Causes of death and comorbidities in hospitalized patients with COVID-19

Infection by the new corona virus strain SARS-CoV-2 and its related syndrome COVID-19 has been associated with more than two million deaths worldwide. Patients of higher age and with preexisting chronic health conditions are at an increased risk of fatal disease outcome. However, detailed information on causes of death and the contribution of pre-existing health conditions to death yet is missing, which can be reliably established by autopsy only. We performed full body autopsies on 26 patients that had died after SARS-CoV-2 infection and COVID-19 at the Charite University Hospital Berlin, Germany, or at associated teaching hospitals. We systematically evaluated causes of death and pre-existing health conditions. Additionally, clinical records and death certificates were evaluated. We report findings on causes of death and comorbidities of 26 decedents that had clinically presented with severe COVID-19. We found that septic shock and multi organ failure was the most common immediate cause of death, often due to suppurative pulmonary infection. Respiratory failure due to diffuse alveolar damage presented as immediate cause of death in fewer cases. Several comorbidities, such as hypertension, ischemic heart disease, and obesity were present in the vast majority of patients. Our findings reveal that causes of death were directly related to COVID-19 in the majority of decedents, while they appear not to be an immediate result of preexisting health conditions and comorbidities. We therefore suggest that the majority of patients had died of COVID-19 with only contributory implications of preexisting health conditions to the mechanism of death

Prognostic value of regulatory T cells and T helper 17 cells in high grade serous ovarian carcinoma

Purpose: In recent years the tumor microenvironment and its interaction with the tumor has emerged into research focus with increased attention to the composition of Tumor-infiltrating lymphocytes. We wanted to quantify the composition of Regulatory T cells (Tregs) and T helper 17 cells (Th17 cells) and their prognostic impact in high-grade serous tubo-ovarian carcinoma. Methods: Tregs and Th17 cells were determined by immunohistochemical analysis of CD25 FoxP3 and ROR gamma t, respectively on tissue microarrays of a cohort of 222 patients with reviewed histology and available clinical data. Expression was analyzed with Qupath for quantification and integration with clinical data enabled calculation of prognostic impact. For validation FOXP3 and RORC mRNA expression levels from 502 patients with HGSC in publicly available datasets were evaluated. Results: An average percentage of 0.93 Tregs and of 0.06 Th17 cells was detected per cells in overall tissue. Optimal cut-offs were determined and higher Tregs were associated with a better overall survival in stroma (p = 0.006), tumor area (p = 0.0012) and overall tissue (p = 0.02). After accounting for well-known prognostic factors age at diagnosis, residual tumor and FIGO stage, this association remained significant for stromal Tregs with overall survival (p = 0.02). Survival analysis for Th17 cells revealed no significant association with survival rates. Moreover, lower Th17/Treg ratios had a positive impact on patient overall survival (p = 0.025 tumor, p = 0.049 stroma and p = 0.016 overall tissue). Conclusion: Our results outline a positive prognostic effect for higher Tregs but not for Th17 in high grade serous tubo-ovarian carcinoma

Aldehyde dehydrogenase 1 A1, thymosin-β15A and poly(ADP-Ribose) polymerase-1 as prognostic markers in ovarian carcinoma

