Over the last few decades, major efforts in cancer research and treatment have intensified. Apart from standard chemotherapy approaches, immunotherapy has gained substantial traction. Personalized immunotherapy has become an important tool for cancer therapy with the discovery of immune checkpoint inhibitors. Traditionally, tumor-associated antigens are used in immunotherapy-based treatments. Nevertheless, these antigens lack specificity and may have increased toxicity. With the advent of next-generation technologies, the identification of new tumor-specific antigens is becoming more important. In colorectal cancer, several tumor-specific antigens were identified and functionally validated. Multiple clinical trials from vaccine-based and adoptive cell therapy utilizing tumor-specific antigens have commenced. Herein, we will summarize the current landscape of tumor-specific antigens particularly in colorectal cancer.
Topoisomerases are nuclear enzymes that regulate topology of DNA by facilitating the temporary cleavage and ligation cycle of DNA. Among all forms of topoisomerases, TOP-IIA is extensively associated with cell proliferation and therefore is an important therapeutic target in diseases that involved cellular proliferation such as cancers. Nearly half of present-day antitumor regimens contain at least one prescription that act as a topoisomerase inhibitor. Generally, tumor cells show divergent expression of TOP-IIA compared to normal cells. The remarkable expression of TOP-IIA in various carcinomas provides a significant biomarker toward understanding the nature of malignancy. TOP-IIA expression and amplification studies help in diagnosing cancer and to observe the disease progression, overall survival (OS) of patients, and response to therapy. This review highlights the research output and analysis in exploring the standing of TOP-IIA in various carcinomas. As some reports show contradiction within the same field of interest, the outline of that may help to induce researchers for further investigation and clarification. To the best of our knowledge, this is the first overview briefly summarizing the prognostic feature of TOP-IIA in various types of cancer.
The angiogenic potential of native skin (NS), keratinocytes single skin equivalent (SSE-K), fibroblasts single skin equivalent (SSE-F) and bilayered skin equivalent secreting angiogenic growth factors such as transforming growth factor beta1 (TGF-beta1), vascular endothelial growth factor (VEGF), keratinocyte growth factor (KGF) and basic fibroblast growth factor (bFGF) in the in vitro systems at 24, 48, 72 hours and 7 days was compared using Enzyme-Linked Immunosorbent Assay (ELISA). Bilayered skin equivalent exhibit highest release of growth factors within 24 hours to 7 days of culture compared to NS, SSE-K and SSE-F. This proved the potential of bilayered skin equivalent in producing and sustaining growth factors release to enhance angiogenesis, fibroblasts proliferation, matrix deposition, migration and growth of keratinocytes.
We report a case of clear cell "sugar" tumour of the lung (CCTL) occurring in a 26-year-old lady. The patient was asymptomatic and the lesion was picked up in the course of a pre-employment medical examination. A well-defined 5 cm nodule in the right lower lobe was detected on routine chest X-Ray. Microscopical examination of the coin lesion showed clear cells containing abundant diastase-sensitive intracytoplasmic glycogen, as demohstrated with periodic acid-Schiff stains. Tumour immunoreactivity for HMB-45 and non-reactivity for cytokeratin support the histological diagnosis. To our knowledge, this is the first reported case of CCTL in Malaysia.
Malignant melanoma involving the gastrointestinal tract is diagnosed antemortem in only a small percentage of patients with the disease. Presenting symptoms are often non-specific, causing a diagnostic problem. The vast majority of such melanomas are metastatic from a cutaneous primary, however there is evidence that the tumour can arise de novo in the gastrointestinal system. We report a 74-year-old man with malignant melanoma with an unusual presentation simulating a symptomatic gastric ulcer. He presented with epigastric pain, haematemesis and melaena. Explorative laparotomy revealed a large ulcerated tumour with several pigmented satellite nodules in the proximal stomach, multiple ileal nodules and widespread nodal and liver metastases. Proximal gastrectomy and limited small bowel resection was performed. Histology revealed the tumour to be composed of nests of epithelioid cells with melanin pigment. The tumour cells showed immunohistochemical positivity for S100 protein and HMB45 antibodies. This report emphasizes that melanoma should be a diagnostic consideration in patients with gastric ulcer.
