Oncolytic α-herpesvirus and myeloid-tropic cytomegalovirus cooperatively enhance systemic antitumor responses.

Oncolytic virotherapy aims to activate host antitumor immunity. In responsive tumors, intratumorally injected herpes simplex viruses (HSVs) have been shown to lyse tumor cells, resulting in local inflammation, enhanced tumor antigen presentation, and boosting of antitumor cytotoxic lymphocytes. In contrast to HSV, cytomegalovirus (CMV) is nonlytic and reprograms infected myeloid cells, limiting their antigen-presenting functions and protecting them from recognition by natural killer (NK) cells. Here, we show that when co-injected into mouse tumors with an oncolytic HSV, mouse CMV (mCMV) preferentially targeted tumor-associated myeloid cells, promoted the local release of proinflammatory cytokines, and enhanced systemic antitumor immune responses, leading to superior control of both injected and distant contralateral tumors. Deletion of mCMV genes m06, which degrades major histocompatibility complex class I (MHC class I), or m144, a viral MHC class I homolog that inhibits NK activation, was shown to diminish the antitumor activity of the HSV/mCMV combination. However, an mCMV recombinant lacking the m04 gene, which escorts MHC class I to the cell surface, showed superior HSV adjuvanticity. CMV is a potentially promising agent with which to reshape and enhance antitumor immune responses following oncolytic HSV therapy.

Physical durability: are bed nets getting any stronger?

BACKGROUND: For at least a decade, concerns have been raised about the physical durability of insecticide-treated nets (ITNs) and their ability to remain in good condition for at least three years. To discover if the resistance to damage (RD) of ITNs has improved or not, the RD scores of ITNs sampled in 2013 and 2020 were compared. METHODS: The RD scores and disaggregated textile performance data for nine ITNs recommended by the WHO pesticide evaluation scheme (WHOPES) measured in 2013 were compared with WHO-prequalified ITNs sampled in 2020. This included assessment of newer ITNs not available in 2013, to determine the extent to which product development has led to performance improvements across all available ITNs in the intervening years. RESULTS: The resistance to damage of ITNs has not generally improved from 2013 to 2020, and in some cases performance is worse. The average RD score of comparable ITNs brands decreased from 40 in 2013 to 36 in 2020. Of the nets available in 2020, only two of the twenty-four ITN products tested achieved an RD score of > 50, while six ITNs had very low RD scores of < 30, highlighting a serious inherent, and literal weakness in many WHO-prequalified ITNs. CONCLUSIONS: The long-term physical durability of ITN products cannot be expected to improve while their resistance to damage remains so low, and major upgrades to the performance standards of textile materials used to make ITNs, as well as incentives to develop stronger ones are urgently required.

The long-lasting enigma of polycytidine (polyC) tract.

Long polycytidine (polyC) tracts varying in length from 50 to 400 nucleotides were first described in the 5'-noncoding region (NCR) of genomes of picornaviruses belonging to the Cardio- and Aphthovirus genera over 50 years ago, but the molecular basis of their function is still unknown. Truncation or complete deletion of the polyC tracts in picornaviruses compromises virulence and pathogenicity but do not affect replicative fitness in vitro, suggesting a role as "viral security" RNA element. The evidence available suggests that the presence of a long polyC tract is required for replication in immune cells, which impacts viral distribution and targeting, and, consequently, pathogenic progression. Viral attenuation achieved by reduction of the polyC tract length has been successfully used for vaccine strategies. Further elucidation of the role of the polyC tract in viral replication cycle and its connection with replication in immune cells has the potential to expand the arsenal of tools in the fight against cancer in oncolytic virotherapy (OV). Here, we review the published data on the biological significance and mechanisms of action of the polyC tract in viral pathogenesis in Cardio- and Aphthoviruses.

MeV-Stealth: A CD46-specific oncolytic measles virus resistant to neutralization by measles-immune human serum.

