Summary Background We previously reported rates of pathological complete responses (51% 95% CI 39−62 per independent central review, the primary endpoint) and major pathological responses (13% per ...independent central review, a secondary endpoint) to neoadjuvant cemiplimab (an anti-PD-1 inhibitor) among 79 patients with locoregionally advanced, resectable cutaneous squamous cell carcinoma. Here, we present follow-up data, including event-free, disease-free, and overall survival. Methods This single-arm, multicentre, phase 2 study included patients aged 18 years or older with resectable stage II–IV (M0) cutaneous squamous cell carcinoma and Eastern Cooperative Oncology Group performance status of 0 or 1. Patients received up to four planned doses of neoadjuvant cemiplimab 350 mg intravenously every 3 weeks followed by curative-intent surgery. After surgery, per investigator discretion, patients received either adjuvant cemiplimab for up to 48 weeks, radiotherapy, or observation alone. Secondary endpoints included in this follow-up analysis are event-free survival, disease-free survival, and overall survival, all summarised using the Kaplan-Meier method. Activity and safety endpoints were analysed for all enrolled patients who received at least one dose of neoadjuvant cemiplimab. In this report, safety data are reported for all patients who received at least one dose of adjuvant cemiplimab. This trial is registered with ClinicalTrials.gov, NCT04154943, has completed enrolment and follow-up is ongoing. Findings Between March 20, 2020, and July 8, 2021, 79 patients were enrolled. Median age was 73 years (IQR 66–81), 67 (85%) patients were male, 12 (15%) were female, 69 (87%) were White, one was Asian (1%), one was other race (1%), and race was not reported for eight (10%). As of data cutoff (Dec 1, 2022), median follow-up was 18·7 months (IQR 15·6–22·1) for all 79 patients. Among 70 patients who had surgery, 65 (93%) had post-surgical management data: 32 (49%) of 65 were observed postoperatively, 16 (25%) received adjuvant cemiplimab, and 17 (26%) received adjuvant radiotherapy. 11 (14%) of 79 patients had event-free survival events, with an estimated 12-month event-free survival of 89% (95% CI 79–94) for all patients. None of 40 patients who had a pathological complete response and one (10%) of ten patients with major pathological response had recurrence. Six (9%) of 70 patients who completed surgery had a disease-free survival event, with an estimated 12-month disease-free survival of 92% (95% CI 82–97). Nine (11%) of 79 patients died, with an estimated 12-month overall survival for all patients of 92% (95% CI 83–96). Four (25%) of 16 patients who received adjuvant cemiplimab treatment had grade 3 adverse events, including one (6%) who had increased blood potassium, one (6%) who had traumatic limb amputation, and two who had serious adverse events (one 6% cardiomyopathy and one 6% hypophysitis). There were no grade 4 adverse events or treatment-related deaths. Interpretation For patients with resectable stage II–IV cutaneous squamous cell carcinoma, neoadjuvant cemiplimab followed by surgery might be a potential treatment option, addressing a substantial unmet need. Funding Regeneron Pharmaceuticals and Sanofi.
Self-reactive B cells not controlled by receptor editing or clonal deletion may become anergic. We report that fully mature human B cells negative for surface IgM and retaining only IgD are ...autoreactive and functionally attenuated (referred to as naive IgD(+)IgM(-) B cells B(ND)). These B(ND) cells typically make up 2.5% of B cells in the peripheral blood, have antibody variable region genes in germline (unmutated) configuration, and, by all current measures, are fully mature. Analysis of 95 recombinant antibodies expressed from the variable genes of single B(ND) cells demonstrated that they are predominantly autoreactive, binding to HEp-2 cell antigens and DNA. Upon B cell receptor cross-linkage, B(ND) cells have a reduced capacity to mobilize intracellular calcium or phosphorylate tyrosines, demonstrating that they are anergic. However, intense stimulation causes B(ND) cells to fully respond, suggesting that these cells could be the precursors of autoantibody secreting plasma cells in autoimmune diseases such as systemic lupus erythematosus or rheumatoid arthritis. This is the first identification of a distinct mature human B cell subset that is naturally autoreactive and controlled by the tolerizing mechanism of functional anergy.
INTRODUCTION
T Cell Receptor mimic (TCRm) antibodies to low-density peptide epitopes from undruggable intracellular proteins presented in the context of major histocompatibility (MHC) molecules are ...therapeutically effective in mouse models of human cancers. CD47 blockade by use of a high affinity SIRPα variant protein (CV1) has been shown to improve the effects of monoclonal antibodies to high-density antigens in tumor models by enhancement of antibody dependent cellular phagocytosis (ADCP). We asked if combination therapy with a TCRm antibody to Preferentially Expressed Antigen in Melanoma (PRAME) could enhance activity of both drugs in vitro and in vivo. Additionally, we explored the role of macrophage-secreted cytokines in the enhanced in vivo activity.
