the subsequent development of invasive aspergillosis (P = 0.38). Overall, the duration of the episodes of active CMV infection and the peak level of CMV DNAemia within the episodes were comparable, irrespective of whether invasive aspergillosis developed subsequently or not (P = 0.99; P = 0.70, respectively). Peak CMV DNA load in patients with proven or probable invasive aspergillosis who died was higher (median, 5,461 copies/ml) than that in those who survived (median 1,179 copies/ml) (P = 0.41). The data argue against the existence of an association between the occurrence of CMV DNAemia and the development of invasive aspergillosis, however, CMV replication, particularly at high levels, might aggravate the prognosis of this disease.

Cytomegalovirus Esophagitis in an Immunocompromised Patient.

Cytomegalovirus (CMV) can present with end-organ disease (EOD), particularly in patients with a CD4 cell count <50/mm3. While EOD in immunocompromised patients commonly presents as CMV retinitis (30%) and CMV colitis (5-10%), CMV esophagitis is rare. CMV is the third most common infectious esophagitis following Candida and Herpes Simplex. CMV esophagitis presents with odynophagia, dysphagia, and abdominal pain. Endoscopic exam may reveal large, linear distal esophageal ulcers. Histopathology or serology studies are diagnostic, though serology may be unreliable in the extremely immunosuppressed. Current treatment consists of antivirals such as ganciclovir and valganciclovir. Esophageal disease due to CMV carries a poor prognosis in the immunocompromised. We present the case of a 56-year-old male with a medical history of HIV/AIDS and stage III rectal squamous cell cancer who presented with shortness of breath, weakness, and chronic diarrhea. His HIV was previously well-controlled on antiretroviral therapy. However, due to his malignancy, he was undergoing treatment with chemotherapy and radiation. Initial labs revealed a CD4 count of 42. His clinical course consisted of Escherichia coli septicemia, new-onset atrial fibrillation with a rapid ventricular response, worsening pneumonia, possible metastasis, progressive diarrhea, and potential oropharyngeal candidiasis. Despite several broad-spectrum antimicrobial regimens, he remained symptomatic with new complaints of dysphagia and odynophagia. Eventually, the appearance of vesicular lesions on the lips and a repeat CD4 count of 13 garnered a suspicion of HSV or CMV. This complicated case highlights the necessity for a high index of suspicion of rare manifestations of CMV EOD in an immunocompromised patient presenting with confounding clinical symptoms and extensive diagnoses.

Multisystem cytomegalovirus end-organ disease in a patient with advanced HIV.

Cytomegalovirus (CMV) infection is common in people living with HIV, but multisystem CMV end-organ disease (EOD) is rare following the introduction of effective antiretroviral therapy. We present the case of a patient with advanced HIV and multisystem manifestations of CMV EOD. Contributions: This case report highlights the potential morbidity and mortality associated with CMV disease in patients with advanced HIV. Clinicians should be vigilant in considering CMV EOD in patients with advanced HIV and visual, neurological and gastointestinal symptoms.

Epidemiological changes in cytomegalovirus end-organ diseases in a developed country: A nationwide, general-population-based study.

BACKGROUND: Cytomegalovirus (CMV) can cause tissue-invasive diseases in various organs after primary infection or through reactivation of latent-to-lytic switch over a lifetime. The number of individuals who are at risk of CMV diseases, such as elderly or immunocompromised patients, is constantly increasing; however, recent epidemiological changes associated with CMV disease have not been fully evaluated. METHODS: We used claims data of about 50 million individuals between 2010 and 2015 from the Korean Health Insurance Review and Assessment Service nationwide database. The code for CMV end-organ diseases in the 'Relieved Co-payment Policy' program matches the ICD-10 code of B25, except for congenital CMV infection and mononucleosis. A 628 cases of CMV and 3140 controls (without CMV disease), matched for age and sex, were selected from this dataset in order to evaluate the effect of adult CMV diseases on all-cause death. RESULTS: The overall unadjusted incidence rate (IR) of CMV end-organ diseases was 0.52/100,000 individuals. The standardized IR, adjusted for age and sex, have continuously increased from 0.32/100,000 in 2010 to 0.75/100,000 in 2015. The overall unadjusted IR in adult population was highest in 70-79 years for six years (0.96/100,000). In the model adjusted for age, sex, immunocompromised status including solid-organ or hematopoietic stem cell transplant recipients, hematologic malignancies, and human immunodeficiency virus diseases, the hazard ratio of case group was 5.2 (95% confidence interval, 3.6-7.4) for all-cause mortality. CONCLUSION: Nationwide data indicates that CMV end-organ disease has steadily increased in the past six years and is associated with higher mortality.

