Human leucocyte antigen genotype association with the development of immune-related adverse events in patients with non-small cell lung cancer treated with single agent immunotherapy

Introduction: Biomarkers that predict the risk of immune-mediated adverse events (irAEs) among patients with non-small cell lung cancer (NSCLC) may reduce morbidity and mortality associated with these treatments. Methods: We carried out high resolution human leucocyte antigen (HLA)-I typing on 179 patients with NSCLC treated with anti-program death (PD)-1/program death ligand (PDL)-1. Toxicity data were collected and graded as per common terminology criteria for adverse event (CTCAE) v5.0. We used 14.8-week for landmark analysis to address lead-time bias to investigate the correlation between HLA-I/II zygosity, supertypes and alleles with irAE. Furthermore, we assessed the association for irAE with clinical benefit rate (CBR), progression-free survival (PFS) and overall survival (OS). Results: Homozygosity at one or more HLA-I loci, but not HLA-II, was associated with a reduced risk of irAE (relative risk (RR) = 0.61, 95% CI 0.33–0.95, P = 0.035) especially pneumonitis or any grade 3 toxicity. Patients with HLA-A03 supertype had a higher risk of developing irAE (RR = 1.42, 95% CI 1.02–2.01, P = 0.039). The occurrence of any irAE was significantly associated with improved CBR (RR = 1.48, P \u3c 0.0001), PFS (HR = 0.45, P = 0.0003) and OS (HR = 0.34, P \u3c 0.0001). Conclusions: Homozygosity at one or more HLA-I loci may serve as biomarker to predict patients who are unlikely to experience severe irAEs among patients with NSCLC and treated with anti-PD1/PDL1, but less likely to derive clinical benefit. Patients with HLA-I homozygous might benefit from additional therapy

Tumour PD-L1 expression in small-cell lung cancer: A systematic review and meta-analysis

: Antibodies against programmed death-1 (PD-1), and its ligand, (PD-L1) have been approved recently for the treatment of small-cell lung cancer (SCLC). Although there are previous reports that addressed PD-L1 detection on tumour cells in SCLC, there is no comprehensive meta-analysis on the prevalence of PD-L1 expression in SCLC. We performed a systematic search of the PubMed, Cochrane Library and EMBASE databases to assess reports on the prevalence of PD-L1 expression and the association between PD-L1 expression and overall survival (OS). This meta-analysis included 27 studies enrolling a total of 2792 patients. The pooled estimate of PD-L1 expression was 26.0% (95% CI 17.0-37.0), (22.0% after removing outlying studies). The effect size was significantly heterogeneous (I2 = 97.4, 95% CI: 95.5-98.5, p \u3c 0.0001).Positive PD-L1 expression was a favourable prognostic factor for SCLC but not statistically significant (HR = 0.86 (95% CI (0.49-1.50), p = 0.5880; I2 = 88.7%, p \u3c 0.0001). Begg\u27s funnel plots and Egger\u27s tests indicated no publication bias across included studies (p \u3e 0.05). Overall, there is heterogeneity in the prevalence of PD-L1 expression in SCLC tumour cells across studies. This is significantly moderated by factors such as immunohistochemistry (IHC) evaluation cut-off values, and assessment of PD-L1 staining patterns as membranous and/or cytoplasmic. There is the need for large size, prospective and multicentre studies with well-defined protocols and endpoints to advance the clinical value of PD-L1 expression in SCLC

Role of serum vascular endothelial growth factor (VEGF) as a potential biomarker of response to immune checkpoint inhibitor therapy in advanced melanoma: results of a pilot study

Background: The development of biomarkers predictive of response to immune checkpoint inhibitor (ICI) therapies in advanced melanoma is an area of great interest in oncology. Our study evaluated the potential role of serum vascular endothelial growth factor (VEGF) as a predictive biomarker of clinical benefit and response to treatment with ICIs. Methods: Pre-treatment peripheral blood samples were obtained from advanced melanoma patients undergoing ICI therapy as monotherapy or in combination at two tertiary care hospitals in Western Australia. Serum VEGF levels were correlated with response to therapy and survival outcomes. Results: Serum VEGF samples were collected from a total of 130 patients treated with ICI therapy (pembrolizumab 73, ipilimumab 15, and ipilimumab/nivolumab combination 42). Median serum VEGF level was significantly higher in the non-responders (82.15 pg/mL) vs. responders (60.40 pg/mL) in the ipilimumab monotherapy cohort (P \u3c 0.0352). However, no difference was seen in VEGF levels between non-responders and responders in pembrolizumab and ipilimumab/nivolumab treated patients. Conclusions: The results of our study confirm previous observations that that high pre-treatment serum VEGF levels in advanced melanoma patients may predict poor response to ipilimumab. However, serum VEGF is not predictive of outcome in patients treated with anti-PD-1 agents alone or in combination with ipilimumab