The frequent overexpression of CD46 in malignant tumors has provided a basis to use vaccine-lineage measles virus (MeV) as an oncolytic virotherapy platform. However, widespread measles seropositivity limits the systemic deployment of oncolytic MeV for the treatment of metastatic neoplasia. Here, we report the development of MeV-Stealth, a modified vaccine MeV strain that exhibits oncolytic properties and escapes antimeasles antibodies in vivo. We engineered this virus using homologous envelope glycoproteins from the closely-related but serologically non-cross reactive canine distemper virus (CDV). By fusing a high-affinity CD46 specific single-chain antibody fragment (scFv) to the CDV-Hemagglutinin (H), ablating its tropism for human nectin-4 and modifying the CDV-Fusion (F) signal peptide we achieved efficient retargeting to CD46. A receptor binding affinity of ~20 nM was required to trigger CD46-dependent intercellular fusion at levels comparable to the original MeV H/F complex and to achieve similar antitumor efficacy in myeloma and ovarian tumor-bearing mice models. In mice passively immunized with measles-immune serum, treatment of ovarian tumors with MeV-Stealth significantly increased overall survival compared with treatment with vaccine-lineage MeV. Our results show that MeV-Stealth effectively targets and lyses CD46-expressing cancer cells in mouse models of ovarian cancer and myeloma, and evades inhibition by human measles-immune serum. MeV-Stealth could therefore represent a strong alternative to current oncolytic MeV strains for treatment of measles-immune cancer patients.

Oncolytic Foamy Virus - generation and properties of a nonpathogenic replicating retroviral vector system that targets chronically proliferating cancer cells.

Nonpathogenic retroviruses of the Spumaretrovirinae subfamily can persist long-term in the cytoplasm of infected cells, completing their lifecycle only after the nuclear membrane dissolves at the time of cell division. Since the targeting of slowly dividing cancer cells remains an unmet need in oncolytic virotherapy we constructed a replication competent Foamy Virus vector (oFV) from the genomes of two chimpanzee Simian Foamy Viruses (PAN1 and PAN2) and inserted a GFP transgene in place of the bel-2 open reading frame. oFV-GFP infected and propagated with slow kinetics in multiple human tumor cell lines, inducing a syncytial cytopathic effect. Infection of growth arrested MRC5 cells was not productive, but oFV genomes persisted in the cytoplasm and the productive viral lifecycle resumed when cell division was later restored. In vivo, the virus propagated extensively in intraperitoneal ovarian cancer xenografts, slowing tumor growth, significantly prolonging survival of the treated mice and sustaining GFP transgene expression for at least 45 days. Our data indicate that oFV is a promising new replication-competent viral and gene delivery platform for efficient targeting of the most fundamental trait of cancer cells, their ability to sustain chronic proliferation.Significance:The infectivity of certain retroviruses is limited to dividing cells, which makes them attractive tools for targeting cancer cell proliferation. Previously developed replication-competent gammaretroviral vectors spread efficiently in rapidly dividing cancer cells, but not in cancer cells that divide more slowly. In contrast to rapidly proliferating transplantable mouse tumors, slow proliferation is a hallmark of human cancers and may have contributed to the clinical failure of the preclinically promising Murine Leukemia Virus vector Toca511 which failed to show efficacy in a phase 3 clinical trial in patients with glioblastoma. The studies presented in our manuscript show that oncolytic Foamy Virus (oFV) vectors are capable of persisting unintegrated in quiescent cells and resuming their life cycle once the cells start dividing again. This property of oFVs, together with their lack of pathogenicity and their ability to catalyze the fusion of infected cancer cells, makes them an attractive platform for further investigation.

Improved Noninvasive In Vivo Tracking of AAV-9 Gene Therapy Using the Perchlorate-Resistant Sodium Iodide Symporter from Minke Whale.