Epitheliale Ovarialkarzinome stellen eine häufige gynäkologische Tumorerkrankung mit schlechter Prognose dar. In der vorliegenden Arbeit wurden daher die Marker Aldehyddehydrogenase 1 A1 (ALDH1A1), Thymosin-β15A (TMSB15A) und Poly(ADP-Ribose)-Polymerase-1 (PARP1), welche sich bereits im Mammakarzinom als Biomarker mit prädiktiver und prognostischer Relevanz erwiesen hatten, hinsichtlich ihres prognostischen Wertes in Ovarialkarzinomen untersucht. Der Schwerpunkt lag in der Untersuchung der Marker in Typ II Ovarialkarzinomen. Untersucht wurden insbesondere Korrelationen mit Überlebensdaten, wichtigen klinisch-pathologischen Faktoren sowie eine Änderung der Markerexpression im Krankheitsverlauf. Hierfür wurden Formalin- fixierte und in Paraffin eingebettete Gewebeproben von insgesamt 209 Patientinnen mit primären epithelialen Ovarialkarzinomen sowie 38 korrespondierende Rezidivtumoren untersucht. Auf mRNA-Ebene wurden die Marker mittels einer quantitativen real-time-PCR untersucht. Auf Proteinebene wurden immunhistochemische Färbungen an Tissue Microarrays durchgeführt. Es konnte gezeigt werden, dass die Proteinexpression des potentiellen Krebsstammzellmarkers ALDH1 einen signifikant ungünstigen Prognosefaktor in Typ II Ovarialkarzinomen darstellt. Darüber hinaus zeigten Tumoren die das ALDH1-Protein und den epidermalen Wachstumsfaktorrezeptor (EGFR) exprimierten die statistisch ungünstigsten Verläufe. Das aktinbindende Protein TMSB15, welches die zytoskelettgesteuerte Lokomotion und somit möglicherweise auch die Metastasierungsfähigkeit der Zelle beeinflussen kann, war lediglich univariat ein signifikant günstiger Prognosemarker. Auf mRNA-Ebene konnte kein Einfluss der TMSB15A-Expression auf das Überleben der Patientinnen beobachtet werden. Die PARP1 ist als wichtiger Mediator des Basenexzisionsweg in die DNA- Reparatur involviert. Es konnte weder auf mRNA- noch auf Proteinebene ein Einfluss von PARP1 auf die Prognose festgestellt werden. Gemessene PARP1-mRNA- und Proteinwerte waren jedoch signifikant in Rezidivtumoren gegenüber den korrespondieren Primärtumoren erhöht. Eine Assoziation von ALDH1 und EGFR wird in der Literatur bereits in BRCA1-defizienten Brustepithelien und tripel negativen Mammakarzinomen beschrieben. Die Ergebnisse der vorliegenden Arbeit weisen darauf hin, dass es sich bei ALDH1- und EGFR-exprimierenden Tumoren um eine besonders aggressive Subgruppe von Typ II Ovarialkarzinomen handelt für die alternative Therapien angezeigt sein könnten. Eine weitere Charakterisierung dieser Subgruppe zusammen mit dem BRCA1-Mutationsstatus wäre daher ein interessanter Aspekt für zukünftige Studien. Höhere PARP1-Level in Rezidivtumoren der untersuchten Patientenproben könnten durch eine erhöhte chromosomale Instabilität und somit erhöhte Aktivität des Basenexzisionsweges in diesen Tumoren erklärt werden. Die Inhibierung von PARP in Zellen, in denen die homologe Rekombination beispielsweise durch eine BRCA-Mutation gestört ist, kann zur selektiven Apoptose führen, weswegen die Wirkung von PARP- Inhibitoren derzeit in klinischen Studien untersucht wird. Dennoch ist unklar, welche Patientengruppen tatsächlich von dieser Therapie profitieren und die PARP1-Expression sollte als potentieller prädiktiver Faktor weiter untersucht werden.Epithelial ovarian carcinoma constitutes a frequent gynecologic malignancy with a poor prognosis. As aldehyde dehydrogenase 1 A1 (ALDH1A1), thymosin-β15A (TMSB15A) and poly(ADP-ribose) polymerase-1 (PARP1) have already proved to be prognostic and predictive biomarkers in breast cancer, the aim of this study was to evaluate these biomarkers in ovarian cancer with emphasis on type II ovarian carcinoma. In this context correlations with survival and clinicopathologic parameters were regarded as well as changes in marker expression during the course of disease. This study included 209 formalin- fixed and paraffin-embedded samples of patients with primary epithelial ovarian carcinoma and additional 38 specimens of corresponding relapse tumors. mRNA levels were quantified by quantitative real-time polymerase chain reaction. To determine the protein expression immunohistochemical staining was performed on tissue microarrays. In this study it was demonstrated that the potential cancer stem cell marker ALDH1 was a significant marker of poor prognosis in type II ovarian carcinoma. Furthermore ALDH1- and epidermal growth factor receptor (EGFR)-protein expressing tumors showed most unfavorable outcome compared to the other expression groups. The actin binding protein TMSB15 which is assumed to be involved in cytoskeleton mediated locomotion and metastasis was a significant favorable prognostic marker merely in univariate analysis. There was no association between TMSB15A mRNA expression and survival. As an important mediator of base excision repair pathway PARP1 is involved in DNA repair. Neither PARP1-mRNA nor cytoplasmatic or nuclear PARP-protein expression were associated with prognosis. However, PARP1 mRNA and nuclear protein expression were upregulated in relapse tumors compared to the corresponding primary tumors. A correlation of ALDH1 and EGFR expression in BRCA1-deficient breast epithelium and triple negative breast cancer has already been described in previous observations. The results of this study indicate that ALDH1 and EGFR expressing tumors represent a highly aggressive subgroup of type II ovarian carcinoma for which alternative treatment options might be indicated. Further characterization of this subgroup in combination with the BRCA mutation status represents an interesting subject for future studies. Higher PARP1 levels in relapse tumors might be explained by a higher chromosomal instability in these tumors. Inhibition of PARP in cells with defective homologous recombination due to BRCA mutation, for example, can lead to selective cell death, which is why the effect of PARP inhibitors is investigated in clinical studies to date. However it is still unclear which patients actually benefit from PARP inhibitor therapy so that the PARP1-expression should be further investigated as a predictive factor