Association of cytomegalovirus diseases with newly developed myocardial infarction and congestive heart failure: data from a national population-based cohort.

Introduction: Anti-cytomegalovirus (CMV) IgG seropositive and/or titer are associated with a higher risk of cardiovascular diseases (CVD). However, it is not clear whether CMV end-organ disease may have a relation with development of CVD or chronic heart diseases. Material and methods: In matched cohort study, the National Health Insurance Database covering 50 million people was used to identify 667 patients with CMV diseases and aged ≥ 20 years between 2010 and 2014. 6,670 control subjects without CMV diseases were matched by age, sex, type 2 diabetes mellitus (DM), hypertension, dyslipidemia, and cohort entry year. Data on CMV disease and heart disease events of myocardial infarction (MI), congestive heart failure (CHF), and atrial fibrillation (AF) were retrieved. Previous events before CMV disease or cohort entry were excluded until January 2006. Subjects were followed until December 2015 in subjects without events and until date of events in subjects with events. Results: The multivariate regression model adjusted by age, sex, low-income status, type 2 DM, hypertension, dyslipidemia, solid organ transplantation, and hematopoietic stem cell transplantation showed a significantly higher incidence rate of MI (odds ratio (OR) = 2.1, 95% confidence intervals (CI): 1.0-4.5) and CHF (OR = 3.8, 95% CI: 2.1-6.8) but not AF (OR = 1.9, 95% CI: 0.9-4.0) in patients with CMV disease. The age group of 40-64 years with CMV disease had the highest risk for new-onset CHF in this regression model (OR = 9.4, 95% CI: 4.12-21.44, p = 0.029). Conclusions: Symptomatic CMV tissue-invasive diseases were associated with a higher risk of new-onset MI and CHF.

Outcomes of Refractory Cytomegalovirus Infection in the First Year after Allogeneic Hematopoietic Cell Transplantation.

Outcomes of refractory (Rf) cytomegalovirus (CMV) infection (CMVi) after hematopoietic cell transplantation (HCT) are poor owing to limited treatment options and treatment related toxicities. Maribavir, an orally bioavailable CMV antiviral, was recently approved for treatment of Rf-CMVi. Real-world studies quantifying the burden of Rf-CMVi prior to maribavir provide a benchmark for evaluating the net value of novel treatments. Here we report the incidence, clinical outcomes, and healthcare resource utilization (HRU) associated with Rf-CMVi in the first year post-HCT in a cohort of CMV-seropositive HCT recipients (R+) who underwent HCT between January 1, 2014, and December 31, 2017, at Memorial Sloan Kettering Cancer Center and were managed exclusively by preemptive therapy. CMVi was defined as CMV viremia treated preemptively. Rf-CMVi was defined as a <1 log10 decrease and CMV viral load >1000 U/mL after ≥14 days of appropriately dosed therapy. Welldays were defined as alive days not hospitalized and off CMV antivirals by 1 year post-HCT. The impact of Rf-CMVi on mortality and HRU was examined in multivariable models. Of the 286 R+ patients, 145 (50.7%) developed CMVi (99 no Rf-CMVi and 46 Rf-CMVi). Compared with the no Rf-CMVi group, the Rf-CMVi group had higher rates of CMV EOD (23.9% versus 10.1%; P = .030), CMV-related mortality (9.5% versus .0%; P = .002), and all-cause mortality (33.3% versus 15.6%; adjusted P = .049). Rf-CMVi was an independent predictor for readmission (adjusted odds ratio [aOR], 3.24; 95% confidence interval [CI], 2.19 to 4.87; P < .0001); CMV-related readmission (aOR, 9.48; 95% CI, 5.83 to 15.80; P < .0001), and decreased well days (adjusted arithmetic mean ratio, .72; 95% CI, .58 to .89; P = .001) in the first year post-HCT. Rf-CMVi is associated with increased mortality and increased HRU at 1 year after HCT. Improved therapies for Rf-CMVi have the potential of improving HCT outcomes and reducing HRU.