The sodium iodide symporter (NIS) is widely used as a reporter gene to noninvasively monitor the biodistribution and durability of vector-mediated gene expression via gamma scintigraphy, single-photon emission computed tomography (SPECT), and positron-emission tomography (PET). However, the approach is limited by background signal due to radiotracer uptake by endogenous NIS-expressing tissues. In this study, using the SPECT tracer pertechnetate (99mTcO4) and the PET tracer tetrafluoroborate (B18F4), in combination with the NIS inhibitor perchlorate, we compared the transport properties of human NIS and minke whale (Balaenoptera acutorostrata scammoni) NIS in vitro and in vivo. Based on its relative resistance to perchlorate, the NIS protein from minke whale appeared to be the superior candidate reporter gene. SPECT and PET imaging studies in nude mice challenged with NIS-encoding adeno-associated virus (AAV)-9 vectors confirmed that minke whale NIS, in contrast to human and endogenous mouse NIS, continues to function as a reliable reporter even when background radiotracer uptake by endogenous NIS is blocked by perchlorate.

The causes of holes and loss of physical integrity in long-lasting insecticidal nets.

BACKGROUND: Long-lasting insecticidal nets (LLINs) are expected to last for at least 3 years, but whilst this may be achieved from an insecticidal perspective, physical protection is frequently compromised much earlier because of the rapid accumulation of holes during use. To understand why LLINs are so susceptible to loss of physical integrity, thousands of hole damage sites in LLINs retrieved from the field in Africa and Asia were forensically studied to identify the persistent underlying causes. METHODS: A total of 525 LLINs consisting of six different brands from five different countries across Africa and Asia were collected from the field after 1 to 3 years in use. More than 42,000 individual sites of hole damage were analysed based on the morphology and size of each individual hole, aided by optical microscopy (OM) and scanning electron microscopy (SEM). The fracture morphology enabled positive identification of the underlying mechanisms of the damage. RESULTS: Across all LLINs and geographical settings, mechanical damage is the primary cause of holes and loss of physical integrity in LLINs (63.14% by frequency and 81.52% by area). Snagging is the single most frequent mechanical damage mechanism, whilst the largest sized holes in LLINs result from seam failure and tearing. Abrasion and hole enlargement are also responsible for a progressive loss in the physical integrity of nets. Collectively, these five modes of mechanical damage can be expected to result from normal use of LLINs by households. Evidence of deliberate cutting, burn holes and rodent damage was observed to a lesser degree, which LLINs are not designed to withstand. CONCLUSIONS: Loss of physical integrity in LLINs is an inevitable consequence of using a vector control product that has an inherently low resistance to mechanical damage during normal use. To improve performance, new specifications based on laboratory textile testing is needed, to assess the resistance of LLIN products to the primary causes of mechanical damage when in use, which are snagging, tearing, abrasion and hole enlargement. Seam construction also needs to meet a revised minimum standard to reduce the risk of a rapid loss of physical integrity during use.

Development of a single resistance to damage metric for mosquito nets related to physical integrity in the field.

BACKGROUND: In common with the majority of personal protective equipment and healthcare products, the ability for long-lasting insecticidal nets (LLINs) to remain in good physical condition during use is a key factor governing fitness for purpose and serviceability. The inherent ability of a product to resist physical deterioration should be known in advance of it being used to ensure it has maximum value to both the end-user and procurer. The objective of this study was to develop a single performance metric of resistance to damage (RD) that can be applied to any LLIN product prior to distribution. METHODS: Algorithms to calculate RD values were developed based on consideration of both human factors and laboratory testing data. Quantitative reference forces applied to LLINs by users during normal use were determined so that aspirational performance levels could be established. The ability of LLINs to resist mechanical damage was assessed based on a new suite of textile tests, reflecting actual mechanisms of physical deterioration during normal household use. These tests quantified the snag strength, bursting strength, abrasion resistance and resistance to hole enlargement. Sixteen different unused LLINs were included in the analysis. The calculated RD values for all LLINs and the corresponding physical integrity data for the same nets retrieved from the field (up to 3 years of use) were then compared. RESULTS: On a RD scale of 0 (lowest resistance) - 100 (highest resistance), only six of the sixteen LLINs achieved an RD value above 50. No current LLIN achieved the aspirational level of resistance to damage (RD = 100), suggesting that product innovation is urgently required to increase the RD of LLINs. LLINs with higher RD values were associated with lower hole damage (PHI) in the field when adjusted for normal use conditions. CONCLUSIONS: The RD value of any LLIN product can be determined prior to distribution based on the developed algorithms and laboratory textile testing data. Generally, LLINs need to achieve higher RD values to improve their ability to resist hole formation during normal use. Innovation in LLIN product design focused on the textile material should be actively encouraged and is urgently needed to close the performance gap.

