COMPOSITIONS AND METHODS FOR MODULATING GENOMIC COMPLEX INTEGRITY INDEX
The present disclosure relates generally to modulation of genomic complexes via modulation (e.g., disruption) based on certain integrity index scores.
1 . A method of disrupting a genomic complex, e.g., anchor sequence mediated conjunction (ASMC), in a mammalian subject, comprising:
administering to a subject a disrupting agent targeted to the genomic complex (e.g., ASMC),
wherein the genomic complex (e.g., ASMC) has, or is identified as having, an IntInch, measured by Formula 2
(
IntInd
i
=
min
(
Frequency
of
genomic
complex
(
e
.
g
.
,
ASMC
)
i
in
cell
sample
95
th
percentile
frequency
of
all
genomic
complexes
(
e
.
g
.
,
ASMCs
)
within
cell
sample
,
1
)
)
,
of between 0.25-0.75 (e.g., 0.3-0.4, 0.4-0.5, 0.5-0.6, or 0.6-0.7), or of between 0.5-1 (e.g., about 0.5-0.6, 0.6-0.7, 0.7-0.8, 0.8-0.9, or 0.9-1.0).
2 . A method of disrupting a genomic complex, e.g., anchor sequence mediated conjunction (ASMC), in a mammalian subject, comprising:
administering to a subject a disrupting agent targeted to the genomic complex (e.g., ASMC),
wherein the genomic complex (e.g., ASMC) is present in a target cell type, and
wherein the genomic complex (e.g., ASMC) is present in less than 9, 8, 7, 6, 5, 4, 3, 2, or 1 reference cell types of Table 2.
3 . The method of claim 2 , wherein the target cell type of is chosen from: neuronal cells, myocytes (e.g., cardiomyocytes), immune cells, endothelial cells, hepatocytes, CD34+ cells, CD3+ cells, and fibroblasts.
4 . A disrupting agent that specifically binds a genomic complex (e.g., anchor sequence-mediated conjunction (ASMC)),
wherein the genomic complex (e.g., ASMC) has, or is identified as having, an IntInch, measured by Formula 2
(
IntInd
i
=
min
(
Frequency
of
genomic
complex
(
e
.
g
.
,
ASMC
)
i
in
cell
sample
95
th
percentile
frequency
of
all
genomic
complexes
(
e
.
g
.
,
ASMCs
)
within
cell
sample
,
1
)
)
,
of between 0.25-0.75 (e.g., 0.3-0.4, 0.4-0.5, 0.5-0.6, or 0.6-0.7), or of between 0.5-1 (e.g., about 0.5-0.6, 0.6-0.7, 0.7-0.8, 0.8-0.9, or 0.9-1.0).
5 . A disrupting agent that specifically binds a genomic complex (e.g., anchor sequence-mediated conjunction (ASMC)),
wherein the genomic complex (e.g., ASMC) is present in a target cell type, and
wherein the genomic complex (e.g., ASMC) is present in less than 9, 8, 7, 6, 5, 4, 3, 2, or 1 reference cell types of Table 2.
6 . The disrupting agent of either of claim 4 or 5 , wherein the disrupting agent comprises a nucleic acid complementary to DNA sequence of the genomic complex (e.g., ASMC).
7 . The method or composition of any of claim 1 or 4 , wherein the IntInd i is measured using ChIA-PET, e.g., against CTCF, e.g., as described in Example 2.
8 . The method of any of claim 2 or 5 wherein genomic complex (e.g., ASMC) presence is measured by ChIA-PET, e.g., against cohesin, e.g., using an assay of Example 1.
9 . The method of any of claim 1 or 4 , wherein the cell sample is a cell line sample or a primary cell sample (e.g., a biopsy sample).
10 . The method of any of the preceding claims, wherein the disrupting agent comprises a DNA-binding moiety that binds specifically to one or more target anchor sequences within a cell and not to non-targeted anchor sequences within the cell with sufficient affinity that it competes with binding of an endogenous nucleating polypeptide within the cell.
11 . The method of any of the preceding claims, wherein the disrupting agent comprises (i) a site-specific targeting moiety and (ii) a deaminating agent.
