Large-Scale Plasma Proteome Epitome Profiling is an Efficient Tool for the Discovery of Cancer Biomarkers.

Current proteomic technologies focus on the quantification of protein levels, while little effort is dedicated to the development of system approaches to simultaneously monitor proteome variability and abundance. Protein variants may display different immunogenic epitopes detectable by monoclonal antibodies. Epitope variability results from alternative splicing, posttranslational modifications, processing, degradation, and complex formation and possesses dynamically changing availability of interacting surface structures that frequently serve as reachable epitopes and often carry different functions. Thus, it is highly likely that the presence of some of the accessible epitopes correlates with function under physiological and pathological conditions. To enable the exploration of the impact of protein variation on the immunogenic epitome first, here, we present a robust and analytically validated PEP technology for characterizing immunogenic epitopes of the plasma. To this end, we prepared mAb libraries directed against the normalized human plasma proteome as a complex natural immunogen. Antibody producing hybridomas were selected and cloned. Monoclonal antibodies react with single epitopes, thus profiling with the libraries is expected to profile many epitopes which we define by the mimotopes, as we present here. Screening blood plasma samples from control subjects (n = 558) and cancer patients (n = 598) for merely 69 native epitopes displayed by 20 abundant plasma proteins resulted in distinct cancer-specific epitope panels that showed high accuracy (AUC 0.826-0.966) and specificity for lung, breast, and colon cancer. Deeper profiling (≈290 epitopes of approximately 100 proteins) showed unexpected granularity of the epitope-level expression data and detected neutral and lung cancer-associated epitopes of individual proteins. Biomarker epitope panels selected from a pool of 21 epitopes of 12 proteins were validated in independent clinical cohorts. The results demonstrate the value of PEP as a rich and thus far unexplored source of protein biomarkers with diagnostic potential.

Comparison of Killing Activity of Micafungin Against Six Candida Species Isolated from Peritoneal and Pleural Cavities in RPMI-1640, 10 and 30% Serum.

Currently echinocandins are recommended in Candida peritonitis and pleuritis. We determined micafungin killing rates (k values) at therapeutic concentrations (0.25-2 mg/L) in RPMI-1640 with and without 10 and 30% serum mimicking in vivo conditions against six Candida species isolated from peritoneal and pleural fluid. In RPMI-1640, micafungin was fungicidal against C. glabrata, C. krusei and C. kefyr within 2.27 ± 10.68, 2.69 ± 10.29 and 3.10 ± 4.41 h, respectively, while was fungistatic against C. albicans, C. tropicalis and C. parapsilosis. In 10% serum, ≥ 0.25, ≥ 0.5, ≥ 0.5 and ≥ 1 mg/L micafungin produced positive k values (killing) for all C. albicans, C. glabrata, C. kefyr and C. krusei, respectively. In 30% serum, 2 mg/L micafungin produced killing against all C. albicans, C. glabrata and C. kefyr isolates, but was ineffective against C. krusei, C. parapsilosis and 2 of 3 C. tropicalis. Micafungin exposure should be increased against non-albicans species to eradicate fungi from peritoneal and pleural cavities.

Postantifungal effect of micafungin against Candida albicans, Candida dubliniensis and Candida africana in the presence and absence of serum.

We compared the micafungin killing rate and postantifungal effect (PAFE) at 4, 16 and 32 mg/L in RPMI- 1640 and in 50% serum against the C. albicans complex. In RPMI-1640 PAFEs were 1.5 - >19.4, 9.7 - >20.1 and 15.9 - >18.5 hours for C. albicans, C. africana and C. dubliniensis, respectively. In 50% serum PAFEs decreased sharply to 0-1.7 hours for all three species; killing rates were always negative. Short growth inhibition without killing in 50% serum suggests that micafungin PAFE has a limited role in the eradication of the C. albicans complex from the bloodstream.

Killing Activity of Micafungin Against Candida albicans, C. dubliniensis and Candida africana in the Presence of Human Serum.