Textile testing to assess the resistance to damage of long-lasting insecticidal nets for malaria control and prevention.

BACKGROUND: LLINs are susceptible to forming holes within a short time in use, compromising their ability to provide long-term physical protection against insect-borne vectors of disease. Mechanical damage is known to be responsible for the majority of holes, with most being the result of snagging, tearing, hole enlargement, abrasion and seam failure, which can readily occur during normal household use. To enable an assessment of the ability of LLINs to resist such damage prior to distribution, a new suite of testing methods was developed to reflect the main damage mechanisms encountered during normal use of LLINs. METHODS: Four existing BS EN and ISO standards used by the textile industry were adapted to determine the ability of LLINs to resist the most common mechanisms of real-world damage experienced in the field. The new suite comprised tests for snag strength (BS 15,598:2008), bursting strength (ISO 13938-2:1999), hole enlargement resistance (BS 3423-38:1998), abrasion resistance (ISO 12947-1:1998) and new guidance around the seam construction of LLINs. Fourteen different LLINs were tested using the new suite of tests to evaluate their resistance to damage. RESULTS: The resistance to mechanical damage of LLINs is not the same, even when the bursting strength values are comparable. Differences in performance between LLINs are directly related to the fabric design specifications, including the knitted structure and constituent yarns. The differences in performance do not primarily relate to what polymer type the LLIN is made from. LLINs made with a Marquisette knitted structure produced the highest snag strength and lowest hole enlargement values. By contrast, LLINs made with a traverse knitted structure exhibited low snag strength values when compared at the same mesh count. CONCLUSIONS: Prequalification of LLINs should consider not only insecticidal performance, but also inherent resistance to mechanical damage. This is critical to ensuring LLINs are fit for purpose prior to distribution, and are capable of remaining in good physical condition for longer. The new suite of test methods enables the performance of LLINs to be assessed and specified in advance of distribution and can be used to establish minimum performance standards. Implementation of these testing methods is therefore recommended.

Correlation of textile 'resistance to damage' scores with actual physical survival of long-lasting insecticidal nets in the field.

BACKGROUND: Attempts have been made to link procurement of long-lasting insecticidal nets (LLIN) not only to the price but also the expected performance of the product. However, to date it has not been possible to identify a specific textile characteristic that predicts physical durability in the field. The recently developed resistance to damage (RD) score could provide such a metric. This study uses pooled data from durability monitoring to explore the usefulness of the RD methodology. METHODS: Data from standardized, 3-year, prospective LLIN durability monitoring for six LLIN brands in 10 locations and four countries involving 4672 campaign LLIN were linked to the RD scores of the respective LLIN brands. The RD score is a single quantitative metric based on a suite of standardized textile tests which in turn build on the mechanisms of damage to a mosquito net. Potential RD values range from 0 to 100 where 100 represents optimal resistance to expected day-to-day stress during reasonable net use. Survival analysis was set so that risk of failure only started when nets were first hung. Cox regression was applied to explore RD effects on physical survival adjusting for known net use environment variables. RESULTS: In a bivariate analysis RD scores showed a linear relationship with physical integrity suggesting that the proportion of LLIN with moderate damage decreased by 3%-points for each 10-point increase of the RD score (p = 0.02, R2 = 0.65). Full adjustment for net care and handling behaviours as well as other relevant determinants and the country of study showed that increasing RD score by 10 points resulted in a 36% reduction of risk of failure to survive in serviceable condition (p < 0.0001). LLINs with RD scores above 50 had an additional useful life of 7 months. CONCLUSIONS: This study provides proof of principle that the RD metric can predict physical durability of LLIN products in the field and could be used to assess new products and guide manufacturers in creating improved products. However, additional validation from other field data, particularly for next generation LLIN, will be required before the RD score can be included in procurement decisions for LLINs.

