Co-culture tumour spheroid experiments are routinely performed to investigate cancer progression ... more Co-culture tumour spheroid experiments are routinely performed to investigate cancer progression and test anti-cancer therapies. Therefore, methods to characterise and interpret coculture spheroid growth are of great interest. However, co-culture spheroid growth is complex. Multiple biological processes occur on overlapping timescales and different cell types within the spheroid may have different characteristics, for example proliferation rate or response to nutrient availability. There is no standard, widely-accepted mathematical model of such complex spatio-temporal growth processes. Typical approaches to analyse these experiments focus on the late-time temporal evolution of spheroid size and overlook early-time spheroid formation, spheroid structure and geometry. Here we make theoretical and practical contributions. We develop a general framework, based on mathematical and statistical modelling, to analyse a series of co-culture experiments. Using a range of different mathematic...
2020 IEEE 17th International Symposium on Biomedical Imaging (ISBI), 2020
Visualization and classification of cell cycle stages in live cells requires the introduction of ... more Visualization and classification of cell cycle stages in live cells requires the introduction of transient or stably expressing fluorescent markers. This is not feasible for all cell types, and can be time consuming to implement. Labelling of living cells also has the potential to perturb normal cellular function. Here we describe a computational strategy to estimate core cell cycle stages without markers by taking advantage of features extracted from informationrich ptychographic time-lapse movies. We show that a deeplearning approach can estimate the cell cycle trajectories of individual human melanoma cells from short 3-frame (~23 minute) snapshots, and can identify cell cycle arrest induced by chemotherapeutic agents targeting melanoma driver mutations.
Three-dimensional (3D) in vitro tumour spheroid experiments are an important tool for studying ca... more Three-dimensional (3D) in vitro tumour spheroid experiments are an important tool for studying cancer progression and potential drug therapies. Standard experiments involve growing and imaging spheroids to explore how different experimental conditions lead to different rates of spheroid growth. These kinds of experiments, however, do not reveal any information about the spatial distribution of the cell cycle within the expanding spheroid. Since 2008, a new experimental technology called fluorescent ubiquitination-based cell cycle indicator (FUCCI), has enabled
Proceedings of the National Academy of Sciences, 2021
Significance The FUCCI reporter system allows live monitoring of the cell cycle via temporal expr... more Significance The FUCCI reporter system allows live monitoring of the cell cycle via temporal expression of fluorescence markers of G0/1 or S/G2/M cell cycle phases. We found transient FUCCI reporter activity in naive neurons but not cell division, suggesting that the postmitotic state of neurons is rather a dynamic process of suppressing the cell cycle than a definite G0 state. Exposing neurons to amyloid-β resulted in death of the majority of neurons without cell cycle contribution. A subset of neurons that entered early stages of cell cycle and maintained this state were protected from amyloid-β-induced cell death. Consistently, we found high FUCCI reporter activity in the brains of mice that form amyloid-β through transgenic expression of the amyloid-β precursor protein.