APOBEC3B protein expression and mRNA analyses in patients with high-grade serous ovarian carcinoma

APOBEC3 enzymes are part of the innate immune system and they are important in retroviral defense. The number of mutations in ovarian cancer increases with rising levels of APOBEC3B mRNA. We could confirm that APOBEC3B mRNA is upregulated in ovarian cancer cell lines and in ovarian cancer tissue. We evaluated APOBEC3B expression in histologically defined subtypes of ovarian cancer to identify its influence on overall survival (OS) and progression-free survival (PFS). Tissue microarrays from 219 patients with highgrade serous (HGSC), 61 with low-grade serous (LGSC), 62 with endometrioid (EC) and 55 with clear cell (CCC) ovarian carcinoma were stained using an antibody against APOBEC3B. Real-time quantitative PCR was performed to detect APOBEC3B mRNA levels in 274 cases of HGSC, in 11 cases of LGSC, in 47 cases of EC and in 29 cases of CCC. Tumor-infiltrating lymphocytes (TILs) have been evaluated in a previous project. APOBEC3B staining was cytoplasmic as well as nuclear and both were positively correlated (P<0.001). In HGSC a trend was detectable for positive cytoplasmic staining as favorable regarding OS (P=0.283) and PFS (P=0.137). High levels of APOBEC3B mRNA were associated with prolonged PFS in HGSC in univariate analyses (P=0.043) and multivariate analyses (HR 0.55; 95% CI 0.35-0.88; P=0.012). APOBEC3B cytoplasmic staining and APOBEC3B mRNA were positively correlated with TILs. APOBEC3B in HGSC is related to an active immune infiltrate. However, there is no evidence for APOBEC3B as a clinically relevant prognostic biomarker

Low Expression of RGS2 Promotes Poor Prognosis in High-Grade Serous Ovarian Cancer

RGS2 regulates G-protein signaling by accelerating hydrolysis of GTP and has been identified as a potentially druggable target in carcinomas. Since the prognosis of patients with high-grade serous ovarian carcinoma (HGSOC) remains utterly poor, new therapeutic options are urgently needed. Previous in vitro studies have linked RGS2 suppression to chemoresistance in HGSOC, but in situ data are still missing. In this study, we characterized the expression of RGS2 and its relation to prognosis in HGSOC on the protein level by immunohistochemistry in 519 patients treated at Charité, on the mRNA level in 299 cases from TCGA and on the single-cell level in 19 cases from publicly available datasets. We found that RGS2 is barely detectable on the mRNA level in both bulk tissue (median 8.2. normalized mRNA reads) and single-cell data (median 0 normalized counts), but variably present on the protein level (median 34.5% positive tumor cells, moderate/strong expression in approximately 50% of samples). Interestingly, low expression of RGS2 had a negative impact on overall survival (p = 0.037) and progression-free survival (p = 0.058) on the protein level in lower FIGO stages and in the absence of residual tumor burden. A similar trend was detected on the mRNA level. Our results indicated a significant prognostic impact of RGS2 protein suppression in HGSOC. Due to diverging expression patterns of RGS2 on mRNA and protein levels, posttranslational modification of RGS2 is likely. Our findings warrant further research to unravel the functional role of RGS2 in HGSOC, especially in the light of new drug discovery

COVID-19: Autopsy findings in six patients between 26 and 46 years of age

Objectives: Studies on coronavirus disease 2019 (COVID-19) usually focus on middle-aged and older adults. However, younger patients may present with severe COVID-19 with potentially fatal outcomes. For optimized, more specialized therapeutic regimens in this particular patient group, a better understanding of the underlying pathomechanisms is of utmost importance. Methods: Our study investigated relevant, pre-existing medical conditions, clinical histories, and autopsy findings, together with SARS-CoV-2-RNA, determined by qPCR, and laboratory data in six COVID-19 decedents aged 50 years or younger, who were autopsied at the Charite University Hospital. Results: From a total of 76 COVID-19 patients who underwent an autopsy at our institution, six (7.9%) were 50 years old or younger. Most of these younger COVID-19 decedents presented with pre-existing medical conditions prior to SARS-CoV-2 infection. These included overweight and obesity, arterial hypertension, asthma, and obstructive sleep apnea, as well as graft-versus-host disease following cancer and bone marrow transplantation. Furthermore, clinical histories and autopsy results revealed a disproportionally high prevalence of thromboembolism and ischemic organ damage in this patient cohort. Histopathology and laboratory results indicated coagulopathies, signs of immune dysregulation, and liver damage. Conclusions: In conclusion, pre-existing health conditions may increase the risk of severe and fatal COVID-19 in younger patients, who may be especially prone to developing thromboembolic complications, immune dysregulation, and liver damage