Comparison of the current renal staging, progression and response criteria to predict renal survival in AL amyloidosis using a Mayo cohort.

Three sets of criteria (International Society of Amyloidosis [ISA], Palladini and Kastritis) were independently developed for staging, progression and response criteria to predict renal survival in patients with AL amyloidosis. We evaluated these criteria using a cohort of 495 newly diagnosed AL amyloidosis patients with renal involvement using time to event competing risk analysis at baseline, 3, 6 and 12 months after treatment. Only Palladini and Kastritis had a staging system and both predicted a higher risk of end stage renal disease (ESRD) in the stage III vs stage I patients but only the Palladini model was predictive for stage II patients. At 3 months, risk of ESRD was significantly higher for Palladini and ISA renal progression (hazard ratio [HR] 2.8 [95% CI: 1.5-5.3, p = .001] and 2.5 [CI: 1.4-4.6, p = .004, respectively]), but renal response was not significantly protective; conversely, the risk of ESRD was not significantly higher for the Kastritis renal progression, but was significantly protective for the Kastritis renal responders (HR 0.38 [95% CI: 0.17-0.84], p = .017). Both progression and response with ISA, Palladini and Kastritis criteria were predictive of ESRD at 6 months and 12 months. While the Palladini staging criteria at baseline, and the ISA and Palladini criteria for progression at 3 months performed better than the Kastritis criteria at baseline and 3 months post-treatment, the Kastritis criteria performed better for response 3 months after treatment. All three sets of criteria performed well at and after 6 months post-treatment. These differences are important when choosing endpoints for clinical trials.

Mapping of Ion and Substrate Binding Sites in Human Sodium Iodide Symporter (hNIS).

The human sodium iodide symporter (hNIS) is a theranostic reporter gene which concentrates several clinically approved SPECT and PET radiotracers and plays an essential role for the synthesis of thyroid hormones as an iodide transporter in the thyroid gland. Development of hNIS mutants which could enhance translocation of the desired imaging ions is currently underway. Unfortunately, it is hindered by lack of understanding of the 3D organization of hNIS and its relation to anion transport. There are no known crystal structures of hNIS in any of its conformational states. Homology modeling can be very effective in such situations; however, the low sequence identity between hNIS and relevant secondary transporters with available experimental structures makes the choice of a template and the generation of 3D models nontrivial. Here, we report a combined application of homology modeling and molecular dynamics refining of the hNIS structure in its semioccluded state. The modeling was based on templates from the LeuT-fold protein family and was done with emphasis on the refinement of the substrate-ion binding pocket. The consensus model developed in this work is compared to available biophysical and biochemical experimental data for a number of different LeuT-fold proteins. Some functionally important residues contributing to the formation of putative binding sites and permeation pathways for the cotransported Na+ ions and I- substrate were identified. The model predictions were experimentally tested by generation of mutant versions of hNIS and measurement of relative (to WT hNIS) 125I- uptake of 35 hNIS variants.

Induced Periosteum-Mimicking Membrane with Cell Barrier and Multipotential Stromal Cell (MSC) Homing Functionalities.

The current management of critical size bone defects (CSBDs) remains challenging and requires multiple surgeries. To reduce the number of surgeries, wrapping a biodegradable fibrous membrane around the defect to contain the graft and carry biological stimulants for repair is highly desirable. Poly(ε-caprolactone) (PCL) can be utilised to realise nonwoven fibrous barrier-like structures through free surface electrospinning (FSE). Human periosteum and induced membrane (IM) samples informed the development of an FSE membrane to support platelet lysate (PL) absorption, multipotential stromal cells (MSC) growth, and the prevention of cell migration. Although thinner than IM, periosteum presented a more mature vascular system with a significantly larger blood vessel diameter. The electrospun membrane (PCL3%-E) exhibited randomly configured nanoscale fibres that were successfully customised to introduce pores of increased diameter, without compromising tensile properties. Additional to the PL absorption and release capabilities needed for MSC attraction and growth, PCL3%-E also provided a favourable surface for the proliferation and alignment of periosteum- and bone marrow derived-MSCs, whilst possessing a barrier function to cell migration. These results demonstrate the development of a promising biodegradable barrier membrane enabling PL release and MSC colonisation, two key functionalities needed for the in situ formation of a transitional periosteum-like structure, enabling movement towards single-surgery CSBD reconstruction.