Defining the Innate Immune Responses for SARS-CoV-2-Human Macrophage Interactions.

Host innate immune response follows severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, and it is the driver of the acute respiratory distress syndrome (ARDS) amongst other inflammatory end-organ morbidities. Such life-threatening coronavirus disease 2019 (COVID-19) is heralded by virus-induced activation of mononuclear phagocytes (MPs; monocytes, macrophages, and dendritic cells). MPs play substantial roles in aberrant immune secretory activities affecting profound systemic inflammation and end-organ malfunctions. All follow the presence of persistent viral components and virions without evidence of viral replication. To elucidate SARS-CoV-2-MP interactions we investigated transcriptomic and proteomic profiles of human monocyte-derived macrophages. While expression of the SARS-CoV-2 receptor, the angiotensin-converting enzyme 2, paralleled monocyte-macrophage differentiation, it failed to affect productive viral infection. In contrast, simple macrophage viral exposure led to robust pro-inflammatory cytokine and chemokine expression but attenuated type I interferon (IFN) activity. Both paralleled dysregulation of innate immune signaling pathways, specifically those linked to IFN. We conclude that the SARS-CoV-2-infected host mounts a robust innate immune response characterized by a pro-inflammatory storm heralding end-organ tissue damage.

The direct and indirect consequences of cytomegalovirus infection and potential benefits of vaccination.

Active infection with cytomegalovirus (CMV) occurs in patients who are immunocompromised and may produce the high viral loads required to cause end-organ disease. Such patients have complex medical histories and many experienced physicians have speculated that CMV may, additionally, contribute to adverse clinical outcomes. In 1989, Dr Bob Rubin coined the term "indirect effects" to describe this potential relationship between virus and patient. Examples include accelerated atherosclerosis in patients after heart transplant or with underlying HIV infection, the number of days patients require ventilation after admission to intensive care units, the development of immunosenescence in the elderly and mortality in many groups of patients, including the general population. It is difficult to distinguish between CMV acting as causal contributor to such diverse pathology or simply having a benign bystander effect. However, recruitment of patients into placebo-controlled randomised trials of antiviral drugs with activity against CMV offers such a potential. This article describes the studies that have been conducted to date and emphasises that mortality after stem cell transplant (not attributed to CMV end-organ disease) has recently become the first proven indirect effect of CMV now that letermovir has significantly reduced non-relapse deaths. The implications for CMV vaccines are then discussed. Vaccines are already predicted to be highly cost-effective if they can reduce CMV end-organ disease. Health planners should now consider that cost effectiveness is likely to be enhanced further through reduction of the indirect effects of CMV. A prototype scheme for assessing this possibility is provided in order to stimulate discussion within the field. This article forms part of an online symposium on the prevention and therapy of DNA virus infections, dedicated to the memory of Mark Prichard.

Systemic Inflammation, Coagulation, and Clinical Risk in the START Trial.

