Perspectives on the integration of Immuno-Oncology Biomarkers and drugs in a Health Care setting.

Immunotherapies, specifically checkpoint inhibitors, are becoming an important component in cancer care with the most application now in melanoma and lung cancer patients. Some drawbacks that converge with this new evolution are the rather low response rates to these drugs and their high cost with a significant economic impact on the health care system. These major challenges can likely be circumvented by implementing a "personalized immuno-oncology" approach to accomplish a selection of optimal responders based on biomarkers. In this paper we first discuss the legal framework for the development of valuable in vitro diagnostics. Based on a case study in lung cancer, the clinical validity and utility requirements of predictive immuno-oncology biomarkers is highlighted and an overview is given on the evolution towards multiplex or omics-based assays together with its challenges and pitfalls. Finally, some initiatives between the public and private sector are pinpointed to sustain the future access to innovative medicines in cancer therapy at a reasonable cost.

An accelerated, clinical-grade protocol to generate high yields of type 1-polarizing messenger RNA-loaded dendritic cells for cancer vaccination.

BACKGROUND: Many efforts have been devoted to improve the performance of dendritic cell (DC)-based cancer vaccines. Ideally, a DC vaccine should induce robust type 1-polarized T-cell responses and efficiently expand antigen (Ag)-specific cytotoxic T-cells, while being applicable regardless of patient human leukocyte antigen (HLA) type. Production time should be short, while maximally being good manufacturing practice (GMP)-compliant. We developed a method that caters to all of these demands and demonstrated the superiority of the resulting product compared with DCs generated using a well-established "classical" protocol. METHODS: Immunomagnetically purified monocytes were cultured in a closed system for 3 days in GMP-compliant serum-free medium and cytokines, and matured for 24 h using monophosphoryl lipid A (MPLA)+ interferon-gamma (IFN-γ). Mature DCs were electroporated with messenger RNA (mRNA) encoding full-length antigen and cryopreserved. "Classical" DCs were cultured for 8 days in flasks, with one round of medium and cytokine supplementation, and matured with tumor necrosis factor alpha (TNF-α) + prostaglandin E2 (PGE2) during the last 2 days. RESULTS: Four-day MPLA/IFN-γ-matured DCs were superior to 8-day TNF-α/PGE2-matured DCs in terms of yield, co-stimulatory/co-inhibitory molecule expression, resilience to electroporation and cryopreservation and type 1-polarizing cytokine and chemokine release after cell thawing. Electroporated and cryopreserved DCs according to our protocol efficiently present epitopes from tumor antigen-encoding mRNA, inducing a strong expansion of antigen-specific CD8+ T-cells with full cytolytic capacity. CONCLUSION: We demonstrate using a GMP-compliant culture protocol the feasibility of generating high yields of mature DCs in a short time, with a superior immunogenic profile compared with 8-day TNF-α/PGE2-matured DCs, and capable of inducing vigorous cytotoxic T-cell responses to antigen from electroporated mRNA. This method is now being applied in our clinical trial program.

Exposing a deadly alliance: novel insights into the biological links between COPD and lung cancer.

Chronic obstructive pulmonary disease (COPD) affects more than 200 million people worldwide and is expected to become the third leading cause of death in 2020. COPD is characterized by progressive airflow limitation, due to a combination of chronic inflammation and remodeling of the small airways (bronchiolitis) and loss of elastic recoil caused by destruction of the alveolar walls (emphysema). Lung cancer is the most important cause of cancer-related death in the world. (Cigarette) smoking is the principal culprit causing both COPD and lung cancer; in addition, exposure to environmental tobacco smoke, biomass fuel smoke, coal smoke and outdoor air pollution have also been associated with an increased incidence of both diseases. Importantly, smokers with COPD--defined as either not fully reversible airflow limitation or emphysema--have a two- to four-fold increased risk to develop lung cancer. In this review, we highlight several of the genetic, epigenetic and inflammatory mechanisms, which link COPD and carcinogenesis in the lungs. Elucidating the biological pathways and networks, which underlie the increased susceptibility of lung cancer in patients with COPD, has important implications for screening, prevention, diagnosis and treatment of these two devastating pulmonary diseases.

Diagnosing COVID-19; towards a feasible COVID-19 rule-out protocol.