Recontamination of Healthcare Surfaces by Repeated Wiping with Biocide-Loaded Wipes: "One Wipe, One Surface, One Direction, Dispose" as Best Practice in the Clinical Environment.

The wiping of high-touch healthcare surfaces made of metals, ceramics and plastics to remove bacteria is an accepted tool in combatting the transmission of healthcare-associated infections (HCAIs). In practice, surfaces may be repeatedly wiped using a single wipe, and the potential for recontamination may be affected by various factors. Accordingly, we studied how the surface to be wiped, the type of fibre in the wipe and how the presence of liquid biocide affected the degree of recontamination. Experiments were conducted using metal, ceramic and plastic healthcare surfaces, and two different wipe compositions (hygroscopic and hydrophilic), with and without liquid biocide. Despite initially high removal efficiencies of >70% during initial wiping, all healthcare surfaces were recontaminated with E. coli, S. aureus and E. faecalis when wiped more than once using the same wipe. Recontamination occurred regardless of the fibre composition of the wipe or the presence of a liquid biocide. The extent of recontamination by E. coli, S. aureus and E. faecalis bacteria also increased when metal healthcare surfaces possessed a higher microscale roughness (<1 µm), as determined by Atomic Force Microscopy (AFM). The high propensity for healthcare surfaces to be re-contaminated following initial wiping suggests that a "One wipe, One surface, One direction, Dispose" policy should be implemented and rigorously enforced.

Substratification of patients with newly diagnosed standard-risk multiple myeloma.

Despite the absence of high-risk cytogenetics and lower International Staging System (ISS) stages, a subset of patients with multiple myeloma (MM) experience poor overall survival (OS). We studied 1461 patients with newly diagnosed MM to identify patient and disease characteristics that predict a high-risk phenotype among standard-risk patients. Fifty-six percent of all patients presented with standard-risk disease. Among them, advanced age, extremes of body mass index, non-hyperdiploid karyotype and abnormal lymphocyte counts were associated with worse OS. Standard-risk patients with 0-1 of these adverse factors (hazard ratio [HR] 0·32, 95% confidence interval [CI] 0·24-0·43, P < 0·001) and 2 adverse factors (HR 0·54, 95% CI 0·41-0·72, P < 0·001) experienced better OS than high-risk patients. Two or more adverse factors were present in 17% of standard-risk patients and were associated with OS comparable to high-risk patients (HR 0·91, 95% CI 0·67-1·24, P = 0·548). Predictive power among standard-risk patients was improved using score groups compared to ISS stages. Patients with standard-risk MM are a heterogeneous group with one in six patients experiencing OS comparable to high-risk disease. Patients at risk can be identified using readily available patient and disease characteristics. These findings emphasize the importance of accurate risk stratification and help explain part of the heterogeneity observed in clinical practice.

Pomalidomide, bortezomib, and dexamethasone for patients with relapsed lenalidomide-refractory multiple myeloma.

