Serial Quantitation of Plasma Microbial Cell-Free DNA Before and After Diagnosis of Pulmonary Invasive Mold Infections After Hematopoietic Cell Transplant.

BACKGROUND: Plasma microbial cell-free DNA sequencing (mcfDNA-Seq) is a noninvasive test for microbial diagnosis of invasive mold infection (IMI). The utility of mcfDNA-Seq for predicting IMI onset and the clinical implications of mcfDNA concentrations are unknown. METHODS: We retrospectively tested plasma from hematopoietic cell transplant (HCT) recipients with pulmonary IMI and ≥1 mold identified by mcfDNA-Seq in plasma collected within 14 days of clinical diagnosis. Samples collected from up to 4 weeks before and 4 weeks after IMI diagnosis were evaluated using mcfDNA-Seq. RESULTS: Thirty-five HCT recipients with 39 IMIs (16 Aspergillus and 23 non-Aspergillus infections) were included. Pathogenic molds were detected in 38%, 26%, 11%, and 0% of samples collected during the first, second, third, and fourth week before clinical diagnosis, respectively. In non-Aspergillus infections, median mcfDNA concentrations in samples collected within 3 days of clinical diagnosis were higher in infections with versus without extrapulmonary spread (4.3 vs 3.3 log10 molecules per microliter [mpm], P = .02), and all patients (8/8) with mcfDNA concentrations >4.0 log10 mpm died within 42 days after clinical diagnosis. CONCLUSIONS: Plasma mcfDNA-Seq can identify pathogenic molds up to 3 weeks before clinical diagnosis of pulmonary IMI. Plasma mcfDNA concentrations may correlate with extrapulmonary spread and mortality in non-Aspergillus IMI.

Anti-HLA antibodies in recipients of CD19 versus BCMA-targeted CAR T-cell therapy.

Antibodies against foreign human leukocyte antigen (HLA) molecules are barriers to successful organ transplantation. B cell-depleting treatments are used to reduce anti-HLA antibodies but have limited efficacy. We hypothesized that the primary source for anti-HLA antibodies is long-lived plasma cells, which are ineffectively targeted by B cell depletion. To study this, we screened for anti-HLA antibodies in a prospectively enrolled cohort of 49 patients who received chimeric antigen receptor T-cell therapy (CARTx), targeting naïve and memory B cells (CD19-targeted, n = 21) or plasma cells (BCMA-targeted, n = 28) for hematologic malignancies. Longitudinal samples were collected before and up to 1 year after CARTx. All individuals were in sustained remission. We identified 4 participants with anti-HLA antibodies before CD19-CARTx. Despite B cell depletion, anti-HLA antibodies and calculated panel reactive antibody scores were stable for 1 year after CD19-CARTx. Only 1 BCMA-CARTx recipient had pre-CARTx low-level anti-HLA antibodies, with no follow-up samples available. These data implicate CD19neg long-lived plasma cells as an important source for anti-HLA antibodies, a model supported by infrequent HLA sensitization in BCMA-CARTx subjects receiving previous plasma cell-targeted therapies. Thus, plasma cell-targeted therapies may be more effective against HLA antibodies, thereby enabling improved access to organ transplantation and rejection management.

Pathogen-Specific Humoral Immunity and Infections in B Cell Maturation Antigen-Directed Chimeric Antigen Receptor T Cell Therapy Recipients with Multiple Myeloma.