We compared killing activity of micafungin in time-kill experiments in RPMI-1640 with and without 50% serum against Candida albicans, Candida dubliniensis and Candida africana reference strains and clinical isolates. Killing rates (k values) were determined for each strain and concentration. In RPMI-1640 MIC ranges were 0.015-0.03, 0.015-0.03 and 0.015 mg/L against C. albicans, C. dubliniensis and C. africana, respectively. In 50% serum MIC values for the three species increased 16- to 64-fold. In RPMI-1640 micafungin was fungicidal against two of three C. albicans isolates at 16 and 32 mg/L within 14.54 h and fungistatic against all C. africana and C. dubliniensis. Fifty per cent serum significantly decreased the growth rate of C. africana, but not of the other two species; weak in vivo replication ability of C. africana was confirmed in murine model. In 50% serum micafungin at 0.25 and 1 mg/L did not inhibit any of the three species (k values were always negative). Micafungin killing rate in 50% serum at 4, 16 and 32 mg/L was significantly decreased for C. albicans, but increased for C. dubliniensis compared to RPMI-1640. Killing activity of micafungin against C. africana was comparable or higher in 50% serum than in RPMI-1640. Although micafungin is a highly protein-bound drug, it was equally effective against the species of the C. albicans complex in 50% serum at therapeutic trough concentration (4 mg/L). Both in vitro and in vivo data confirmed the low virulence of C. africana compared to the two sibling species.

The role of immunotherapy in early-stage and metastatic NSCLC.

In the past decade we have seen new advances and thus remarkable progress in the therapeutic options for non-small cell lung cancer (NSCLC). Among cytostatic therapies with new approaches in molecularly targeted therapies, we see new developments in a wide range of applications for immunotherapies. In this review we discuss the new potential modalities for the use of immune checkpoint inhibitors (ICIs) in the frontlines, including in early-stage (perioperative) and metastatic settings. The perioperative use of ICIs in both neoadjuvant and adjuvant settings may show benefits for patients. In early-stage NSCLC (from stage IIB and above) a multimodality approach is recommended as the gold standard for the treatment. After surgical resection platinum-based adjuvant chemotherapy has been the standard of care for many years. Based on the benefit of disease-free survival, the approval of adjuvant atezolizumab and adjuvant pembrolizumab was a significant breakthrough. In the metastatic setting, the use of immune checkpoint inhibitors with chemotherapy, regardless of PD-L1 expression or ICI alone (PD-L1 expression equal to or greater than 50%) also improves overall survival and progression-free survival.

Decreasing incidence and mortality of lung cancer in Hungary between 2011 and 2021 revealed by robust estimates reconciling multiple data sources.

Objective: Hungary has repeatedly been shown to have the highest cancer-related mortality and incidence in Europe. Despite lung cancer being the most abundant malignant diagnosis in Hungary, numerous concerns have been raised recently regarding the bias inherent to reported incidence estimates. Re-analysis of reimbursement claims has been suggested previously by our group as an alternative approach, offering revised figures of lung cancer incidence between 2011 and 2016. Leveraging on this methodology, we aimed at updating Hungarian lung cancer incidence estimates with an additional 5 years (2017-2021), including years affected by the COVID-19 pandemic. Additionally, we also attempted to improve the robustness of estimates by taking additional characteristics of the patient pathway into account. Methods: Lung cancer patients between 2011 and 2021 were identified based on reimbursement-associated ICD-10 codes, histology codes and time patterns. Multiple query architectures were tested for sensitivity and compared to official estimates of the Hungarian National Cancer Registry (HNCR). Epidemiological trends were estimated by Poisson-regression, corrected for age and sex. Results: A total of 89,948 lung cancer patients diagnosed in Hungary between 2011 and 2021 have been identified by our study. In 2019 alone, 7,887 patients were diagnosed according to our optimized query. ESP2013 standardized rate was estimated between 92.5/100,000 (2011) and 78.4/100,000 (2019). In 2019, standardized incidence was 106.8/100,000 for men and 59.7/100,000 for women. Up until the COVID-19 pandemic, lung cancer incidence was decreasing by 3.18% (2.1%-4.3%) yearly in men, while there was no significant decrease in women. Young age groups (40-49 and 50-59) featured the largest improvement, but women aged 60-79 are at an increasing risk for developing lung cancer. The COVID-19 pandemic resulted in a statistically significant decrease in lung cancer incidence, especially in the 50-59 age group (both sexes). Conclusion: Our results show that using an optimized approach, re-analysis of reimbursement claims yields robust estimates of lung cancer incidence. According to this approach, the incidence rate of male lung cancer is declining in Hungary, in concordance with the trend observed for lung cancer mortality. Among women aged 60-79, the incidence of lung cancer has risen, requiring more attention in the near future.