12 . The method of any of the preceding claims, wherein the disrupting agent comprises (i) a fusion polypeptide comprising an enzymatically inactive Cas polypeptide and a deaminating agent, or a nucleic acid encoding the fusion polypeptide; and (ii) a guide RNA, wherein the guide RNA targets the fusion polypeptide to an anchor sequence comprised by the ASMC.
13 . The method of any of the preceding claims, wherein the disrupting agent comprises (i) a site-specific targeting moiety and (ii) an epigenetic modifying agent, e.g., wherein the epigenetic modifying agent is selected from a DNA methylase, DNA demethylase, histone methyltransferase, a histone deacetylase, or any combination thereof.
14 . The method of any of the preceding claims, wherein the disrupting agent comprises (i) a fusion polypeptide comprising an enzymatically inactive Cas polypeptide and an epigenetic modifying agent, or a nucleic acid encoding the fusion polypeptide; and (ii) a guide RNA, wherein the guide RNA targets the fusion polypeptide to an anchor sequence comprised by the genomic complex (e.g., ASMC).
15 . The method of any of the preceding embodiments, wherein the disrupting agent comprises a fusion polypeptide comprising a TAL effector molecule and an epigenetic modifying agent, or a nucleic acid encoding the fusion polypeptide, wherein the TAL effector molecule targets the fusion polypeptide to an anchor sequence comprised by the genomic complex (e.g., ASMC).
16 . The method of any of the preceding embodiments, wherein the disrupting agent comprises a fusion polypeptide comprising a Zn finger molecule and an epigenetic modifying agent, or a nucleic acid encoding the fusion polypeptide, wherein the Zn finger molecule targets the fusion polypeptide to an anchor sequence comprised by the genomic complex (e.g., ASMC).
17 . The method of any of the preceding claims, wherein the IntInd i as measured by Formula 2 in a cell of the subject, is reduced to less than 0.3-0.4, 0.4-0.5, 0.5-0.6, 0.7-0.8, or 0.8-0.9.
18 . The method of any of the preceding claims, wherein the IntInd i as measured by Formula 2 in a cell of the subject, is reduced by at least 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7. 0.8, or 0.9.
19 . The method of any of the preceding claims, which further comprises, after administration of the disrupting agent, obtaining a value for (e.g., measuring) the IntInd i as measured by Formula 2 of the genomic complex (e.g., ASMC).
20 . The method of claim 19 , which further comprises, responsive to the value for the IntInd i as measured by Formula 2, administering one or more additional doses of the disrupting agent to the mammalian subject, or administering one or more different therapies.
21 . The method of claim 20 , which comprises administering the one or more additional doses of the disrupting agent to the mammalian subject until the IntInd i as measured by Formula 2 in a cell of the subject, is less than 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1.
22 . The method of the preceding claims, which further comprises, after administration of the disrupting agent, determining obtaining a value for (e.g., measuring) expression of a gene associated with (e.g., situated at least partially within) the genomic complex (e.g., ASMC).
23 . The method of claim 22 , which further comprises, responsive to the value for the expression of the gene, administering one or more additional doses of the disrupting agent to the mammalian subject, or administering one or more different therapies.
24 . The method of any of the preceding claims, wherein the genomic complex (e.g., ASMC) comprises a gene, an anchor sequence, or two anchor sequences listed in Table 4 or 5.
25 . The method of any of the preceding claims, wherein the genomic complex (e.g., ASMC) is bound by a polypeptide selected from CTCF, cohesin, YY1, USF1, TAF3, or ZNF143.
26 . The method of any of the preceding claims wherein the genomic complex (e.g., ASMC) is a type 1 or type 2 ASMC.
27 . The method of any of the preceding claims wherein disruption of the genomic complex (e.g., ASMC) results in upregulation of expression of a gene situated at least partly within the genomic complex (e.g., ASMC).
28 . The method of any of claims 1 - 26 , wherein disruption of the genomic complex (e.g., ASMC) results in downregulation of expression of a gene situated at least partly within the genomic complex (e.g., ASMC).
29 . The method of claim 28 , wherein the IntInd i as measured by Formula 2 of the ASMC in the cell is at least 0.5, 0.55, 0.6, 0.65, 0.7, 0.75, 0.8, 0.85, or 0.9.