Spindle cell melanoma (SCM) is a rare histological subtype of malignant melanoma composed of spin... more Spindle cell melanoma (SCM) is a rare histological subtype of malignant melanoma composed of spindled neoplastic cells. Due to its rarity, population-based clinical and epidemiological characterization is necessary. We searched the National Cancer Database (NCDB) for all cases of SCM confirmed histologically diagnosed from 2004-2016 and excluded those with missing survival information (n¼3906). Kaplan-Meier (KM) and Cox proportional-hazards models were used to analyze the epidemiology and overall survival (OS) of SCM. The median age of diagnosis was 66. Five-year and 10-year OS were 65.0% and 49.6%, respectively. Males represented 64.3% of patients. Head and neck (39.6%) was the most common primary site. Surgery alone (80.2%) was the most common treatment. The mean Breslow thickness was 3.47 mm (SD AE3.00 mm) with 31.1% with ulceration. 79.4% of staged cases presented with American Joint Commission on Cancer (AJCC) Stage I or Stage II disease. 44.4% of cases were treated at academic/research institutions followed by community programs (36.1%) and integrated network cancer programs (INCP) (12.7%). Age (HR¼1.05; CI 95%[1.04-1.06]), black race (HR¼3.0; CI 95%[1.60-5.62]), Breslow thickness greater than 9.8 mm (HR¼1.38; CI 95%[1.04-1.82]), ulceration (HR¼1.51; CI 95%[1.30-1.75]) CDCC score greater than 2 (HR¼2.28; CI 95%[1.72-3.02]), and facility locations in the East South Central (HR¼1.60; CI 95%[1.19-2.15]) and West South Central United States (HR¼1.47; CI 95%[1.06-2.05]) were independently associated with decreased OS. Females (HR¼0.85; CI 95%[0.72-0.99]), primary tumors on the trunk (HR¼0.735; CI 95%[0.61-0.89]) and upper extremities (HR¼0.71; CI 95%[0.55-0.92]), Stage I disease (HR¼0.36; CI 95% [0.13-0.99]), and treatment at INCP (HR¼0.76; CI 95%[0.68-1.42]) are independently associated with increased OS. Our results suggest that when accounting for patient and tumor characteristics, facility type and location significantly impact OS of patients with SCM.
The go-or-grow hypothesis states that adherent cells undergo reversible phenotype switching betwe... more The go-or-grow hypothesis states that adherent cells undergo reversible phenotype switching between migratory and proliferative states, with cells in the migratory state being more motile than cells in the proliferative state. Here we examine go-or-grow in 2-D in vitro assays using melanoma cells with fluorescent cell-cycle indicators and cell cycle-inhibiting drugs. We analyse the experimental data using single-cell tracking to calculate mean diffusivities, and compare motility between cells in different cell-cycle phases and in cell-cycle arrest. Unequivocally, our analysis does not support the go-or-grow hypothesis. We present clear evidence that cell motility is independent of the cell-cycle phase, and non-proliferative arrested cells have the same motility as cycling cells.
Malignant melanoma is one of the most difficult cancers to treat due to its resistance to chemoth... more Malignant melanoma is one of the most difficult cancers to treat due to its resistance to chemotherapy. Despite recent successes with BRAF inhibitors and immune checkpoint inhibitors, many patients do not respond or become resistant to these drugs. Hence, alternative treatments are still required. Due to the importance of the BCL-2-regulated apoptosis pathway in cancer development and drug resistance, it is of interest to establish which proteins are most important for melanoma cell survival, though the outcomes of previous studies have been conflicting. To conclusively address this question, we tested a panel of established and early passage patient-derived cell lines against several BH3-mimetic drugs designed to target individual or subsets of pro-survival BCL-2 proteins, alone and in combination, in both 2D and 3D cell cultures. None of the drugs demonstrated significant activity as single agents, though combinations targeting MCL-1 plus BCL-XL, and to a lesser extent BCL-2, show...
Soluble adenylyl cyclase (sAC) is a non-canonical, pH-sensitive source of cAMP in mammalian cells... more Soluble adenylyl cyclase (sAC) is a non-canonical, pH-sensitive source of cAMP in mammalian cells. We previously demonstrated that loss of sAC elevates melanosomal pH and increases melanin production, both in vitro and in vivo. An ultrastructural analysis of melanosomes following loss of sAC suggested that sAC inhibition induced melanosome maturation. Since melanosome maturation and changes in pH are associated with melanosome protein processing, we have begun to explore whether sAC activity influences the expression of melanosome proteins. RNAseq analysis of wild-type and knockout sAC melanocytes did not reveal any overt changes in melanosome protein mRNA. However, Western analysis suggests that post-translational modification of melanosome proteins is affected by loss of sAC. The observed change in protein processing is consistent with previous studies using drugs that alter melanosome pH. Thus, in addition to altering pigmentation, sAC regulation of melanosome pH affects the melanosome protein profile.