This phase 1/2 trial evaluated the maximum tolerated doses, safety, and efficacy of pomalidomide, bortezomib, and dexamethasone (PVD) combination in patients with relapsed lenalidomide-refractory multiple myeloma (MM). In phase 1, dose level 1 consisted of pomalidomide (4 mg by mouth on days 1 to 21), IV or subcutaneous bortezomib (1.0 mg/m2 on days 1, 8, 15, and 22), and dexamethasone (40 mg by mouth on days 1, 8, 15, and 22) given every 28 days. Bortezomib was increased to 1.3 mg/m2 for dose level 2 and adopted in the phase 2 expansion cohort. We describe the results of 50 patients. Objective response rate was 86% (95% confidence interval [CI], 73-94) among all evaluable patients (stringent complete response, 12%; complete response, 10%; very good partial response, 28%; and partial response, 36%) and 100% among high-risk patients. Within a median follow-up of 42 months, 20% remain progression free, 66% are alive, and 4% remain on treatment. Median progression-free survival was 13.7 months (95% CI, 9.6-17.7). The most common toxicities were neutropenia (96%), leukopenia (84%), thrombocytopenia (82%), anemia (74%), and fatigue (72%); however, the majority of these were grade 1 or 2. The most common grade ≥3 toxicities included neutropenia (70%), leukopenia (36%), and lymphopenia (20%). Deep vein thrombosis occurred in 5 patients. In conclusion, PVD is a highly effective combination in lenalidomide-refractory MM patients. Weekly administration of bortezomib enhanced tolerability and convenience. Toxicities are manageable, mostly consisting of mild cytopenias with no significant neuropathy. This trial was registered at www.clinicaltrials.gov as #NCT01212952.

Peripheral blood biomarkers of early immune reconstitution in newly diagnosed multiple myeloma.

Peripheral blood biomarkers of tumor microenvironment and immune surveillance are independent prognostic factors in multiple myeloma. The timing and prognostic impact of immune reconstitution has been studied after autologous hematopoietic stem cell transplantation, less is known about its significance in newly diagnosed multiple myeloma. We studied absolute lymphocyte (ALC) and absolute monocyte (AMC) counts at the time of treatment initiation and 1 month thereafter in 771 newly diagnosed patients. Two hundred and thirty-four patients (31%) had evidence of immune dysregulation at baseline (abnormal biomarkers). Eighty-seven of these patients (37%) recovered normal biomarkers at 1 month (early immune reconstitution). The absence of immune dysregulation at baseline (compared to the presence thereof) was associated with better overall survival (HR 0.77, 95% CI 0.61-0.97, P = 0.025, n = 771). The absence of immune dysregulation at 1 month (compared to the persistence or development thereof) was associated with better overall survival (HR 0.63, 95% CI 0.50-0.80, P < 0.001, n = 771). Early immune reconstitution (compared to the persistence or development of immune dysregulation) was associated with better overall survival (HR 0.62, 95% CI 0.43-0.92, P = 0.016, n = 771). Cytogenetic high-risk disease was negatively, and treatment with immunomodulators positively, associated with early immune reconstitution. The presence or development of immune dysregulation in newly diagnosed multiple myeloma is an independent risk factor. The favorable impact of early immune reconstitution suggests immune dysregulation to be a potentially modifiable risk factor that may be exploited for therapeutic benefit.

Prognostic value of minimal residual disease and polyclonal plasma cells in myeloma patients achieving a complete response to therapy.

Achievement of a complete response has been associated with improved outcomes in patients with multiple myeloma. Recently, increasing application of minimal residual disease (MRD) assessment has shown that MRD negativity is a powerful prognostic factor for survival outcomes. We wanted to examine the impact of the polyclonal plasma cell (pPC) compartment among patients in complete response (CR) but are MRD positive. This is a retrospective cohort study where 460 myeloma patients were identified who met criteria for CR and had multicolor flow cytometry performed on the bone marrow (BM). Monoclonal and pPCs were estimated during MRD testing. Final outcomes including overall survival (OS) and time to next treatment (TTNT) were compared among the groups. The median OS for the entire cohort was not reached (95% CI; 63 mos, NR) and the median TTNT was 31 months (95% CI; 27,36). Among the MRDneg group, median TTNT was 37.6 months vs 23 months for MRDpos patients (P < .001); the median OS was not reached for either group, but there was a trend toward better survival for MRDneg patients. Among the MRDpos group, median percentage of pPCs was 65% (2.5-98.5), and those with >95% pPCs had a significantly better TTNT (NR vs 23 months; P = .02) and a trend toward better OS. We conclude that achievement of MRD negativity predicts for better response durability and trend toward improved OS and an increased proportion of pPC predicts for better outcomes within those who have residual tumor cells highlighting the importance of marrow normalization.

Antigenic Drift Defines a New D4 Subgenotype of Measles Virus.