Chimeric antigen receptor (CAR) T cell therapy targeting B cell maturation antigen (BCMA-CARTx) is an emerging treatment for relapsed or refractory multiple myeloma (R/R MM). Here we characterize the epidemiology of infections, risk factors for infection, and pathogen-specific humoral immunity in patients receiving BCMA-CARTx for R/R MM. We performed a retrospective cohort study in 32 adults with R/R MM enrolled in 2 single-institution phase 1 clinical trials of BCMA-CARTx administered after lymphodepleting chemotherapy alone (n = 22) or with a gamma secretase inhibitor (GSI). We tested serum before and up to approximately 180 days after BCMA-CARTx for measles-specific IgG and for any viral-specific IgG using a systematic viral epitope scanning assay to describe the kinetics of total and pathogen-specific IgG levels pre- and post-BCMA-CARTx. We identified microbiologically documented infections to determine infection incidence and used Poisson regression to explore risk factors for infections within 180 days after BCMA-CARTx. Most individuals developed severe neutropenia, lymphopenia, and hypogammaglobulinemia after BCMA-CARTx. Grade ≥3 cytokine release syndrome (CRS; Lee criteria) occurred in 16% of the participants; 50% of the participants received corticosteroids and/or tocilizumab. Before BCMA-CARTx, 28 of 32 participants (88%) had an IgG <400 mg/dL, and only 5 of 27 (19%) had seropositive measles antibody titers. After BCMA-CARTx, all participants had an IgG <400 mg/dL and declining measles antibody titers; of the 5 individuals with baseline seropositive levels, 2 remained above the seroprotective threshold post-treatment. Participants with IgG MM (n = 13) had significantly fewer antibodies to a panel of viral antigens compared with participants with non-IgG MM (n = 6), both before and after BCMA-CARTx. In the first 180 days after BCMA-CARTx, 17 participants (53%) developed a total of 23 infections, of which 13 (57%) were mild-to-moderate viral infections. Serious infections were more frequent in the first 28 days post-treatment. Infections appeared to be more common in individuals with higher-grade CRS. Individuals with R/R MM have substantial deficits in humoral immunity. These data demonstrate the importance of plasma cells in maintaining long-lived pathogen-specific antibodies and suggest that BCMA-CARTx recipients need ongoing surveillance for late-onset infections. Most infections were mild-moderate severity viral infections. The incidence of early infection appears to be lower than has been reported after CD19-directed CARTx for B cell neoplasms, possibly due to differences in patient and disease characteristics and regimen-related toxicities.

Humoral immunogenicity of the seasonal influenza vaccine before and after CAR-T-cell therapy: a prospective observational study.

Recipients of chimeric antigen receptor-modified T (CAR-T) cell therapies for B cell malignancies have profound and prolonged immunodeficiencies and are at risk for serious infections, including respiratory virus infections. Vaccination may be important for infection prevention, but there are limited data on vaccine immunogenicity in this population. We conducted a prospective observational study of the humoral immunogenicity of commercially available 2019-2020 inactivated influenza vaccines in adults immediately prior to or while in durable remission after CD19-, CD20-, or B cell maturation antigen-targeted CAR-T-cell therapy, as well as controls. We tested for antibodies to all four vaccine strains using neutralization and hemagglutination inhibition (HAI) assays. Antibody responses were defined as at least fourfold titer increases from baseline. Seroprotection was defined as a HAI titer ≥40. Enrolled CAR-T-cell recipients were vaccinated 14-29 days prior to (n=5) or 13-57 months following therapy (n=13), and the majority had hypogammaglobulinemia and cellular immunodeficiencies prevaccination. Eight non-immunocompromised adults served as controls. Antibody responses to ≥1 vaccine strain occurred in 2 (40%) individuals before CAR-T-cell therapy and in 4 (31%) individuals vaccinated after CAR-T-cell therapy. An additional 1 (20%) and 6 (46%) individuals had at least twofold increases, respectively. One individual vaccinated prior to CAR-T-cell therapy maintained a response for >3 months following therapy. Across all tested vaccine strains, seroprotection was less frequent in CAR-T-cell recipients than in controls. There was evidence of immunogenicity even among individuals with low immunoglobulin, CD19+ B cell, and CD4+ T-cell counts. These data support consideration for vaccination before and after CAR-T-cell therapy for influenza and other relevant pathogens such as SARS-CoV-2, irrespective of hypogammaglobulinemia or B cell aplasia. However, relatively impaired humoral vaccine immunogenicity indicates the need for additional infection-prevention strategies. Larger studies are needed to refine our understanding of potential correlates of vaccine immunogenicity, and durability of immune responses, in CAR-T-cell therapy recipients.

Humoral immunogenicity of the seasonal influenza vaccine before and after CAR-T-cell therapy.

