Adipose cells promote resistance of breast cancer cells to trastuzumab-mediated antibody-dependent cellular cytotoxicity.

INTRODUCTION: Trastuzumab has been used in the treatment of human epidermal growth factor receptor 2 (HER2)-expressing breast cancer, but its efficacy is limited by de novo or acquired resistance. Although many mechanisms have been proposed to explain resistance to trastuzumab, little is known concerning the role of the tumor microenvironment. Given the importance of antibody-dependent cellular cytotoxicity (ADCC) in the antitumor effect of trastuzumab and the abundance of adipose tissue in the breast, we investigated the impact of adipocytes on ADCC. METHODS: We set up a coculture system to study the effect of adipocytes on ADCC in vitro. The results were validated in vivo in a mouse xenograft model. RESULTS: We found that adipocytes, as well as preadipocytes, inhibited trastuzumab-mediated ADCC in HER2-expressing breast cancer cells via the secretion of soluble factors. The inhibition of ADCC was not due to titration or degradation of the antibody. We found that adipose cells decreased the secretion of interferon-γ by natural killer cells, but did not alter natural killer cells' cytotoxicity. Preincubation of breast cancer cells with the conditioned medium derived from adipocytes reduced the sensitivity of cancer cells to ADCC. Using a transcriptomic approach, we found that cancer cells undergo major modifications when exposed to adipocyte-conditioned medium. Importantly, breast tumors grafted next to lipomas displayed resistance to trastuzumab in mouse xenograft models. CONCLUSIONS: Collectively, our findings underline the importance of adipose tissue in the resistance to trastuzumab and suggest that approaches targeting the adipocyte-cancer cell crosstalk may help sensitize cancer cells to trastuzumab-based therapy.

Hospital-acquired pneumonia due to Burkholderia cepacia in a thalassemia pregnancy with postoperative eclampsia: A case report.

There are limited case reports on individuals infected with Burkholderia cepacia who do not have typical risk factors, particularly pregnant women with beta-thalassemia. A 34-year-old pregnant female with beta-thalassemia trait and hypertension was admitted to the hospital. The patient was diagnosed with eclampsia and underwent a cesarean section. After two days following the surgery, the patient experienced hospitality-acquired pneumonia. B.cepacia was isolated from blood cultures, and antibiotic susceptibility testing indicated sensitivity to trimethoprim/sulfamethoxazole and levofloxacin. The patient responded to antibiotic treatment. These findings highlight the importance of prompt diagnosis and appropriate treatment in managing B.cepacia infections in pregnant beta-thalassemia patients.

Challenges in diagnosis and management of central airway obstruction: Case series.

Central airway obstruction often presents with respiratory symptoms identical to those of common respiratory diseases. Diagnosis of central airway obstruction is challenging in clinical practice, especially misdiagnosed as asthma or chronic obstructive pulmonary disease in case of "normal" chest X-ray. Here, we reported five central airway obstruction cases: the first and fourth cases misdiagnosed as asthma, the second masquerading as chronic obstructive pulmonary disease exacerbation, the third diagnosed incorrectly with non-resolving pneumonia, and the fifth misdiagnosed as chronic obstructive pulmonary disease. We then analyzed diagnostic clues potentially useful to differentiate central airway obstruction from asthma/chronic obstructive pulmonary disease. A multidisciplinary approach to manage central airway obstruction is essential, particularly selecting judiciously the method of respiratory support due to the high risk of completed airway collapse or obstruction during interventional period.

The First Report of Candida Auris Infection in Vietnam.

Infection caused by Candida auris ha C. auris s rapidly become a global health threat. C. auris created a significant healthcare burden due to various complicating factors, including misidentification by commercial identification methods, potent antifungal resistance, high mortality rates and the possibility of nosocomial outbreaks through direct contact. In Vietnam, there are currently no clinical reports on C. auris infections. Here, we present four clinical cases of C. auris infections in the Department of Pulmonary Medicine of Cho Ray Hospital in southern Vietnam. Through this report, we aim to highlight the attention to the emergence of C. auris in Vietnam. Further research on C. auris infections is warranted, focusing on newly observed clinical characteristics present in all cases in this report, including hypoalbuminaemia and corticosteroid usage. Moreover, one case of resistance to amphotericin B has been identified, possibly due to prior exposure to this antifungal agent. LEARNING POINTS: Further research on Candida auris infections is warranted, focusing on newly observed clinical features present in all cases in this report, including hypoalbuminaemia and corticosteroid use during hospitalisation.While Candida auris remains susceptible to commonly used antifungal drugs, one case of resistance to amphotericin B has been documented, possibly due to prior exposure to this antifungal agent.