Squamous cell carcinoma-related antigen (SCC-Ag), first described by Kato and Torigoe in 1977, has been cited by various workers as a serological marker for some epithelial neoplasms. The most well-studied is its association with carcinoma of the uterine cervix. In January 1989, we embarked on a prospective, multivariate study at the University Hospital, Kuala Lumpur to assess the usefulness of serologically assaying SCC-Ag (using the Abbott RIA diagnostic kit) in our patients with carcinoma of the uterine cervix. We were also interested to ascertain whether SCC-Ag is a 'general' marker for all histological types of cervical carcinoma or specific for squamous carcinoma. From the time of commencement to June 1990, 35 newly-diagnosed and histologically-proven cases were entered into the study. Of these, 4 were keratinising squamous carcinoma, 18 large cell non-keratinising carcinoma, 3 adenosquamous carcinoma, 7 adenocarcinoma and 3 carcinoma-in-situ. Our preliminary results show that all keratinising squamous carcinoma and 1/3 each of large cell non-keratinising carcinoma, adenosquamous carcinoma and carcinoma-in-situ had positive pre-therapy serum SCC-Ag levels (i.e greater than 2 ng/ml, 2 ng/ml being an arbitrarily selected 'cut-off' value). In contrast, no adenocarcinoma was serologically positive. In addition, keratinising squamous carcinoma had the highest mean pre-therapy serum SCC-Ag level. The results imply that serum SCC-Ag is related to the (1) presence of squamous and not glandular differentiation and (2) degree of squamous differentiation.
To assess the diagnostic utility of glypican-3 (GPC-3) in comparison to Hep Par 1 in the diagnosis of liver tumours, a cross-sectional study involving 66 resected liver tumours were tested for the protein expression of these markers by immunohistochemistry using monoclonal antibodies. Of the 66 cases, 26 (39.4%) were hepatocellular carcinoma (HCC), 4 (6.1%) were intrahepatic cholangiocarcinoma and 36 (54.5%) were metastatic tumours. Hep Par 1 and GPC-3 expressions in HCC were 24/26 (92.3%) and 19/26 (73.1%) respectively. In contrast, of non-HCC cases, only 2/40 cases (5.0%) expressed Hep Par 1, including a metastatic colorectal adenocarcinoma and a metastatic gastric adenocarcinoma. GPC-3 was expressed in 3/40 cases (7.5%), i.e. a metastatic adenocarcinoma of unknown origin, a metastatic gastric adenocarcinoma and an intrahepatic cholangiocarcinoma. The sensitivity and specificity for Hep Par 1 were 92.3% and 95% respectively while that of GPC-3 was 73.1% and 92.5% respectively. GPC-3 is a useful marker in the diagnosis of HCC. However it is not superior to Hep Par 1 in its sensitivity and specificity. We recommend that it is utilized together with Hep Par 1 as a panel in the diagnosis of HCC.
Breast cancer is one of the most common malignancies among woman. Decades of scientific study have linked the
overexpression of ERBB2 antigen to aggressive tumors. To target aggressive breast cancer, chimeric antigen receptor
(CAR) technology can be utilized. For this, human T-cells are transduced with a gene sequence encoding a CAR that is
specific for tumor-associated antigens (TAAs). These genetically-engineered CAR transduced T-cells (CAR-T cells) are
able to target the tumor antigen without the need for major histocompatibility complex (MHC) recognition, rendering
it a potentially universal immunotherapeutic option. However, efficient transduction of therapeutic gene into human
T-cells and further cell expansion are challenging. In this study, we reported a successful optimization of a transduction
protocol using spinoculation on CD3+ T-cells with different concentrations of lentiviral plasmid encoding the CAR gene.
CD3+T-cells were isolated from the peripheral blood mononuclear cells (PBMCs). The constructed CAR gene was inserted
into a lentiviral plasmid containing the green fluorescent protein (GFP) tag and lentiviral particles were produced. These
lentiviral particles were used to transduce activated T-cells by spinoculation. T-cells were activated using Dynabeadconjugated
CD3/CD28 human T-cell activator and interleukin-2 (IL-2) before transduction. CD3+ T-cells were selected
and GFP expression, which indicated transduction, was observed. Future studies will focus on in vitro and in vivo models
to determine the efficiency of CAR-T cells in specifically targeting ERBB2-expressing cells.