Relative Frequency of Paradoxical Growth and Trailing Effect with Caspofungin, Micafungin, Anidulafungin, and the Novel Echinocandin Rezafungin against Candida Species.

Rezafungin is a next-generation echinocandin that has favorable pharmacokinetic properties. We compared the occurrence of paradoxical growth (PG) and trailing effect (TE) characteristics to echinocadins with rezafungin, caspofungin, micafungin and anidulafungin using 365 clinical Candida isolates belonging to 13 species. MICs were determined by BMD method according to CLSI (M27 Ed4). Disconnected growth (PG plus TE) was most frequent with caspofungin (49.6%), followed by anidulafungin (33.7%), micafungin (25.7%), while it was least frequent with rezafungin (16.9%). PG was relatively common in the case of caspofungin (30.1%) but was rare in the case of rezafungin (3.0%). C. tropicalis, C. albicans, C. orthopsilosis and C. inconspicua exhibited PG most frequently with caspofungin, micafungin or anidulafungin. PG never occurred in the case of C. krusei isolates. Against C. tropicalis and C. albicans, echinocandins frequently showed PG after 24 h followed by TE after 48 h. All four echinocandins exhibited TE for the majority of C. auris and C. dubliniensis isolates. Disconnected growth was common among Candida species and was echinocandin- and species-dependent. In contrast to earlier echinocandins, PG was infrequently found with rezafungin.

Poor in vivo efficacy of caspofungin, micafungin and amphotericin B against wild-type Candida krusei clinical isolates does not correlate with in vitro susceptibility results.

We determined micafungin, caspofungin and amphotericin B (AMB) minimum inhibitory concentration (MICs) and killing rates in RPMI-1640 and in RPMI-1640 with 50% serum against three Candida krusei bloodstream isolates. MIC ranges in RPMI-1640 were 0.125-0.25, 0.25 and 0.125-0.5 mg/L, in RPMI-1640 with 50% serum, MICs were 64-128-, 8- and 4-16-fold higher, respectively. In RPMI-1640 micafungin and caspofungin at 1, 4, 16 and 32 mg/L as well as AMB at 2 mg/L were fungicidal against all isolates in ≤3.96, ≤4.42 and 14.96 h, respectively. In RPMI-1640 with 50% serum, caspofungin was fungicidal for all isolates only at 32 mg/L, micafungin and AMB were fungistatic. In neutropenic mice, 5 mg/kg caspofungin and 1 mg/kg AMB were ineffective against two of the three isolates. Thus, in vivo efficacy of echinocandins and AMB is weak or absent against C. krusei. Prescribers treating C. krusei infections with echinocandins should watch out for clinical resistance and therapeutic failure.

Decreased Killing Activity of Micafungin Against Candida guilliermondii, Candida lusitaniae, and Candida kefyr in the Presence of Human Serum.