Primary Pulmonary T-Cell Lymphoma Mimicking Asthma and Community-Acquired Pneumonia: A Rare Case Report.

Primary pulmonary T-cell lymphoma (PPTL) is a rare disease. Diagnosing PPTL is challenging due to non-specific clinical symptoms and imaging. A 32-year-old female presented with persistent fever, cough, and dyspnoea. The symptoms were initially treated as asthma and community-acquired pneumonia without improvement. Chest computed tomography (CT) revealed bilateral consolidations with a CT angiogram sign, and flexible bronchoscopy showed infiltrative lesions causing bronchial stenosis. Histopathological examination of the tissue biopsy identified T-cell lymphoma through immunohistochemical staining positive for CD3. This case highlights the importance of considering differential diagnoses such as PPTL in patients with atypical presentations of asthma or non-resolving pneumonia. This case also demonstrates the diagnostic utility of flexible bronchoscopy in identifying airway obstruction due to malignant cells, which can mimic asthma. LEARNING POINTS: Primary pulmonary T-cell lymphoma can manifest as atypical asthma and non-resolving pneumonia, making early diagnosis challenging.Malignant aetiologies, including primary pulmonary T-cell lymphoma, should be considered in cases of bilateral consolidations that do not respond to antibiotics and present CT angiogram signs.Histopathology remains the gold standard in primary pulmonary T-cell lymphoma diagnosis, wherein flexible bronchoscopy should be employed as a minimally invasive first-line approach for tissue biopsy.

The Role and Associated Factors of Liquid-Based Cytology of Bronchoalveolar Lavage Fluid in Lung Cancer Diagnosis: A Prospective Study.

Background Liquid-based cytology (LBC) has shown advantages over conventional smears (CS), but previous applications in bronchoalveolar lavage (BAL) fluid have produced inconsistent results. This study compared LBC and CS for diagnosing lung cancer using BAL fluid. Methodology A prospective study was conducted on 92 patients suspected of having lung cancer. All patients underwent bronchoscopy and had a final diagnosis confirmed by histopathology of lesions tissue through biopsy. The study aimed to assess the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of the two cytological methods, in a pair-wise fashion. In addition, the study evaluated the correlation of factors, such as the volume of fluid used in LBC and bronchoscopy lesion morphology, with the sensitivity of LBC. Results The study involved 78 participants who were diagnosed with lung cancer. The sensitivity, specificity, PPV, and NPV of LBC were 25.7%, 100%, 100%, and 19.4%, respectively, whereas those of CS were 15.4%, 85.7%, 85.7%, and 15.4%, respectively. Although the sensitivity of LBC was higher than that of CS, the difference was not statistically significant (p=0.096, McNemar test). Furthermore, the median fluid volume performed during LBC in patients with positive results was significantly higher than in those with negative results (p=0.001, Mann-Whitney U test). Conclusions The application of LBC to BAL fluid has demonstrated similar and potentially superior diagnostic accuracy compared to CS in detecting lung cancer. It is recommended that further investigation be undertaken to examine the relationship between the volume of fluid utilized during the LBC process and its diagnostic accuracy to enhance its sensitivity.

Chronic TCR-MHC (self)-interactions limit the functional potential of TCR affinity-increased CD8 T lymphocytes.

BACKGROUND: Affinity-optimized T cell receptor (TCR)-engineered lymphocytes targeting tumor antigens can mediate potent antitumor responses in cancer patients, but also bear substantial risks for off-target toxicities. Most preclinical studies have focused on T cell responses to antigen-specific stimulation. In contrast, little is known on the regulation of T cell responsiveness through continuous TCR triggering and consequent tonic signaling. Here, we addressed the question whether increasing the TCR affinity can lead to chronic interactions occurring directly between TCRs and MHC-(self) molecules, which may modulate the overall functional potency of tumor-redirected CD8 T cells. For this purpose, we developed two complementary human CD8 T cell models (i.e. HLA-A2 knock-in and knock-out) engineered with incremental-affinity TCRs to the HLA-A2/NY-ESO-1 tumor antigen. METHODS: The impact of HLA-A2 recognition, depending on TCR affinity, was assessed at the levels of the TCR/CD3 complex, regulatory receptors, and signaling, under steady-state conditions and in kinetic studies. The quality of CD8 T cell responses was further evaluated by gene expression and multiplex cytokine profiling, as well as real-time quantitative cell killing, combined with co-culture assays. RESULTS: We found that HLA-A2 per se (in absence of cognate peptide) can trigger chronic activation followed by a tolerance-like state of tumor-redirected CD8 T cells with increased-affinity TCRs. HLA-A2pos but not HLA-A2neg T cells displayed an activation phenotype, associated with enhanced upregulation of c-CBL and multiple inhibitory receptors. T cell activation preceded TCR/CD3 downmodulation, impaired TCR signaling and functional hyporesponsiveness. This stepwise activation-to-hyporesponsive state was dependent on TCR affinity and already detectable at the upper end of the physiological affinity range (KD ≤ 1 µM). Similar findings were made when affinity-increased HLA-A2neg CD8 T cells were chronically exposed to HLA-A2pos-expressing target cells. CONCLUSIONS: Our observations indicate that sustained interactions between affinity-increased TCR and self-MHC can directly adjust the functional potential of T cells, even in the absence of antigen-specific stimulation. The observed tolerance-like state depends on TCR affinity and has therefore potential implications for the design of affinity-improved TCRs for adoptive T cell therapy, as several engineered TCRs currently used in clinical trials share similar affinity properties.