Endometrial cancer is the most commonly diagnosed gynecologic malignancy worldwide; yet the tumor microenvironment, especially the fibroblast cells surrounding the cancer cells, is poorly understood. We established four primary cultures of fibroblasts from human endometrial cancer tissues (cancer-associated fibroblasts, CAFs) using antibody-conjugated magnetic bead isolation. These relatively homogenous fibroblast cultures expressed fibroblast markers (CD90, vimentin and alpha-smooth muscle actin) and hormonal (estrogen and progesterone) receptors. Conditioned media collected from CAFs induced a dose-dependent proliferation of both primary cultures and cell lines of endometrial cancer in vitro (175%) when compared to non-treated cells, in contrast to those from normal endometrial fibroblast cell line (51%) (P<0.0001). These effects were not observed in fibroblast culture derived from benign endometrial hyperplasia tissues, indicating the specificity of CAFs in affecting endometrial cancer cell proliferation. To determine the mechanism underlying the differential fibroblast effects, we compared the activation of PI3K/Akt and MAPK/Erk pathways in endometrial cancer cells following treatment with normal fibroblasts- and CAFs-conditioned media. Western blot analysis showed that the expression of both phosphorylated forms of Akt and Erk were significantly down-regulated in normal fibroblasts-treated cells, but were up-regulated/maintained in CAFs-treated cells. Treatment with specific inhibitors LY294002 and U0126 reversed the CAFs-mediated cell proliferation (P<0.0001), suggesting for a role of these pathways in modulating endometrial cancer cell proliferation. Rapamycin, which targets a downstream molecule in PI3K pathway (mTOR), also suppressed CAFs-induced cell proliferation by inducing apoptosis. Cytokine profiling analysis revealed that CAFs secrete higher levels of macrophage chemoattractant protein (MCP)-1, interleukin (IL)-6, IL-8, RANTES and vascular endothelial growth factor (VEGF) than normal fibroblasts. Our data suggests that in contrast to normal fibroblasts, CAFs may exhibit a pro-tumorigenic effect in the progression of endometrial cancer, and PI3K/Akt and MAPK/Erk signaling may represent critical regulators in how endometrial cancer cells respond to their microenvironment.
Pigmentation is a readily scorable and quantitative human phenotype, making it an excellent model for studying multifactorial traits and diseases. Convergent human evolution from the ancestral state, darker skin, towards lighter skin colors involved divergent genetic mechanisms in people of European vs. East Asian ancestry. It is striking that the European mechanisms result in a 10-20-fold increase in skin cancer susceptibility while the East Asian mechanisms do not. Towards the mapping of genes that contribute to East Asian pigmentation there is need for one or more populations that are admixed for ancestral and East Asian ancestry, but with minimal European contribution. This requirement is fulfilled by the Senoi, one of three indigenous tribes of Peninsular Malaysia collectively known as the Orang Asli. The Senoi are thought to be an admixture of the Negrito, an ancestral dark-skinned population representing the second of three Orang Asli tribes, and regional Mongoloid populations of Indo-China such as the Proto-Malay, the third Orang Asli tribe. We have calculated skin reflectance-based melanin indices in 492 Orang Asli, which ranged from 28 (lightest) to 75 (darkest); both extremes were represented in the Senoi. Population averages were 56 for Negrito, 42 for Proto-Malay, and 46 for Senoi. The derived allele frequencies for SLC24A5 and SLC45A2 in the Senoi were 0.04 and 0.02, respectively, consistent with greater South Asian than European admixture. Females and individuals with the A111T mutation had significantly lighter skin (p = 0.001 and 0.0039, respectively). Individuals with these derived alleles were found across the spectrum of skin color, indicating an overriding effect of strong skin lightening alleles of East Asian origin. These results suggest that the Senoi are suitable for mapping East Asian skin color genes.
Nasopharyngeal carcinoma (NPC) is highly prevalent in South East Asia and China. The poor outcome is due to late presentation, recurrence, distant metastasis and limited therapeutic options. For improved treatment outcome, immunotherapeutic approaches focusing on dendritic and autologous cytotoxic T-cell based therapies have been developed, but cost and infrastructure remain barriers for implementing these in low-resource settings. As our prior observations had found that four-jointed box 1 (FJX1), a tumor antigen, is overexpressed in NPCs, we investigated if short 9-20 amino acid sequence specific peptides matching to FJX1 requiring only intramuscular immunization to train host immune systems would be a better treatment option for this disease. Thus, we designed 8 FJX1-specific peptides and implemented an assay system to first, assess the binding of these peptides to HLA-A2 molecules on T2 cells. After, ELISPOT assays were used to determine the peptides immunogenicity and ability to induce potential cytotoxicity activity towards cancer cells. Also, T-cell proliferation assay was used to evaluate the potential of MHC class II peptides to stimulate the expansion of isolated T-cells. Our results demonstrate that these peptides are immunogenic and peptide stimulated T-cells were able to induce peptide-specific cytolytic activity specifically against FJX1-expressing cancer cells. In addition, we demonstrated that the MHC class II peptides were capable of inducing T-cell proliferation. Our results suggest that these peptides are capable of inducing specific cytotoxic cytokines secretion against FJX1-expressing cancer cells and serve as a potential vaccine-based therapy for NPC patients.