Hemicolectomy versus appendectomy for patients with appendiceal neuroendocrine tumours 1–2 cm in size: a retrospective, Europe-wide, pooled cohort study

Background: Awareness of the potential global overtreatment of patients with appendiceal neuroendocrine tumours (NETs) of 1–2 cm in size by performing oncological resections is increasing, but the rarity of this tumour has impeded clear recommendations to date. We aimed to assess the malignant potential of appendiceal NETs of 1–2 cm in size in patients with or without right-sided hemicolectomy. Methods: In this retrospective cohort study, we pooled data from 40 hospitals in 15 European countries for patients of any age and Eastern Cooperative Oncology Group performance status with a histopathologically confirmed appendiceal NET of 1–2 cm in size who had a complete resection of the primary tumour between Jan 1, 2000, and Dec 31, 2010. Patients either had an appendectomy only or an appendectomy with oncological right-sided hemicolectomy or ileocecal resection. Predefined primary outcomes were the frequency of distant metastases and tumour-related mortality. Secondary outcomes included the frequency of regional lymph node metastases, the association between regional lymph node metastases and histopathological risk factors, and overall survival with or without right-sided hemicolectomy. Cox proportional hazards regression was used to estimate the relative all-cause mortality hazard associated with right-sided hemicolectomy compared with appendectomy alone. This study is registered with ClinicalTrials.gov, NCT03852693. Findings: 282 patients with suspected appendiceal tumours were identified, of whom 278 with an appendiceal NET of 1–2 cm in size were included. 163 (59%) had an appendectomy and 115 (41%) had a right-sided hemicolectomy, 110 (40%) were men, 168 (60%) were women, and mean age at initial surgery was 36·0 years (SD 18·2). Median follow-up was 13·0 years (IQR 11·0–15·6). After centralised histopathological review, appendiceal NETs were classified as a possible or probable primary tumour in two (1%) of 278 patients with distant peritoneal metastases and in two (1%) 278 patients with distant metastases in the liver. All metastases were diagnosed synchronously with no tumour-related deaths during follow-up. Regional lymph node metastases were found in 22 (20%) of 112 patients with right-sided hemicolectomy with available data. On the basis of histopathological risk factors, we estimated that 12·8% (95% CI 6·5 –21·1) of patients undergoing appendectomy probably had residual regional lymph node metastases. Overall survival was similar between patients with appendectomy and right-sided hemicolectomy (adjusted hazard ratio 0·88 [95% CI 0·36–2·17]; p=0·71). Interpretation: This study provides evidence that right-sided hemicolectomy is not indicated after complete resection of an appendiceal NET of 1–2 cm in size by appendectomy, that regional lymph node metastases of appendiceal NETs are clinically irrelevant, and that an additional postoperative exclusion of metastases and histopathological evaluation of risk factors is not supported by the presented results. These findings should inform consensus best practice guidelines for this patient cohort. Funding: Swiss Cancer Research foundation

Hemicolectomy versus appendectomy for patients with appendiceal neuroendocrine tumours 1-2 cm in size: a retrospective, Europe-wide, pooled cohort study.

