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International Journal of Bioprinting Lumen-forming colorectal cancer organoids

It is widely known that organoids can only be fabricated moieties to obtain a biofunctional IKVAV-derived peptide
within 3D matrices that mimic the mechanical and (IDP) sequence [19,20] .
biochemical properties of extracellular matrix (ECM); to We expect that the interruption of the polarity gradient
this end, researchers mostly depend on naturally derived of the tetrapeptide might adversely affect the assembly,
hydrogels, such as decellularized ECM, laminin, fibrinogen, thereby reducing the mechanical properties of the hydrogel.
and collagen [7,8] . For instance, a gelatin methacryloyl In order to overcome this, we hypothesize that by mixing
(GelMA)-based bioresin was used in a volumetric 3D printer a parent SAP together with its biofunctional version, we
to generate a construct that contains liver organoids to be can create a backbone to sustain the biochemical cues in
deployed as metabolic “biofactories” . The bioresin had to the matrix. A similar strategy has been used before, where
[9]
be formulated together with a refractive index matching two self-assembling peptides derived from RADA16 were
compound in order to tune the printing performance of biofunctionalized with IKVAV and arginylglycylaspartic
GelMA. Cell polarization and hepatic differentiation were acid (RGD) motifs . This approach used oppositely
[21]
observed after 10 days of culture. On the other hand, charged RADA peptides to induce self-assembly. A
researchers have generated vascularized neural organoids molecular model, in which a non-biofunctional RADA16
by using commercial gelatin/fibrinogen-based bioinks . “core” stabilizes the random coils formed by the functional
[10]
Interestingly, the reported method allows for the motifs, was proposed. Moreover, IKVAV is known as a self-
simultaneous differentiation of vascular endothelium and assembling peptide itself, capable of forming nanofibers
neurons from pluripotent cell lines while retaining their that promote cell growth (in particular, neuron growth and
identity during co-culture. From the coupling of this new development) [22,23] .
method with multi-material bioprinting to produce integer
vascularized organoids, researchers were able to reproduce The proposed strategy would guarantee a constant
the geometry of a human dorsal forebrain coronal section. concentration of a small molecule of interest in the matrix,
even during cell proliferation, whereas a free molecule
Despite the positive outcomes of these techniques, the
use of these biomaterials carries several disadvantages, such would diffuse, and its effect in cells would diminish over
time. Furthermore, the use of a biofunctional peptide will
as immunogenicity, batch-to-batch variability, xenogenic reassure a homogeneous distribution of the biofunctional
material, and presence of unwanted growth factors . signal in 3D constructs. With regard to the biofunctional
[11]
Moreover, these materials lack mechanical tunability part of this new peptide, we presume that the introduction
and have shown poor performance as bioinks, for which of a laminin-derived motif (IKVAV, active site in LMα1)
they need to be chemically modified to be light-reactive. into a self-assembling peptide will have an effect on cell
Similarly, supplemented synthetic materials have shown behavior similar to that of the motif itself.
promising discovery as tunable matrices for organoids .
[12]
Still, all these options require ultraviolet (UV) crosslinking, Given that colorectal organoids secrete laminin in
which introduces risk to the embedded cellular structures. order to maintain structural integrity , we speculate that
[24]
In this work, the biofunctionalization of a known self- this bioactive peptide can be employed as an alternative
assembling peptide (SAP) was proposed to design a novel material for organoid-like structures in bioprinted
laminin-like bioink. scaffolds. In addition, laminin receptors are highly
expressed in colorectal cancer cells . By introducing the
[25]
Self-assembling peptides are amphiphilic molecules that presence of a cell-interacting motif, like laminin-111, we
can spontaneously aggregate into complex 3D structures expect to obtain a minimalist matrix that mimics the ECM
such as nanofibers . Given their similarities to ECMs, characteristics that allow organoid formation.
[13]
nanofiber-forming SAPs have shown to be an alternative
for 3D cell culture. SAPs, developed by the Hauser Group, This biofunctional SAP will allow us to retain the
have shown to be biocompatible, biodegradable, and non- mechanical properties [14,26-29] as well as the tunability
immunogenic; additionally, they can gelate to fibrillar and printability of our previously reported SAP, while
hydrogels under controlled physiological conditions and incorporating a known cue for cell–matrix interactions.
specific concentrations [14-18] . Moreover, SAPs provide Here, we investigated the effect of this motif on cell
compositional control and simplicity in synthesis. These proliferation and lumen formation as an early organoid
SAPs can be used as bioinks in a dual extrusion setting formation signal. We also evaluated the bioprintability
to generate 3D constructs of up to 4 cm in height . In of this modified tetrapeptide hydrogel by investigating
[14]
order to provide cues for the growth of colorectal cancer the cell viability and proliferation of a printed construct
cells, we supported a previously developed SAP sequence with cells for possible tissue engineering applications.
(IIFK) with laminin-derived IKVAV motif and a newly Both manual and bioprinted construct containing IDP
synthesized peptide containing both IIFK and IKVAV demonstrated clear advantages, particularly in organoid

Volume 9 Issue 1 (2023)olume 9 Issue 1 (2023)
V 161 https://doi.org/10.18063/ijb.v9i1.633