# Anti-Cancer Drug Screening with Microfluidic Technology
## 4 Combinatorial Drug Screening on a Microscale Sample
### 3D Cell Cultures  
[PERSON] et al. developed a PDMS-based microfluidic device by co-culturing carcinoma cells with human umbilical vein endothelial cells (HUVECs) for anti-invasive and anti-metastatic combinatorial drug screening [66]. [PERSON] et al. engineered a hydrogel-based microfluidic chip for glioblastoma multiforme (GBM) brain cancer [67]. They cultured U87 cells to form spheroids in microwells and performed combinatorial treatment of pitavastatin and irinotecanin. They found that dual drug administration is more effective than administration of individual drugs. This is a powerful platform for high-throughput screening and prolonged drug release and can be used in the future with small biopsy-derived tissue samples. Fabrication of photo-polymerizable poly(ethylene) glycol diacrylate (PEGDA)-based microfluidics is easier than polydimethylsiloxane (PDMS). Moreover, PEGDA blocks protein absorption and better mimics the 3D tumor microenvironment. [PERSON] et al. developed a high-throughput pairwise drug-combination screening device for arbitrary numbers of drugs [68]. In this system, a logarithmic concentration gradient is generated between each drug pair. Furthermore, for inlets arrangement, a sudoku puzzle is solved to guarantee the adjacent drug pairs cover all the possible combinations. As a proof-of-concept, 7-drug combinatorial treatments were tested over 1032 cancer spheroids of pancreatic cancer patient-derived xenograft cell lines on this three-layer chip. This study benefits precision medicine with high-throughput screening due to low sample consumption and fast analysis. [PERSON] et al. recently evaluated the effect of anti-inflammatory combinatorial treatment of doxorubicin and aspirin on cell lines and patient-derived CTC clusters of breast cancer [69]. This microfluidic device could be a valuable clinical decision-support tool for tumor relapse prevention  
[PERSON] et al. developed an automated microfluidic platform based on the platform of [PERSON] et al. [60] using pneumatic valves. The combinatorial treatment of doxorubicin and paclitaxel on a thin-gel 3D cell-culture from MDA-MB-231 and MCF-7 breast cancer cell lines was tested [70]. They assembled two air-controlled channel layers, two drug channel layers, and two porous membrane layers on the top and bottom of a cell culture layer. Thus, the cell culture chamber layer is sandwiched between two layers of porous PDMS membranes. [PERSON] et al. [57] developed an automated microfluidic platform for dynamic and combinatorial drug screening of tumor organoids. They cultured organoids from different samples, including MDA-MB-231 cell lines, patient-derived pancreatic tumor, and human-derived colon tissue samples. Utilizing this device, 200 different conditions (i.e., 20 different subsets of 10 individual chambers) can be tested. They evaluated individual, combinatorial, and sequential drug screens of chemotherapeutic drugs and found that the temporally modified treatments are more effective than static ones in vitro. All the presented drug screening platforms for cancer therapy have been summarized in Table 1.  
\begin{table}
\begin{tabular}{c c c c} \hline \hline \multicolumn{5}{c}{**Monodrug Screening on a Microscale Sample**} \\ \hline
**Sample Type** & **Tested Drugs** & **Number of Dosages Tested** & **Ref.