Association of computed tomography measures of muscle and adipose tissue and progressive changes throughout treatment with clinical endpoints in patients with advanced lung cancer treated with immune checkpoint inhibitors

To investigate the association between skeletal muscle mass and adiposity measures with disease-free progression (DFS) and overall survival (OS) in patients with advanced lung cancer receiving immunotherapy, we retrospectively analysed 97 patients (age: 67.5 ± 10.2 years) with lung cancer who were treated with immunotherapy between March 2014 and June 2019. From computed tomography scans, we assessed the radiological measures of skeletal muscle mass, and intramuscular, subcutaneous and visceral adipose tissue at the third lumbar vertebra. Patients were divided into two groups based on specific or median values at baseline and changes throughout treatment. A total number of 96 patients (99.0 %) had disease progression (median of 11.3 months) and died (median of 15.4 months) during follow-up. Increases of 10 % in intramuscular adipose tissue were significantly associated with DFS (HR: 0.60, 95 % CI: 0.38 to 0.95) and OS (HR: 0.60, 95 % CI: 0.37 to 0.95), while increases of 10 % in subcutaneous adipose tissue were associated with DFS (HR: 0.59, 95 % CI: 0.36 to 0.95). These results indicate that, although muscle mass and visceral adipose tissue were not associated with DFS or OS, changes in intramuscular and subcutaneous adipose tissue can predict immunotherapy clinical outcomes in patients with advanced lung cancer

Powering single-cell genomics to unravel circulating tumour cell subpopulations in non-small cell lung cancer patients

Background Circulating tumour cells (CTCs) are attractive “liquid biopsy” candidates that could provide insights into the different phenotypes of tumours present within a patient. The epithelial-to-mesenchymal transition (EMT) of CTCs is considered a critical step in tumour metastasis; however, it may confound traditional epithelial feature-based CTC isolation and detection. We applied single-cell copy number alteration (CNA) analysis for the identification of genomic alterations to confirm the neoplastic nature of circulating cells with only mesenchymal phenotypes. Methods We isolated CTCs from blood samples collected from 46 NSCLC patients using the Parsortix system. Enriched cells were subjected to immunofluorescent staining for CTC identification using a multi-marker panel comprising both epithelial and mesenchymal markers. A subset of isolated CTCs was subjected to whole genome amplification (WGA) and low-pass whole-genome sequencing (LP-WGS) for the analysis of copy number alterations (CNAs). Results CTCs were detected in 16/46 (34.8%) patients, inclusive of CK+/EpCAM+ CTCs (3/46, 6.5%) and Vim+ CTCs (13/46, 28.3%). Clusters of Vim+ cells were detected in 8 samples, which constitutes 50% of the total number of NSCLC patients with CTCs. No patients had detectable hybrid CK+/EpCAM+/Vim+ cells. All of the tested CK+/EpCAM+ CTCs and 7/8 Vim+ CTCs or CTC clusters carried CNAs confirming their neoplastic nature. Notably, the Vim+ cluster with no CNAs was characterised by spindle morphology and, therefore, defined as normal mesenchymal circulating cells. Conclusion Our results revealed that CK-negative, vimentin-expressing cells represent a large proportion of CTCs detected in NSCLC patients, which are likely missed by standard epithelial-marker-dependent CTC categorisation

Prognostic value of HLA-I homozygosity in patients with non-small cell lung cancer treated with single agent immunotherapy

Background We aimed to assess the impact of genomic human leukocyte antigen (HLA)-I/II homozygosity on the survival benefit of patients with unresectable locally advanced, metastatic non-small lung cancer treated by single-agent programmed cell death protein-1/programmed death ligand 1 (PD1/PDL1) inhibitors. Methods We collected blood from 170 patients with advanced lung cancer treated with immunotherapy at two major oncology centers in Western Australia. Genomic DNA was extracted from white blood cells and used for HLA-I/II high-resolution typing. HLA-I/II homozygosity was tested for association with survival outcomes. Univariable and multivariable Cox regression models were constructed to determine whether HLA homozygosity was an independent prognostic factor affecting Overall Survival (OS) and Progression Free Survival (PFS). We also investigated the association between individual HLA-A and -B supertypes with OS. Results Homozygosity at HLA-I loci, but not HLA-II, was significantly associated with shorter OS (HR=2.17, 95% CI 1.13 to 4.17, p=0.02) in both univariable and multivariable analysis. The effect of HLA-I homozygosity in OS was particularly relevant for patients with tumors expressing PDL1 ≥ 50% (HR=3.93, 95% CI 1.30 to 11.85, p \u3c 0.001). The adverse effect of HLA-I homozygosity on PFS was only apparent after controlling for interactions between PDL1 status and HLA-I genotype (HR=2.21, 95% CI 1.04 to 4.70, p=0.038). The presence of HLA-A02 supertype was the only HLA-I supertype to be associated with improved OS (HR=0.56, 95% CI 0.34 to 0.93, p=0.023). Conclusion Our results suggest that homozygosity at ≥ 1 HLA-I loci is associated with short OS and PFS in patients with advanced non-small cell lung cancer with PDL1 ≥ 50% treated with single-agent immunotherapy. Carriers of HLA-A02 supertype reported better survival outcomes in this cohort of patients