METHODS
We performed in vitro ADCP assays with human acute myeloid and acute lymphoid leukemia cell lines containing antigens of interest using the two agents alone and in combination. We performed therapy experiments in NSG mice using the same leukemia cell lines transformed with a luciferase vector and measured tumor burden through bioluminescent imaging. Survival was measured. We examined cell-surface expression of epitopes of interest and HLA on cell lines in vitro after incubation with IFNγ and TNFα using flow cytometry and performed in vitro ADCP assays with the leukemia cell lines after pretreatment with IFNγ.
RESULTS
CV1 and TCRm antibody showed additive effects in vitro with a statistically significant increase in phagocytosis in both antigen positive cell lines with combination therapy versus single agent therapy. CV1 and TCRm antibody showed greater than additive therapeutic effects in vivo with a 3-log reduction in leukemia burden relative to control untreated mice and a 5-10 fold reduction relative to single agent groups. After therapy was stopped, mice treated with the combination had statistically significant increases in survival (p<0.0001). IFNγ and TNFα led to up-regulation of cell surface HLA-A*02:01. Additionally, the cytokines led to up-regulation of the PRAME derived epitope of interest. Pretreatment of human leukemia cell lines with IFNγ led to statistically significant increases in ADCP in vitro.
CONCLUSIONS
The elimination of anti-phagocytic signal produced by CD47 blockade with the high affinity SIRPα variant CV1, combined with the pro-phagocytic signal of Fc receptor engaging TCRm was effective even with an ultra-low density epitope (700-3000 sites per cell) in vitro and in vivo. A greater than additive effect was seen in two tumor models. These results support the potency of this drug combination. The large enhancement in activity in vivo vs. in vitro may be explained by macrophage-released cytokines leading to increased presentation of epitopes of interest and increased tumor kill.
Liu:Eureka Therapeutics: Employment, Equity Ownership, Patents & Royalties. Garcia:Alexo Therapeutics: Equity Ownership.
Abstract 2111▪▪This icon denotes a clinically relevant abstract
The acute painful crisis is the clinical hallmark of sickle cell disease (SCD). Recently, time-to-opioid administration (TTO) has been ...suggested as quality-of-care (QOC) measure for SCD patients with an acute painful crisis (Wang,Pediatrics,128:484,2011). We sought to determine whether TTO was associated with outcomes of emergency department (ED) visits for acute painful crisis.
We conducted a retrospective cohort study of ED visits for acute painful crisis to Children’s Medical Center Dallas between 1/1/2008 and 12/31/2010. Potential subjects were identified by query of hospital administrative records using all ICD-9CM codes for SCD (282.41, 282.42, and 282.6x) as either the primary or a secondary diagnosis. We included patients with any SCD genotype between 5 and 18 years of age. Records of all potential subjects were reviewed to determine whether the episode represented an acute painful crisis, defined as the recent onset of acute pain not explained by other mechanisms. We excluded patients with confounding sources of pain (e.g., headache and gallstones), patients transferred from other medical centers, and patients who had a surgical procedure within 2 weeks of presentation. We also excluded patients who received a simple transfusion in the preceding 30 days or were part of a chronic transfusion program in the preceding 6 months. The primary predictor variable was TTO in minutes, analyzed continuously and in quartiles. The primary outcome variable was hospital admission with secondary outcomes of change in pain scores 1 & 2, area under the curve (AUC) for pain scores at 4 hours (pain score AUC), and length of stay in the ED (ED LOS) in minutes. Univariate mixed regression (logistic for admission, linear for other outcome variables), including patient random effects, was used to test for association between TTO and other relevant covariates with the outcome variables. Variables with P-values <0.10 in the univariate regression models were subsequently evaluated in multivariate mixed models.
From 2,866 ED visits in 2008–2010, 177 subjects were identified with 416 presentations for acute painful crisis treated with IV opioids. 47% of subjects were female and 65% had HbSS/HbSβ0. The primary outcome, inpatient admission, occurred in 53% of ED visits. The median TTO was 86 minutes (intraquartile range (IQR) 54 – 130 minutes) and was not different between those admitted and not admitted to the hospital (not admitted - 86.5 minutes, (IQR 56.5, 126.5); admitted - 85.5 minutes, (IQR 50.5, 130)). TTO (both as a continuous and categorical variable) was not associated with inpatient admission in either univariate or multivariate analyses. In multivariate analyses with secondary outcomes, TTO was associated with change in pain scores 1 & 2, pain score AUC at 4 hours, and ED LOS. After adjusting for age and initial pain score, decreased TTO was independently associated with a decrease in pain scores 1 & 2 (referent - TTO Quartile 4 (>131 minutes) vs Quartile1 (<56 minutes), β coefficient = 0.49, 95% Confidence Interval (CI) 0.08, 0.89, p=0.019). After adjusting for age, decreased TTO was independently associated with a decreased pain score AUC (referent - TTO Quartile 4 vs Quartile 1, β coefficient = −81, 95% CI −153, −9, p=0.027). Finally, after adjusting for admission status, decreased TTO was independently associated with decreased ED LOS (referent – TTO Quartile 4 vs Quartile 1, β coefficient = −131, 95% CI −191, −72, p=<0.001).