Wisteria floribunda agglutinin-positive Mac-2 binding protein (WFA+-M2BP) has been suggested to be useful for the assessment of disease severity in non-alcoholic fatty liver disease (NAFLD). Consecutive adult NAFLD patients who had a liver biopsy were included. Serum WFA+-M2BP level was measured using a lectin-antibody sandwich immunoassay using a chemiluminescence enzyme immunoassay machine (HISCL-5000, Sysmex, Kobe, Japan). The measured levels were indexed using the following equation: Cut-off index (COI) = ([WFA+-M2BP]sample-[WFA+-M2BP]NC) / ([WFA+-M2BP]PC-[WFA+-M2BP]NC), where PC = positive control and NC = negative control. Histopathological examination of liver biopsy specimen was reported according to Non-Alcoholic Steatohepatitis (NASH) Clinical Research Network Scoring System. Data for 220 cases were analyzed. The AUROC of the COI for the diagnosis of NASH was 0.65. The AUROC of the COI for the diagnosis of steatosis grade ≥2 and 3 was 0.64 and 0.53, respectively. The AUROC of the COI for the diagnosis of lobular inflammation grade ≥1, ≥2 and 3 was 0.57, 0.68 and 0.59, respectively. The AUROC of the COI for the diagnosis of hepatocyte ballooning grade ≥1 and 2 was 0.64 and 0.65, respectively. The AUROC of the COI for the diagnosis of fibrosis stage ≥1, ≥2, ≥3 and 4 was 0.61, 0.71, 0.74 and 0.84, respectively. Out of the 220 cases, 152 cases were the same 76 patients who had a repeat liver biopsy after 48 weeks of intervention. The AUROC of the change in the COI to detect improvement in steatosis, lobular inflammation, hepatocyte ballooning and fibrosis was 0.57, 0.54, 0.59 and 0.52, respectively. In conclusion, serum WFA+-M2BP was most useful for the diagnosis of significant fibrosis, advanced fibrosis and cirrhosis in NAFLD patients. However, it was less useful for differentiating NASH from non-NASH, and for diagnosis and follow-up of the individual histopathological components of NASH.
CD44 is a cell adhesion molecule that plays an important role in the cascade of metastasis and progression of human malignant tumours. A large family of variants or isoforms, generated by alternative splicing of a single gene, has been reported to be involved in the malignant process by conferring metastatic potential to non-metastatic cells. The objective of this study was to compare the expression of CD44 standard molecule with the International Neuroblastoma Pathology Classification (INPC) for neuroblastic tumours, a histological grading system based on the Shimada system for predicting the clinical outcome in neuroblastic tumours.
Both zinc and the α-subunit of hypoxia-inducible factor (HIF-1α) play important roles in the remodelling of mammary gland tissues. In the present study, we examined the level and the transcriptional activity of HIF-1α in mammary cells upon zinc treatment. In MCF-7 mammary adenocarcinoma and MCF-10A mammary epithelial cell lines, the toxicity levels of zinc differ. Interestingly, both cell lines overexpress HIF-1α following zinc treatment. As it was evident from an up-regulation of its specific target gene CA9 that encodes carbonic anhydrase IX, the stabilized HIF-1α translocated to the nucleus and was transcriptionally active. Hence, we conclude that zinc causes normoxic accumulation of transcriptionally active HIF-1α by interfering with its post-translational regulation.