A retrospective study for prognostic significance of type II diabetes mellitus and hemoglobin A1c levels in non-small cell lung cancer patients treated with pembrolizumab

Background: Diabetes mellitus (DM) is common and recognized as a risk factor for developing non-small cell lung cancer (NSCLC) while the prognostic evaluation is still controversial. As immunotherapy is widely used in clinical practice, its efficacy and survival should be investigated in patients with DM. Methods: We retrospectively recruited 266 locally advanced and metastatic NSCLC patients who received pembrolizumab alone or in combination with chemotherapy. Patients\u27 clinicopathological data, including age, history of DM, hemoglobin A1c (HbA1c), genetic tumor profiling, and survival data were collected. Associations between clinical characteristics and survival were evaluated by univariate and multivariate analyses. Results: In this cohort, 15.04 % (40/266) of the patients had a history of DM. Fifty-nine (22.2 %) patients had a HbA1c level ≥ 6.5 %. A total of 169 (63.5 %) patients received 1st-line therapy, and 97 (36.5 %) received 2nd- or subsequent-line therapy. Patients with high ( ≥ 6.5 %) HbA1c and lower ( \u3c 35 g/L) albumin levels at baseline had worse survivals, and epidermal growth factor receptor (EGFR) mutants significantly associated with worse outcomes at normal HbA1c ( \u3c 6.5%) levels (all P \u3c 0.05). Among the 1st-line therapy patients, a higher HbA1c level ( ≥ 6.5 %) at baseline indicated a worse overall survival (OS) (2-year survival rate: 31.25 % vs. 27.03 %, P = 0.045), tumor protein p53 (TP53) alternations and high programmed death-ligand 1 (PD-L1) expression ( ≥ 50 %) were significantly associated with better outcomes (P \u3c 0.05). For 2nd- or subsequent-line patients, EGFR mutants and non-squamous carcinomas (non-SCs) indicated worse survivals, and the normal peripheral blood markers of the carcinoembryonic antigen (CEA), C-reactive protein (CRP), albumin levels were favorable prognostic factors for survivals. In non-SCs, Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations, high PD-L1 expression, and normal alkaline phosphatase (ALP) levels favored better progression-free survival (PFS), while EGFR mutants indicated poor PFS (P \u3c 0.05). Conclusions: Among patients treated with 1st-line immunotherapy, a higher HbA1c level ( ≥ 6.5 %) indicated dismal OS, while history of DM, baseline blood glucose levels, and glucose changes during the treatment process were not significantly associated with any of the outcomes

Detection of clinical progression through plasma ctDNA in metastatic melanoma patients: A comparison to radiological progression

Background The validity of circulating tumour DNA (ctDNA) as an indicator of disease progression compared to medical imaging in patients with metastatic melanoma requires detailed evaluation. Methods Here, we carried out a retrospective ctDNA analysis of 108 plasma samples collected at the time of disease progression. We also analysed a validation cohort of 66 metastatic melanoma patients monitored prospectively after response to systemic therapy. Results ctDNA was detected in 62% of patients at the time of disease progression. For 67 patients that responded to treatment, the mean ctDNA level at progressive disease was significantly higher than at the time of response (P \u3c 0.0001). However, only 30 of these 67 (45%) patients had a statistically significant increase in ctDNA by Poisson test. A validation cohort of 66 metastatic melanoma patients monitored prospectively indicated a 56% detection rate of ctDNA at progression, with only two cases showing increased ctDNA prior to radiological progression. Finally, a correlation between ctDNA levels and metabolic tumour burden was only observed in treatment naïve patients but not at the time of progression in a subgroup of patients failing BRAF inhibition (N = 15). Conclusions These results highlight the low efficacy of ctDNA to detect disease progression in melanoma when compared mainly to standard positron emission tomography imaging

Evaluation of Meropenem, Imipenem and Ertapenem Impregnated MacConkey Agar Plates for the Detection of Carbapenem Resistant Enterobacteriaceae