The fat and the bad: Mature adipocytes, key actors in tumor progression and resistance.

Growing evidence has raised the important roles of adipocytes as an active player in the tumor microenvironment. In many tumors adipocytes are in close contact with cancer cells. They secrete various factors that can mediate local and systemic effects. The adipocyte-cancer cell crosstalk leads to phenotypical and functional changes of both cell types, which can further enhance tumor progression. Moreover, obesity, which is associated with an increase in adipose mass and an alteration of adipose tissue, has been established as a risk factor for cancer incidence and cancer-related mortality. In this review, we summarize the mechanisms of the adipocyte-cancer cell crosstalk in both obese and lean conditions as well as its impact on cancer cell growth, local invasion, metastatic spread and resistance to treatments. Better characterization of cancer-associated adipocytes and the key molecular events in the adipocyte-cancer cell crosstalk will provide insights into tumor biology and suggest efficient therapeutic opportunities.

Fine-Tuning of Optimal TCR Signaling in Tumor-Redirected CD8 T Cells by Distinct TCR Affinity-Mediated Mechanisms.

Redirecting CD8 T cell immunity with self/tumor-specific affinity-matured T cell receptors (TCRs) is a promising approach for clinical adoptive T cell therapy, with the aim to improve treatment efficacy. Despite numerous functional-based studies, little is known about the characteristics of TCR signaling (i.e., intensity, duration, and amplification) and the regulatory mechanisms underlying optimal therapeutic T cell responses. Using a panel of human SUP-T1 and primary CD8 T cells engineered with incremental affinity TCRs against the cancer-testis antigen NY-ESO-1, we found that upon activation, T cells with optimal-affinity TCRs generated intense and sustained proximal (CD3ζ, LCK) signals associated with distal (ERK1/2) amplification-gain and increased function. In contrast, in T cells with very high affinity TCRs, signal initiation was rapid and strong yet only transient, resulting in poor MAPK activation and low proliferation potential even at high antigen stimulation dose. Under resting conditions, the levels of surface TCR/CD3ε, CD8ß, and CD28 expression and of CD3ζ phosphorylation were significantly reduced in those hyporesponsive cells, suggesting the presence of TCR affinity-related activation thresholds. We also show that SHP phosphatases were involved along the TCR affinity gradient, but displayed spatially distinct regulatory roles. While PTPN6/SHP-1 phosphatase activity controlled TCR signaling initiation and subsequent amplification by counteracting CD3ζ and ERK1/2 phosphorylation, PTPN11/SHP-2 augmented MAPK activation without affecting proximal TCR signaling. Together, our findings indicate that optimal TCR signaling can be finely tuned by TCR affinity-dependent SHP-1 and SHP-2 activity, and this may readily be determined at the TCR/CD3 complex level. We propose that these TCR affinity-associated regulations represent potential protective mechanisms preventing high affinity TCR-mediated autoimmune diseases.

TCR-ligand dissociation rate is a robust and stable biomarker of CD8+ T cell potency.