BACKGROUND Awareness of the potential global overtreatment of patients with appendiceal neuroendocrine tumours (NETs) of 1-2 cm in size by performing oncological resections is increasing, but the rarity of this tumour has impeded clear recommendations to date. We aimed to assess the malignant potential of appendiceal NETs of 1-2 cm in size in patients with or without right-sided hemicolectomy. METHODS In this retrospective cohort study, we pooled data from 40 hospitals in 15 European countries for patients of any age and Eastern Cooperative Oncology Group performance status with a histopathologically confirmed appendiceal NET of 1-2 cm in size who had a complete resection of the primary tumour between Jan 1, 2000, and Dec 31, 2010. Patients either had an appendectomy only or an appendectomy with oncological right-sided hemicolectomy or ileocecal resection. Predefined primary outcomes were the frequency of distant metastases and tumour-related mortality. Secondary outcomes included the frequency of regional lymph node metastases, the association between regional lymph node metastases and histopathological risk factors, and overall survival with or without right-sided hemicolectomy. Cox proportional hazards regression was used to estimate the relative all-cause mortality hazard associated with right-sided hemicolectomy compared with appendectomy alone. This study is registered with ClinicalTrials.gov, NCT03852693. FINDINGS 282 patients with suspected appendiceal tumours were identified, of whom 278 with an appendiceal NET of 1-2 cm in size were included. 163 (59%) had an appendectomy and 115 (41%) had a right-sided hemicolectomy, 110 (40%) were men, 168 (60%) were women, and mean age at initial surgery was 36·0 years (SD 18·2). Median follow-up was 13·0 years (IQR 11·0-15·6). After centralised histopathological review, appendiceal NETs were classified as a possible or probable primary tumour in two (1%) of 278 patients with distant peritoneal metastases and in two (1%) 278 patients with distant metastases in the liver. All metastases were diagnosed synchronously with no tumour-related deaths during follow-up. Regional lymph node metastases were found in 22 (20%) of 112 patients with right-sided hemicolectomy with available data. On the basis of histopathological risk factors, we estimated that 12·8% (95% CI 6·5 -21·1) of patients undergoing appendectomy probably had residual regional lymph node metastases. Overall survival was similar between patients with appendectomy and right-sided hemicolectomy (adjusted hazard ratio 0·88 [95% CI 0·36-2·17]; p=0·71). INTERPRETATION This study provides evidence that right-sided hemicolectomy is not indicated after complete resection of an appendiceal NET of 1-2 cm in size by appendectomy, that regional lymph node metastases of appendiceal NETs are clinically irrelevant, and that an additional postoperative exclusion of metastases and histopathological evaluation of risk factors is not supported by the presented results. These findings should inform consensus best practice guidelines for this patient cohort. FUNDING Swiss Cancer Research foundation

Hemicolectomy versus appendectomy for patients with appendiceal neuroendocrine tumours 1–2 cm in size: a retrospective, Europe-wide, pooled cohort study

Background: Awareness of the potential global overtreatment of patients with appendiceal neuroendocrine tumours (NETs) of 1–2 cm in size by performing oncological resections is increasing, but the rarity of this tumour has impeded clear recommendations to date. We aimed to assess the malignant potential of appendiceal NETs of 1–2 cm in size in patients with or without right-sided hemicolectomy. Methods: In this retrospective cohort study, we pooled data from 40 hospitals in 15 European countries for patients of any age and Eastern Cooperative Oncology Group performance status with a histopathologically confirmed appendiceal NET of 1–2 cm in size who had a complete resection of the primary tumour between Jan 1, 2000, and Dec 31, 2010. Patients either had an appendectomy only or an appendectomy with oncological right-sided hemicolectomy or ileocecal resection. Predefined primary outcomes were the frequency of distant metastases and tumour-related mortality. Secondary outcomes included the frequency of regional lymph node metastases, the association between regional lymph node metastases and histopathological risk factors, and overall survival with or without right-sided hemicolectomy. Cox proportional hazards regression was used to estimate the relative all-cause mortality hazard associated with right-sided hemicolectomy compared with appendectomy alone. This study is registered with ClinicalTrials.gov, NCT03852693. Findings: 282 patients with suspected appendiceal tumours were identified, of whom 278 with an appendiceal NET of 1–2 cm in size were included. 163 (59%) had an appendectomy and 115 (41%) had a right-sided hemicolectomy, 110 (40%) were men, 168 (60%) were women, and mean age at initial surgery was 36·0 years (SD 18·2). Median follow-up was 13·0 years (IQR 11·0–15·6). After centralised histopathological review, appendiceal NETs were classified as a possible or probable primary tumour in two (1%) of 278 patients with distant peritoneal metastases and in two (1%) 278 patients with distant metastases in the liver. All metastases were diagnosed synchronously with no tumour-related deaths during follow-up. Regional lymph node metastases were found in 22 (20%) of 112 patients with right-sided hemicolectomy with available data. On the basis of histopathological risk factors, we estimated that 12·8% (95% CI 6·5 –21·1) of patients undergoing appendectomy probably had residual regional lymph node metastases. Overall survival was similar between patients with appendectomy and right-sided hemicolectomy (adjusted hazard ratio 0·88 [95% CI 0·36–2·17]; p=0·71). Interpretation: This study provides evidence that right-sided hemicolectomy is not indicated after complete resection of an appendiceal NET of 1–2 cm in size by appendectomy, that regional lymph node metastases of appendiceal NETs are clinically irrelevant, and that an additional postoperative exclusion of metastases and histopathological evaluation of risk factors is not supported by the presented results. These findings should inform consensus best practice guidelines for this patient cohort. Funding: Swiss Cancer Research foundation