Recipients of chimeric antigen receptor-modified T (CAR-T) cell therapies for B-cell malignancies are immunocompromised and at risk for serious infections. Vaccine immunogenicity is unknown in this population. We conducted a prospective observational study of the humoral immunogenicity of 2019-2020 inactivated influenza vaccines (IIV) in children and adults immediately prior to (n=7) or 13-57 months after (n=15) CD19-, CD20-, or BCMA-targeted CAR-T-cell therapy, as well as controls (n=8). Individuals post-CAR-T-cell therapy were in remission. We tested for antibodies to 4 vaccine strains at baseline and ≥1 time point after IIV using neutralization and hemagglutination inhibition assays. An antibody response was defined as a ≥4-fold titer increase from baseline at the first post-vaccine time point. Baseline A(H1N1) titers in the CAR-T cohorts were significantly lower compared to controls. Antibody responses to ≥1 vaccine strain occurred in 2 (29%) individuals before CAR-T-cell therapy; one individual maintained a response for >3 months post-CAR-T-cell therapy. Antibody responses to ≥1 vaccine strain occurred in 6 (40%) individuals vaccinated after CAR-T-cell therapy. An additional 2 (29%) and 6 (40%) individuals had ≥2-fold increases (at any time) in the pre- and post-CAR-T cohorts, respectively. There were no identified clinical or immunologic predictors of antibody responses. Neither severe hypogammaglobulinemia nor B-cell aplasia precluded antibody responses. These data support consideration for vaccination before and after CAR-T-cell therapy for influenza and other relevant pathogens such as SARS-CoV-2, irrespective of hypogammaglobulinemia or B-cell aplasia. Larger studies are needed to determine correlates of vaccine immunogenicity and durability in CAR-T-cell therapy recipients. KEY POINTS: Influenza vaccination was immunogenic pre- and post-CAR-T-cell therapy, despite hypogammaglobulinemia and B-cell aplasia.Vaccination with inactivated vaccines can be considered before CAR-T-cell therapy and in individuals with remission after therapy.

Antibodies against vaccine-preventable infections after CAR-T cell therapy for B cell malignancies.

BACKGROUNDLittle is known about pathogen-specific humoral immunity after chimeric antigen receptor-modified T (CAR-T) cell therapy for B cell malignancies.METHODSWe conducted a prospective cross-sectional study of CD19-targeted or B cell maturation antigen-targeted (BCMA-targeted) CAR-T cell therapy recipients at least 6 months posttreatment and in remission. We measured pathogen-specific IgG against 12 vaccine-preventable infections and the number of viral and bacterial epitopes to which IgG was detected ("epitope hits") using a serological profiling assay. The primary outcome was the proportion of participants with IgG levels above a threshold correlated with seroprotection for vaccine-preventable infections.RESULTSWe enrolled 65 children and adults a median of 20 months after CD19- (n = 54) or BCMA- (n = 11) CAR-T cell therapy. Among 30 adults without IgG replacement therapy (IGRT) in the prior 16 weeks, 27 (90%) had hypogammaglobulinemia. These individuals had seroprotection to a median of 67% (IQR, 59%-73%) of tested infections. Proportions of participants with seroprotection per pathogen were comparable to population-based studies, but most individuals lacked seroprotection to specific pathogens. Compared with CD19-CAR-T cell recipients, BCMA-CAR-T cell recipients were half as likely to have seroprotection (prevalence ratio, 0.47; 95% CI, 0.18-1.25) and had fewer pathogen-specific epitope hits (mean difference, -90 epitope hits; 95% CI, -157 to -22).CONCLUSIONSeroprotection for vaccine-preventable infections in adult CD19-CAR-T cell recipients was comparable to the general population. BCMA-CAR-T cell recipients had fewer pathogen-specific antibodies. Deficits in both groups support the need for vaccine and immunoglobulin replacement therapy studies.FUNDINGSwiss National Science Foundation (Early Postdoc Mobility grant P2BSP3_188162), NIH/National Cancer Institute (NIH/NCI) (U01CA247548 and P01CA018029), NIH/NCI Cancer Center Support Grants (P30CA0087-48 and P30CA015704-44), American Society for Transplantation and Cellular Therapy, and Juno Therapeutics/BMS.