INTRODUCTION: We present the results of the COVID-19 rule-out protocol at Ghent University Hospital, a step-wise testing approach which included repeat NFS SARS-CoV-2 rRT-PCR, respiratory multiplex RT-PCR, low-dose chest CT and bronchoscopy with BAL to confirm or rule-out SARS-CoV-2 infection in patients admitted with symptoms suggestive of COVID-19. RESULTS: Between 19 March 2020 and 30 April 2020, 455 non-critically ill patients with symptoms suspect for COVID-19 were admitted. The initial NFS for SARS-CoV-2 rRT-PCR yielded 66.9%, the second NFS 25.4% and bronchoscopy with BAL 5.9% of total COVID-19 diagnoses. In the BAL fluid, other respiratory pathogens were detected in 65% (13/20) of the COVID-19 negative patients and only in 1/7 COVID-19 positive patients. Retrospective antibody testing at the time around BAL sampling showed a positive IgA or IgG in 42.9 % of the COVID-19 positive and 10.5% of the COVID-19 negative group. Follow-up serology showed 100% COVID-19 positivity in the COVID-19 positive group and 100% IgG negativity in the COVID-19 negative group. CONCLUSION: In our experience, bronchoscopy with BAL can have an added value to rule-in or rule-out COVID-19 in patients with clinical and radiographical high-likelihood of COVID-19 and repeated negative NFS testing. Furthermore, culture and respiratory multiplex PCR on BAL fluid can aid to identify alternative microbial etiological agents in this group. Retrospective analysis of antibody development in this selected group of patients suggests that the implementation of serological assays in the routine testing protocol will decrease the need for invasive procedures like bronchoscopy.

Specific migratory dendritic cells rapidly transport antigen from the airways to the thoracic lymph nodes.

Antigen transport from the airway mucosa to the thoracic lymph nodes (TLNs) was studied in vivo by intratracheal instillation of fluorescein isothiocyanate (FITC)-conjugated macromolecules. After instillation, FITC(+) cells with stellate morphology were found deep in the TLN T cell area. Using flow cytometry, an FITC signal was exclusively detected in CD11c(med-hi)/major histocompatibility complex class II (MHCII)(hi) cells, representing migratory airway-derived lymph node dendritic cells (AW-LNDCs). No FITC signal accumulated in lymphocytes and in a CD11c(hi)MHCII(med) DC group containing a CD8 alpha(hi) subset (non-airway-derived [NAW]-LNDCs). Sorted AW-LNDCs showed long MHCII(bright) cytoplasmic processes and intracytoplasmatic FITC(+) granules. The fraction of FITC(+) AW-LNDCs peaked after 24 h and had reached baseline by day 7. AW-LNDCs were depleted by 7 d of ganciclovir treatment in thymidine kinase transgenic mice, resulting in a strong reduction of FITC-macromolecule transport into the TLNs. Compared with intrapulmonary DCs, AW-LNDCs had a mature phenotype and upregulated levels of MHCII, B7-2, CD40, and intracellular adhesion molecule (ICAM)-1. In addition, sorted AW-LNDCs from FITC-ovalbumin (OVA)-instilled animals strongly presented OVA to OVA-TCR transgenic T cells. These results validate the unique sentinel role of airway DCs, picking up antigen in the airways and delivering it in an immunogenic form to the T cells in the TLNs.

Treatment of extensive-stage small cell lung carcinoma: current status and future prospects.

Small cell lung cancer (SCLC) is an aggressive lung tumour strongly associated with cigarette smoking, with patients often presenting with metastatic disease at the time of diagnosis. Although SCLC is very chemoradiosensitive and high response rates are obtained with treatment, relapse rates are high and the prognosis remains very poor. In limited-stage SCLC, the overall survival rate has been significantly improved by adding dose-hyperfractionated thoracic radiotherapy and prophylactic cranial irradiation to systemic chemotherapy. In contrast, little progress has been made in the treatment of extensive-stage SCLC (ES-SCLC), apart from the recently documented survival gain by the addition of prophylactic cranial irradiation. First-line therapy in ES-SCLC currently consists of chemotherapy, combining a platinum drug with either etoposide or irinotecan as a possible alternative. New treatments are needed in order to improve the prognosis of ES-SCLC, as median survival with current standard treatment is still only 9-10 months from diagnosis. The present review focuses on the management of ES-SCLC, with special attention to the development of new treatment options.

[Other thoracic cancers. Small-cell lung cancer: an update]. / Le carcinome bronchique à petites cellules: traitement de la maladie disséminée.

Small cell lung cancer (SCLC) is an aggressive tumor with a strong association to heavy tobacco smoking. The majority of cases is metastasized at initial clinical presentation. SCLC is an extremely chemosensitive neoplasm and the combination of a platinum-derivative with etoposide represents the chemotherapeutic standard in terms of effectiveness and toxicity profile. Recently, extended stages of SCLC have benefited from the addition of prophylactic pancranial irradiation, resulting in a considerable gain in survival. Unfortunately, with its high rate of relapse, SCLC remains a tumor with a dismal prognosis, while the road towards new effective therapies is paved with failures and disillusion. Nevertheless, new chemotherapeutic agents and several targeted molecular therapies are being evaluated, in an effort to hopefully achieve the "holy Grail" of treatment in SCLC: that is a long term consolidation of a promising initial response.