For acute painful crisis, TTO in the ED setting has been suggested as a QOC measure. While TTO is not associated with our primary outcome, inpatient admission, it is independently associated three other important outcomes: change in pain scores 1 & 2, pain score AUC, and ED LOS. The association of the process measure, TTO, with these outcomes increases the appeal for its use in broader sickle cell populations and should lead to necessary steps to reduce TTO in the ED setting. Subsequent studies should investigate TTO in other SCD populations including adults, examine determinants of prolonged TTO, and study interventions designed to reduce TTO in various ED settings.
No relevant conflicts of interest to declare.
Preferentially expressed antigen in melanoma (PRAME) is a cancer-testis antigen that is expressed in many cancers and leukemias. In healthy tissue, PRAME expression is limited to the testes and ...ovaries, making it a highly attractive cancer target. PRAME is an intracellular protein that cannot currently be drugged. After proteasomal processing, the PRAME.sup.300-309 peptide ALYVDSLFFL (ALY) is presented in the context of human leukocyte antigen HLA-A.sup.*02:01 molecules for recognition by the T cell receptor (TCR) of cytotoxic T cells. Here, we have described Pr20, a TCR mimic (TCRm) human IgG1 antibody that recognizes the cell-surface ALY peptide/HLA-A2 complex. Pr20 is an immunological tool and potential therapeutic agent. Pr20 bound to PRAME.sup.+HLA-A2.sup.+ cancers. An afucosylated Fc form (Pr20M) directed antibody-dependent cellular cytotoxicity against PRAME.sup.+HLA-A2.sup.+ leukemia cells and was therapeutically effective against mouse xenograft models of human leukemia. In sometumors, Pr20 binding markedly increased upon IFN-gamma treatment, mediated by induction of the immunoproteasome catalytic subunit beta5i. The immunoproteasome reduced internal destructive cleavages within the ALY epitope compared with the constitutive proteasome. The data provide rationale for developing TCRm antibodies as therapeutic agents for cancer, offer mechanistic insight on proteasomal regulation of tumor-associated peptide/HLA antigen complexes, and yield possible therapeutic solutions to target antigens with ultra-low surface presentation.
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Background: The natural history of sickle cell disease (SCD) includes premature death. Advancements in care, such as penicillin prophylaxis, have made death during childhood uncommon. Prior ...survival estimates for adults with SCD have been limited by survival bias (subjects enrolled as adults) or the limitations of administrative data. No prior study has estimated survival for adults with SCD from a newborn cohort. Herein, we report an continued observation of subjects in the Dallas Newborn Cohort (DNC)1.
Methods: The DNC is an inception cohort of patients with SCD defined by: (1) diagnosis by Texas newborn screening after October 31, 1983; (2) evaluation in the pediatric sickle cell program at UT Southwestern (UTSW) / Children’s Medical Center Dallas (CMCD); and (3) confirmation of SS, SC, Sβ0 or Sβ+ genotype. Patients followed at CMCD are transitioned to adult care before the 19th birthday. To identify transitioned patients or those lost to follow-up (LTFU), we reviewed the medical records at Parkland Memorial Hospital (PMH) and UTSW University Hospital (UH), the public and private hospitals affiliated with CMCD and the most common care-sites for transitioned patients. In addition, for patients no longer actively followed at CMCD, we queried the National Death Index (NDI). The NDI is a centralized database of death records obtained from all state vital statistic bureaus in the US and is maintained by the National Center for Health Statistics. As of our query in April 2014, deaths occurring from 1/1/79 - 12/31/11 were available in the NDI. Probabilistic matching is performed by the NDI to identify subjects using identifiers such as name, social security number, birthdate, gender, state of birth, and race. Patients who were LTFU or transitioned to adult care were censored at 12/31/11 or the date of their last clinical encounter, whichever occurred latest. Patients actively followed at CMCD, PMH, or UH were censored at 12/31/13.
In addition to descriptive statistics, we constructed survival curves using the Kaplan-Meier method to evaluate overall survival and survival by genotype. Also, Cox Proportional Hazard modeling was used to assess genotype, baseline hemoglobin, baseline reticulocyte count (%), and baseline oxygen saturation as potential predictors of early mortality. To test the sensitivity of the NDI to identify deceased individuals, we submitted 36 known deaths from the cohort.