Liquid Biopsy for Invasive Mold Infections in Hematopoietic Cell Transplant Recipients With Pneumonia Through Next-Generation Sequencing of Microbial Cell-Free DNA in Plasma.

BACKGROUND: Noninvasive diagnostic options are limited for invasive mold infections (IMIs). We evaluated the performance of a plasma microbial cell-free DNA sequencing (mcfDNA-Seq) test for diagnosing pulmonary IMI after hematopoietic cell transplant (HCT). METHODS: We retrospectively assessed the diagnostic performance of plasma mcfDNA-Seq next-generation sequencing in 114 HCT recipients with pneumonia after HCT who had stored plasma obtained within 14 days of diagnosis of proven/probable Aspergillus IMI (n = 51), proven/probable non-Aspergillus IMI (n = 24), possible IMI (n = 20), and non-IMI controls (n = 19). Sequences were aligned to a database including >400 fungi. Organisms above a fixed significance threshold were reported. RESULTS: Among 75 patients with proven/probable pulmonary IMI, mcfDNA-Seq detected ≥1 pathogenic mold in 38 patients (sensitivity, 51% [95% confidence interval {CI}, 39%-62%]). When restricted to samples obtained within 3 days of diagnosis, sensitivity increased to 61%. McfDNA-Seq had higher sensitivity for proven/probable non-Aspergillus IMI (sensitivity, 79% [95% CI, 56%-93%]) compared with Aspergillus IMI (sensitivity, 31% [95% CI, 19%-46%]). McfDNA-Seq also identified non-Aspergillus molds in an additional 7 patients in the Aspergillus subgroup and Aspergillus in 1 patient with possible IMI. Among 19 non-IMI pneumonia controls, mcfDNA-Seq was negative in all samples, suggesting a high specificity (95% CI, 82%-100%) and up to 100% positive predictive value (PPV) with estimated negative predictive values (NPVs) of 81%-99%. The mcfDNA-Seq assay was complementary to serum galactomannan index testing; in combination, they were positive in 84% of individuals with proven/probable pulmonary IMI. CONCLUSIONS: Noninvasive mcfDNA-Seq had moderate sensitivity and high specificity, NPV, and PPV for pulmonary IMI after HCT, particularly for non-Aspergillus species.

Fertility desires and preferences for safer conception strategies among people receiving care for HIV at a publicly-funded clinic in Seattle, WA.

Understanding fertility desires and preferences for HIV prevention among individuals living with HIV, including the potential use of pre-exposure prophylaxis (PrEP) by HIV uninfected partners, can inform the delivery of safer conception counseling to reduce the risk of HIV transmission during pregnancy attempts. Men and women, predominantly heterosexual, engaged in HIV care in Seattle, WA, self-administered a questionnaire and we abstracted antiretroviral therapy (ART) status and HIV viral levels from medical records. We summarized participants' sexual behavior, fertility desires, and preferences for safer conception strategies and used log-binomial regression to identify demographic, sexual, and behavioral factors associated with perceived acceptability of PrEP for HIV uninfected partners during pregnancy attempts. 52% of the 150 participants were female and the mean age was 48 years (range 23-74). 94.7% of participants were using ART and 79.3% had HIV viral load < 40 copies/mL. 22.2% of men and 34.6% of women reported that a healthcare provider had initiated discussion about fertility desires. 28.7% of participants were reproductive-age and desired children. Among sexually active reproductive-age participants with fertility desires, 56.3% reported inconsistent condom use and 62.5% did not report using effective birth control. 74.4% of reproductive age participants with fertility desires perceived that PrEP would be acceptable to an HIV uninfected partner and there were no significant predictors of PrEP acceptability. Nearly one third of reproductive-aged individuals living with HIV expressed fertility desires, highlighting a need for safer conception counseling in this setting. PrEP and ART were favored safer conception strategies.