Translocation t(8;10)(q12;p14) in lymphoproliferative disorders.

Cytogenetic studies were performed in two patients with a B-cell lymphoproliferative disease in progressive phase characterized by leukocytosis and important splenomegaly. In both patients an identical chromosome marker derived from a t(8;10)(q12;p14) translocation was found. This chromosome anomaly was associated with other abnormalities characteristic for lymphoid malignancies, i.e. a structural rearrangement of the long arm of chromosome 11 and a 14q+ in both patients. This new t(8;10) translocation may be a marker of accelerated phase in lymphoid disorders.

Indirect stimulation of B-cell proliferation in vitro by T cells, as evidenced by cytogenetic analysis of PHA-stimulated cell cultures of B-cell lymphomas.

In a retrospective study of non-Hodgkin's lymphomas, the 14q+ marker was found in at least one of the samples examined from 17 patients with B-cell lymphoproliferative diseases (LPD). In the PHA-stimulated cultures, the marker was found in each sample in 10%-100% of the cells. An indirect stimulation, as indicated by a 3H-thymidine incorporation and IG secretion, of normal B cells by a T-cell mitogen, such as PHA, has been recently documented. This phenomenon is confirmed by our chromosome analysis, which demonstrated characteristic chromosome changes in PHA-stimulated cultures of patients with B-cell malignancies and indicated that the phenomenon can be observed not only in normal B cells but also in malignant B cells.

Preliminary data on the in vitro proliferation pattern and karyotypic characteristics in cells of patients with ANLL.

Preliminary results of an in vitro study of the proliferation characteristics and karyotypes in the cells of a series of 20 patients with ANLL are presented. The leukemic cells exhibited in vitro proliferation patterns of essentially three types, all of which were very different from the proliferation pattern of normal bone marrow. Correlation with the karyotypic findings did not confirm that the cell cycle may be longer in aneuploid cells. To the contrary, a substantial part of the ANLL presented a karyotype that was normal by routine banding technique standards, in spite of the presence of a strikingly abnormal proliferation pattern of the dividing cells. These observations are important for a better understanding of the significance of chromosome anomalies in the development of leukemia, as well as for the clinician who, in the absence of chromosome markers, may use the proliferation pattern in the monitoring of leukemic patients.

Reciprocal translocation t(6;9)(p21;q33): a new characteristic chromosome anomaly in myeloid leukemias.

Three patients with myeloproliferative disorders, two acute myeloid leukemias and one chronic myelocytic leukemia, were found to present a t(6;9)(p21;q33) as the sole anomaly in the leukemic cells. It is likely that this translocation is to be added to the steadily growing list of characteristic chromosome changes in human malignancy.

High incidence of chromosome abnormalities in IgG3 myeloma.

Chromosomes were studied in 33 untreated myeloma patients, and results were correlated with the class of Ig secreted by the myeloma cells. A high incidence of clonal karyotypic anomalies seemed to be present in IgG3 myeloma patients, in whom the disease was advanced at diagnosis and rapidly progressing. Among the chromosome anomalies, the t(11;14)(q14;q32) was particularly prominent, and this chromosome anomaly, in analogy with the Ph1 chromosome, may characterize a family of lymphoproliferative disorders.

The 5q-anomaly.