The measles virus hemagglutinin (MeV-H) protein is the main target of protective neutralizing antibodies. Using a panel of monoclonal antibodies (MAbs) that recognize known major antigenic sites in MeV-H, we identified a D4 genotype variant that escapes neutralization by MAbs targeting the neutralizing epitope (NE) antigenic site. By site-directed mutagenesis, L249P was identified as the critical mutation disrupting the NE in this genotype D4 variant. Forty-two available D4 genotype gene sequences were subsequently analyzed and divided into 2 groups according to the presence or absence of the L249P MeV-H mutation. Further analysis of the MeV-N gene sequences of these 2 groups confirmed that they represent clearly definable, sequence-divergent D4 subgenotypes, which we named subgenotypes D4.1 and D4.2. The subgenotype D4.1 MeVs were isolated predominantly in Kenya and Ethiopia, whereas the MAb-resistant subgenotype D4.2 MeVs were isolated predominantly in France and Great Britain, countries with higher vaccine coverage rates. Interestingly, D4.2 subgenotype viruses showed a trend toward diminished susceptibility to neutralization by human sera pooled from approximately 60 to 80 North American donors. Escape from MAb neutralization may be a powerful epidemiological surveillance tool to monitor the evolution of new MeV subgenotypes.IMPORTANCE Measles virus is a paradigmatic RNA virus, as the antigenic composition of the vaccination has not needed to be updated since its discovery. The vaccine confers protection by inducing neutralizing antibodies that interfere with the function of the hemagglutinin protein. Viral strains are indistinguishable serologically, although characteristic nucleotide sequences differentiate 24 genotypes. In this work, we describe a distant evolutionary branch within genotype D4. Designated subgenotype D4.2, this virus is distinguishable by neutralization with vaccine-induced monoclonal antibodies that target the neutralizing epitope (NE). The subgenotype D4.2 viruses have a higher predominance in countries with intermediary levels of vaccine coverage. Our studies demonstrate that subgenotype D4.2 lacks epitopes associated with half of the known antigenic sites, which significantly impacts our understanding of measles virus evolution.

Immunovirotherapy with vesicular stomatitis virus and PD-L1 blockade enhances therapeutic outcome in murine acute myeloid leukemia.

Patients with relapsed acute myeloid leukemia (AML) have limited therapeutic options. Vesicular stomatitis virus (VSV)-interferon ß (IFNß)-sodium iodide symporter (NIS) is an oncolytic VSV encoding IFNß and the NIS reporter. Syngeneic AML C1498 tumors responded to IV therapy with VSV-murine IFNß (mIFNß)-NIS in a dose-dependent manner. Imaging for NIS expression showed robust virus infection within the tumors. Virus infection did not increase programmed death ligand 1 (PD-L1) on tumor cells. Combining VSV-mIFNß-NIS with anti-PD-L1 antibody (Ab) therapy enhanced antitumor activity compared with treatment with virus alone or Ab alone; this enhancement was not significant at higher VSV-mIFNß-NIS doses. Systemic VSV therapy reduced systemic C1498-green fluorescent protein (GFP) tumor burden in the blood, bone marrow, spleen, and liver of mice with AML. Combination VSV-mIFNß-NIS and anti-PD-L1 Ab therapy significantly enhanced the survival of these mice with no evidence of toxicity, compared with isotype control, anti-PD-L1, or virus alone. There was an increase in tumor-infiltrating CD4 and CD8 cells. Single-agent VSV-mIFNß-NIS virotherapy induced both VSV-specific and GFP-specific CD8 T cells as determined by IFN-γ enzyme-linked immunospot, pentamer, and intracellular IFN-γ staining assays. Both of these responses were further enhanced by addition of anti-PD-L1 Ab. Depletion of CD8 or natural killer cells, but not CD4 cells, resulted in loss of antitumor activity in the VSV/anti-PD-L1 group. Clinical samples from chronic myelomonocytic leukemia and acute myelomonocytic leukemia appear to be especially susceptible to VSV. Overall, our studies show that oncolytic virotherapy combined with immune checkpoint blockade is a promising approach to AML therapy.