Results: The DNC is now comprised of 1214 patients with 16,636 years of follow-up. The cohort is 49.2% female with a mean age, and duration of follow-up, of 13.2 years (Range 0.1 – 30.0 years). SCD genotypes included 60.5% HbSS, 30.6% HbSC, 6.8% HbSβ+, and 2.1% HbSβ0. Of the 1214 patients in the cohort, 511 were active in the pediatric program, 195 were in the pediatric age range but had moved or were LTFU, 238 were adults with 1 or more visits at PMH or UH, 234 were adults with no subsequent identifiable clinical encounters, and 36 were deceased.
Of 36 known deaths submitted to the NDI, 34 were identified by their databases (94.4% sensitivity). Of the 606 patients who were LTFU or aged out of the pediatric program, only 10 were reported to have died by the NDI. One of them was confirmed to be erroneous because the patient had clinical encounters after the NDI-reported date of death. In addition to these 9 deaths in the NDI, 8 additional patients were known to have died as of 12/31/13, bringing the total number of deaths to 53 (4.3%). Of these, 21 (39.6%) have been identified since the last cohort update in 2007.
Overall survival in the DNC to age 25 years was 91.4% (95% CI: 88.4%, 93.7%) for all members (Figure), 87.7% (95% CI: 83.3%, 91%) for those with HbSS or HbSβ0, and 98.6% (95% CI: 96.7%, 99.4%) for those with HbSC or HbSβ+. In univariate analysis, early mortality was associated with SS or Sβ0 genotypes (compared to HbSC or HbSβ+), lower baseline hemoglobin, and low baseline SpO2 but not gender or reticulocytes. In multivariate analysis, early mortality was associated with only lower baseline hemoglobin (Hazard Ratio 0.76 95% CI 0.59, 0.98, P=0.037).
Conclusions: These are the first modern estimates of survival into adulthood with an unbiased newborn cohort of SCD patients. Early mortality is associated with low baseline hemoglobin. Survival to age 20 remains excellent at 95% but the mortality rate appears to accelerate in early adulthood.
Reference: 1. Quinn et al. Blood. 115(17):3447-52, 2010.
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No relevant conflicts of interest to declare.
Cemiplimab in combination with radiation therapy, cyclophosphamide, and granulocyte macrophage colony-stimulating factor (GM-CSF) did not demonstrate efficacy above what can be achieved with other ...PD-1 inhibitor monotherapies in patients with Refractory and metastatic head and neck squamous cell carcinoma. The safety profile of cemiplimab combination therapy was consistent with previously reported safety profiles of cemiplimab monotherapy. No new safety signal was observed.
Refractory and metastatic (R/M) head and neck squamous cell carcinoma (HNSCC) generally does not respond to PD-1 inhibitor monotherapy. Cemiplimab is a human anti-PD-1 monoclonal antibody. An expansion cohort enrolled patients with R/M HNSCC in a phase I study combining cemiplimab plus radiation therapy (RT), cyclophosphamide, and granulocyte macrophage colony-stimulating factor (GM-CSF).
Patients with R/M HNSCC refractory to at least first-line therapy and for whom palliative RT is clinically indicated received cemiplimab plus RT, cyclophosphamide, and GM-CSF. The co-primary objectives were the safety, tolerability, and efficacy of cemiplimab plus RT, cyclophosphamide and GM-CSF in 15 patients (pts) with R/M HNSCC.
Fifteen patients were enrolled. Patients discontinued treatment due to progression of disease. The most common treatment-emergent adverse events (TEAEs) of any grade (Gr) were fatigue (40.0%), constipation (26.7%), and asthenia, dyspnea, maculo-papular rash, and pneumonia (each 20%). The only Gr ≥3 TEAE that occurred in 2 pts was pneumonia (13.3%). By investigator-assessment, there was 1 partial response (6.7%); disease control rate was 40.0% (95% CI: 16.3-67.7; 5 with stable disease); 7 patients had progressive disease and 2 were not evaluable. Median progression-free survival by investigator-assessment was 1.8 months (95% CI: 1.7-4.7).
The regimen demonstrated tolerability but not efficacy above that which can be achieved with anti-PD-1 inhibitor monotherapy for R/M HNSCC.
The discovery of lymphocytes bearing two light chains in mice carrying self-reactive antibody transgenes has challenged the "one lymphocyte-one antibody" rule. However, the extent and nature of ...allelically included cells in normal mice is unknown. We show that 10% of mature B cells coexpress both Igkappa alleles. These cells are not the result of failure in allelic exclusion per se, but arise through receptor editing. We find that under physiological conditions, editing occurs both by deletion and by inclusion with equal probability. In addition, we demonstrate that B lymphocytes carrying two B-cell receptors are recruited to germinal center reactions, and thus fully participate in humoral immune responses. Our data measure the scope of allelic inclusion and provide a mechanism whereby autoreactive B cells might "escape" central tolerance.
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