Conventional monotherapies only benefit a minority of melanoma patients while combined immunother... more Conventional monotherapies only benefit a minority of melanoma patients while combined immunotherapy exhibited extremely high rates of treatment-related adverse events. Herein, we created a multifunctional and immunogenic nanoplatform, AuNR@mSiO 2 @DOX-Cu x S-PEG, which integrating photothermal properties of gold nanoparticles, photodynamic properties of CuxS and chemotherapy into a single nanoplatform. Upon near-infrared laser irradiation (NIR), the Cu x S were uncapped and triggered chemotherapy drugs release into tumor environment. The properties and cytotoxicity to melanoma cells were explored in vitro. In addition, the anti-tumor effect and molecular mechanism of inducing tumor-specific immune responses were also explored in vitro and in vivo. The nanoplatform was stabilized, facilely synthesized and exhibited a high drug payload up to 36.37AE2.74%. Compared to control groups, the nanoplatform could forcefully inhibit tumor growth in situ (p<0.001) and lung metastasis with lower time and power intensity. Furthermore, it could promote maturation of dendritic cells and induce the infiltration of both CD8 + and CD4 + T lymphocytes in tumor in vivo. Mechanistically, the nanoplatform not only can eradicate cancer cells by inducing cell apoptosis and/or necrosis directly, but also induce a strong tumor-specific immune response by resulting in calreticulin exposure on the surface of cancer cell (p<0.01) and released highmobility group box 1 (HMGB1) protein (p<0.01). Our study developed a promising NIRtriggered multifunctional and immunogenic nanocomposite, which could induce anti-tumor immunity and would be a potential strategy for melanoma in advanced stage. 828 Dermatopathologist-level classification of skin cancer with deep neural networks at multi-magnification
Most BRAF‐mutant melanoma patients experience a fulminate relapse after several months of treatme... more Most BRAF‐mutant melanoma patients experience a fulminate relapse after several months of treatment with BRAF/MEK inhibitors. To improve therapeutic efficacy, natural plant‐derived compounds might be considered as potent additives. Here, we show that magnolol, a constituent of Magnolia officinalis, induced G1 arrest, apoptosis and cell death in BRAF‐ and NRAS‐mutant melanoma cells at low concentration, with no effect in BRAF‐ and NRAS wild‐type melanoma cells and human keratinocytes. This was confirmed in a 3D spheroid model. The apoptosis‐inducing effect of magnolol was completely rescued by activating Akt suggesting a mechanism relying primarily on Akt signaling. Magnolol significantly downregulated the PI3K/Akt pathway which led to a global decrease of the active histone mark H3K4me3. Alongside, the repressive histone mark H3K9me3 was increased as a response to DNA damage. Magnolol‐induced alterations of histone modifications are reversible upon activation of the Akt pathway. Mag...
While previous studies identify gender differences in melanoma, limited research on the phenomeno... more While previous studies identify gender differences in melanoma, limited research on the phenomenon exists. We aim to investigate melanoma prognostic factors amongst males versus females.In this retrospective chart review, 1,156 adults diagnosed with melanoma between 2006-2016 at the University of Colorado were included. Breslow's depth, mitotic rate, ulceration status, and location were extracted between March and August 2016. Cochran-Armitage trend tests and cumulative logistic regression were used to examine the association between gender and Breslow's depth, univariately and after adjusting for potential confounders. In univariate analysis, males were significantly more likely to present with lesions with higher Breslow's depths (p for trend¼0.005). In models adjusted for age, melanoma subtype, and location, males were marginally more likely to present with lesions with higher Breslow's depths (cumulative OR: 1.261, 95% CI: 0.988-1.611, p¼0.060). Males were also marginally more likely to present with lesions with higher mitotic rates, after further adjustments for all other prognostic factors (cumulative OR: 1.244, 95% CI: 0.979-1.580, p¼0.074). Differences in mitotic rates among melanomas in males versus females, even after adjustments for all other prognostic factors, suggests that biological differences may contribute to the female advantage.
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Papers by Nikolas Haass