Currently, echinocandins are first-line drugs for treatment of invasive candidiasis. However, data on how serum influences killing activity of echinocandins against uncommon Candida species are limited. Therefore, the killing activity of micafungin in RPMI-1640 and in 50% serum was compared against Candida guilliermondii, Candida lusitaniae, and Candida kefyr. Minimum inhibitory concentration (MIC) ranges in RPMI-1640 were 0.5-1, 0.12-0.25, and 0.06-0.12 mg/L, respectively. In 50% serum, MICs increased 32- to 256-fold. In RPMI-1640 ≥ 0.25, ≥4, and 32 mg/L micafungin was fungicidal against all four C. kefyr (≤4.04 hours), two of three C. lusitaniae (≤16.10 hours), and two of three C. guilliermondii (≤12.30 hours), respectively. In 50% serum, all three species grew at ≤4 mg/L. Micafungin at 16-32 mg/L was fungicidal against all C. kefyr isolates (≤3.03 hours) and at 32 mg/L was fungistatic against one of three C. lusitaniae isolates. Two C. lusitaniae isolates and all three C. guilliermondii grew at all tested concentrations. Adding human serum to susceptibility test media drew attention to loss of fungicidal or fungistatic activity of micafungin in the presence of serum proteins, which is not predicted by MICs in case of C. kefyr and C. lusitaniae in RPMI-1640. Our results strongly suggest that micafungin and probably other echinocandins should be used with caution against rare Candida species.

Killing Rates of Caspofungin in 50 Percent Serum Correlate with Caspofungin Efficacy Against Candida albicans in a Neutropenic Murine Model.

Previous studies suggested that caspofungin dose escalation against Candida species is more beneficial than currently used lower daily doses. Thus, we determined in vitro and in vivo activity of caspofungin against six wild-type C. albicans clinical isolates, the ATCC 10231 strain and an echinocandin resistant strain. MIC ranges of clinical isolates in RPMI-1640 with and without 50% serum were 0.125-0.25 and 0.015-0.06 mg/L, respectively. Two and three isolates showed paradoxical growth in MIC and time-kill tests, respectively, in RPMI-1640 but not in 50% serum. Caspofungin killing rate (k) in RPMI-1640 at 1 mg/L was higher than at 16 and 32 mg/L for all isolates (p<0.001). Killing rates for five of six isolates were concentration independent between 1-32 mg/L in 50% serum (p>0.05 for all comparisons), but for one isolate k value at 32 mg/L was significantly lower than at 1-16 mg/L. Although k values at 1-32 mg/L showed a great variability in 50% serum (the lowest and highest k value ranges were 0.085-0.109 and 0.882-0.985 1/h, respectively), daily 3, 5 and 15 mg/kg caspofungin was effective in a neutropenic murine model against all isolates, without significant differences between the effective doses. This study confirms that paradoxical growth does not affect the in vivo efficacy of caspofungin. We demonstrated that dose escalation did not increase the efficacy of caspofungin against C. albicans either in vitro or in vivo. These results are in concordance with the clinical experience that efficacy of echinocandins does not increase at larger doses.

Patterns of erythropoiesis-stimulating agent use for chemotherapy-induced anemia in lung cancer: results of a retrospective Hungarian real-life clinical data analysis.

OBJECTIVE: Lung cancer carries a relatively high risk of chemotherapy-induced anemia, one of the most frequent hematological complications. Previous data show a lack of optimal anemia correction in patients with chemotherapy-induced anemia. This paper analyzes real-life data considering the prevalence and severity of chemotherapy-induced anemia, together with the frequency and efficacy of erythropoietin treatment of anemia in Hungarian lung cancer patients. RESEARCH DESIGN AND METHODS: Data of 482 patients with histological or cytological confirmed lung cancer receiving chemotherapy were collected retrospectively between 1 January and 31 December, 2008. In all, 83 (17%) of them developed chemotherapy-induced moderate to severe anemia (44.6% male, 55.4% female; mean age 70 ± 8.6 years; NSCLC 67.5%, small cell lung cancer 32.5%). RESULTS: More than 50% of the patients suffering from moderate to severe chemotherapy-induced anemia (hemoglobin below 10 g/dl) did not receive erythropoietin treatment. Chemotherapy had to be postponed due to anemia in 32.26% of the patients receiving erythropoietin supplementation, while this was seen in 41.94% of the group without erythropoietin treatment (p < 0.05). In patients not receiving erythropoietin, the severity of anemia increased, while erythropoietin treated patients maintained acceptable hemoglobin levels after the end of the chemotherapy. CONCLUSIONS: The data draws attention to the fact that nowadays chemotherapy-induced anemia is not treated according to current guidelines in many lung cancer cases in Hungary.