A deletion of the long arm of chromosome #5 (5q-) occurs nonrandomly in human malignancies. As a rule, the deletion is interstitial; the distal breakpoint by conventional techniques is usually in band q32, the proximal breakpoints in q12 or q14. Variant breakpoints occur in less than 10% of all cases. As the sole anomaly, 5q- is characteristically found in refractory anemia with or without excess of blasts. It can occur as the sole anomaly in de novo or secondary acute nonlymphocytic leukemia, but is usually accompanied in those disorders by other chromosome changes that are also nonrandomly distributed. In addition, it can be found in lymphoproliferative disorders, and occasionally, also in solid tumors. The 5q- myelodysplastic syndrome typically occurs in older age groups, particularly in females. Characteristic features are macrocytic anemia, normal or elevated platelets in the presence of megakaryocytic anomalies, and a mild clinical course. In cases with 5q- only, transformation into ANLL occurs rarely. Additional chromosome anomalies and male sex are prognostically unfavorable signs. Sex ratio is also at the disadvantage of females in de novo 5q- ANLL, and the latter disorder can occur without being preceded by a myelodysplastic phase. A myelodysplastic phase usually precedes 5q- secondary leukemia, in males as well as in females, and additional chromosome anomalies, especially of chromosome #7, are almost invariably present in those cases. We conclude that 5q- is the most frequently occurring single chromosome anomaly in secondary leukemia. Furthermore, the resemblance between de novo and secondary 5q- MDS and ANLL is striking; clinically, as well as cytogenetically, they are indistinguishable, suggesting that all de novo cases may be due to environmental (chemical) carcinogens. Response to treatment and prognosis are very poor with current therapeutic regimens in de novo as well as in secondary 5q- ANLL. Morphologically, these ANLLs fall into all FAB categories. There is considerable evidence to show that the 5q- anomaly occurs in a myeloid precursor stem cell. The occasional occurrence in lymphoid malignancies, of B cell as well as T cell type, suggests that, as in Ph-positive disorders, a common progenitor stem cell may be affected in 5q- also. The 5q- lymphoid malignancies, however, are much more rare; it is not clear at the present time whether or not a 5q- counterpart of Ph-positive ALL exists, and mixed lymphoid-myeloid 5q- disorders have not yet been documented.(ABSTRACT TRUNCATED AT 400 WORDS)

Interstitial 9q- deletions in hematologic malignancies.

Sixteen patients with interstitial deletions of the long arm of chromosome #9 (9q-). were studied. From our observations and the findings of ten other cases in the literature, it can be deduced that the anomaly is almost exclusively found in myeloproliferative disorders and that it rarely occurs as the sole anomaly; however, in more than one-third of the cases, it was associated with a t(8;21) and occurred as a secondary event. The deletion appears to be interstitial, the breakpoints are somewhat variable, and the region carrying the abl oncogene was never involved.

11q-chromosome is associated with abnormal iron stores in myelodysplastic syndromes.

Nine cases of myelodysplastic syndrome with a deletion of the long arm of chromosome #11 (11q-) showed ringed sideroblasts, and three of which had an acquired sideroblastic anemia according to the criteria of the FAB classification. In contrast, among four cases of myelodysplastic syndromes with translocation of extra material to the long arm of chromosome #11 (11q+), only one showed bone marrow sideroblasts. These results strongly indicate that an 11q- chromosome is a marker of iron overload in myelodysplastic syndromes. Within the cases of 11q- associated with sideroblastosis, two cytogenetically different anomalies (i.e., terminal or interstitial deletions) were delineated.

Pentasomy 21 characterizing spontaneously regressing congenital acute leukemia.

Pentasomy 21 was found to characterize the proliferating cells in a case of transient congenital acute leukemia (or congenital acute leukemia) with spontaneous remission. The patient was phenotypically normal, and cytogenetically no evidence could be found for the existence of a mosaic with a normal cell line and one with more than two No 21 chromosomes. The importance of these findings is discussed.

5q- anomaly in a patient with disseminated teratoma.

A 5q- anomaly associated with other chromosome anomalies was found in the infiltrated bone marrow of a patient with a highly malignant teratoma originally located in the mediastinum. There was no evidence of a second malignancy, and it is likely that the 5q- anomaly was, indeed, associated with the malignant teratoma cells.

3q-, 3q+ anomaly in malignant proliferations in humans.

Anomalies of both No. 3 chromosomes, of the t(3q-; 3q+) type can be observed in human malignancy as reported previously. It is our experience that this anomaly is found predominantly in myeloproliferative disorders, as a rather rare event, though occurring more frequently than similar exchanges between other homologous chromosomes. Previous claims about a relationship between this anomaly and thrombocytosis could not be confirmed, but the features found in a few patients indicate that further research should be undertaken to clarify this point.

Anomalies of the long arm of chromosome 11 in human myelo- and lymphoproliferative disorders. I. Acute nonlymphocytic leukemia.

In a series of 365 consecutive ANLL cases of which 45.1% had abnormal karyotypes, 13 cases were detected with a structural abnormality of the long arm of chromosome 11. Besides one isochromosome 11q, there were six deletions and six translocations. Of these 12 patients, seven had acute monocytic leukemia (FAB-type M5), two had an M4, two had an M2, and one case of secondary leukemia had an M3-like disorder. Similar results with regard to the type of leukemia were obtained upon analysis of 41 cases of ANLL with an 11q anomaly described in the literature. This study confirms that a high proportion of acute monocytic leukemias and a lesser proportion of acute myelomonocytic leukemias are characterized by an 11q anomaly, mostly involving bands q22 and/or q23. Acute monocytic leukemia with an 11q structural anomaly appears to have a poor prognosis.