Despite influencing many aspects of T cell biology, the kinetics of T cell receptor (TCR) binding to peptide-major histocompatibility molecules (pMHC) remain infrequently determined in patient monitoring or for adoptive T cell therapy. Using specifically designed reversible fluorescent pMHC multimeric complexes, we performed a comprehensive study of TCR-pMHC off-rates combined with various functional assays on large libraries of self/tumor- and virus-specific CD8+ T cell clones from melanoma patients and healthy donors. We demonstrate that monomeric TCR-pMHC dissociation rates accurately predict the extent of cytotoxicity, cytokine production, polyfunctionality, cell proliferation, activating/inhibitory receptor expression, and in vivo antitumor potency of naturally occurring antigen-specific CD8+ T cells. Our data also confirm the superior binding avidities of virus-specific T cells as compared with self/tumor-specific T cell clonotypes (n > 300). Importantly, the TCR-pMHC off-rate is a more stable and robust biomarker of CD8+ T cell potency than the frequently used functional assays/metrics that depend on the T cell's activation state, and therefore show major intra- and interexperimental variability. Taken together, our data show that the monomeric TCR-pMHC off-rate is highly useful for the ex vivo high-throughput functional assessment of antigen-specific CD8+ T cell responses and a strong candidate as a biomarker of T cell therapeutic efficacy.

Effect of kinase inhibitors on the therapeutic properties of monoclonal antibodies.

Targeted therapies of malignancies currently consist of therapeutic monoclonal antibodies and small molecule kinase inhibitors. The combination of these novel agents raises the issue of potential antagonisms. We evaluated the potential effect of 4 kinase inhibitors, including the Bruton tyrosine kinase inhibitor ibrutinib, and 3 PI3K inhibitors idelalisib, NVP-BEZ235 and LY294002, on the effects of the 3 monoclonal antibodies, rituximab and obinutuzumab (directed against CD20) and trastuzumab (directed against HER2). We found that ibrutinib potently inhibits antibody-dependent cell-mediated cytotoxicity exerted by all antibodies, with a 50% inhibitory concentration of 0.2 microM for trastuzumab, 0.5 microM for rituximab and 2 microM for obinutuzumab, suggesting a lesser effect in combination with obinutuzumab than with rituximab. The 4 kinase inhibitors were found to inhibit phagocytosis by fresh human neutrophils, as well as antibody-dependent cellular phagocytosis induced by the 3 antibodies. Conversely co-administration of ibrutinib with rituximab, obinutuzumab or trastuzumab did not demonstrate any inhibitory effect of ibrutinib in vivo in murine xenograft models. In conclusion, some kinase inhibitors, in particular, ibrutinib, are likely to exert inhibitory effects on innate immune cells. However, these effects do not compromise the antitumor activity of monoclonal antibodies in vivo in the models that were evaluated.

Bone marrow stem cells.

The "mesenchymal stem cells (MSCs)" are cells adherent in the bone marrow, which can be isolated to induce differentiation. In contrast to the "embryonic stem cells" whose goal is to develop a new organism, the "MSC adult stem cells" can participate in tissue growth and repair throughout postnatal life. Addition of 5-azacytidine to MSCs in vitro induces the gradual increase in cellular size and begins spontaneous beatings, thereafter differentiating into cardiomyocytes. The "Methods" and "Protocols" to induce structural and functional maturations of MSCs, thus to achieve "Cellular Cardiomyoplasty," are described. With appropriate media, differentiations of MSCs to various kinds of cells such as chondrocytes, osteocytes, and adipocytes are also achievable.

Preclinical activity of the type II CD20 antibody GA101 (obinutuzumab) compared with rituximab and ofatumumab in vitro and in xenograft models.

We report the first preclinical in vitro and in vivo comparison of GA101 (obinutuzumab), a novel glycoengineered type II CD20 monoclonal antibody, with rituximab and ofatumumab, the two currently approved type I CD20 antibodies. The three antibodies were compared in assays measuring direct cell death (AnnexinV/PI staining and time-lapse microscopy), complement-dependent cytotoxicity (CDC), antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cell-mediated phagocytosis (ADCP), and internalization. The models used for the comparison of their activity in vivo were SU-DHL4 and RL xenografts. GA101 was found to be superior to rituximab and ofatumumab in the induction of direct cell death (independent of mechanical manipulation required for cell aggregate disruption formed by antibody treatment), whereas it was 10 to 1,000 times less potent in mediating CDC. GA101 showed superior activity to rituximab and ofatumumab in ADCC and whole-blood B-cell depletion assays, and was comparable with these two in ADCP. GA101 also showed slower internalization rate upon binding to CD20 than rituximab and ofatumumab. In vivo, GA101 induced a strong antitumor effect, including complete tumor remission in the SU-DHL4 model and overall superior efficacy compared with both rituximab and ofatumumab. When rituximab-pretreated animals were used, second-line treatment with GA101 was still able to control tumor progression, whereas tumors escaped rituximab treatment. Taken together, the preclinical data show that the glyoengineered type II CD20 antibody GA101 is differentiated from the two approved type I CD20 antibodies rituximab and ofatumumab by its overall preclinical activity, further supporting its clinical investigation.