A method is described for the electrochemical determination of squamous cell carcinoma (SCC) antigen, and by testing the effect of 30 nm gold nanoparticles (GNPs). Three comparative studies were performed in the presence and absence of GNPs, and with agglomerated GNPs. The divalent ion Ca(II) was used to induce a strong agglomeration of GNPs, as confirmed by colorimetry and voltammetry. Herein, colorimetry was used to test the best amount of salt needed to aggregate the GNPs. Despite, voltammetry was used to determine the status of biomolecules on the sensor. The topography of the surface of ZnO-coated interdigitated electrodes was analyzed by using 3D-nano profilometry, scanning electron microscopy, atomic force microscopy and high-power microscopy. The interaction between SCC antigen and antibody trigger vibrations on the sensor and cause dipole moment, which was measured using a picoammeter with a linear sweep from 0 to 2 V at 0.01 V step voltage. The sensitivity level was 10 fM by 3σ calculation for the dispersed GNP-conjugated antigen. This indicates a 100-fold enhancement compared to the condition without GNP conjugation. However, the sensitivity level for agglomerated GNPs conjugated antibody was not significant with 100 fM sensitivity. Specificity was tested for other proteins in serum, namely blood clotting factor IX, C-reactive protein, and serum albumin. The SCC antigen was quantified in spiked serum and gave recoveries that ranged between 80 and 90%. Graphical abstractSchematic representation of SCC (squamous cell carcinoma) antigen determination using divalent ion induced agglomerated GNPs. Sensitivity increment depends on the occurrence of more SCC antigen and antibody binding event via GNPs integration. Notably, lower detection limit was achieved at femto molar with proper orientation of biological molecules.
Discovery of tumor antigenic epitopes is important for cancer vaccine development. Such epitopes can be designed and modified to become more antigenic and immunogenic in order to overcome immunosuppression towards the native tumor antigen. In silico-guided modification of epitope sequences allows predictive discrimination of those that may be potentially immunogenic. Therefore, only candidates predicted with high antigenicity will be selected, constructed, and tested in the lab. Here, we described the employment of in silico tools using a multiparametric approach to assess both potential T-cell epitopes (MHC class I/II binding) and B-cell epitopes (hydrophilicity, surface accessibility, antigenicity, and linear epitope). A scoring and ranking system based on these parameters was developed to shortlist potential mimotope candidates for further development as peptide cancer vaccines.
Introduction: Oral squamous cell carcinoma (OSCC) is a major health problem worldwide. The overall survival rate remains at 50% despite numerous studies and various treatment modalities in OSCC. The presence of lymph node metastasis in OSCC is well established as an independent prognostic factor. This present study aims to investigate the association of four tumour antigens; FJX-1, GNα12, IFITM3 and MAGED4B with the sociodemographic and clinicopathological parameters of OSCC. The potential use of these markers as a prognostic indicator of patient sur-vival and lymph node metastasis in OSCC was explored. Methods: 35 cases of OSCC with available formalin-fixed paraffin-embedded (FFPE) specimens involving the tongue, buccal mucosa, gingiva, alveolus and floor of mouth were evaluated by immunohistochemistry for FJX-1, GNA12, IFITM3 and MAGED4B expression. Assessment of the expression of these tumour antigens was based on the cellular sub-site, intensity and percentage of staining in the OSCC samples. Results: The expression of all four tumour markers were expressed in all samples (n=35) but none statistically associated with any clinicopathological or socio-demographic parameters. Survival analysis using Kaplan-Meier test showed high expression of GNA12, IFITM3 and MAGED4B individually with poor prognosis in OSCC patients. A combination of markers, GNA12 and MAGED4B demonstrated a significant association with pa-tient survival in OSCC (p=0.014). Multivariate analysis after adjustment for selected socio-demographic factors (age, gender, risk habits and sub-sites of the oral cavity) revealed that high expression of both MAGED4B and GNA12 remained as an independent prognostic factor for poor prognosis in OSCC (HRR =5.231, 95% CI 1.601,17.084; p=0.006). Conclusion: We concluded that high combined expression of both marker (Gα12 and mAGED4B) might be used as an independent prognostic indicator in OSCC.
Head and neck cancers have been reported to have high immune infiltration scores, and clinical benefits of the anti-PD1 checkpoint inhibitor have been demonstrated in recurrent and metastatic cancers. Recent genetic signa-tures of the immune compartment have provided insights to delineate immune-active and -exhausted subtypes, to understand the immune status of OSCC patients that could further drive the development of novel immunotherapies. Vaccination with tumour-associated antigens is an approach to improve tumour recognition which could result in the eradication of cancer cells. Here, I would describe our efforts in developing antigen-specific vaccines for head and neck cancer. Using the B6.Cg-Tg(HLA-A/H2-D)2Enge/J mice bearing established tumours overexpressing the tumour antigens, we demonstrated that the vaccine delayed tumour growth, and in combination with anti-PD1, completely eliminated the tumour. The vaccine increased the expression of PD1 in T cells, and vaccinated animals showed increased antigen-specific responses by the ELISPOT assay. In summary, our data show that antigen-specific vaccine works synergistically with anti-PD1 and could be a promising therapeutic agent for head and neck cancer.