BACKGROUND: The Strategic Timing of AntiRetroviral Treatment (START) trial demonstrated that immediate (at CD4+ >500 cells/µL) vs deferred (to CD4+ <350 cells/µL or AIDS) antiretroviral therapy (ART) initiation reduced risk for AIDS and serious non-AIDS (SNA). We investigated associations of inflammation, coagulation, and vascular injury biomarkers with AIDS, SNA or death, and the effect of immediate ART initiation. METHODS: Biomarkers were measured from stored plasma prior to randomization and at month 8. Associations of baseline biomarkers with event risk were estimated with Cox regression, pooled across groups, adjusted for age, gender, and treatment group, and stratified by region. Mean changes over 8 months were estimated and compared between the immediate and deferred ART arms using analysis of covariance models, adjusted for levels at entry. RESULTS: Baseline biomarker levels were available for 4299 START participants (92%). Mean follow-up was 3.2 years. Higher levels of IL-6 and D-dimer were the only biomarkers associated with risk for AIDS, SNA or death, as well as the individual components of SNA and AIDS events (HRs ranged 1.37-1.41 per 2-fold higher level), even after adjustment for baseline CD4+ count, HIV RNA level, and other biomarkers. At month 8, biomarker levels were lower in the immediate arm by 12%-21%. CONCLUSIONS: These data, combined with evidence from prior biomarker studies, demonstrate that IL-6 and D-dimer consistently predict clinical risk across a broad spectrum of CD4 counts for those both ART-naïve and treated. Research is needed to identify disease-modifying treatments that target inflammation beyond the effects of ART.

Cytomegalovirus viraemia in HIV exposed and infected infants: prevalence and clinical utility for diagnosing CMV pneumonia.

BACKGROUND: Human cytomegalovirus (HCMV) is an important pathogen in HIV exposed infants with pneumonia. However, the diagnosis of HCMV pneumonia in this setting is challenging due to limited access to bronchoscopy, lung biopsy and direct sampling of the lower respiratory tract. HCMV viraemia is more accessible, but their diagnostic performance in this context has not been studied. OBJECTIVE: To describe the prevalence of HCMV viraemia and evaluate its clinical utility in HIV exposed infants. STUDY DESIGN: In this cross-sectional study, we performed qualitative and quantitative PCR to detect HCMV viraemia in HIV exposed asymptomatic infants and in infants with severe pneumonia in the Western Cape province of South Africa. RESULTS: 283 asymptomatic HIV exposed infants and 142 HIV exposed infants with severe pneumonia were studied. Infants with pneumonia had a higher prevalence of HCMV viraemia compared to asymptomatic infants (68% vs 24% OR 6.7, 95% CI 4.2-10.8). This increased prevalence remained significant (OR 4.3 95% CI 2.6-7.0) after adjusting for HIV infection. Of the infants with pneumonia, the level of HCMV viraemia was significantly higher in a subset of infants diagnosed with HCMV pneumonia (median HCMV viral load 4.6 vs 2.5 log copies/ml p<0.001). Receiver operator characteristic (ROC) analysis showed the area under the curve was 0.78 (95% CI 0.71-0.86) and a threshold of 4.1 log copies/ml was able to correctly identify 70% of HCMV pneumonia cases. CONCLUSION: Prevalence and level of HCMV viraemia in sub-Saharan HIV-exposed and infected infants peaks at 3-4 months of age. Quantitative HCMV PCR may be useful in diagnosing HCMV pneumonia.

Clinical Development of a Cytomegalovirus DNA Vaccine: From Product Concept to Pivotal Phase 3 Trial.

2013 marks a milestone year for plasmid DNA vaccine development as a first-in-class cytomegalovirus (CMV) DNA vaccine enters pivotal phase 3 testing. This vaccine consists of two plasmids expressing CMV antigens glycoprotein B (gB) and phosphoprotein 65 (pp65) formulated with a CRL1005 poloxamer and benzalkonium chloride (BAK) delivery system designed to enhance plasmid expression. The vaccine's planned initial indication under investigation is for prevention of CMV reactivation in CMV-seropositive (CMV⁺) recipients of an allogeneic hematopoietic stem cell transplant (HCT). A randomized, double-blind placebo-controlled phase 2 proof-of-concept study provided initial evidence of the safety of this product in CMV⁺ HCT recipients who underwent immune ablation conditioning regimens. This study revealed a significant reduction in viral load endpoints and increased frequencies of pp65-specific interferon-γ-producing T cells in vaccine recipients compared to placebo recipients. The results of this endpoint-defining trial provided the basis for defining the primary and secondary endpoints of a global phase 3 trial in HCT recipients. A case study is presented here describing the development history of this vaccine from product concept to initiation of the phase 3 trial.