Anti-cancer drugs  

Hundreds of compounds are currently undergoing clinical trials for evaluating their effectiveness in fighting cancer, and thousands more are being developed and investigated during the preclinical drug development stages. Anti-cancer compounds span from cytotoxic compounds, immunotherapies, hormonal agents to molecular targeted therapies, and can take in the form of small molecule compounds, antibodies, cell therapies or vaccines.  

Preceding clinical studies that need to demonstrate the clinical benefits of the drug in the target population are preclinical studies. Preclinical evaluation of novel cancer therapies starts from fundamental research, involving target identification and validation to early preclinical research to demonstrate the compounds’ overall safety and toxicity profile. Efficacy and safety are typically evaluated in an exploratory manner, depending on the drug’s mechanism of action.  

In vivo research normally starts with the most suitable selection of an oncology model, in line with the objectives and goals of the study (i.e., the use of more complex orthotopic models that recapitulate tumoral microenvironment, to simpler xenograft studies). The tumoral response to a novel compound can be determined by assessing the implanted tumor volume using in vivo imaging techniques such as ultrasound, bioluminescent and fluorescent imaging. In order to provide deeper insights into the tumor’s microenvironment, flow cytometry proves to be a powerful technique. However, as flow cytometry is traditionally considered to be a bulk method, histopathology emerges as a more suitable method to provide deeper insights into the tumor’s microenvironment, offering the advantage of a preserved tissue morphology.  


The histopathological assessment of tumor tissue samples represents one of key endpoints of many studies conducted during the preclinical stage. Deeper insights into the spatial tumor microenvironment, proliferation, histologic malignancy grades, angiogenesis, inflammation, fibrosis, intra-tumoral drug levels, target engagement and efficacy can be evaluated using various histopathology techniques.  

Most people associate histopathology with classical gold-standard readouts such as H&E and IHC. However, the field of histopathology is also keeping up with the newest research trends to provide advanced high-throughput multiplexing techniques similar to -omics techniques such as RNA sequencing. Therefore, it may be tempting to include the most novel state-of-the-art readouts such as multiplex IF offering users benefits such as large-scale cell phenotyping. However, not every project will benefit from this technique. On the contrary, many projects, are sufficient with H&E or single IHC stainings to determine characteristics such as:  

  • Proliferation index 
  • Characterization of tumor morphology related to malignancy (pleomorphism, chromatic appearance, nucleus-to-cytoplasm ratio, etc.,)  
  • Tumor invasion, metastasis 
  • Immune cell infiltration 
  • Angiogenesis  
  • Determining general compound toxicology characteristics 
  • Pleomorphism  

How to select the most suitable technique  

To help you select the most suitable and cost-effective technique applicable for your research, we have compiled the most commonly used techniques and their most common applications in oncology research (non-exhaustive) in the table below.  

Table summarising of main histopathology techniques and their applicability in research.  

Histopathology technique   Study type Why should you use this technique?  Common markers
  • Tumor engraftment evaluation (rodent models of cancer) 
  • Efficacy 
  • Toxicology 
  • Subcellular structures assessment
  • Gold standard for tumor diagnosis and subtyping  
  • Automated tissue staining allows for high-volume studies 
  • Toxicity studies 
  • Histologic malignancy characteristics 
  • Metastasis 
  • Invasion 
Special Stain
  • Hematopoietic tumor studies  
  • Diagnosis of tumor cell type 
  • Giemsa stain 
  • Target confirmation  
  • Tumor microenvironment  
  • Immunotherapy target identification  
  • Co-expression of markers 
  • Tumor microenvironment and immune cell interplay  
  • Angiogenesis (CD31 and CD44 
  • Proliferation (Ki67) 
  • Immune cell infiltration (CD3, CD4, CD8)
  • Tumor histologic differentiation (CK, Vimentin, Ca19-9 etc,.) 
  • Anaplasia (CEA) 


  • Compound distribution (fluorescently labeled compounds) 
  • Immunotherapy target identification
  • Identification of sensitive targets 
  • Detection of GFP-labelled cells 
  • Detection of BrdU in proliferating cells  
Multiplex IHC and IF
  • Tumor/microenvironment interaction 
  • Immune cell phenotyping 
  • Studying cell-cell interactions 
  • Localizing cells in different areas 
  • Studying cellular phenotypes and mapping them in a tissue context 
  • Immune cell markers (CD3, CD68, PAX5 etc,.) 
  • Tumor-associated fibroblasts activity  
  • Probing DNA and RNA in tissues


  • Biodistribution studies 
  • Cancer-initiating infectious agents 
  • Labelling vaccine particles with RNAScope™ ISH 
  • Detection of viruses causing neoplasms (Herpes virus, EBV, etc.,) 


Working with Connected-Pathology 

Rigorous histopathology analysis during the preclinical stage is critical in confirming not only the efficacy of the novel anti-cancer compound, but also to demonstrate the overall safety and toxicity profile. However, without the necessary pathology expertise and experience, proper histopathology analyses take significant time and resources. In the field of oncology, preclinical researchers need to ensure target specificity, selection of correct preclinical in vitro and in vivo models to answer their research questions and unbiased approach to testing their novel compound. All of this takes a significant amount of time, leaving little time and expertise to focus on analyses that are routine for expert histopathologists.  

Connected-Pathology proves to be a reliable outsourcing histopathology partner, allowing you to benefit from:  

  • Whole-slide scanning provides competitive leading times as our veterinary pathologists to view images in high-quality. 
  • Our in-house designed viewer, I-Connect, gives you not only access to the full image directory but also the ability to directly communicate with our veterinary pathologists. 
  • Expert in assay development solutions, Connected-Pathology works with you to validate novel biomarkers. 
  • Our automated devices enable us to perform large-scale studies from H&E to IHC/IF and more complex methods.
  • Our expert veterinary and clinical pathologists assist you with interpreting study findings and taking the next step.  

Histopathology readouts are applicable and can be tailored to all preclinical oncology models:  

  • Cell-line (CDX) and patient-derived xenograft (PDX) tumor models 
  • Syngeneic tumor models 
  • Orthotopic tumor models 
  • Metastatic tumor models
  • Humanized models
  • Cancer cell lines
  • Patient-derived 3D models