Background: Rapid detection of carbapenem resistant bacteria, in particular, members of the Enterobacteriaceae family (CRE), is of utmost importance for the management of infected or colonized patients. Methods: Three carbapenems; meropenem, imipenem and ertapenem, with two different concentrations (0.5 mg/ml and 1.0 mg/ml), were impregnated in MacConkey agar. The carbapenem impregnated MacConkey agar plates; ([Mac-Mem], [Mac-Imp] and [Mac-Ert]), were then evaluated for the detection of carbapenem resistant Gram-negative bacteria in particular the blaKPC producing Enterobacteriaceae. The Limit of Detection (LOD) of the plates was determined after counting the colonies that grew on the plates after serial logarithmic dilutions of ten. Carbapenem resistant Gram-negative bacteria were prepared in normal saline, inoculated on the different plates and incubated at 35oC for 18-24 hours. The specificity and the shelf-life of the plates were determined by challenging the plates with six ESBL positive members of the Enterobacteriaceae family (K. pneumoniae, Salmonella species, Shigella species, E. coli, Proteus species and Citrobacter species) and one Enterobacter species with the blaAmpC phenotype. Finally, the MacConkey agar plates impregnated with 0.5 mg/ml meropenem were further challenged by incorporating them in the routine Caritas Baby Hospital active surveillance program for the detection of carbapenem resistant bacteria. Results: Of the three carbapenems impregnated plates, Mac-Ert plates gave the lowest number of colony forming units (CFU’s) detected regardless of the concentration of the antibiotic used. This was followed by the Mac-Mem plates which showed an LOD of less than 200 CFU’s for most of the blaKPC positive bacteria tested at both antibiotic concentrations. The worst performance was noted for the Mac-Imp plate regardless of the antibiotic concentration used as a number of carbapenem resistant bacterial strains failed to grow on the plate. The Mac-Mem plates showed the best specificity as none of the ESBL and blaAmpC positive isolates grew on the plates at either antibiotic concentration tested after 18-24hours incubation in ambient air at 35oC. On the other hand, the Mac-Ert plates failed to inhibit the growth of the Citrobacter species tested at both antibiotic concentrations and the Proteus species tested at the 0.5µg/ml antibiotic concentration. The Mac-Imp plates showed poor specificity as both concentrations failed to inhibit the growth of the Proteus, Enterobacter and Citrobacter species evaluated after 18-24 hours incubation in ambient air at 35oC. Of all the plates tested, the 0.5 µg/ml Mac-Mem agar had the best shelf-life of up to one month at 4-8oC. Conclusions: The high specificity and the good selectivity, in addition to the long shelf-life allowed the 0.5µg/ml Mac-Mem agar to be used as a cost effective selective medium for the isolation of carbapenem resistant Gram-negative bacteria, in particular the blaKPC producing members of the Enterobacteriaceae family

Emergence of Klebsiella pneumoniae Carbapenemase (blaKPC-2) in members of the Enterobacteriaceae family in Palestine

Abstract Background: The global spread of carbapenem resistant Enterobacteriaceae (CRE) has limited the physicians’ antimicrobial treatment options of infected patients. CRE’s which carry the Klebsiella pneumonia Carbapenemase (blaKPC) resistance mechanism have been rapidly spreading in many parts of the world, and have been responsible for high patients’ morbidity and mortality. Methods: Two protocols recommended by the Center for Disease Control and Prevention (CDC) were followed to detect CRE’s in Palestine. In addition, the antimicrobial sensitivity patterns for several antibiotic classes were determined for the isolated CRE’s by the disc diffusion method according to the clinical and laboratory standard institute (CLSI) M100-S22 guidelines. The Minimal Inhibitory Concentrations (MICs) of the carbapenem, ertapenem, imipenem and meropenem were determined for all the CRE’s by E-test. The isolates β-lactam resistance mechanisms were further investigated by analyzing 31 different types of β–lactamase genes by polymerase chain reaction (PCR). Results: Four bacterial isolates, 3 Enterobacter cloacae and 1 Klebsiella pneumoniae, were determined to be non-susceptible to one or all of the carbapenems (ertapenem, imipenem and meropenem) tested. All isolates which carried the blaKPC-2 gene showed an extreme drug resistance profile. These isolates were resistant to all β-lactam antibiotics, co-trimoxazole and gentamicin, while susceptible to only amikacin and colistin sulfate. Different combination of plasmid encoded b–lactamase genes (blaTEM, blaSHV, blaOXA-1, blaMIR-1, blaGES-23 and blaKPC-2) were present in these isolates. Of interest, was the isolation of the first E. cloacae strains co-producing the blaKPC-2 and a novel blaGES-23 β-lactamase. Conclusions: The presence of all these plasmid encoded b-lactamase in Palestine is alarming and mandates actions to be taken to control antibiotics usage and the activation of hospital infection control programs in order to prevent the spread of these extremely drug resistant bacteria